Anthony Oro, MD, PhD

All Publications

• Basal-to-inflammatory transition and tumor resistance via crosstalk with a pro-inflammatory stromal niche. Nature communications Li, N. Y., Zhang, W., Haensel, D., Jussila, A. R., Pan, C., Gaddam, S., Plevritis, S. K., Oro, A. E. 2024; 15 (1): 8134

  Abstract

  Cancer-associated inflammation is a double-edged sword possessing both pro- and anti-tumor properties through ill-defined tumor-immune dynamics. While we previously identified a carcinoma tumor-intrinsic resistance pathway, basal-to-squamous cell carcinoma transition, here, employing a multipronged single-cell and spatial-omics approach, we identify an inflammation and therapy-enriched tumor state we term basal-to-inflammatory transition. Basal-to-inflammatory transition signature correlates with poor overall patient survival in many epithelial tumors. Basal-to-squamous cell carcinoma transition and basal-to-inflammatory transition occur in adjacent but distinct regions of a single tumor: basal-to-squamous cell carcinoma transition arises within the core tumor nodule, while basal-to-inflammatory transition emerges from a specialized inflammatory environment defined by a tumor-associated TREM1 myeloid signature. TREM1 myeloid-derived cytokines IL1 and OSM induce basal-to-inflammatory transition in vitro and in vivo through NF-κB, lowering sensitivity of patient basal cell carcinoma explant tumors to Smoothened inhibitor treatment. This work deepens our knowledge of the heterogeneous local tumor microenvironment and nominates basal-to-inflammatory transition as a drug-resistant but targetable tumor state driven by a specialized inflammatory microenvironment.

  View details for DOI 10.1038/s41467-024-52394-3

  View details for PubMedID 39289380

  View details for PubMedCentralID 7613740

• A scalable and cGMP-compatible autologous organotypic cell therapy for Dystrophic Epidermolysis Bullosa. Nature communications Neumayer, G., Torkelson, J. L., Li, S., McCarthy, K., Zhen, H. H., Vangipuram, M., Mader, M. M., Gebeyehu, G., Jaouni, T. M., Jacków-Malinowska, J., Rami, A., Hansen, C., Guo, Z., Gaddam, S., Tate, K. M., Pappalardo, A., Li, L., Chow, G. M., Roy, K. R., Nguyen, T. M., Tanabe, K., McGrath, P. S., Cramer, A., Bruckner, A., Bilousova, G., Roop, D., Tang, J. Y., Christiano, A., Steinmetz, L. M., Wernig, M., Oro, A. E. 2024; 15 (1): 5834

  Abstract

  We present Dystrophic Epidermolysis Bullosa Cell Therapy (DEBCT), a scalable platform producing autologous organotypic iPS cell-derived induced skin composite (iSC) grafts for definitive treatment. Clinical-grade manufacturing integrates CRISPR-mediated genetic correction with reprogramming into one step, accelerating derivation of COL7A1-edited iPS cells from patients. Differentiation into epidermal, dermal and melanocyte progenitors is followed by CD49f-enrichment, minimizing maturation heterogeneity. Mouse xenografting of iSCs from four patients with different mutations demonstrates disease modifying activity at 1 month. Next-generation sequencing, biodistribution and tumorigenicity assays establish a favorable safety profile at 1-9 months. Single cell transcriptomics reveals that iSCs are composed of the major skin cell lineages and include prominent holoclone stem cell-like signatures of keratinocytes, and the recently described Gibbin-dependent signature of fibroblasts. The latter correlates with enhanced graftability of iSCs. In conclusion, DEBCT overcomes manufacturing and safety roadblocks and establishes a reproducible, safe, and cGMP-compatible therapeutic approach to heal lesions of DEB patients.

  View details for DOI 10.1038/s41467-024-49400-z

  View details for PubMedID 38992003

  View details for PubMedCentralID PMC11239819

• The manatee variational autoencoder model for predicting gene expression alterations caused by transcription factor perturbations. Scientific reports Yang, Y., Seninge, L., Wang, Z., Oro, A., Stuart, J. M., Ding, H. 2024; 14 (1): 11794

  Abstract

  We present the Manatee variational autoencoder model to predict transcription factor (TF) perturbation-induced transcriptomes. We demonstrate that the Manatee in silico perturbation analysis recapitulates target transcriptomic phenotypes in diverse cellular lineage transitions. We further propose the Manatee in silico screening analysis for prioritizing TF combinations targeting desired transcriptomic phenotypes.

  View details for DOI 10.1038/s41598-024-62620-z

  View details for PubMedID 38782963

  View details for PubMedCentralID PMC11116378

• Skin basal cell carcinomas assemble a pro-tumorigenic spatially organized and self-propagating Trem2+ myeloid niche. Nature communications Haensel, D., Daniel, B., Gaddam, S., Pan, C., Fabo, T., Bjelajac, J., Jussila, A. R., Gonzalez, F., Li, N. Y., Chen, Y., Hou, J., Patel, T., Aasi, S., Satpathy, A. T., Oro, A. E. 2023; 14 (1): 2685

  Abstract

  Cancer immunotherapies have revolutionized treatment but have shown limited success as single-agent therapies highlighting the need to understand the origin, assembly, and dynamics of heterogeneous tumor immune niches. Here, we use single-cell and imaging-based spatial analysis to elucidate three microenvironmental neighborhoods surrounding the heterogeneous basal cell carcinoma tumor epithelia. Within the highly proliferative neighborhood, we find that TREM2+ skin cancer-associated macrophages (SCAMs) support the proliferation of a distinct tumor epithelial population through an immunosuppression-independent manner via oncostatin-M/JAK-STAT3 signaling. SCAMs represent a unique tumor-specific TREM2+ population defined by VCAM1 surface expression that is not found in normal homeostatic skin or during wound healing. Furthermore, SCAMs actively proliferate and self-propagate through multiple serial tumor passages, indicating long-term potential. The tumor rapidly drives SCAM differentiation, with intratumoral injections sufficient to instruct naive bone marrow-derived monocytes to polarize within days. This work provides mechanistic insights into direct tumor-immune niche dynamics independent of immunosuppression, providing the basis for potential combination tumor therapies.

  View details for DOI 10.1038/s41467-023-37993-w

  View details for PubMedID 37164949

• GRHL2 and AP2a coordinate early surface ectoderm lineage commitment during development. iScience Collier, A. E., Piekos, S. N., Liu, A., Pattison, J. M., Felix, F., Bailetti, A. A., Sedov, E., Gaddam, S., Zhen, H., Oro, A. E. 2023; 26 (3): 106125

  Abstract

  Ectodermal dysplasias including skin abnormalities and cleft lip/palate result from improper surface ectoderm (SE) patterning. However, the connection between SE gene regulatory networks and disease remains poorly understood. Here, we dissect human SE differentiation with multiomics and establish GRHL2 as a key mediator of early SE commitment, which acts by skewing cell fate away from the neural lineage. GRHL2 and master SE regulator AP2a balance early cell fate output, with GRHL2 facilitating AP2a binding to SE loci. In turn, AP2a restricts GRHL2 DNA binding away from de novo chromatin contacts. Integration of these regulatory sites with ectodermal dysplasia-associated genomic variants annotated within the Biomedical Data Commons identifies 55 loci previously implicated in craniofacial disorders. These include ABCA4/ARHGAP29 and NOG regulatory regions where disease-linked variants directly affect GRHL2/AP2a binding and gene transcription. These studies elucidate the logic underlying SE commitment and deepen our understanding of human oligogenic disease pathogenesis.

  View details for DOI 10.1016/j.isci.2023.106125

  View details for PubMedID 36843855

• LY6D marks pre-existing resistant basosquamous tumor subpopulations. Nature communications Haensel, D., Gaddam, S., Li, N. Y., Gonzalez, F., Patel, T., Cloutier, J. M., Sarin, K. Y., Tang, J. Y., Rieger, K. E., Aasi, S. Z., Oro, A. E. 2022; 13 (1): 7520

  Abstract

  Improved response to canonical therapies requires a mechanistic understanding of dynamic tumor heterogeneity by identifying discrete cellular populations with enhanced cellular plasticity. We have previously demonstrated distinct resistance mechanisms in skin basal cell carcinomas, but a comprehensive understanding of the cellular states and markers associated with these populations remains poorly understood. Here we identify a pre-existing resistant cellular population in naive basal cell carcinoma tumors marked by the surface marker LY6D. LY6D+ tumor cells are spatially localized and possess basal cell carcinoma and squamous cell carcinoma-like features. Using computational tools, organoids, and spatial tools, we show that LY6D+ basosquamous cells represent a persister population lying on a central node along the skin lineage-associated spectrum of epithelial states with local environmental and applied therapies determining the kinetics of accumulation. Surprisingly, LY6D+ basosquamous populations exist in many epithelial tumors, such as pancreatic adenocarcinomas, which have poor outcomes. Overall, our results identify the resistant LY6D+ basosquamous population as an important clinical target and suggest strategies for future therapeutic approaches to target them.

  View details for DOI 10.1038/s41467-022-35020-y

  View details for PubMedID 36473848

  View details for PubMedCentralID PMC9726704

• Gibbin mesodermal regulation patterns epithelial development. Nature Collier, A., Liu, A., Torkelson, J., Pattison, J., Gaddam, S., Zhen, H., Patel, T., McCarthy, K., Ghanim, H., Oro, A. E. 2022

  Abstract

  Proper ectodermal patterning during human development requires previously identified transcription factors such as GATA3 and p63, as well as positional signalling from regional mesoderm1-6. However, the mechanism by which ectoderm and mesoderm factors act to stably pattern gene expression and lineage commitment remains unclear. Here we identify the protein Gibbin,encoded by the Xia-Gibbs AT-hook DNA-binding-motif-containing 1 (AHDC1) disease gene7-9, as a key regulator of early epithelial morphogenesis. We find that enhancer- or promoter-bound Gibbin interacts with dozens of sequence-specific zinc-finger transcription factors and methyl-CpG-binding proteins to regulate the expression of mesoderm genes. The loss of Gibbin causes an increase in DNA methylation at GATA3-dependent mesodermal genes, resulting in a loss of signalling between developing dermal and epidermal cell types. Notably, Gibbin-mutant human embryonic stem-cell-derived skin organoids lack dermal maturation, resulting in p63-expressing basal cells that possess defective keratinocyte stratification. In vivo chimeric CRISPR mouse mutants reveal a spectrum of Gibbin-dependent developmental patterning defects affecting craniofacial structure, abdominal wall closure and epidermal stratification that mirror patient phenotypes. Our results indicate that the patterning phenotypes seen in Xia-Gibbs and related syndromes derive from abnormal mesoderm maturation as a result of gene-specific DNA methylation decisions.

  View details for DOI 10.1038/s41586-022-04727-9

  View details for PubMedID 35585237

• Biomedical Data Commons (BMDC) prioritizes B-lymphocyte non-coding genetic variants in Type 1 Diabetes. PLoS computational biology Piekos, S. N., Gaddam, S., Bhardwaj, P., Radhakrishnan, P., Guha, R. V., Oro, A. E. 2021; 17 (9): e1009382

  Abstract

  The repurposing of biomedical data is inhibited by its fragmented and multi-formatted nature that requires redundant investment of time and resources by data scientists. This is particularly true for Type 1 Diabetes (T1D), one of the most intensely studied common childhood diseases. Intense investigation of the contribution of pancreatic β-islet and T-lymphocytes in T1D has been made. However, genetic contributions from B-lymphocytes, which are known to play a role in a subset of T1D patients, remain relatively understudied. We have addressed this issue through the creation of Biomedical Data Commons (BMDC), a knowledge graph that integrates data from multiple sources into a single queryable format. This increases the speed of analysis by multiple orders of magnitude. We develop a pipeline using B-lymphocyte multi-dimensional epigenome and connectome data and deploy BMDC to assess genetic variants in the context of Type 1 Diabetes (T1D). Pipeline-identified variants are primarily common, non-coding, poorly conserved, and are of unknown clinical significance. While variants and their chromatin connectivity are cell-type specific, they are associated with well-studied disease genes in T-lymphocytes. Candidates include established variants in the HLA-DQB1 and HLA-DRB1 and IL2RA loci that have previously been demonstrated to protect against T1D in humans and mice providing validation for this method. Others are included in the well-established T1D GRS2 genetic risk scoring method. More intriguingly, other prioritized variants are completely novel and form the basis for future mechanistic and clinical validation studies The BMDC community-based platform can be expanded and repurposed to increase the accessibility, reproducibility, and productivity of biomedical information for diverse applications including the prioritization of cell type-specific disease alleles from complex phenotypes.

  View details for DOI 10.1371/journal.pcbi.1009382

  View details for PubMedID 34543288

• c-FOS drives reversible basal to squamous cell carcinoma transition. Cell reports Kuonen, F., Li, N. Y., Haensel, D., Patel, T., Gaddam, S., Yerly, L., Rieger, K., Aasi, S., Oro, A. E. 2021; 37 (1): 109774

  Abstract

  While squamous transdifferentiation within subpopulations of adenocarcinomas represents an important drug resistance problem, its underlying mechanism remains poorly understood. Here, using surface markers of resistant basal cell carcinomas (BCCs) and patient single-cell and bulk transcriptomic data, we uncover the dynamic roadmap of basal to squamous cell carcinoma transition (BST). Experimentally induced BST identifies activator protein 1 (AP-1) family members in regulating tumor plasticity, and we show that c-FOS plays a central role in BST by regulating the accessibility of distinct AP-1 regulatory elements. Remarkably, despite prominent changes in cell morphology and BST marker expression, we show using inducible model systems that c-FOS-mediated BST demonstrates reversibility. Blocking EGFR pathway activation after c-FOS induction partially reverts BST in vitro and prevents BST features in both mouse models and human tumors. Thus, by identifying the molecular basis of BST, our work reveals a therapeutic opportunity targeting plasticity as a mechanism of tumor resistance.

  View details for DOI 10.1016/j.celrep.2021.109774

  View details for PubMedID 34610301

• The Sky's the LEMit: New insights into nuclear structure regulation of transcription factor activity. Current opinion in cell biology Mirza, A. N., Gonzalez, F., Ha, S. K., Oro, A. E. 2020; 68: 173–80

  Abstract

  The nucleoskeleton has been associated with partitioning the genome into active and inactive compartments that dictate local transcription factor (TF) activity. However, recent data indicatethat the nucleoskeleton and TFs reciprocally influence each other in dynamic TF trafficking pathways through the functions of LEM proteins. While the conserved peripheral recruitment of TFs by LEM proteins has been viewed as a mechanism of repressing transcription, a diversity of release mechanisms from the lamina suggest this compartment serves as a refuge for nuclear TF accumulation for rapid mobilization and signal stability. Detailed mechanisms suggest that TFs toggle between nuclear lamina refuge and nuclear matrix lamin-LEM protein complexes at sites of active transcription. In this review we will highlight emerging LEM functions acting at the interface of chromatin and nucleoskeleton to create TF trafficking networks.

