All Publications


  • Sialylated glycoproteins suppress immune cell killing by binding to Siglec-7 and Siglec-9 in prostate cancer. The Journal of clinical investigation Wen, R. M., Stark, J. C., Marti, G. E., Fan, Z., Lyu, A., Garcia Marques, F. J., Zhang, X., Riley, N. M., Totten, S. M., Bermudez, A., Nolley, R., Zhao, H., Fong, L., Engleman, E. G., Pitteri, S. J., Bertozzi, C. R., Brooks, J. D. 2024

    Abstract

    Prostate cancer is the second leading cause of male cancer death in the U.S. Current immune checkpoint inhibitor-based immunotherapies have improved survival for many malignancies; however, they have failed to prolong survival for prostate cancer. Siglecs (sialic acid-binding immunoglobulin-like lectins) are expressed on immune cells and regulate immune responses and function. Siglec-7 and Siglec-9 contribute to immune evasion by interacting with their ligands. However, the role of Siglec-7/9 receptors and their ligands in prostate cancer remains poorly understood. Here, we find that Siglec-7 and Siglec-9 are associated with poor prognosis in prostate cancer patients, and are highly expressed in myeloid cells, including macrophages, in prostate tumor tissues. Siglecs-7 and -9 ligands were expressed in prostate cancer cells and human prostate tumor tissues. Blocking the interactions between Siglec-7/9 and sialic acids inhibited prostate cancer xenograft growth and increased immune cell infiltration in humanized mice in vivo. Using a CRISPRi screen and mass spectrometry, we identified CD59 as a candidate Siglec-9 ligand in prostate cancer. The identification of Siglecs-7 and -9 as potential therapeutic targets, including CD59/Siglec-9 axis, opens up opportunities for immune-based interventions in prostate cancer.

    View details for DOI 10.1172/JCI180282

    View details for PubMedID 39436703

  • Ubiquitin C-terminal hydrolase L1 is a regulator of tumor growth and metastasis in double-negative prostate cancer. American journal of clinical and experimental urology Liu, S., Garcia-Marques, F. J., Shen, M., Bermudez, A., Pitteri, S. J., Stoyanova, T. 2024; 12 (5): 306-322

    Abstract

    Prostate cancer is the second leading cause of cancer-related deaths among men worldwide. With heavy androgen deprivation therapies, prostate cancer may shift to androgen receptor negative and neuroendocrine negative subtype of castration resistant prostate cancer, defined as double-negative prostate cancer. Double-negative prostate cancer is associated with poor prognosis and disease mortality. The molecular mechanisms underlying the emergence of double-negative prostate cancer remain poorly understood. Here, we demonstrate that Ubiquitin C-Terminal Hydrolase L1 (UCH-L1), is negatively correlated with androgen receptor levels in prostate cancer patients. UCH-L1 plays a functional role in tumorigenesis and metastasis in double-negative prostate cancer. Knock-down of UCH-L1 decreases double-negative prostate cancer colony formation in vitro and tumor growth in vivo. Moreover, decrease of UCH-L1 significantly delays cell migration in vitro and spontaneous metastasis and metastatic colonization in vivo. Proteomic analysis revealed that mTORC1 signaling, androgen response signaling and MYC targets are the top three decreased pathways upon UCH-L1 decrease. Further, treatment with LDN-57444, a UCH-L1 small molecule inhibitor, impairs double-negative prostate cancer cell colony formation, migration in vitro, and metastatic colonization in vivo. Our study reveals that UCH-L1 is an important regulator of double-negative prostate cancer tumor growth and progression, providing a promising therapeutic target for this subtype of metastatic prostate cancer.

    View details for DOI 10.62347/JNBR1463

    View details for PubMedID 39584005

    View details for PubMedCentralID PMC11578776

  • Identification of a 5-gene signature panel for the prediction of prostate cancer progression. British journal of cancer Shen, M., García-Marqués, F., Muruganantham, A., Liu, S., White, J. R., Bermudez, A., Rice, M. A., Thompson, K., Chen, C. L., Hung, C. N., Zhang, Z., Huang, T. H., Liss, M. A., Pienta, K. J., Pitteri, S. J., Stoyanova, T. 2024

    Abstract

    Despite nearly 100% 5-year survival for localised prostate cancer, the survival rate for metastatic prostate cancer significantly declines to 32%. Thus, it is crucial to identify molecular indicators that reflect the progression from localised disease to metastatic prostate cancer.To search for molecular indicators associated with prostate cancer metastasis, we performed proteomic analysis of rapid autopsy tissue samples from metastatic prostate cancer (N = 8) and localised prostate cancer (N = 2). Then, we utilised multiple independent, publicly available prostate cancer patient datasets to select candidates that also correlate with worse prostate cancer clinical prognosis.We identified 154 proteins with increased expressions in metastases relative to localised prostate cancer through proteomic analysis. From the subset of these candidates that correlate with prostate cancer recurrence (N = 28) and shorter disease-free survival (N = 37), we identified a 5-gene signature panel with improved performance in predicting worse clinical prognosis relative to individual candidates.Our study presents a new 5-gene signature panel that is associated with worse clinical prognosis and is elevated in prostate cancer metastasis on both protein and mRNA levels. Our 5-gene signature panel represents a potential modality for the prediction of prostate cancer progression towards the onset of metastasis.

    View details for DOI 10.1038/s41416-024-02854-w

    View details for PubMedID 39402324

    View details for PubMedCentralID 9275764

  • A Multiplexed Quantitative Proteomics Approach to the Human Plasma Protein Signature. Biomedicines Nunez, E., Gomez-Serrano, M., Calvo, E., Bonzon-Kulichenko, E., Trevisan-Herraz, M., Rodriguez, J. M., Garcia-Marques, F., Magni, R., Lara-Pezzi, E., Martin-Ventura, J. L., Camafeita, E., Vazquez, J. 2024; 12 (9)

    Abstract

    Despite the plasma proteome being able to provide a unique insight into the health and disease status of individuals, holding singular promise as a source of protein biomarkers that could be pivotal in the context of personalized medicine, only around 100 proteins covering a few human conditions have been approved as biomarkers by the US Food and Drug Administration (FDA) so far. Mass spectrometry (MS) currently has enormous potential for high-throughput analysis in clinical research; however, plasma proteomics remains challenging mainly due to the wide dynamic range of plasma protein abundances and the time-consuming procedures required. We applied a new MS-based multiplexed proteomics workflow to quantitate proteins, encompassing 67 FDA-approved biomarkers, in >1300 human plasma samples from a clinical cohort. Our results indicate that this workflow is suitable for large-scale clinical studies, showing good accuracy and reproducibility (coefficient of variation (CV) < 20 for 90% of the proteins). Furthermore, we identified plasma signature proteins (stable in time on an individual basis), stable proteins (exhibiting low biological variability and high temporal stability), and highly variable proteins (with low temporal stability) that can be used for personalized health monitoring and medicine.

    View details for DOI 10.3390/biomedicines12092118

    View details for PubMedID 39335631

  • Establishing and characterizing the molecular profiles, cellular features, and clinical utility of a patient-derived xenograft model using benign prostatic tissues. Laboratory investigation; a journal of technical methods and pathology Polasko, A. L., Zhang, D., Ramraj, A., Chiu, C. L., Garcia-Marques, F. J., Bermudez, A., Kapp, K., Peterson, E., Qiu, Z., Pollack, A. S., Zhao, H., Pollack, J. R., Pitteri, S. J., Brooks, J. D. 2024: 102129