  View details for DOI 10.1016/j.ceb.2020.10.006

  View details for PubMedID 33227657

• AP-1 and TGFSS cooperativity drives non-canonical Hedgehog signaling in resistant basal cell carcinoma. Nature communications Yao, C. D., Haensel, D., Gaddam, S., Patel, T., Atwood, S. X., Sarin, K. Y., Whitson, R. J., McKellar, S., Shankar, G., Aasi, S., Rieger, K., Oro, A. E. 2020; 11 (1): 5079

  Abstract

  Tumor heterogeneity and lack of knowledge about resistant cell states remain a barrier to targeted cancer therapies. Basal cell carcinomas (BCCs) depend on Hedgehog (Hh)/Gli signaling, but can develop mechanisms of Smoothened (SMO) inhibitor resistance. We previously identified a nuclear myocardin-related transcription factor (nMRTF) resistance pathway that amplifies noncanonical Gli1 activity, but characteristics and drivers of the nMRTF cell state remain unknown. Here, we use single cell RNA-sequencing of patient tumors to identify three prognostic surface markers (LYPD3, TACSTD2, and LY6D) which correlate with nMRTF and resistance to SMO inhibitors. The nMRTF cell state resembles transit-amplifying cells of the hair follicle matrix, with AP-1 and TGFSS cooperativity driving nMRTF activation. JNK/AP-1 signaling commissions chromatin accessibility and Smad3 DNA binding leading to a transcriptional program of RhoGEFs that facilitate nMRTF activity. Importantly, small molecule AP-1 inhibitors selectively target LYPD3+/TACSTD2+/LY6D+ nMRTF human BCCs ex vivo, opening an avenue for improving combinatorial therapies.

  View details for DOI 10.1038/s41467-020-18762-5

  View details for PubMedID 33033234

• TFAP2C-and p63-Dependent Networks Sequentially Rearrange Chromatin Landscapes to Drive Human Epidermal Lineage Commitment CELL STEM CELL Li, L., Wang, Y., Torkelson, J. L., Shankar, G., Pattison, J. M., Zhen, H. H., Fang, F., Duren, Z., Xin, J., Gaddam, S., Melo, S. P., Piekos, S. N., Li, J., Liaw, E. J., Chen, L., Li, R., Wernig, M., Wong, W. H., Chang, H. Y., Oro, A. E. 2019; 24 (2): 271-+

  View details for DOI 10.1016/j.stem.2018.12.012

  View details for Web of Science ID 000458027300013

• Loss of primary cilia drives switching from Hedgehog to Ras/MAPK pathway in resistant basal cell carcinoma. The Journal of investigative dermatology Kuonen, F., Huskey, N. E., Shankar, G., Jaju, P., Whitson, R. J., Rieger, K. E., Atwood, S. X., Sarin, K. Y., Oro, A. E. 2019

  Abstract

  Basal cell carcinomas (BCCs) rely on Hedgehog (HH) pathway growth signal amplification by the microtubule-based organelle, the primary cilium. Despite naive tumors responsiveness to Smoothened inhibitors (Smoi), resistance in advanced tumors remains frequent. While the resistant BCCs usually maintain HH pathway activation, squamous cell carcinomas with Ras/MAPK pathway activation also arise, with the molecular basis of tumor type and pathway selection still obscure. Here we identify the primary cilium as a critical determinant controlling tumor pathway switching. Strikingly, Smoi-resistant BCCs possess an increased mutational load in ciliome genes, resulting in reduced primary cilia and HH pathway activation compared to naive or Gorlin patient BCCs. Gene set enrichment analysis of resistant BCCs with a low HH pathway signature reveals increased Ras/MAPK pathway activation. Tissue analysis confirms an inverse relationship between primary cilia presence and Ras/MAPK activation, and primary cilia removal in BCCs potentiates Ras/MAPK pathway activation. Moreover, activating Ras in HH-responsive cell lines confers resistance to both canonical (vismodegib) and non-canonical (aPKC and MRTF inhibitors) HH pathway inhibitors, while conferring sensitivity to MAPK inhibitors. Our results provide insights into BCC treatment and identify the primary cilium as an important lineage gatekeeper, preventing HH to Ras/MAPK pathway switching.

  View details for PubMedID 30707899

• TFAP2C- and p63-Dependent Networks Sequentially Rearrange Chromatin Landscapes to Drive Human Epidermal Lineage Commitment. Cell stem cell Li, L., Wang, Y., Torkelson, J. L., Shankar, G., Pattison, J. M., Zhen, H. H., Fang, F., Duren, Z., Xin, J., Gaddam, S., Melo, S. P., Piekos, S. N., Li, J., Liaw, E. J., Chen, L., Li, R., Wernig, M., Wong, W. H., Chang, H. Y., Oro, A. E. 2019

  Abstract

  Tissue development results from lineage-specific transcription factors (TFs) programming a dynamic chromatin landscape through progressive cell fate transitions. Here, we define epigenomic landscape during epidermal differentiation of human pluripotent stem cells (PSCs) and create inference networks that integrate gene expression, chromatin accessibility, and TF binding to define regulatory mechanisms during keratinocyte specification. We found two critical chromatin networks during surface ectoderm initiation and keratinocyte maturation, which are driven by TFAP2C and p63, respectively. Consistently, TFAP2C, but not p63, is sufficient to initiate surface ectoderm differentiation, and TFAP2C-initiated progenitor cells are capable of maturing into functional keratinocytes. Mechanistically, TFAP2C primes the surface ectoderm chromatin landscape and induces p63 expression and binding sites, thus allowing maturation factor p63 to positively autoregulate its own expression and close a subset of the TFAP2C-initiated surface ectoderm program. Our work provides a general framework to infer TF networks controlling chromatin transitions that will facilitate future regenerative medicine advances.

  View details for PubMedID 30686763

• LAP2 Proteins Chaperone GLI1 Movement between the Lamina and Chromatin to Regulate Transcription CELL Mirza, A. N., McKellar, S. A., Urman, N. M., Brown, A. S., Hollmig, T., Aasi, S. Z., Oro, A. E. 2019; 176 (1-2): 198-+

  View details for DOI 10.1016/j.cell.2018.10.054

  View details for Web of Science ID 000455410800018

• MTSS1/Src family kinase dysregulation underlies multiple inherited ataxias. Proceedings of the National Academy of Sciences of the United States of America Brown, A. S., Meera, P., Altindag, B., Chopra, R., Perkins, E. M., Paul, S., Scoles, D. R., Tarapore, E., Magri, J., Huang, H., Jackson, M., Shakkottai, V. G., Otis, T. S., Pulst, S. M., Atwood, S. X., Oro, A. E. 2018

  Abstract

  The genetically heterogeneous spinocerebellar ataxias (SCAs) are caused by Purkinje neuron dysfunction and degeneration, but their underlying pathological mechanisms remain elusive. The Src family of nonreceptor tyrosine kinases (SFK) are essential for nervous system homeostasis and are increasingly implicated in degenerative disease. Here we reveal that the SFK suppressor Missing-in-metastasis (MTSS1) is an ataxia locus that links multiple SCAs. MTSS1 loss results in increased SFK activity, reduced Purkinje neuron arborization, and low basal firing rates, followed by cell death. Surprisingly, mouse models for SCA1, SCA2, and SCA5 show elevated SFK activity, with SCA1 and SCA2 displaying dramatically reduced MTSS1 protein levels through reduced gene expression and protein translation, respectively. Treatment of each SCA model with a clinically approved Src inhibitor corrects Purkinje neuron basal firing and delays ataxia progression in MTSS1 mutants. Our results identify a common SCA therapeutic target and demonstrate a key role for MTSS1/SFK in Purkinje neuron survival and ataxia progression.

  View details for PubMedID 30530649

• Retinoic acid and BMP4 cooperate with p63 to alter chromatin dynamics during surface epithelial commitment NATURE GENETICS Pattison, J. M., Melo, S. P., Piekos, S. N., Torkelson, J. L., Bashkirova, E., Mumbach, M. R., Rajasingh, C., Zhen, H., Li, L., Liaw, E., Alber, D., Rubin, A. J., Shankar, G., Bao, X., Chang, H. Y., Khavari, P. A., Oro, A. E. 2018; 50 (12): 1658-+

  View details for DOI 10.1038/s41588-018-0263-0

  View details for Web of Science ID 000451434400012

• LAP2 Proteins Chaperone GLI1 Movement between the Lamina and Chromatin to Regulate Transcription. Cell Mirza, A. N., McKellar, S. A., Urman, N. M., Brown, A. S., Hollmig, T., Aasi, S. Z., Oro, A. E. 2018

  Abstract

  Understanding transcription factor navigation through the nucleus remains critical for developing targeted therapeutics. The GLI1 transcription factor must maintain maximal Hedgehog pathway output in basal cell carcinomas (BCCs), and we have previously shown that resistant BCCs increase GLI1 deacetylation through atypical protein kinase Ciota/lambda (aPKC) andHDAC1. Here we identify a lamina-associated polypeptide 2 (LAP2) isoform-dependent nuclear chaperoning system that regulates GLI1 movement between the nuclear lamina and nucleoplasm to achieve maximal activation. LAP2beta forms a two-site interaction with the GLI1 zinc-finger domain and acetylation site, stabilizing an acetylation-dependent reserve on the inner nuclear membrane (INM). By contrast, the nucleoplasmic LAP2alpha competes with LAP2beta for GLI1 while scaffolding HDAC1 to deacetylate the secondary binding site. aPKC functions to promote GLI1 association with LAP2alpha, promoting egress off the INM. GLI1 intranuclear trafficking by LAP2 isoforms represents a powerful signal amplifier in BCCs with implications for zinc finger-based signal transduction and therapeutics.

  View details for PubMedID 30503211

• Retinoic acid and BMP4 cooperate with p63 to alter chromatin dynamics during surface epithelial commitment. Nature genetics Pattison, J. M., Melo, S. P., Piekos, S. N., Torkelson, J. L., Bashkirova, E., Mumbach, M. R., Rajasingh, C., Zhen, H. H., Li, L., Liaw, E., Alber, D., Rubin, A. J., Shankar, G., Bao, X., Chang, H. Y., Khavari, P. A., Oro, A. E. 2018

  Abstract

  Human embryonic stem cell (hESC) differentiation promises advances in regenerative medicine1-3, yet conversion of hESCs into transplantable cells or tissues remains poorly understood. Using our keratinocyte differentiation system, we employ a multi-dimensional genomics approach to interrogate the contributions of inductive morphogens retinoic acid and bone morphogenetic protein 4 (BMP4) and the epidermal master regulator p63 (encoded by TP63)4,5 during surface ectoderm commitment. In contrast to other master regulators6-9, p63 effects major transcriptional changes only after morphogens alter chromatin accessibility, establishing an epigenetic landscape for p63 to modify. p63 distally closes chromatin accessibility and promotes accumulation of H3K27me3 (trimethylated histone H3 lysine 27). Cohesin HiChIP10 visualizations of chromosome conformation show that p63 and the morphogens contribute to dynamic long-range chromatin interactions, as illustrated by TFAP2C regulation11. Our study demonstrates the unexpected dependency of p63 on morphogenetic signaling and provides novel insights into how a master regulator can specify diverse transcriptional programs based on the chromatin landscape induced by exposure to specific morphogens.

  View details for PubMedID 30397335

• Frequent basal cell cancer development is a clinical marker for inherited cancer susceptibility JCI INSIGHT Cho, H. G., Kuo, K. Y., Li, S., Bailey, I., Aasi, S., Chang, A. S., Oro, A. E., Tang, J. Y., Sarin, K. Y. 2018; 3 (15)

  Abstract

  Innate DNA repair mechanisms play a critical role in protecting skin keratinocytes from UV mutagenesis and skin cancer development. We hypothesized that individuals who develop frequent skin cancers may harbor germline defects in DNA repair genes and have increased predisposition to internal malignancies. We enrolled 61 patients with unusually frequent basal cell carcinoma (BCC) development, seen at Stanford Hospital and Clinics from January 2005 until December 2015, for germline analysis of 29 DNA repair genes. In parallel, a case-control retrospective review was performed to interrogate the association of malignancies with frequent BCC development in a large US medical insurance claims database (Truven), which included 13,264 individuals with 6 or more BCCs from 2007 to 2011. 19.7% of the frequent BCC cohort harbored pathogenic mutations in DNA repair genes: APC, BARD1, BRCA1, BRCA2, CDH1, CHEK2, MLH1, MSH2, MSH6, MUTYH, NBN, and PALB2. Individuals with 6 or more BCCs had an increased risk of other malignancies, with a 3.5-fold increase in the frequent BCC cohort and a 3.2-fold increase in the Truven database. Individuals who developed frequent BCCs have an increased prevalence of germline mutations in DNA repair genes and increased malignancy risk. Our data implicate frequent BCC development as an external marker of inherited cancer risk.

  View details for DOI 10.1172/jci.insight.122744

  View details for Web of Science ID 000441201300022

  View details for PubMedID 30089731

• Noncanonical hedgehog pathway activation through SRF-MKL1 promotes drug resistance in basal cell carcinomas. Nature medicine Whitson, R. J., Lee, A. n., Urman, N. M., Mirza, A. n., Yao, C. Y., Brown, A. S., Li, J. R., Shankar, G. n., Fry, M. A., Atwood, S. X., Lee, E. Y., Hollmig, S. T., Aasi, S. Z., Sarin, K. Y., Scott, M. P., Epstein, E. H., Tang, J. Y., Oro, A. E. 2018; 24 (3): 271–81

  Abstract

  Hedgehog pathway-dependent cancers can escape Smoothened (SMO) inhibition through mutations in genes encoding canonical hedgehog pathway components; however, around 50% of drug-resistant basal cell carcinomas (BCCs) lack additional variants of these genes. Here we use multidimensional genomics analysis of human and mouse drug-resistant BCCs to identify a noncanonical hedgehog activation pathway driven by the transcription factor serum response factor (SRF). Active SRF along with its coactivator megakaryoblastic leukemia 1 (MKL1) binds DNA near hedgehog target genes and forms a previously unknown protein complex with the hedgehog transcription factor glioma-associated oncogene family zinc finger-1 (GLI1), causing amplification of GLI1 transcriptional activity. We show that cytoskeletal activation through Rho and the formin family member Diaphanous (mDia) is required for SRF-MKL-driven GLI1 activation and for tumor cell viability. Remarkably, nuclear MKL1 staining served as a biomarker in tumors from mice and human subjects to predict tumor responsiveness to MKL inhibitors, highlighting the therapeutic potential of targeting this pathway. Thus, our study illuminates, for the first time, cytoskeletal-activation-driven transcription as a personalized therapeutic target for combatting drug-resistant malignancies.

  View details for PubMedID 29400712

  View details for PubMedCentralID PMC5839965

• Soil Primes the Seed: Epigenetic Landscape Drives Tumor Behavior. Cell stem cell Whitson, R. J., Oro, A. E. 2017; 20 (2): 149-150

  Abstract

  Whether invasive tumor phenotypes like EMT arise from oncogenic drivers or from priming of the pre-tumor cell of origin remains unknown. In this issue of Cell Stem Cell, Latil et al. (2017) show that the pre-tumor niche establishes a chromatin state predisposing squamous cell carcinomas to undergo EMT and metastasis, suggesting that the pre-tumor epigenome has prognostic value.