    Abstract

    Benign Prostatic Hyperplasia (BPH) is a common condition marked by the enlargement of the prostate gland, which often leads to significant urinary symptoms and a decreased quality of life. The development of clinically relevant animal models is crucial for understanding the pathophysiology of BPH and improving treatment options. This study aims to establish a patient-derived xenograft (PDX) model using benign prostatic tissues to explore the molecular and cellular mechanisms of BPH. PDXs were generated by implanting fresh BPH (transition zone) and paired normal (peripheral zone) prostate tissue from eight patients under the renal capsule of immunodeficient male mice. Tissue weight, architecture, cellular proliferation, apoptosis, prostate-specific marker expression, and molecular profiles of PDXs were assessed after 1 week, 1 month, 2 months, or 3 months of implantation by immunohistochemistry, ELISA, transcriptomics, and proteomics. Responses to finasteride, a standard-of-care therapy, were evaluated. PDXs maintained the histological and molecular characteristics of the parental human tissues. BPH, but not normal PDXs, demonstrated significant increases in weight and cellular proliferation, particularly at 1 month. Molecular profiling revealed specific gene and protein expression patterns correlating with BPH pathophysiology. Specifically, an increased immune and stress response was observed at 1 week, followed by increased expression of proliferation markers and BPH-specific stromal signaling molecules, such as BMP5 and CXCL13, at 1 month. Graft stabilization to pre-implant characteristics was apparent between 2 and 3 months. Treatment with finasteride reduced proliferation, increased apoptosis, and induced morphological changes consistent with therapeutic responses observed in human BPH. Our PDX model recapitulates the morphological, histological, and molecular features of human BPH, offering a significant advancement in modeling the complex interactions of cell types in BPH microenvironments. These PDXs respond to therapeutic intervention as expected, providing a valuable tool for preclinical testing of new therapeutics which will improve the well-being for BPH patients.

    View details for DOI 10.1016/j.labinv.2024.102129

    View details for PubMedID 39222914

  • Inhibition of protein translational machinery in triple-negative breast cancer as a promising therapeutic strategy. Cell reports. Medicine Dheeraj, A., Garcia Marques, F. J., Tailor, D., Bermudez, A., Resendez, A., Pandrala, M., Grau, B., Kumar, P., Haley, C. B., Honkala, A., Kujur, P., Jeffrey, S. S., Pitteri, S., Malhotra, S. V. 2024: 101552

    Abstract

    Y-box binding protein-1 (YB-1) is a proto-oncogenic protein associated with protein translation regulation. It plays a crucial role in the development and progression of triple-negative breast cancer (TNBC). In this study, we describe a promising approach to inhibit YB-1 using SU056, a small-molecule inhibitor. SU056 physically interacts with YB-1 and reduces its expression, which helps to restrain the progression of TNBC. Proteome profiling analysis indicates that the inhibition of YB-1 by SU056 can alter the proteins that regulate protein translation, an essential process for cancer cell growth. Preclinical studies on human cells, mice, and patient-derived xenograft tumor models show the effectiveness of SU056. Moreover, toxicological studies have shown that SU056 treatment and dosing are well tolerated without any adverse effects. Overall, our study provides a strong foundation for the further development of SU056 as a potential treatment option for patients with TNBC by targeting YB-1.

    View details for DOI 10.1016/j.xcrm.2024.101552

    View details for PubMedID 38729158

  • Identification and characterization of intact glycopeptides in human urine. Scientific reports Garcia-Marques, F., Fuller, K., Bermudez, A., Shamsher, N., Zhao, H., Brooks, J. D., Flory, M. R., Pitteri, S. J. 2024; 14 (1): 3716

    Abstract

    Glycoproteins in urine have the potential to provide a rich class of informative molecules for studying human health and disease. Despite this promise, the urine glycoproteome has been largely uncharacterized. Here, we present the analysis of glycoproteins in human urine using LC-MS/MS-based intact glycopeptide analysis, providing both the identification of protein glycosites and characterization of the glycan composition at specific glycosites. Gene enrichment analysis reveals differences in biological processes, cellular components, and molecular functions in the urine glycoproteome versus the urine proteome, as well as differences based on the major glycan class observed on proteins. Meta-heterogeneity of glycosylation is examined on proteins to determine the variation in glycosylation across multiple sites of a given protein with specific examples of individual sites differing from the glycosylation trends in the overall protein. Taken together, this dataset represents a potentially valuable resource as a baseline characterization of glycoproteins in human urine for future urine glycoproteomics studies.

    View details for DOI 10.1038/s41598-024-53299-3

    View details for PubMedID 38355753

    View details for PubMedCentralID PMC10866872

  • UCHL1 is a potential molecular indicator and therapeutic target for neuroendocrine carcinomas. Cell reports. Medicine Liu, S., Chai, T., Garcia-Marques, F., Yin, Q., Hsu, E. C., Shen, M., Shaw Toland, A. M., Bermudez, A., Hartono, A. B., Massey, C. F., Lee, C. S., Zheng, L., Baron, M., Denning, C. J., Aslan, M., Nguyen, H. M., Nolley, R., Zoubeidi, A., Das, M., Kunder, C. A., Howitt, B. E., Soh, H. T., Weissman, I. L., Liss, M. A., Chin, A. I., Brooks, J. D., Corey, E., Pitteri, S. J., Huang, J., Stoyanova, T. 2024: 101381

    Abstract

    Neuroendocrine carcinomas, such as neuroendocrine prostate cancer and small-cell lung cancer, commonly have a poor prognosis and limited therapeutic options. We report that ubiquitin carboxy-terminal hydrolase L1 (UCHL1), a deubiquitinating enzyme, is elevated in tissues and plasma from patients with neuroendocrine carcinomas. Loss of UCHL1 decreases tumor growth and inhibits metastasis of these malignancies. UCHL1 maintains neuroendocrine differentiation and promotes cancer progression by regulating nucleoporin, POM121, and p53. UCHL1 binds, deubiquitinates, and stabilizes POM121 to regulate POM121-associated nuclear transport of E2F1 and c-MYC. Treatment with the UCHL1 inhibitor LDN-57444 slows tumor growth and metastasis across neuroendocrine carcinomas. The combination of UCHL1 inhibitors with cisplatin, the standard of care used for neuroendocrine carcinomas, significantly delays tumor growth in pre-clinical settings. Our study reveals mechanisms of UCHL1 function in regulating the progression of neuroendocrine carcinomas and identifies UCHL1 as a therapeutic target and potential molecular indicator for diagnosing and monitoring treatment responses in these malignancies.

    View details for DOI 10.1016/j.xcrm.2023.101381

    View details for PubMedID 38244540

  • Guanylate-binding protein 1 modulates proteasomal machinery in ovarian cancer. iScience Tailor, D., Garcia-Marques, F. J., Bermudez, A., Pitteri, S. J., Malhotra, S. V. 2023; 26 (11): 108292

    Abstract

    Guanylate-binding protein 1 (GBP1) is known as an interferon-γ-induced GTPase. Here, we used genetically modified ovarian cancer (OC) cells to study the role of GBP1. The data generated show that GBP1 inhibition constrains the clonogenic potential of cancer cells. In vivo studies revealed that GBP1 overexpression in tumors promotes tumor progression and reduces median survival, whereas GBP1 inhibition delayed tumor progression with longer median survival. We employed proteomics-based thermal stability assay (CETSA) on GBP1 knockdown and overexpressed OC cells to study its molecular functions. CETSA results show that GBP1 interacts with many members of the proteasome. Furthermore, GBP1 inhibition sensitizes OC cells to paclitaxel treatment via accumulated ubiquitinylated proteins where GBP1 inhibition decreases the overall proteasomal activity. In contrast, GBP1-overexpressing cells acquired paclitaxel resistance via boosted cellular proteasomal activity. Overall, these studies expand the role of GBP1 in the activation of proteasomal machinery to acquire chemoresistance.