  View details for DOI 10.1016/j.stem.2017.01.007

  View details for PubMedID 28157493

• Combined inhibition of atypical PKC and histone deacetylase 1 is cooperative in basal cell carcinoma treatment. JCI insight Mirza, A. N., Fry, M. A., Urman, N. M., Atwood, S. X., Roffey, J. n., Ott, G. R., Chen, B. n., Lee, A. n., Brown, A. S., Aasi, S. Z., Hollmig, T. n., Ator, M. A., Dorsey, B. D., Ruggeri, B. R., Zificsak, C. A., Sirota, M. n., Tang, J. Y., Butte, A. n., Epstein, E. n., Sarin, K. Y., Oro, A. E. 2017; 2 (21)

  Abstract

  Advanced basal cell carcinomas (BCCs) circumvent Smoothened (SMO) inhibition by activating GLI transcription factors to sustain the high levels of Hedgehog (HH) signaling required for their survival. Unfortunately, there is a lack of efficacious therapies. We performed a gene expression-based drug repositioning screen in silico and identified the FDA-approved histone deacetylase (HDAC) inhibitor, vorinostat, as a top therapeutic candidate. We show that vorinostat only inhibits proliferation of BCC cells in vitro and BCC allografts in vivo at high dose, limiting its usefulness as a monotherapy. We leveraged this in silico approach to identify drug combinations that increase the therapeutic window of vorinostat and identified atypical PKC Ɩ/ʎ (aPKC) as a HDAC costimulator of HH signaling. We found that aPKC promotes GLI1-HDAC1 association in vitro, linking two positive feedback loops. Combination targeting of HDAC1 and aPKC robustly inhibited GLI1, lowering drug doses needed in vitro, in vivo, and ex vivo in patient-derived BCC explants. We identified a bioavailable and selective small-molecule aPKC inhibitor, bringing the pharmacological blockade of aPKC and HDAC1 into the realm of clinical possibility. Our findings provide a compelling rationale and candidate drugs for combined targeting of HDAC1 and aPKC in HH-dependent cancers.

  View details for PubMedID 29093271

• Safety and efficacy of the JAK inhibitor tofacitinib citrate in patients with alopecia areata. JCI insight Kennedy Crispin, M., Ko, J. M., Craiglow, B. G., Li, S., Shankar, G., Urban, J. R., Chen, J. C., Cerise, J. E., Jabbari, A., Winge, M. C., Marinkovich, M. P., Christiano, A. M., Oro, A. E., King, B. A. 2016; 1 (15)

  Abstract

  Alopecia areata (AA) is an autoimmune disease characterized by hair loss mediated by CD8(+) T cells. There are no reliably effective therapies for AA. Based on recent developments in the understanding of the pathomechanism of AA, JAK inhibitors appear to be a therapeutic option; however, their efficacy for the treatment of AA has not been systematically examined.This was a 2-center, open-label, single-arm trial using the pan-JAK inhibitor, tofacitinib citrate, for AA with >50% scalp hair loss, alopecia totalis (AT), and alopecia universalis (AU). Tofacitinib (5 mg) was given twice daily for 3 months. Endpoints included regrowth of scalp hair, as assessed by the severity of alopecia tool (SALT), duration of hair growth after completion of therapy, and disease transcriptome.Of 66 subjects treated, 32% experienced 50% or greater improvement in SALT score. AA and ophiasis subtypes were more responsive than AT and AU subtypes. Shorter duration of disease and histological peribulbar inflammation on pretreatment scalp biopsies were associated with improvement in SALT score. Drug cessation resulted in disease relapse in 8.5 weeks. Adverse events were limited to grade I and II infections. An AA responsiveness to JAK/STAT inhibitors score was developed to segregate responders and nonresponders, and the previously developed AA disease activity index score tracked response to treatment.At the dose and duration studied, tofacitinib is a safe and effective treatment for severe AA, though it does not result in a durable response. Transcriptome changes reveal unexpected molecular complexity within the disease.ClinicalTrials.gov NCT02197455 and NCT02312882.This work was supported by the US Department of Veterans Affairs Office of Research and Development, National Institute of Arthritis and Musculoskeletal and Skin Diseases National Institutes of Health grant R01 AR47223 and U01 AR67173, the National Psoriasis Foundation, the Swedish Society of Medicine, the Fernström Foundation, the Locks of Love Foundation, the National Alopecia Areata Foundation, and the Ranjini and Ajay Poddar Resource Fund for Dermatologic Diseases Research.

  View details for PubMedID 27699252

• Smoothened variants explain the majority of drug resistance in Basal cell carcinoma. Cancer cell Atwood, S. X., Sarin, K. Y., Whitson, R. J., Li, J. R., Kim, G., Rezaee, M., Ally, M. S., Kim, J., Yao, C., Chang, A. L., Oro, A. E., Tang, J. Y. 2015; 27 (3): 342-353

  Abstract

  Advanced basal cell carcinomas (BCCs) frequently acquire resistance to Smoothened (SMO) inhibitors through unknown mechanisms. Here we identify SMO mutations in 50% (22 of 44) of resistant BCCs and show that these mutations maintain Hedgehog signaling in the presence of SMO inhibitors. Alterations include four ligand binding pocket mutations defining sites of inhibitor binding and four variants conferring constitutive activity and inhibitor resistance, illuminating pivotal residues that ensure receptor autoinhibition. In the presence of a SMO inhibitor, tumor cells containing either class of SMO mutants effectively outcompete cells containing the wild-type SMO. Finally, we show that both classes of SMO variants respond to aPKC-ι/λ or GLI2 inhibitors that operate downstream of SMO, setting the stage for the clinical use of GLI antagonists.

  View details for DOI 10.1016/j.ccell.2015.02.002

  View details for PubMedID 25759020

• Human COL7A1-corrected induced pluripotent stem cells for the treatment of recessive dystrophic epidermolysis bullosa SCIENCE TRANSLATIONAL MEDICINE Sebastiano, V., Zhen, H. H., Derafshi, B. H., Bashkirova, E., Melo, S. P., Wang, P., Leung, T. L., Siprashvili, Z., Tichy, A., Li, J., Ameen, M., Hawkins, J., Lee, S., Li, L., Schwertschkow, A., Bauer, G., Lisowski, L., Kay, M. A., Kim, S. K., Lane, A. T., Wernig, M., Oro, A. E. 2014; 6 (264)

  Abstract

  Patients with recessive dystrophic epidermolysis bullosa (RDEB) lack functional type VII collagen owing to mutations in the gene COL7A1 and suffer severe blistering and chronic wounds that ultimately lead to infection and development of lethal squamous cell carcinoma. The discovery of induced pluripotent stem cells (iPSCs) and the ability to edit the genome bring the possibility to provide definitive genetic therapy through corrected autologous tissues. We generated patient-derived COL7A1-corrected epithelial keratinocyte sheets for autologous grafting. We demonstrate the utility of sequential reprogramming and adenovirus-associated viral genome editing to generate corrected iPSC banks. iPSC-derived keratinocytes were produced with minimal heterogeneity, and these cells secreted wild-type type VII collagen, resulting in stratified epidermis in vitro in organotypic cultures and in vivo in mice. Sequencing of corrected cell lines before tissue formation revealed heterogeneity of cancer-predisposing mutations, allowing us to select COL7A1-corrected banks with minimal mutational burden for downstream epidermis production. Our results provide a clinical platform to use iPSCs in the treatment of debilitating genodermatoses, such as RDEB.

  View details for DOI 10.1126/scitranslmed.3009540

  View details for Web of Science ID 000345595200003

  View details for PubMedCentralID PMC4428910

• Epigenetic targeting of Hedgehog pathway transcriptional output through BET bromodomain inhibition NATURE MEDICINE Tang, Y., Gholamin, S., Schubert, S., Willardson, M. I., Lee, A., Bandopadhayay, P., Bergthold, G., Masoud, S., Nguyen, B., Vue, N., Balansay, B., Yu, F., Oh, S., Woo, P., Chen, S., Ponnuswami, A., Monje, M., Atwood, S. X., Whitson, R. J., Mitra, S., Cheshier, S. H., Qi, J., Beroukhim, R., Tang, J. Y., Wechsler-Reya, R., Oro, A. E., Link, B. A., Bradner, J. E., Cho, Y. 2014; 20 (7): 732-740

  Abstract

  Hedgehog signaling drives oncogenesis in several cancers, and strategies targeting this pathway have been developed, most notably through inhibition of Smoothened (SMO). However, resistance to Smoothened inhibitors occurs by genetic changes of Smoothened or other downstream Hedgehog components. Here we overcome these resistance mechanisms by modulating GLI transcription through inhibition of bromo and extra C-terminal (BET) bromodomain proteins. We show that BRD4 and other BET bromodomain proteins regulate GLI transcription downstream of SMO and suppressor of fused (SUFU), and chromatin immunoprecipitation studies reveal that BRD4 directly occupies GLI1 and GLI2 promoters, with a substantial decrease in engagement of these sites after treatment with JQ1, a small-molecule inhibitor targeting BRD4. Globally, genes associated with medulloblastoma-specific GLI1 binding sites are downregulated in response to JQ1 treatment, supporting direct regulation of GLI activity by BRD4. Notably, patient- and GEMM (genetically engineered mouse model)-derived Hedgehog-driven tumors (basal cell carcinoma, medulloblastoma and atypical teratoid rhabdoid tumor) respond to JQ1 even when harboring genetic lesions rendering them resistant to Smoothened antagonists. Altogether, our results reveal BET proteins as critical regulators of Hedgehog pathway transcriptional output and nominate BET bromodomain inhibitors as a strategy for treating Hedgehog-driven tumors with emerged or a priori resistance to Smoothened antagonists.

  View details for Web of Science ID 000338689500015

• Somatic Correction of Junctional Epidermolysis Bullosa by a Highly Recombinogenic AAV Variant. Molecular therapy : the journal of the American Society of Gene Therapy Melo, S. P., Lisowski, L., Bashkirova, E., Zhen, H. H., Chu, K., Keene, D. R., Marinkovich, M. P., Kay, M. A., Oro, A. E. 2014; 22 (4): 725-733

  Abstract

  Definitive correction of disease causing mutations in somatic cells by homologous recombination (HR) is an attractive therapeutic approach for the treatment of genetic diseases. However, HR-based somatic gene therapy is limited by the low efficiency of gene targeting in mammalian cells and replicative senescence of primary cells ex vivo, forcing investigators to explore alternative strategies such as retro- and lentiviral gene transfer, or genome editing in induced pluripotent stem cells. Here, we report correction of mutations at the LAMA3 locus in primary keratinocytes derived from a patient affected by recessive inherited Herlitz junctional epidermolysis bullosa (H-JEB) disorder using recombinant adenoassociated virus (rAAV)-mediated HR. We identified a highly recombinogenic AAV serotype, AAV-DJ, that mediates efficient gene targeting in keratinocytes at clinically relevant frequencies with a low rate of random integration. Targeted H-JEB patient cells were selected based on restoration of adhesion phenotype, which eliminated the need for foreign sequences in repaired cells, enhancing the clinical use and safety profile of our approach. Corrected pools of primary cells assembled functional laminin-332 heterotrimer and fully reversed the blistering phenotype both in vitro and in skin grafts. The efficient targeting of the LAMA3 locus by AAV-DJ using phenotypic selection, together with the observed low frequency of off-target events, makes AAV-DJ based somatic cell targeting a promising strategy for ex vivo therapy for this severe and often lethal epithelial disorder.

  View details for DOI 10.1038/mt.2013.290

  View details for PubMedID 24390279

• "Atypical" regulation of Hedgehog-dependent cancers. Cancer cell Atwood, S. X., Oro, A. E. 2014; 25 (2): 133-134

  Abstract

  Growing evidence indicates targeting PKCι may be effective in treating Hedgehog-dependent cancers. In this issue of Cancer Cell, Justilien and colleagues present the surprising finding that PKCι promotes Hedgehog ligand production and lung squamous cell carcinoma growth through SOX2, rather than the canonical transcription factor GLI.

  View details for DOI 10.1016/j.ccr.2014.01.027

  View details for PubMedID 24525228

• State-dependent signaling by Cav1.2 regulates hair follicle stem cell function. Genes & development Yucel, G., Altindag, B., Gomez-Ospina, N., Rana, A., Panagiotakos, G., Lara, M. F., Dolmetsch, R., Oro, A. E. 2013; 27 (11): 1217-1222

  Abstract

  The signals regulating stem cell activation during tissue regeneration remain poorly understood. We investigated the baldness associated with mutations in the voltage-gated calcium channel (VGCC) Cav1.2 underlying Timothy syndrome (TS). While hair follicle stem cells express Cav1.2, they lack detectable voltage-dependent calcium currents. Cav1.2(TS) acts in a dominant-negative manner to markedly delay anagen, while L-type channel blockers act through Cav1.2 to induce anagen and overcome the TS phenotype. Cav1.2 regulates production of the bulge-derived BMP inhibitor follistatin-like1 (Fstl1), derepressing stem cell quiescence. Our findings show how channels act in nonexcitable tissues to regulate stem cells and may lead to novel therapeutics for tissue regeneration.

  View details for DOI 10.1101/gad.216556.113

  View details for PubMedID 23752588

  View details for PubMedCentralID PMC3690395

• GLI activation by atypical protein kinase C ?/? regulates the growth of basal cell carcinomas. Nature Atwood, S. X., Li, M., Lee, A., Tang, J. Y., Oro, A. E. 2013; 494 (7438): 484-488

  Abstract

  Growth of basal cell carcinomas (BCCs) requires high levels of hedgehog (HH) signalling through the transcription factor GLI. Although inhibitors of membrane protein smoothened (SMO) effectively suppress HH signalling, early tumour resistance illustrates the need for additional downstream targets for therapy. Here we identify atypical protein kinase C ι/λ (aPKC-ι/λ) as a novel GLI regulator in mammals. aPKC-ι/λ and its polarity signalling partners co-localize at the centrosome and form a complex with missing-in-metastasis (MIM), a scaffolding protein that potentiates HH signalling. Genetic or pharmacological loss of aPKC-ι/λ function blocks HH signalling and proliferation of BCC cells. Prkci is a HH target gene that forms a positive feedback loop with GLI and exists at increased levels in BCCs. Genome-wide transcriptional profiling shows that aPKC-ι/λ and SMO control the expression of similar genes in tumour cells. aPKC-ι/λ functions downstream of SMO to phosphorylate and activate GLI1, resulting in maximal DNA binding and transcriptional activation. Activated aPKC-ι/λ is upregulated in SMO-inhibitor-resistant tumours and targeting aPKC-ι/λ suppresses signalling and growth of resistant BCC cell lines. These results demonstrate that aPKC-ι/λ is critical for HH-dependent processes and implicates aPKC-ι/λ as a new, tumour-selective therapeutic target for the treatment of SMO-inhibitor-resistant cancers.

  View details for DOI 10.1038/nature11889

  View details for PubMedID 23446420

• GLI activation by atypical protein kinase C iota/lambda regulates the growth of basal cell carcinomas NATURE Atwood, S. X., Li, M., Lee, A., Tang, J. Y., Oro, A. E. 2013; 494 (7438): 484-488

  Abstract

  Growth of basal cell carcinomas (BCCs) requires high levels of hedgehog (HH) signalling through the transcription factor GLI. Although inhibitors of membrane protein smoothened (SMO) effectively suppress HH signalling, early tumour resistance illustrates the need for additional downstream targets for therapy. Here we identify atypical protein kinase C ι/λ (aPKC-ι/λ) as a novel GLI regulator in mammals. aPKC-ι/λ and its polarity signalling partners co-localize at the centrosome and form a complex with missing-in-metastasis (MIM), a scaffolding protein that potentiates HH signalling. Genetic or pharmacological loss of aPKC-ι/λ function blocks HH signalling and proliferation of BCC cells. Prkci is a HH target gene that forms a positive feedback loop with GLI and exists at increased levels in BCCs. Genome-wide transcriptional profiling shows that aPKC-ι/λ and SMO control the expression of similar genes in tumour cells. aPKC-ι/λ functions downstream of SMO to phosphorylate and activate GLI1, resulting in maximal DNA binding and transcriptional activation. Activated aPKC-ι/λ is upregulated in SMO-inhibitor-resistant tumours and targeting aPKC-ι/λ suppresses signalling and growth of resistant BCC cell lines. These results demonstrate that aPKC-ι/λ is critical for HH-dependent processes and implicates aPKC-ι/λ as a new, tumour-selective therapeutic target for the treatment of SMO-inhibitor-resistant cancers.

  View details for DOI 10.1038/nature11889

  View details for Web of Science ID 000315661500040

• Hedgehog pathway inhibition and the race against tumor evolution JOURNAL OF CELL BIOLOGY Atwood, S. X., Chang, A. L., Oro, A. E. 2013: 45-49

  View details for DOI 10.1083/jcb.201207140

  View details for Web of Science ID 000313944700021

• Efficacy and Safety of Vismodegib in Advanced Basal-Cell Carcinoma NEW ENGLAND JOURNAL OF MEDICINE Sekulic, A., Migden, M. R., Oro, A. E., Dirix, L., Lewis, K. D., Hainsworth, J. D., Solomon, J. A., Yoo, S., Arron, S. T., Friedlander, P. A., Marmur, E., Rudin, C. M., Chang, A. L., Low, J. A., Mackey, H. M., Yauch, R. L., Graham, R. A., Reddy, J. C., Hauschild, A. 2012; 366 (23): 2171-2179

  Abstract

  Alterations in hedgehog signaling are implicated in the pathogenesis of basal-cell carcinoma. Although most basal-cell carcinomas are treated surgically, no effective therapy exists for locally advanced or metastatic basal-cell carcinoma. A phase 1 study of vismodegib (GDC-0449), a first-in-class, small-molecule inhibitor of the hedgehog pathway, showed a 58% response rate among patients with advanced basal-cell carcinoma.In this multicenter, international, two-cohort, nonrandomized study, we enrolled patients with metastatic basal-cell carcinoma and those with locally advanced basal-cell carcinoma who had inoperable disease or for whom surgery was inappropriate (because of multiple recurrences and a low likelihood of surgical cure, or substantial anticipated disfigurement). All patients received 150 mg of oral vismodegib daily. The primary end point was the independently assessed objective response rate; the primary hypotheses were that the response rate would be greater than 20% for patients with locally advanced basal-cell carcinoma and greater than 10% for those with metastatic basal-cell carcinoma.In 33 patients with metastatic basal-cell carcinoma, the independently assessed response rate was 30% (95% confidence interval [CI], 16 to 48; P=0.001). In 63 patients with locally advanced basal-cell carcinoma, the independently assessed response rate was 43% (95% CI, 31 to 56; P<0.001), with complete responses in 13 patients (21%). The median duration of response was 7.6 months in both cohorts. Adverse events occurring in more than 30% of patients were muscle spasms, alopecia, dysgeusia (taste disturbance), weight loss, and fatigue. Serious adverse events were reported in 25% of patients; seven deaths due to adverse events were noted.Vismodegib is associated with tumor responses in patients with locally advanced or metastatic basal-cell carcinoma. (Funded by Genentech; Erivance BCC ClinicalTrials.gov number, NCT00833417.).

  View details for Web of Science ID 000304863400006

  View details for PubMedID 22670903

  View details for PubMedCentralID PMC5278761

• Translocation Affecting Sonic Hedgehog Genes in Basal-Cell Carcinoma NEW ENGLAND JOURNAL OF MEDICINE Gomez-Ospina, N., Chang, A. L., Qu, K., Oro, A. E. 2012; 366 (23): 2233-2234

  View details for Web of Science ID 000304863400029

  View details for PubMedID 22670922

  View details for PubMedCentralID PMC3839666

• Shh maintains dermal papilla identity and hair morphogenesis via a Noggin-Shh regulatory loop GENES & DEVELOPMENT Woo, W., Zhen, H. H., Oro, A. E. 2012; 26 (11): 1235-1246

  Abstract

  During hair follicle morphogenesis, dermal papillae (DPs) function as mesenchymal signaling centers that cross-talk with overlying epithelium to regulate morphogenesis. While the DP regulates hair follicle formation, relatively little is known about the molecular basis of DP formation. The morphogen Sonic hedgehog (Shh) is known for regulating hair follicle epithelial growth, with excessive signaling resulting in basal cell carcinomas. Here, we investigate how dermal-specific Shh signaling contributes to DP formation and hair growth. Using a Cre-lox genetic model and RNAi in hair follicle reconstitution assays, we demonstrate that dermal Smoothened (Smo) loss of function results in the loss of the DP precursor, the dermal condensate, and a stage 2 hair follicle arrest phenotype reminiscent of Shh(-/-) skin. Surprisingly, dermal Smo does not regulate cell survival or epithelial proliferation. Rather, molecular screening and immunostaining studies reveal that dermal Shh signaling controls the expression of a subset of DP-specific signature genes. Using a hairpin/cDNA lentiviral system, we show that overexpression of the Shh-dependent gene Noggin, but not Sox2 or Sox18, can partially rescue the dermal Smo knockdown hair follicle phenotype by increasing the expression of epithelial Shh. Our findings suggest that dermal Shh signaling regulates specific DP signatures to maintain DP maturation while maintaining a reciprocal Shh-Noggin signaling loop to drive hair follicle morphogenesis.

  View details for DOI 10.1101/gad.187401.112

  View details for Web of Science ID 000304767000010

  View details for PubMedID 22661232

  View details for PubMedCentralID PMC3371411

• Efficacy and safety of the hedgehog pathway inhibitor vismodegib in patients with advanced basal cell carcinoma (BCC): ERIVANCE BCC study update Sekulic, A., Migden, M. R., Oro, A. E., Lewis, K. D., Hainsworth, J. D., Yoo, S., Dirix, L., Hou, J., Yue, H., Hauschild, A., ERIVANCE BCC Study Investigators AMER SOC CLINICAL ONCOLOGY. 2012

  View details for Web of Science ID 000318009800685

• SnapShot: Hair Follicle Stem Cells CELL Woo, W., Oro, A. E. 2011; 146 (2): 334-U159

  View details for DOI 10.1016/j.cell.2011.07.001

  View details for Web of Science ID 000293013000015

  View details for PubMedID 21784251

• MIM and Cortactin Antagonism Regulates Ciliogenesis and Hedgehog Signaling DEVELOPMENTAL CELL Bershteyn, M., Atwood, S. X., Woo, W., Li, M., Oro, A. E. 2010; 19 (2): 270-283

  Abstract

  The primary cilium is critical for transducing Sonic hedgehog (Shh) signaling, but the mechanisms of its transient assembly are poorly understood. Previously we showed that the actin regulatory protein Missing-in-Metastasis (MIM) regulates Shh signaling, but the nature of MIM's role was unknown. Here we show that MIM is required at the basal body of mesenchymal cells for cilia maintenance, Shh responsiveness, and de novo hair follicle formation. MIM knockdown results in increased Src kinase activity and subsequent hyperphosphorylation of the actin regulator Cortactin. Importantly, inhibition of Src or depletion of Cortactin compensates for the cilia defect in MIM knockdown cells, whereas overexpression of Src or phospho-mimetic Cortactin is sufficient to inhibit ciliogenesis. Our results suggest that MIM promotes ciliogenesis by antagonizing Src-dependent phosphorylation of Cortactin and describe a mechanism linking regulation of the actin cytoskeleton with ciliogenesis and Shh signaling during tissue regeneration.

  View details for DOI 10.1016/j.devcel.2010.07.009

  View details for Web of Science ID 000281090000012

  View details for PubMedID 20708589

  View details for PubMedCentralID PMC3108505

• I-BAR protein antagonism of endocytosis mediates directional sensing during guided cell migration JOURNAL OF CELL BIOLOGY Quinones, G. A., Jin, J., Oro, A. E. 2010; 189 (2): 353-367

  Abstract

  Although directed cellular migration facilitates the coordinated movement of cells during development and repair, the mechanisms regulating such migration remain poorly understood. Missing-in-metastasis (MIM) is a defining member of the inverse Bin/Amphiphysin/Rvs domain (I-BAR) subfamily of lipid binding, cytoskeletal regulators whose levels are altered in a number of cancers. Here, we provide the first genetic evidence that an I-BAR protein regulates directed cell migration in vivo. Drosophila MIM (dmim) is involved in Drosophila border cell migration, with loss of dmim function resulting in a lack of directional movement by the border cell cluster. In vivo endocytosis assays combined with genetic analyses demonstrate that the dmim product regulates directed cell movement by inhibiting endocytosis and antagonizing the activities of the CD2-associated protein/cortactin complex in these cells. These studies demonstrate that DMIM antagonizes pro-endocytic components to facilitate polarity and localized guidance cue sensing during directional cell migration.

  View details for DOI 10.1083/jcb.200910136

  View details for Web of Science ID 000276825200017

  View details for PubMedID 20385776

  View details for PubMedCentralID PMC2856902

• Basal-to-inflammatory transition and tumor resistance via crosstalk with a proinflammatory stromal niche Nature Communications Li, N. Y., Zhang, W., Haensel, D., Jussila, A., Pan, C., Gaddam, S., Plevritis, S., Oro, A. 2024; 15

  View details for DOI 10.1038/s41467-024-52394-3

• Integrated single-cell chromatin and transcriptomic analyses of human scalp identify gene-regulatory programs and critical cell types for hair and skin diseases. Nature genetics Ober-Reynolds, B., Wang, C., Ko, J. M., Rios, E. J., Aasi, S. Z., Davis, M. M., Oro, A. E., Greenleaf, W. J. 2023

  Abstract

  Genome-wide association studies have identified many loci associated with hair and skin disease, but identification of causal variants requires deciphering of gene-regulatory networks in relevant cell types. We generated matched single-cell chromatin profiles and transcriptomes from scalp tissue from healthy controls and patients with alopecia areata, identifying diverse cell types of the hair follicle niche. By interrogating these datasets at multiple levels of cellular resolution, we infer 50-100% more enhancer-gene links than previous approaches and show that aggregate enhancer accessibility for highly regulated genes predicts expression. We use these gene-regulatory maps to prioritize cell types, genes and causal variants implicated in the pathobiology of androgenetic alopecia (AGA), eczema and other complex traits. AGA genome-wide association studies signals are enriched in dermal papilla regulatory regions, supporting the role of these cells as drivers of AGA pathogenesis. Finally, we train machine learning models to nominate single-nucleotide polymorphisms that affect gene expression through disruption of transcription factor binding, predicting candidate functional single-nucleotide polymorphism for AGA and eczema.

  View details for DOI 10.1038/s41588-023-01445-4

  View details for PubMedID 37500727

  View details for PubMedCentralID 4006068

• Acquisition of drug resistance in basal cell nevus syndrome tumors through basal to squamous cell carcinoma transition. bioRxiv : the preprint server for biology Jussila, A. R., Haensel, D., Gaddam, S., Oro, A. E. 2023

  Abstract

  While basal cell carcinomas (BCCs) arise from ectopic hedgehog pathway activation and can be treated with pathway inhibitors, sporadic BCCs display high resistance rates while tumors arising in Gorlin syndrome patients with germline Patched ( PTCH1 ) mutations are uniformly suppressed by inhibitor therapy. In rare cases, Gorlin syndrome patients on long-term inhibitor therapy will develop individual resistant tumor clones that rapidly progress, but the basis of this resistance remains unstudied. Here we report a case of an SMO i -resistant tumor arising in a Gorlin patient on suppressive SMO i for nearly a decade. Using a combination of multi-omics and spatial transcriptomics, we define the tumor populations at the cellular and tissue level to conclude that Gorlin tumors can develop resistance to SMO i through the previously described basal to squamous cell carcinoma transition (BST). Intriguingly, through spatial whole exome genomic analysis, we nominate PCYT2, ETNK1, and the phosphatidylethanolamine biosynthetic pathway as novel genetic suppressors of BST resistance. These observations provide a general framework for studying tumor evolution and provide important clinical insight into mechanisms of resistance to SMO i for not only Gorlin syndrome but sporadic BCCs as well.

  View details for DOI 10.1101/2023.07.26.550719

  View details for PubMedID 37546976

• The HHIP-AS1 lncRNA promotes tumorigenicity through stabilization of dynein complex 1 in human SHH-driven tumors. Nature communications Bartl, J., Zanini, M., Bernardi, F., Forget, A., Blumel, L., Talbot, J., Picard, D., Qin, N., Cancila, G., Gao, Q., Nath, S., Koumba, I. M., Wolter, M., Kuonen, F., Langini, M., Beez, T., Munoz, C., Pauck, D., Marquardt, V., Yu, H., Souphron, J., Korsch, M., Molders, C., Berger, D., Gobbels, S., Meyer, F., Scheffler, B., Rotblat, B., Diederichs, S., Ramaswamy, V., Suzuki, H., Oro, A., Stuhler, K., Stefanski, A., Fischer, U., Leprivier, G., Willbold, D., Steger, G., Buell, A., Kool, M., Lichter, P., Pfister, S. M., Northcott, P. A., Taylor, M. D., Borkhardt, A., Reifenberger, G., Ayrault, O., Remke, M. 2022; 13 (1): 4061

  Abstract

  Most lncRNAs display species-specific expression patterns suggesting that animal models of cancer may only incompletely recapitulate the regulatory crosstalk between lncRNAs and oncogenic pathways in humans. Among these pathways, Sonic Hedgehog (SHH) signaling is aberrantly activated in several human cancer entities. We unravel that aberrant expression of the primate-specific lncRNA HedgeHog Interacting Protein-AntiSense 1 (HHIP-AS1) is a hallmark of SHH-driven tumors including medulloblastoma and atypical teratoid/rhabdoid tumors. HHIP-AS1 is actively transcribed from a bidirectional promoter shared with SHH regulator HHIP. Knockdown of HHIP-AS1 induces mitotic spindle deregulation impairing tumorigenicity in vitro and in vivo. Mechanistically, HHIP-AS1 binds directly to the mRNA of cytoplasmic dynein 1 intermediate chain 2 (DYNC1I2) and attenuates its degradation by hsa-miR-425-5p. We uncover that neither HHIP-AS1 nor the corresponding regulatory element in DYNC1I2 are evolutionary conserved in mice. Taken together, we discover an lncRNA-mediated mechanism that enables the pro-mitotic effects of SHH pathway activation in human tumors.

  View details for DOI 10.1038/s41467-022-31574-z

  View details for PubMedID 35831316

• A conserved YAP/Notch/REST network controls the neuroendocrine cell fate in the lungs. Nature communications Shue, Y. T., Drainas, A. P., Li, N. Y., Pearsall, S. M., Morgan, D., Sinnott-Armstrong, N., Hipkins, S. Q., Coles, G. L., Lim, J. S., Oro, A. E., Simpson, K. L., Dive, C., Sage, J. 2022; 13 (1): 2690

  Abstract

  The Notch pathway is a conserved cell-cell communication pathway that controls cell fate decisions. Here we sought to determine how Notch pathway activation inhibits the neuroendocrine cell fate in the lungs, an archetypal process for cell fate decisions orchestrated by Notch signaling that has remained poorly understood at the molecular level. Using intratumoral heterogeneity in small-cell lung cancer as a tractable model system, we uncovered a role for the transcriptional regulators REST and YAP as promoters of the neuroendocrine to non-neuroendocrine transition. We further identified the specific neuroendocrine gene programs repressed by REST downstream of Notch in this process. Importantly, we validated the importance of REST and YAP in neuroendocrine to non-neuroendocrine cell fate switches in both developmental and tissue repair processes in the lungs. Altogether, these experiments identify conserved roles for REST and YAP in Notch-driven inhibition of the neuroendocrine cell fate in embryonic lungs, adult lungs, and lung cancer.

  View details for DOI 10.1038/s41467-022-30416-2

  View details for PubMedID 35577801

• Is hypoimmunogenic stem cell therapy safe in times of pandemics? Stem cell reports Matheus, F., Raveh, T., Oro, A. E., Wernig, M., Drukker, M. 2022

  Abstract

  The manipulation of human leukocyte antigens (HLAs) and immune modulatory factors in "universal" human pluripotent stem cells (PSCs) holds promise for immunological tolerance without HLA matching. This paradigm raises concerns should "universal" grafts become virally infected. Furthermore, immunological manipulation might functionally impair certain progeny, such as hematopoietic stem cells. We discuss the risks and benefits of hypoimmunogenic PSCs, and the need to further advance HLA matching and autologous strategies.

  View details for DOI 10.1016/j.stemcr.2022.02.014

  View details for PubMedID 35334219

• Phase II Open-Label, Single-Arm Trial to Investigate the Efficacy and Safety of Topical Remetinostat Gel in Patients with Basal Cell Carcinoma. Clinical cancer research : an official journal of the American Association for Cancer Research Kilgour, J. M., Shah, A., Urman, N. M., Eichstadt, S., Do, H. N., Bailey, I., Mirza, A., Li, S., Oro, A. E., Aasi, S. Z., Sarin, K. Y. 2021

  Abstract

  PURPOSE: The mainstay of treatment for basal cell carcinoma (BCC) is surgical excision, which can result in significant associated morbidity, particularly for patients with recurrent tumors. We previously conducted a drug repositioning screen using molecular data from human BCCs and identified histone deacetylase (HDAC) inhibitors as a potential treatment for BCC. Here we conduct the first proof-of-principle study of a topical pan-HDAC inhibitor, remetinostat, in human BCC.PATIENTS AND METHODS: We conducted a phase II, open-label, single-arm, single-institution trial of a topical HDAC inhibitor. Participants with at least one BCC were recruited. All participants applied 1% remetinostat gel three times daily for 6 weeks, with measurements of tumor diameter conducted at baseline and week 8. Surgical excision of the remaining tumor was conducted at the end of the study and microscopic evaluation was performed.RESULTS: Thirty-three per-protocol tumors from 25 participants were included in the analysis. The overall response rate, defined as the proportion of tumors achieving more than 30% decrease in the longest diameter from baseline to week 8, was 69.7% [90% confidence interval (CI), 54%-82.5%]. On pathologic examination, 54.8% of tumors demonstrated complete resolution. Pharmacodynamic analysis demonstrated similar levels of acetylated histone H3 in skin tissue before and after treatment, however, phosphorylation was increased. No systemic adverse events were reported.CONCLUSIONS: The HDAC inhibitor remetinostat is a well-tolerated and effective topical treatment for reducing BCC disease burden in a clinically significant manner. This provides in-human validation of HDAC inhibitors as a therapy for BCC.

  View details for DOI 10.1158/1078-0432.CCR-21-0560

  View details for PubMedID 34362809

• C-FOS drives reversible basal to squamous cell carcinoma transition Kuonen, F., Li, N., Haensel, D., Patel, T., Gaddam, S., Yerly, L., Rieger, K., Aasi, S., Oro, A. ELSEVIER SCIENCE INC. 2021: S11

  View details for Web of Science ID 000641872800065

• Starve a cold, and perhaps a cancer. Nature cell biology Haensel, D., Oro, A. E. 2020

  View details for DOI 10.1038/s41556-020-0543-7

  View details for PubMedID 32587343

• Receptor protein tyrosine phosphatases control Purkinje neuron firing. Cell cycle (Georgetown, Tex.) Brown, A. S., Meera, P., Quinones, G., Magri, J., Otis, T. S., Pulst, S. M., Oro, A. E. 2019: 1–7

  Abstract

  Spinocerebellar ataxias (SCA) are a genetically heterogeneous family of cerebellar neurodegenerative diseases characterized by abnormal firing of Purkinje neurons and degeneration. We recently demonstrated the slowed firing rates seen in several SCAs share a common etiology of hyper-activation of the Src family of non-receptor tyrosine kinases (SFKs). However, the lack of clinically available neuroactive SFK inhibitors lead us to investigate alternative mechanisms to modulate SFK activity. Previous studies demonstrate that SFK activity can be enhanced by the removal of inhibitory phospho-marks by receptor-protein-tyrosine phosphatases (RPTPs). In this Extra View we show that MTSS1 inhibits SFK activity through the binding and inhibition of a subset of the RPTP family members, and lowering RPTP activity in cerebellar slices with peptide inhibitors increases the suppressed Purkinje neuron basal firing rates seen in two different SCA models. Together these results identify RPTPs as novel effectors of Purkinje neuron basal firing, extending the MTSS1/SFK regulatory circuit we previously described and expanding the therapeutic targets for SCA patients.

  View details for DOI 10.1080/15384101.2019.1695995

  View details for PubMedID 31876231

• Loss of Primary Cilia Drives Switching from Hedgehog to Ras/MAPK Pathway in Resistant Basal Cell Carcinoma JOURNAL OF INVESTIGATIVE DERMATOLOGY Kuonen, F., Huskey, N. E., Shankar, G., Jaju, P., Whitson, R. J., Rieger, K. E., Atwood, S. X., Sarin, K. Y., Oro, A. E. 2019; 139 (7): 1439–48

  View details for DOI 10.1016/j.jid.2018.11.035

  View details for Web of Science ID 000472234100017

• Genetic Mutations Underlying Phenotypic Plasticity in Basosquamous Carcinoma Journal of INVESTIGATIVE DERMATOLOGY | Original Article | Carcinogenesis/Tumorigenesis Chiang, A., Tan, C. Z., Kuonen, F., Hodgkinson, L. M., Chiang, F., Cho, R. J., South, A. P., Tang, J. Y., Chang, A. S., Rieger, K. E., Oro, A. E., Sarin, K. Y. 2019; 139 (11): 2263-2271. E5

  View details for DOI 10.1016/j.jid.2019.03.1163

• MTSS1/Src family kinase dysregulation underlies multiple inherited ataxias PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Brown, A. S., Meera, P., Altindag, B., Chopra, R., Perkins, E. M., Paul, S., Scoles, D. R., Tarapore, E., Magri, J., Huang, H., Jackson, M., Shakkottai, V. G., Otis, T. S., Pulst, S. M., Atwood, S. X., Oro, A. E. 2018; 115 (52): E12407-E12416

  View details for DOI 10.1073/pnas.1816177115

  View details for Web of Science ID 000454302600035

• TGF beta, Fibronectin and Integrin alpha 5 beta 1 Promote Invasion in Basal Cell Carcinoma JOURNAL OF INVESTIGATIVE DERMATOLOGY Kuonen, F., Surbeck, I., Sarin, K. Y., Dontenwill, M., Ruegg, C., Gilliet, M., Oro, A. E., Gaide, O. 2018; 138 (11): 2432-2442

  View details for DOI 10.1016/j.jid.2018.04.029

  View details for Web of Science ID 000447794000024

• Actin polymerization controls cilia-mediated signaling JOURNAL OF CELL BIOLOGY Drummond, M. L., Li, M., Tarapore, E., Nguyen, T. L., Barouni, B. J., Cruz, S., Tan, K. C., Oro, A. E., Atwood, S. X. 2018; 217 (9): 3255–66

  Abstract

  Primary cilia are polarized organelles that allow detection of extracellular signals such as Hedgehog (Hh). How the cytoskeleton supporting the cilium generates and maintains a structure that finely tunes cellular response remains unclear. Here, we find that regulation of actin polymerization controls primary cilia and Hh signaling. Disrupting actin polymerization, or knockdown of N-WASp/Arp3, increases ciliation frequency, axoneme length, and Hh signaling. Cdc42, a potent actin regulator, recruits both atypical protein pinase C iota/lambda (aPKC) and Missing-in-Metastasis (MIM) to the basal body to maintain actin polymerization and restrict axoneme length. Transcriptome analysis implicates the Src pathway as a major aPKC effector. aPKC promotes whereas MIM antagonizes Src activity to maintain proper levels of primary cilia, actin polymerization, and Hh signaling. Hh pathway activation requires Smoothened-, Gli-, and Gli1-specific activation by aPKC. Surprisingly, longer axonemes can amplify Hh signaling, except when aPKC is disrupted, reinforcing the importance of the Cdc42-aPKC-Gli axis in actin-dependent regulation of primary cilia signaling.

  View details for PubMedID 29945904

  View details for PubMedCentralID PMC6122990

• TGF-beta, Fibronectin and Integrin alpha5beta1 Promote Invasion in Basal Cell Carcinoma. The Journal of investigative dermatology Kuonen, F., Surbeck, I., Sarin, K. Y., Dontenwill, M., Ruegg, C., Gilliet, M., Oro, A. E., Gaide, O. 2018

  Abstract

  Basal cell carcinoma (BCC) is the most frequent human cancer and is becoming an important health problem in an ageing population. Based on their clinical and histological characteristics, thick BCC are typically divided into low-risk nodular and high-risk infiltrative subtypes, although the underlying mechanisms are poorly understood. We have identified molecular mechanisms that explain the aggressiveness of high-risk infiltrative BCC, with a potential direct clinical impact. In this study, we first show that fibroblasts, TGFbeta and fibronectin are found preferentially in infiltrative human BCC. This allowed us to develop in vivo models for the study of infiltrative BCC, which in turn let us confirm the role of TGFbeta in inducing peritumoral fibronectin deposition and tumor infiltration. We then show that fibronectin promotes adhesion and migration of BCC cell lines through integrin alpha5beta1-mediated phosphorylation of focal adhesion kinase (FAK). Fittingly, inhibition of integrin alpha5beta1 and phospho-FAK both prevent fibronectin-induced migration of BCC cells in vitro as well as BCC infiltration in vivo. Altogether our results open important insights into the pathogenesis of aggressive infiltrative BCC, and identify integrin alpha5beta1 or FAK inhibition as promising strategies for the treatment of advanced BCC.

  View details for PubMedID 29758283

• Modulation of the Hedgehog signaling pathway in models of basal cell carcinoma by ATP-competitive PKCi inhibitors Roffey, J., Dillon, C., Oro, A. E., Mirza, A. N., Sarin, K. Y., Aasi, S. Z., Parker, P. J., Riou, P., Barton, C., Patel, B., Turnbull, A., Stanway, E., Fowler, K., Ott, G., Ator, M. AMER ASSOC CANCER RESEARCH. 2018

  View details for DOI 10.1158/1535-7163.TARG-17-LB-B32

  View details for Web of Science ID 000590008800478

• Tumor-Derived Suppressor of Fused Mutations Reveal Hedgehog Pathway Interactions PLOS ONE Urman, N. M., Mirza, A., Atwood, S. X., Whitson, R. J., Sarin, K. Y., Tang, J. Y., Oro, A. E. 2016; 11 (12)

  Abstract

  The Hedgehog pathway is a potent regulator of cellular growth and plays a central role in the development of many cancers including basal cell carcinoma (BCC). The majority of BCCs arise from mutations in the Patched receptor resulting in constitutive activation of the Hedgehog pathway. Secondary driver mutations promote BCC oncogenesis and occur frequently due to the high mutational burden resulting from sun exposure of the skin. Here, we uncover novel secondary mutations in Suppressor of Fused (SUFU), the major negative regulator of the Hedgehog pathway. SUFU normally binds to a Hedgehog transcriptional activator, GLI1, in order to prevent it from initiating transcription of Hedgehog target genes. We sequenced tumor-normal pairs from patients with early sporadic BCCs. This resulted in the discovery of nine mutations in SUFU, which were functionally investigated to determine whether they help drive BCC formation. Our results show that four of the SUFU mutations inappropriately activate the Hedgehog pathway, suggesting they may act as driver mutations for BCC development. Indeed, all four of the loss of function SUFU variants were found to disrupt its binding to GLI, leading to constitutive pathway activation. Our results from functional characterization of these mutations shed light on SUFU's role in Hedgehog signaling, tumor progression, and highlight a way in which BCCs can arise.

  View details for DOI 10.1371/journal.pone.0168031

  View details for Web of Science ID 000391222000030

  View details for PubMedID 28030567

  View details for PubMedCentralID PMC5193403

• Safety and efficacy of the JAK inhibitor tofacitinib citrate in patients with alopecia areata JCI INSIGHT Crispin, M., Ko, J. M., Craiglow, B. G., Li, S., Shankar, G., Urban, J. R., Chen, J. C., Cerise, J. E., Jabbari, A., Winge, M. G., Marinkovich, M., Christiano, A. M., Oro, A. E., King, B. A. 2016; 1 (15)

  View details for DOI 10.1172/jci.insight.89776

  View details for Web of Science ID 000387123200013

• Factors That May Promote an Effective Local Research Environment JOURNAL OF INVESTIGATIVE DERMATOLOGY Wang, K., Lee, C. S., Marinkovich, M., Chang, H. Y., Oro, A. E., Khavari, P. A. 2016; 136 (8): 1529–31

  View details for PubMedID 27450496

• Update to an open-label clinical trial of vismodegib as neoadjuvant before surgery for high-risk basal cell carcinoma (BCC) JOURNAL OF THE AMERICAN ACADEMY OF DERMATOLOGY Kwon, G. P., Ally, M., Bailey-Healy, I., Oro, A. E., Kim, J., Chang, A., Aasi, S., Tang, J. Y. 2016; 75 (1): 213–15

  View details for PubMedID 27317518

• Effects of Combined Treatment With Arsenic Trioxide and Itraconazole in Patients With Refractory Metastatic Basal Cell Carcinoma. JAMA dermatology Ally, M. S., Ransohoff, K., Sarin, K., Atwood, S. X., Rezaee, M., Bailey-Healy, I., Kim, J., Beachy, P. A., Chang, A. L., Oro, A., Tang, J. Y., Colevas, A. D. 2016; 152 (4): 452-456

  Abstract

  Tumor resistance is an emerging problem for Smoothened (SMO) inhibitor-treated metastatic basal cell carcinoma (BCC). Arsenic trioxide and itraconazole antagonize the hedgehog (HH) pathway at sites distinct from those treated by SMO inhibitors.To determine whether administration of intravenous arsenic trioxide and oral itraconazole in patients with metastatic BCC is associated with a reduction in GLI1 messenger RNA expression in tumor and/or normal skin biopsy samples.Five men with metastatic BCC who experienced relapse after SMO inhibitor treatment underwent intravenous arsenic trioxide treatment for 5 days, every 28 days, and oral itraconazole treatment on days 6 to 28. Data were collected from April 10 to November 14, 2013. Follow-up was completed on October 3, 2015, and data were analyzed from June 5 to October 6, 2015.The primary outcome was the change in messenger RNA levels of the GLI family zinc finger 1 (GLI1) gene (HH-pathway target gene) in biopsy specimens of normal skin or BCC before and after treatment. Secondary objectives were evaluation of tumor response and tolerability.Of the 5 patients (mean [SD] age, 52 [9] years; age range, 43-62 years), 3 completed 3 cycles of treatment and 2 discontinued treatment early owing to disease progression or adverse events. Adverse effects included grade 2 transaminitis and grade 4 leukopenia with a grade 3 infection. Overall, arsenic trioxide and itraconazole reduced GLI1 messenger RNA levels by 75% from baseline (P < .001). The best overall response after 3 treatment cycles was stable disease in 3 patients.Targeting the HH pathway with sequential arsenic trioxide and itraconazole treatment is a feasible treatment for metastatic BCC. Although some patients experienced stable disease for 3 months, none had tumor shrinkage, which may be owing to transient GLI1 suppression with sequential dosing. Continuous dosing may be required to fully inhibit the HH pathway and achieve clinical response.

  View details for DOI 10.1001/jamadermatol.2015.5473

  View details for PubMedID 26765315

• An Investigator-Initiated Open-Label Trial of Sonidegib in Advanced Basal Cell Carcinoma Patients Resistant to Vismodegib CLINICAL CANCER RESEARCH Danial, C., Sarin, K. Y., Oro, A. E., Chang, A. L. 2016; 22 (6): 1325-1329

  Abstract

  To assess the tumor response to the smoothened (SMO) inhibitor, sonidegib (LDE225), in patients with an advanced basal cell carcinoma (BCC) resistant to treatment with vismodegib (GDC0449).Nine patients with an advanced BCC that was previously resistant to treatment with vismodegib were given sonidegib in this investigational, open-label study. Tumor response was determined using the response evaluation criteria in solid tumors. SMO mutations were identified using biopsy samples from the target BCC location.The median duration of treatment with sonidegib was 6 weeks (range, 3-58 weeks). Five patients experienced progressive disease with sonidegib. Three patients experienced stable disease and discontinued sonidegib either due to adverse events (n = 1) or due to election for surgery (n = 2). The response of one patient was not evaluable. SMO mutations with in vitro data suggesting resistance to Hh pathway inhibition were identified in 5 patients, and none of these patients experienced responses while on sonidegib.Patients with advanced BCCs that were previously resistant to treatment with vismodegib similarly demonstrated treatment resistance with sonidegib. Patients who have developed treatment resistance to an SMO inhibitor may continue to experience tumor progression in response to other SMO inhibitors. Clin Cancer Res; 22(6); 1325-9. ©2015 AACR.

  View details for DOI 10.1158/1078-0432.CCR-15-1588

  View details for Web of Science ID 000373358900006

  View details for PubMedID 26546616

  View details for PubMedCentralID PMC4794361

• Effects of combined treatment with arsenic trioxide and itraconazole in patients with refractory metastatic basal cell carcinoma JAMA Dermatology Sarin, K. Y., Ally, M. S., Ransohoff, K. J., Atwood, S. X., Rezaee, M. 2016: 1–5

  Abstract

  Tumor resistance is an emerging problem for Smoothened (SMO) inhibitor-treated metastatic basal cell carcinoma (BCC). Arsenic trioxide and itraconazole antagonize the hedgehog (HH) pathway at sites distinct from those treated by SMO inhibitors.To determine whether administration of intravenous arsenic trioxide and oral itraconazole in patients with metastatic BCC is associated with a reduction in GLI1 messenger RNA expression in tumor and/or normal skin biopsy samples.Five men with metastatic BCC who experienced relapse after SMO inhibitor treatment underwent intravenous arsenic trioxide treatment for 5 days, every 28 days, and oral itraconazole treatment on days 6 to 28. Data were collected from April 10 to November 14, 2013. Follow-up was completed on October 3, 2015, and data were analyzed from June 5 to October 6, 2015.The primary outcome was the change in messenger RNA levels of the GLI family zinc finger 1 (GLI1) gene (HH-pathway target gene) in biopsy specimens of normal skin or BCC before and after treatment. Secondary objectives were evaluation of tumor response and tolerability.Of the 5 patients (mean [SD] age, 52 [9] years; age range, 43-62 years), 3 completed 3 cycles of treatment and 2 discontinued treatment early owing to disease progression or adverse events. Adverse effects included grade 2 transaminitis and grade 4 leukopenia with a grade 3 infection. Overall, arsenic trioxide and itraconazole reduced GLI1 messenger RNA levels by 75% from baseline (P < .001). The best overall response after 3 treatment cycles was stable disease in 3 patients.Targeting the HH pathway with sequential arsenic trioxide and itraconazole treatment is a feasible treatment for metastatic BCC. Although some patients experienced stable disease for 3 months, none had tumor shrinkage, which may be owing to transient GLI1 suppression with sequential dosing. Continuous dosing may be required to fully inhibit the HH pathway and achieve clinical response.

  View details for DOI 10.1001/jamadermatol.2015.5473

• Rolling the Genetic Dice: Neutral and Deleterious Smoothened Mutations in Drug-Resistant Basal Cell Carcinoma. journal of investigative dermatology Atwood, S. X., Sarin, K. Y., Li, J. R., Yao, C. Y., Urman, N. M., Chang, A. L., Tang, J. Y., Oro, A. E. 2015; 135 (8): 2138-2141

  View details for DOI 10.1038/jid.2015.115

  View details for PubMedID 25801792

• Vismodegib for advanced basal cell carcinoma: Duration of response after vismodegib discontinuation and response to vismodegib retreatment upon disease progression Sekulic, A., Hainsworth, J. D., Lewis, K. D., Oro, A. E., Gesierich, A., Mortier, L., Dirix, L., Bernard, S., Dreno, B., Murrell, D. F., Williams, S., Hou, J., Fisher, D. MOSBY-ELSEVIER. 2015: AB195

  View details for Web of Science ID 000360942901499

• An investigator-initiated open-label clinical trial of vismodegib as a neoadjuvant to surgery for high-risk basal cell carcinoma. Journal of the American Academy of Dermatology Ally, M. S., Aasi, S., Wysong, A., Teng, C., Anderson, E., Bailey-Healy, I., Oro, A., Kim, J., Chang, A. L., Tang, J. Y. 2014; 71 (5): 904-911 e1

  Abstract

  Vismodegib is an oral hedgehog-pathway inhibitor approved for advanced basal cell carcinoma (BCC). Although most BCCs are amenable to surgery, excision of large tumors in aesthetically sensitive sites may compromise function or cosmesis.We sought to evaluate the reduction in BCC surgical defect area after 3 to 6 months of neoadjuvant vismodegib.This was an open-label, single-arm intervention trial with a primary outcome of change in target-tumor surgical defect area pre- and post-vismodegib (150 mg/d). Secondary outcomes were change in tumor area and tolerability.Eleven of 15 enrolled patients, aged 39 to 100 years, completed the trial. Thirteen target tumors were excised after a mean of 4±2 months of vismodegib. In all, 29% (4 of 14 patients) could not complete more than 3 months because of vismodegib-related side effects. The mean baseline target-tumor diameter was 3.2 cm, and 10 of 13 tumors occurred on the face. Overall, vismodegib reduced the surgical defect area by 27% (95% confidence interval -45.7% to -7.9%; P=.006) from baseline. Vismodegib was not effective in patients who received less than 3 months. Over a mean follow-up of 11.5 (range 4-21) months for all tumors, only 1 tumor recurred at 17 months post-Mohs micrographic surgery.Short follow-up time and no placebo control are limitations.Neoadjuvant vismodegib appears to reduce surgical defect area when taken for 3 months or longer for nonrecurrent BCCs in functionally sensitive locations. Further studies with larger sample sizes and long-term follow-up are warranted.

  View details for DOI 10.1016/j.jaad.2014.05.020

  View details for PubMedID 24929884

• Advanced treatment for basal cell carcinomas. Cold Spring Harbor perspectives in medicine Atwood, S. X., Whitson, R. J., Oro, A. E. 2014; 4 (7)

  Abstract

  Basal cell carcinomas (BCCs) are very common epithelial cancers that depend on the Hedgehog pathway for tumor growth. Traditional therapies such as surgical excision are effective for most patients with sporadic BCC; however, better treatment options are needed for cosmetically sensitive or advanced and metastatic BCC. The first approved Hedgehog antagonist targeting the membrane receptor Smoothened, vismodegib, shows remarkable effectiveness on both syndromic and nonsyndromic BCCs. However, drug-resistant tumors frequently develop, illustrating the need for the development of next-generation Hedgehog antagonists targeting pathway components downstream from Smoothened. In this article, we will summarize available BCC treatment options and discuss the development of next-generation antagonists.

  View details for DOI 10.1101/cshperspect.a013581

  View details for PubMedID 24985127

• Epigenetic targeting of Hedgehog pathway transcriptional output through BET bromodomain inhibition. Nature medicine Tang, Y., Gholamin, S., Schubert, S., Willardson, M. I., Lee, A., Bandopadhayay, P., Bergthold, G., Masoud, S., Nguyen, B., Vue, N., Balansay, B., Yu, F., Oh, S., Woo, P., Chen, S., Ponnuswami, A., Monje, M., Atwood, S. X., Whitson, R. J., Mitra, S., Cheshier, S. H., Qi, J., Beroukhim, R., Tang, J. Y., Wechsler-Reya, R., Oro, A. E., Link, B. A., Bradner, J. E., Cho, Y. 2014; 20 (7): 732-740

  Abstract

  Hedgehog signaling drives oncogenesis in several cancers, and strategies targeting this pathway have been developed, most notably through inhibition of Smoothened (SMO). However, resistance to Smoothened inhibitors occurs by genetic changes of Smoothened or other downstream Hedgehog components. Here we overcome these resistance mechanisms by modulating GLI transcription through inhibition of bromo and extra C-terminal (BET) bromodomain proteins. We show that BRD4 and other BET bromodomain proteins regulate GLI transcription downstream of SMO and suppressor of fused (SUFU), and chromatin immunoprecipitation studies reveal that BRD4 directly occupies GLI1 and GLI2 promoters, with a substantial decrease in engagement of these sites after treatment with JQ1, a small-molecule inhibitor targeting BRD4. Globally, genes associated with medulloblastoma-specific GLI1 binding sites are downregulated in response to JQ1 treatment, supporting direct regulation of GLI activity by BRD4. Notably, patient- and GEMM (genetically engineered mouse model)-derived Hedgehog-driven tumors (basal cell carcinoma, medulloblastoma and atypical teratoid rhabdoid tumor) respond to JQ1 even when harboring genetic lesions rendering them resistant to Smoothened antagonists. Altogether, our results reveal BET proteins as critical regulators of Hedgehog pathway transcriptional output and nominate BET bromodomain inhibitors as a strategy for treating Hedgehog-driven tumors with emerged or a priori resistance to Smoothened antagonists.

  View details for DOI 10.1038/nm.3613

  View details for PubMedID 24973920

• OVERCOMING ACQUIRED AND A PRIORI RESISTANCE TO SMO ANTAGONISTS THROUGH EPIGENETIC REGULATION OF HEDGEHOG PATHWAY TRANSCRIPTIONAL OUTPUT Tang, Y., Schubert, S., Brian Nguyen, Masoud, S., Gholamin, S., Lee, A., Willardson, M., Bandopadhayay, P., Bergthold, G., Atwood, S., Whitson, R., Cheshier, S., Qi, J., Beroukhim, R., Tang, J., Wechsler-Reya, R., Oro, A., Link, B., Bradner, J., Cho, Y. OXFORD UNIV PRESS INC. 2014: 81–82

  View details for Web of Science ID 000337924200309

• Vismodegib for advanced basal cell carcinoma: Duration of response after vismodegib discontinuation and response to vismodegib retreatment upon disease progression. Sekulic, A., Hainsworth, J. D., Lewis, K. D., Oro, A. E., Gesierich, A., Mortier, L., Dirix, L., Bernard, S., Dreno, B., Murrell, D. F., Williams, S., Hou, J., Fisher, D., Hauschild, A. AMER SOC CLINICAL ONCOLOGY. 2014

  View details for DOI 10.1200/jco.2014.32.15_suppl.9081

  View details for Web of Science ID 000358613204366

• Vismodegib in the treatment of patients with metastatic basal cell carcinoma (mBCC) and distant metastases: Survival in the pivotal phase II and phase I studies. Lewis, K. D., Sekulic, A., Hauschild, A., Migden, M., Oro, A. E., LoRusso, P., Rudin, C. M., Dirix, L., Solomon, J. A., Hainsworth, J. D., Williams, S., Hou, J., Von Hoff, D. D. AMER SOC CLINICAL ONCOLOGY. 2014

  View details for DOI 10.1200/jco.2014.32.15_suppl.9012

  View details for Web of Science ID 000358613204298

• Partial Proteasome Inhibitors Induce Hair Follicle Growth by Stabilizing ß-Catenin. Stem cells Yucel, G., Van Arnam, J., Means, P. C., Huntzicker, E., Altindag, B., Lara, M. F., Yuan, J., Kuo, C., Oro, A. E. 2014; 32 (1): 85-92

  Abstract

  The activation of tissue stem cells from their quiescent state represents the initial step in the complex process of organ regeneration and tissue repair. While the identity and location of tissue stem cells are becoming known, how key regulators control the balance of activation and quiescence remains mysterious. The vertebrate hair is an ideal model system where hair cycling between growth and resting phases is precisely regulated by morphogen signaling pathways, but how these events are coordinated to promote orderly signaling in a spatial and temporal manner remains unclear. Here, we show that hair cycle timing depends on regulated stability of signaling substrates by the ubiquitin-proteasome system. Topical application of partial proteasomal inhibitors (PaPIs) inhibits epidermal and dermal proteasome activity throughout the hair cycle. PaPIs prevent the destruction of the key anagen signal β-catenin, resulting in more rapid hair growth and dramatically shortened telogen. We show that PaPIs induce excess β-catenin, act similarly to the GSK3β antagonist LiCl, and antagonize Dickopf-related protein-mediated inhibition of anagen. PaPIs thus represent a novel class of hair growth agents that act through transiently modifying the balance of stem cell activation and quiescence pathways. Stem Cells 2014;32:85-92.

  View details for DOI 10.1002/stem.1525

  View details for PubMedID 23963711

• Epigenetic targeting of hedgehog pathway transcriptional output. Neuro-oncology Cho, Y. J., Tang, Y. n., Schubert, S. n., Willardson, M. n., Bandopadhayay, P. n., Bergthold, G. n., Nguyen, B. n., Masoud, S. n., Vue, N. n., Balansay, B. n., Gholamin, S. n., Cheshier, S. H., Atwood, S. X., Whitson, R. J., Lee, A. n., Tang, J. Y., Qi, J. n., Beroukhim, R. n., Wechsler-Reya, R. n., Oro, A. E., Link, B. n., Bradner, J. E., Cho, Y. J. 2014; 16 Suppl 3: iii25

  Abstract

  (blind field)We used ligand and genetic activation of the Hedgehog pathway to study the effects of BET bromodomain inhibition on Hedgehog pathway transcriptional output. Furthermore, we studied the in vitro and in vivo efficacy of BET bromodomain inhibitors using tumor cells generated from genetically engineered mouse (GEM) and patient derived xenograft models of Hedgehog driven tumors, including a panel of tumors resistant to the current FDA-approved Smoothened antagonists.We show that knockdown of BRD4 or treatment with the BET bromodomain inhibitor, JQ1, dramatically inhibits transcription of GLI1, GLI2 and other Hedgehog target genes upon ligand-mediated or genetic activation of the Hedgehog pathway. We confirm the inhibitory effect of JQ1 occurs downstream of SMO and SUFU and verify by chromatin immunoprecipitation that BRD4 directly occupies the GLI1 and GLI2 promoters with a substantial decrease in the engagement of these genomic sites upon treatment with JQ1. We observe a corresponding downregulation of genes associated with medulloblastoma-specific GLI1 binding sites upon exposure to JQ1, confirming the direct regulation of GLI1 by BET bromodomain proteins. Finally, in patient- and GEM-derived cells of Hedgehog-driven cancer (basal cell carcinoma, medulloblastoma and atypical teratoid/rhabdoid tumor), we show that JQ1 decreases Hh pathway output and proliferation, even in cells resistant to Smoothened inhibitors.These results expand the role of BET bromodomain inhibitors to targeting Hedgehog-driven cancers and highlight a strategy that overcomes the limitation of Hedgehog pathway inhibitors currently in clinical use.Pediatrics.

  View details for DOI 10.1093/neuonc/nou208.9

  View details for PubMedID 25165259

• Augmenting Endogenous Wnt Signaling Improves Skin Wound Healing PLOS ONE Whyte, J. L., Smith, A. A., Liu, B., Manzano, W. R., Evans, N. D., Dhamdhere, G. R., Fang, M. Y., Chang, H. Y., Oro, A. E., Helms, J. A. 2013; 8 (10)

  Abstract

  Wnt signaling is required for both the development and homeostasis of the skin, yet its contribution to skin wound repair remains controversial. By employing Axin2(LacZ/+) reporter mice we evaluated the spatial and temporal distribution patterns of Wnt responsive cells, and found that the pattern of Wnt responsiveness varies with the hair cycle, and correlates with wound healing potential. Using Axin2(LacZ/LacZ) mice and an ear wound model, we demonstrate that amplified Wnt signaling leads to improved healing. Utilizing a biochemical approach that mimics the amplified Wnt response of Axin2(LacZ/LacZ) mice, we show that topical application of liposomal Wnt3a to a non-healing wound enhances endogenous Wnt signaling, and results in better skin wound healing. Given the importance of Wnt signaling in the maintenance and repair of skin, liposomal Wnt3a may have widespread application in clinical practice.

  View details for DOI 10.1371/journal.pone.0076883

  View details for Web of Science ID 000326029300061

  View details for PubMedID 24204695

  View details for PubMedCentralID PMC3799989

• AAV-Mediated Direct Somatic Genetic Correction of Epidermolysis Bullosa Melo, S., Lisowski, L., Bashkirova, L., Kay, M. A., Oro, A. E. NATURE PUBLISHING GROUP. 2013: S106

  View details for Web of Science ID 000319858400276

• Rapid Genetic Analysis of Epithelial-Mesenchymal Signaling During Hair Regeneration JOVE-JOURNAL OF VISUALIZED EXPERIMENTS Woo, W., Atwood, S. X., Zhen, H. H., Oro, A. E. 2013

  Abstract

  Hair follicle morphogenesis, a complex process requiring interaction between epithelia-derived keratinocytes and the underlying mesenchyme, is an attractive model system to study organ development and tissue-specific signaling. Although hair follicle development is genetically tractable, fast and reproducible analysis of factors essential for this process remains a challenge. Here we describe a procedure to generate targeted overexpression or shRNA-mediated knockdown of factors using lentivirus in a tissue-specific manner. Using a modified version of a hair regeneration model, we can achieve robust gain- or loss-of-function analysis in primary mouse keratinocytes or dermal cells to facilitate study of epithelial-mesenchymal signaling pathways that lead to hair follicle morphogenesis. We describe how to isolate fresh primary mouse keratinocytes and dermal cells, which contain dermal papilla cells and their precursors, deliver lentivirus containing either shRNA or cDNA to one of the cell populations, and combine the cells to generate fully formed hair follicles on the backs of nude mice. This approach allows analysis of tissue-specific factors required to generate hair follicles within three weeks and provides a fast and convenient companion to existing genetic models.

  View details for DOI 10.3791/4344

  View details for Web of Science ID 000209226500006

  View details for PubMedCentralID PMC3622109

• Rapid genetic analysis of epithelial-mesenchymal signaling during hair regeneration. Journal of visualized experiments : JoVE Woo, W., Atwood, S. X., Zhen, H. H., Oro, A. E. 2013

  Abstract

  Hair follicle morphogenesis, a complex process requiring interaction between epithelia-derived keratinocytes and the underlying mesenchyme, is an attractive model system to study organ development and tissue-specific signaling. Although hair follicle development is genetically tractable, fast and reproducible analysis of factors essential for this process remains a challenge. Here we describe a procedure to generate targeted overexpression or shRNA-mediated knockdown of factors using lentivirus in a tissue-specific manner. Using a modified version of a hair regeneration model, we can achieve robust gain- or loss-of-function analysis in primary mouse keratinocytes or dermal cells to facilitate study of epithelial-mesenchymal signaling pathways that lead to hair follicle morphogenesis. We describe how to isolate fresh primary mouse keratinocytes and dermal cells, which contain dermal papilla cells and their precursors, deliver lentivirus containing either shRNA or cDNA to one of the cell populations, and combine the cells to generate fully formed hair follicles on the backs of nude mice. This approach allows analysis of tissue-specific factors required to generate hair follicles within three weeks and provides a fast and convenient companion to existing genetic models.

  View details for DOI 10.3791/4344

  View details for PubMedID 23486463

  View details for PubMedCentralID PMC3622109

• Initial Assessment of Tumor Regrowth After Vismodegib in Advanced Basal Cell Carcinoma ARCHIVES OF DERMATOLOGY Chang, A. S., Oro, A. E. 2012; 148 (11): 1324–25

  View details for DOI 10.1001/archdermatol.2012.2354

  View details for Web of Science ID 000311365700023

  View details for PubMedID 22910979

  View details for PubMedCentralID PMC3777384

• Hedgehog pathway inhibition and the race against tumor evolution JOURNAL OF CELL BIOLOGY Atwood, S. X., Chang, A. L., Oro, A. E. 2012; 199 (2): 193-197

  Abstract

  Dependence of basal cell carcinomas and medulloblastomas on the Hedgehog pathway provides an opportunity for targeted or "personalized" therapy. The recent effectiveness and FDA approval of the first Smoothened inhibitors validates this class of agents, but has revealed drug-resistant tumor variants that bypass Smoothened inhibition. Here, we summarize the effectiveness of Hedgehog pathway inhibitors and highlight promising areas for the development of next generation drug antagonists for Hedgehog-dependent cancers.

  View details for DOI 10.1083/jcb.201207140

  View details for Web of Science ID 000309982400002

  View details for PubMedID 23071148

  View details for PubMedCentralID PMC3471227

• A perivascular stem cell niche in the hair follicle 75th Annual Meeting of the Society-for-Investigative-Dermatology Xiao, Y., Woo, W., Terunuma, A., Oro, A. E., Vogel, J. C., Brownell, I. NATURE PUBLISHING GROUP. 2012: S79–S79

  View details for Web of Science ID 000302866900466

• A pivotal multicenter trial evaluating efficacy and safety of the hedgehog pathway inhibitor vismodegib (GDC-0449) in patients with advanced basal cell carcinoma Sekulic, A., Oro, A. E., Solomon, J. A., Hainsworth, J. D., Lewis, K., Dirix, L., Migden, M. R. MOSBY-ELSEVIER. 2012: AB149

  View details for Web of Science ID 000302319800593

• Neuropilins are positive regulators of Hedgehog signal transduction GENES & DEVELOPMENT Hillman, R. T., Feng, B. Y., Ni, J., Woo, W., Milenkovic, L., Gephart, M. G., Teruel, M. N., Oro, A. E., Chen, J. K., Scott, M. P. 2011; 25 (22): 2333-2346

  Abstract

  The Hedgehog (Hh) pathway is essential for vertebrate embryogenesis, and excessive Hh target gene activation can cause cancer in humans. Here we show that Neuropilin 1 (Nrp1) and Nrp2, transmembrane proteins with roles in axon guidance and vascular endothelial growth factor (VEGF) signaling, are important positive regulators of Hh signal transduction. Nrps are expressed at times and locations of active Hh signal transduction during mouse development. Using cell lines lacking key Hh pathway components, we show that Nrps mediate Hh transduction between activated Smoothened (Smo) protein and the negative regulator Suppressor of Fused (SuFu). Nrp1 transcription is induced by Hh signaling, and Nrp1 overexpression increases maximal Hh target gene activation, indicating the existence of a positive feedback circuit. The regulation of Hh signal transduction by Nrps is conserved between mammals and bony fish, as we show that morpholinos targeting the Nrp zebrafish ortholog nrp1a produce a specific and highly penetrant Hh pathway loss-of-function phenotype. These findings enhance our knowledge of Hh pathway regulation and provide evidence for a conserved nexus between Nrps and this important developmental signaling system.

  View details for DOI 10.1101/gad.173054.111

  View details for PubMedID 22051878

• In Vivo Imaging of Human and Mouse Skin with a Handheld Dual-Axis Confocal Fluorescence Microscope JOURNAL OF INVESTIGATIVE DERMATOLOGY Ra, H., Piyawattanametha, W., Gonzalez-Gonzalez, E., Mandella, M. J., Kino, G. S., Solgaard, O., Leake, D., Kaspar, R. L., Oro, A., Contag, C. H. 2011; 131 (5): 1061-1066

  Abstract

  Advancing molecular therapies for the treatment of skin diseases will require the development of new tools that can reveal spatiotemporal changes in the microanatomy of the skin and associate these changes with the presence of the therapeutic agent. For this purpose, we evaluated a handheld dual-axis confocal (DAC) microscope that is capable of in vivo fluorescence imaging of skin, using both mouse models and human skin. Individual keratinocytes in the epidermis were observed in three-dimensional image stacks after topical administration of near-infrared (NIR) dyes as contrast agents. This suggested that the DAC microscope may have utility in assessing the clinical effects of a small interfering RNA (siRNA)-based therapeutic (TD101) that targets the causative mutation in pachyonychia congenita (PC) patients. The data indicated that (1) formulated indocyanine green (ICG) readily penetrated hyperkeratotic PC skin and normal callused regions compared with nonaffected areas, and (2) TD101-treated PC skin revealed changes in tissue morphology, consistent with reversion to nonaffected skin compared with vehicle-treated skin. In addition, siRNA was conjugated to NIR dye and shown to penetrate through the stratum corneum barrier when topically applied to mouse skin. These results suggest that in vivo confocal microscopy may provide an informative clinical end point to evaluate the efficacy of experimental molecular therapeutics.

  View details for DOI 10.1038/jid.2010.401

  View details for Web of Science ID 000289789900014

  View details for PubMedID 21191407

• Cell Migration: GSK3 beta Steers the Cytoskeleton's Tip CELL Yucel, G., Oro, A. E. 2011; 144 (3): 319-321

  Abstract

  Directed cell migration polarizes the cytoskeleton, allowing the cell to move toward migratory cues. In this issue, Wu et al. (2011) demonstrate that the glycogen synthase kinase 3β (GSK3β) controls microtubule architecture and polarized movement of skin stem cells during wound healing in mammals by regulating the microtubule crosslinking protein ACF7.

  View details for DOI 10.1016/j.cell.2011.01.023

  View details for Web of Science ID 000286973100002

  View details for PubMedID 21295692

• Laminin-511 and integrin beta-1 in hair follicle development and basal cell carcinoma formation BMC DEVELOPMENTAL BIOLOGY DeRouen, M. C., Zhen, H., Tan, S. H., Williams, S., Marinkovich, M. P., Oro, A. E. 2010; 10

  Abstract

  Initiation of the hair follicle placode and its subsequent growth, maturation and cycling in post-natal skin requires signaling interactions between epithelial cells and adjacent dermal cells and involves Shh signaling via the primary cilium. Previous reports have implicated laminins in hair follicle epithelial invagination.Here we use a human BCC model system and mouse mutants to re-evaluate the role of laminin-511 in epithelial invagination in the skin. Blocking laminin 511 and 332 in BCCs maintains primary cilia and Shh signalling, but prevents invagination. Similarly, in laminin-511 and dermal beta-1 integrin mutants, dermal papilla development and primary cilia formation are normal. Dermal beta-1 integrin mutants have normal hair follicle development.Our data provides support for a primary role of laminin-511 promoting hair follicle epithelial downgrowth without affecting dermal primary cilia and Shh target gene induction.

  View details for DOI 10.1186/1471-213X-10-112

  View details for Web of Science ID 000284878300001

  View details for PubMedID 21067603

  View details for PubMedCentralID PMC2995472

• BAR domain competition during directional cellular migration CELL CYCLE Quinones, G. A., Oro, A. E. 2010; 9 (13): 2522-2528

  Abstract

  While directed cellular migration facilitates the coordinated movement of cells during development and tissue repair, the precise mechanisms regulating the interplay between the extracellular environment, the actin cytoskeleton, and the overlying plasma membrane remain inadequately understood. The BAR domain family of lipid binding, actin cytoskeletal regulators are gaining greater appreciation for their role in these critical processes. BAR domain proteins are involved as both positive and negative regulators of endocytosis, membrane plasticity, and directional cell migration. This review focuses on the functional relationship between different classes of BAR domain proteins and their role in guiding cell migration through regulation of the endocytic machinery. Competition for key signaling substrates by positive and negative BAR domain endocytic regulators appears to mediate control of directional cell migration, and may have wider applicability to other trafficking functions associated with development and carcinogenesis.

  View details for DOI 10.4161/cc.9.13.12123

  View details for Web of Science ID 000281205400024

  View details for PubMedID 20581461

  View details for PubMedCentralID PMC2990792

• The Primary Cilium: A Small Yet Mighty Organelle JOURNAL OF INVESTIGATIVE DERMATOLOGY DeRouen, M. C., Oro, A. E. 2009; 129 (2): 264-265

  Abstract

  Primary cilia are small, cylindrical membrane protrusions on most vertebrate cell types. Despite an unassuming appearance, this organelle has proven to be a veritable nexus of tightly regulated cell-signal reception between the epithelium and mesenchyme. In this issue, Lehman et al. describe the necessity of Ift88 and intraflagellar transport for signal reception of the sonic hedgehog pathway in the dermal papilla of developing hair follicles.

  View details for DOI 10.1038/jid.2008.404

  View details for Web of Science ID 000262655600003

  View details for PubMedID 19148215

• Laminin-511 is an epithelial message promoting dermal papilla development and function during early hair morphogenesis GENES & DEVELOPMENT Gao, J., DeRouen, M. C., Chen, C., Nguyen, M., Nguyen, N. T., Ido, H., Harada, K., Sekiguchi, K., Morgan, B. A., Miner, J. H., Oro, A. E., Marinkovich, M. P. 2008; 22 (15): 2111-2124

  Abstract

  Hair morphogenesis takes place through reciprocal epithelial and mesenchymal signaling; however, the mechanisms controlling signal exchange are poorly understood. Laminins are extracellular proteins that play critical roles in adhesion and signaling. Here we demonstrate the mechanism of how laminin-511 controls hair morphogenesis. Dermal papilla (DP) from laminin-511 mutants showed developmental defects by E16.5, including a failure to maintain expression of the key morphogen noggin. This maintenance was critical as exogenous introduction of noggin or sonic hedgehog (Shh) produced downstream from noggin was sufficient to restore hair follicle development in lama5(-/-) (laminin-511-null) skin. Hair development required the beta1 integrin binding but not the heparin binding domain of laminin-511. Previous studies demonstrated that Shh signaling requires primary cilia, microtubule-based signaling organelles. Laminin-511 mutant DP showed decreased length and structure of primary cilia in vitro and in vivo. Laminin-511, but not laminin-111, restored primary cilia formation in lama5(-/-) mesenchyme and triggered noggin expression in an Shh- and PDGF-dependent manner. Inhibition of laminin-511 receptor beta1 integrin disrupted DP primary cilia formation as well as hair development. These studies show that epithelial-derived laminin-511 is a critical early signal that directs ciliary function and DP maintenance as a requirement for hair follicle downgrowth.

  View details for DOI 10.1101/gad.1689908

  View details for PubMedID 18676816

• Controlling hair follicle signaling pathways through polyubiquitination JOURNAL OF INVESTIGATIVE DERMATOLOGY Huntzicker, E. G., Oro, A. E. 2008; 128 (5): 1081-1087

  Abstract

  Hair follicle development and maintenance require precise reciprocal signaling interactions between the epithelium and underlying dermis. Three major developmental signaling pathways, Wnt, Sonic hedgehog, and NF-kappaB/Edar, are indispensable for this process and, when aberrantly activated, can lead to skin and appendage neoplasms. Recent data point to protein polyubiquitination as playing a central role in regulating the timing, duration, and location of signaling. Here we review how polyubiquitination regulates the stability and interaction of key signaling components that control hair follicle development and regeneration.

  View details for DOI 10.1038/sj.jid.5700957

  View details for Web of Science ID 000255250400006

  View details for PubMedID 18408747

  View details for PubMedCentralID PMC2724002

• Role of the Basement Membrane Zone in Skin Development. journal of investigative dermatology Oro, A. E. 2008; 128: E11-2

  View details for DOI 10.1038/skinbio.6250016

  View details for PubMedID 26794064

• Dermatology in the postgenomic era - Harnessing human variation for personalized medicine ARCHIVES OF DERMATOLOGY Oro, A. E. 2008; 144 (3): 389-391

  View details for DOI 10.1001/archderm.144.3.389

  View details for Web of Science ID 000254045100014

  View details for PubMedID 18347297

  View details for PubMedCentralID PMC3777396

• A new role for an old friend: NFAT and stem cell quiescence CELL STEM CELL Oro, A. E. 2008; 2 (2): 104-106

  Abstract

  NFAT proteins are calcium-regulated transcription factors that play a critical role during the timing and activation of many vertebrate tissues. A recent paper in Cell (Horsley et al., 2008) demonstrates a role of the calcineurin-NFAT-CDK4 pathway in maintaining hair follicle stem cell quiescence.

  View details for DOI 10.1016/j.stem.2008.01.008

  View details for Web of Science ID 000253301900002

  View details for PubMedID 18371427

• Role of the basement membrane zone in skin development. journal of investigative dermatology Oro, A. E. 2008; 128 (E2): E11-2

  View details for DOI 10.1038/skinbio.6250016

  View details for PubMedID 21233816

• The primary cilia, a 'Rab-id' transit system for hedgehog signaling CURRENT OPINION IN CELL BIOLOGY Oro, A. E. 2007; 19 (6): 691-696

  Abstract

  Intense focus has been centered around how the primary cilia transduces the hedgehog (Hh) signal from smoothened (Smo) to the Gli transcription factors. New data indicate that ligand and signaling lipids help regulate small GTPase-dependent accumulation and activity of signaling components.

  View details for DOI 10.1016/j.ceb.2007.10.008

  View details for Web of Science ID 000252143000014

  View details for PubMedID 18061425

• Bone morphogenetic protein antagonist gremlin 1 is widely expressed by cancer-associated stromal cells and can promote tumor cell proliferation PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA Sneddon, J. B., Zhen, H. H., Montgomery, K., van de Rijn, M., Tward, A. D., West, R., Gladstone, H., Chang, H. Y., Morganroth, G. S., Oro, A. E., Brown, P. O. 2006; 103 (40): 14842-14847

  Abstract

  Although tissue microenvironments play critical roles in epithelial development and tumorigenesis, the factors mediating these effects are poorly understood. In this work, we used a genomic approach to identify factors produced by cells in the microenvironment of basal cell carcinoma (BCC) of the skin, one of the most common human cancers. The global gene expression programs of stromal cell cultures derived from human BCCs showed consistent, systematic differences from those derived from nontumor skin. The gene most consistently expressed at a higher level in BCC tumor stromal cells compared with those from nontumor skin was GREMLIN 1, which encodes a secreted antagonist of the bone morphogenetic protein (BMP) pathway. BMPs and their antagonists are known to play a crucial role in stem and progenitor cell biology as regulators of the balance between expansion and differentiation. Consistent with the hypothesis that BMP antagonists might have a similar role in cancer, we found GREMLIN 1 expression in the stroma of human BCC tumors but not in normal skin in vivo. Furthermore, BMP 2 and 4 are expressed by BCC cells. Ex vivo, BMP inhibits, and Gremlin 1 promotes, proliferation of cultured BCC cells. We further found that GREMLIN 1 is expressed by stromal cells in many carcinomas but not in the corresponding normal tissue counterparts that we examined. Our data suggest that BMP antagonists may be important constituents of tumor stroma, providing a favorable microenvironment for cancer cell survival and expansion in many cancers.

  View details for DOI 10.1073/pnas.0606857103

  View details for PubMedID 17003113

• Dual degradation signals control Gli protein stability and tumor formation GENES & DEVELOPMENT Huntzicker, E. G., Estay, I. S., Zhen, H., Lokteva, L. A., Jackson, P. K., Oro, A. E. 2006; 20 (3): 276-281

  Abstract

  Regulated protein destruction controls many key cellular processes with aberrant regulation increasingly found during carcinogenesis. Gli proteins mediate the transcriptional effects of the Sonic hedgehog pathway, which is implicated in up to 25% of human tumors. Here we show that Gli is rapidly destroyed by the proteasome and that mouse basal cell carcinoma induction correlates with Gli protein accumulation. We identify two independent destruction signals in Gli1, D(N) and D(C), and show that removal of these signals stabilizes Gli1 protein and rapidly accelerates tumor formation in transgenic animals. These data argue that control of Gli protein accumulation underlies tumorigenesis and suggest a new avenue for antitumor therapy.

  View details for DOI 10.1101/gad.1380906

  View details for Web of Science ID 000235267000002

  View details for PubMedID 16421275

  View details for PubMedCentralID PMC1361699

• Mammalian variations on a theme: a Smo and Sufu surprise DEVELOPMENTAL CELL Oro, A. E. 2006; 10 (2): 156-158

  Abstract

  Hedgehog signaling plays a critical role during development and tumorigenesis. While much mechanistic insight has come from pathway investigations in the fruit fly, recent studies suggest a distinct mammalian strategy for signaling from Smoothened to Gli through the novel protein Suppressor of Fused that may have therapeutic implications.

  View details for DOI 10.1016/j.devcel.2006.01.006

  View details for Web of Science ID 000235480500004

  View details for PubMedID 16459294

• Conditional telomerase induction causes proliferation of hair follicle stem cells NATURE Sarin, K. Y., Cheung, P., Gilison, D., Lee, E., Tennen, R. I., Wang, E., Artandi, M. K., Oro, A. E., Artandi, S. E. 2005; 436 (7053): 1048-1052

  Abstract

  TERT, the protein component of telomerase, serves to maintain telomere function through the de novo addition of telomere repeats to chromosome ends, and is reactivated in 90% of human cancers. In normal tissues, TERT is expressed in stem cells and in progenitor cells, but its role in these compartments is not fully understood. Here we show that conditional transgenic induction of TERT in mouse skin epithelium causes a rapid transition from telogen (the resting phase of the hair follicle cycle) to anagen (the active phase), thereby facilitating robust hair growth. TERT overexpression promotes this developmental transition by causing proliferation of quiescent, multipotent stem cells in the hair follicle bulge region. This new function for TERT does not require the telomerase RNA component, which encodes the template for telomere addition, and therefore operates through a mechanism independent of its activity in synthesizing telomere repeats. These data indicate that, in addition to its established role in extending telomeres, TERT can promote proliferation of resting stem cells through a non-canonical pathway.

  View details for DOI 10.1038/nature03836

  View details for Web of Science ID 000231263900057

  View details for PubMedID 16107853

  View details for PubMedCentralID PMC1361120

• Receptor tyrosine phosphatase-dependent cytoskeletal remodeling by the hedgehog-responsive gene MIM/BEG4 66th Annual Meeting of the Society-for-Investigative-Dermatology Gonzalez-Quevedo, R., Shoffer, M., Horng, L., Oro, A. E. NATURE PUBLISHING GROUP. 2005: A103–A103

  View details for Web of Science ID 000228179901160

• Receptor tyrosine phosphatase-dependent cytoskeletal remodeling by the hedgehog-responsive gene MIM/BEG4 JOURNAL OF CELL BIOLOGY Gonzalez-Quevedo, R., Shoffer, M., Horng, L., Oro, A. E. 2005; 168 (3): 453-463

  Abstract

  During development, dynamic remodeling of the actin cytoskeleton allows the precise placement and morphology of tissues. Morphogens such as Sonic hedgehog (Shh) and local cues such as receptor protein tyrosine phosphatases (RPTPs) mediate this process, but how they regulate the cytoskeleton is poorly understood. We previously identified Basal cell carcinoma-enriched gene 4 (BEG4)/Missing in Metastasis (MIM), a Shh-inducible, Wiskott-Aldrich homology 2 domain-containing protein that potentiates Gli transcription (Callahan, C.A., T. Ofstad, L. Horng, J.K. Wang, H.H. Zhen, P.A. Coulombe, and A.E. Oro. 2004. Genes Dev. 18:2724-2729). Here, we show that endogenous MIM is induced in a patched1-dependent manner and regulates the actin cytoskeleton. MIM functions by bundling F-actin, a process that requires self-association but is independent of G-actin binding. Cytoskeletal remodeling requires an activation domain distinct from sequences required for bundling in vitro. This domain associates with RPTPdelta and, in turn, enhances RPTPdelta membrane localization. MIM-dependent cytoskeletal changes can be inhibited using a soluble RPTPdelta-D2 domain. Our data suggest that the hedgehog-responsive gene MIM cooperates with RPTP to induce cytoskeletal changes.

  View details for DOI 10.1083/jcb.200409078

  View details for Web of Science ID 000226925500011

  View details for PubMedID 15684034

  View details for PubMedCentralID PMC2171717

• MIM/BEG4, a Sonic hedgehog-responsive gene that potentiates Gli-dependent transcription GENES & DEVELOPMENT Callahan, C. A., Ofstad, T., Horng, L., Wang, J. K., Zhen, H. H., Coulombe, P. A., Oro, A. E. 2004; 18 (22): 2724-2729

  Abstract

  Sonic hedgehog (Shh) signaling plays a critical role during development and carcinogenesis. While Gli family members govern the transcriptional output of Shh signaling, little is known how Gli-mediated transcriptional activity is regulated. Here we identify the actin-binding protein Missing in Metastasis (MIM) as a new Shh-responsive gene. Together, Gli1 and MIM recapitulate Shh-mediated epidermal proliferation and invasion in regenerated human skin. MIM is part of a Gli/Suppressor of Fused complex and potentiates Gli-dependent transcription using domains distinct from those used for monomeric actin binding. These data define MIM as both a Shh-responsive gene and a new member of the pathway that modulates Gli responses during growth and tumorigenesis.

  View details for DOI 10.1101/gad.1221804

  View details for Web of Science ID 000225170900004

  View details for PubMedID 15545630

  View details for PubMedCentralID PMC528890

• Laminin-10 is crucial for hair morphogenesis EMBO JOURNAL Li, J., Tzu, J., Chen, Y., Zhang, Y. P., Nguyen, N. T., Gao, J., Bradley, M., Keene, D. R., Oro, A. E., Miner, J. H., Marinkovich, M. P. 2003; 22 (10): 2400-2410

  Abstract

  The role of the extracellular matrix in cutaneous morphogenesis is poorly understood. Here, we describe the essential role of laminin-10 (alpha5beta1gamma1) in hair follicle development. Laminin-10 was present in the basement membrane of elongating hair germs, when other laminins were downregulated, suggesting a role for laminin-10 in hair development. Treatment of human scalp xenografts with antibodies to laminin-10, or its receptor beta1 integrin, produced alopecia. E16.5 Lama5 -/- mouse skin, lacking laminin-10, contained fewer hair germs compared with controls, and after transplantation, Lama5 -/- skin showed a failure of hair germ elongation followed by complete hair follicle regression. Lama5 -/- skin showed defective basement membrane assembly, without measurable increases in anoikis. Instead, Lama5 -/- skin showed decreased expression of early hair markers including sonic hedgehog and Gli1, implicating laminin-10 in developmental signaling. Intriguingly, treatment of Lama5 -/- skin with purified laminin-10 corrected basement membrane defects and restored hair follicle development. We conclude that laminin-10 is required for hair follicle development and report the first use of exogenous protein to correct a cutaneous developmental defect.

  View details for PubMedID 12743034

• Hair cycle regulation of Hedgehog signal reception DEVELOPMENTAL BIOLOGY Oro, A. E., Higgins, K. 2003; 255 (2): 238-248

  Abstract

  Proper patterning of self-renewing organs, like the hair follicle, requires exquisite regulation of growth signals. Sonic hedgehog (Shh) signaling in skin controls the growth and morphogenesis of hair follicle epithelium in part through regulating the Gli transcription factors. While ectopic induction of Shh target genes leads to hair follicle tumors, such as basal cell carcinomas, how Shh signaling normally functions during the cyclic process of hair development is unknown. Here, we show that, during the hair cycle, Shh expression and the ability of skin cells to respond to Shh signaling is spatially and temporally regulated. Induction of Shh target genes normally occurs only in the anagen hair follicle in response to expression of Shh. However, in patched1 heterozygous mice, putative tumor precursors form with concomitant induction of Shh target gene transcription only during anagen in follicular and interfollicular keratinocytes. Ectopic production of Gli1 accumulates Gli protein and induces Shh target genes and epithelial tumors at anagen but not other stages, pointing to a restricted competence occurring at the level of Gli protein accumulation. Delivery and reception of growth signals among multipotent cells are restricted in time and space to facilitate cyclic pattern formation.

  View details for DOI 10.1016/S0012-1606(02)00042-8

  View details for Web of Science ID 000181998400005

  View details for PubMedID 12648487

• Ultraviolet and ionizing radiation enhance the growth of BCCs and trichoblastomas in patched heterozygous knockout mice NATURE MEDICINE Aszterbaum, M., Epstein, J., Oro, A., Douglas, V., LeBoit, P. E., Scott, M. P., Epstein, E. H. 1999; 5 (11): 1285-1291

  Abstract

  Basal cell carcinomas, the commonest human skin cancers, consistently have abnormalities of the hedgehog signaling pathway and often have PTCH gene mutations. We report here that Ptch+/- mice develop primordial follicular neoplasms resembling human trichoblastomas, and that exposure to ultraviolet radiation or ionizing radiation results in an increase in the number and size of these tumors and a shift in their histologic features so that they more closely resemble human basal cell carcinoma. The mouse basal cell carcinomas and trichoblastoma-like tumors resemble human basal cell carcinomas in their loss of normal hemidesmosomal components, presence of p53 mutations, frequent loss of the normal remaining Ptch allele, and activation of hedgehog target gene transcription. The Ptch mutant mice provide the first mouse model, to our knowledge, of ultraviolet and ionizing radiation-induced basal cell carcinoma-like tumors, and also demonstrate that Ptch inactivation and hedgehog target gene activation are essential for basal cell carcinoma tumorigenesis.

  View details for Web of Science ID 000086550600039

  View details for PubMedID 10545995

• Regulation of growth by the patched tumor suppressor gene. Wechsler-Reya, R., Milenkovic, L., Goodrich, L. V., Oro, A. E., Higgins, K. M., Scott, M. P. ACADEMIC PRESS INC. 1998: 166

  View details for Web of Science ID 000074281900049

• Hedgehog/patched signaling in animal development and disease Scott, M. P., Goodrich, L. V., Higgins, K. M., Johnson, R. L., Milenkovic, L., Oro, A. E. FEDERATION AMER SOC EXP BIOL. 1997: A1302–A1302

  View details for Web of Science ID 000073305603086

• Induction of features of basal cell epithelioma in transgenic human skin expressing sonic Hedgehog in vivo. Fan, H., Oro, A. E., Scott, M. P., Khavari, P. A. NATURE PUBLISHING GROUP. 1997: 38–38

  View details for Web of Science ID A1997WP04000063

• MOLECULAR ANALYSIS OF THE INHERITANCE AND STABILITY OF THE MITOCHONDRIAL GENOME OF AN INBRED LINE OF MAIZE THEORETICAL AND APPLIED GENETICS Oro, A. E., Newton, K. J., Walbot, V. 1985; 70 (3): 287-293

  View details for Web of Science ID A1985AKY2100012