    View details for DOI 10.1016/j.isci.2023.108292

    View details for PubMedID 38026225

    View details for PubMedCentralID PMC10665831

  • The role of GCNT1 mediated O-glycosylation in aggressive prostate cancer. Scientific reports Hodgson, K., Orozco-Moreno, M., Scott, E., Garnham, R., Livermore, K., Thomas, H., Zhou, Y., He, J., Bermudez, A., Garcia Marques, F. J., Bastian, K., Hysenaj, G., Archer Goode, E., Heer, R., Pitteri, S., Wang, N., Elliott, D. J., Munkley, J. 2023; 13 (1): 17031

    Abstract

    Prostate cancer is the most common cancer in men and a major cause of cancer related deaths worldwide. Nearly all affected men develop resistance to current therapies and there is an urgent need to develop new treatments for advanced disease. Aberrant glycosylation is a common feature of cancer cells implicated in all of the hallmarks of cancer. A major driver of aberrant glycosylation in cancer is the altered expression of glycosylation enzymes. Here, we show that GCNT1, an enzyme that plays an essential role in the formation of core 2 branched O-glycans and is crucial to the final definition of O-glycan structure, is upregulated in aggressive prostate cancer. Using in vitro and in vivo models, we show GCNT1 promotes the growth of prostate tumours and can modify the glycome of prostate cancer cells, including upregulation of core 2 O-glycans and modifying the O-glycosylation of secreted glycoproteins. Furthermore, using RNA sequencing, we find upregulation of GCNT1 in prostate cancer cells can alter oncogenic gene expression pathways important in tumour growth and metastasis. Our study highlights the important role of aberrant O-glycosylation in prostate cancer progression and provides novel insights regarding the mechanisms involved.

    View details for DOI 10.1038/s41598-023-43019-8

    View details for PubMedID 37813880

    View details for PubMedCentralID 8945091

  • Lineage plasticity in SCLC generates non-neuroendocrine cells primed for vasculogenic mimicry. Journal of thoracic oncology : official publication of the International Association for the Study of Lung Cancer Pearsall, S. M., Williamson, S. C., Humphrey, S., Hughes, E., Morgan, D., García Marqués, F. J., Awanis, G., Carroll, R., Burks, L., Shue, Y. T., Bermudez, A., Frese, K. K., Galvin, M., Carter, M., Priest, L., Kerr, A., Zhou, C., Oliver, T. G., Humphries, J. D., Humphries, M. J., Blackhall, F., Cannell, I. G., Pitteri, S. J., Hannon, G. J., Sage, J., Dive, C., Simpson, K. L. 2023

    Abstract

    Vasculogenic mimicry (VM), the process of tumor cell trans-differentiation to endow endothelial-like characteristics supporting de novo vessel formation, is associated with poor prognosis in several tumor types, including small cell lung cancer (SCLC). In genetically engineered mouse models (GEMMs) of SCLC, NOTCH and MYC co-operate to drive a neuroendocrine (NE) to non-NE phenotypic switch and co-operation between NE and non-NE cells is required for metastasis. Here, we define the phenotype of VM-competent cells and molecular mechanisms underpinning SCLC VM using circulating tumor cell-derived explant (CDX) models and GEMMs.We analysed perfusion within VM vessels and their association with NE and non-NE phenotypes using multiplex immunohistochemistry in CDX, GEMMs and patient biopsies. VM-proficient cell subpopulations in ex vivo cultures were molecularly profiled by RNA sequencing and mass spectrometry. We evaluated their 3D structure and defined collagen-integrin interactions.We show that VM vessels are present in 23/25 CDX models, 2 GEMMs and in 20 patient biopsies of SCLC. Perfused VM vessels support tumor growth and only Notch-active non-NE cells are VM-competent in vivo and ex vivo, expressing pseudohypoxia, blood vessel development and extracellular matrix (ECM) organization signatures. On Matrigel, VM-primed non-NE cells re-model ECM into hollow tubules in an integrin β1-dependent process.We identify VM as an exemplar of functional heterogeneity and plasticity in SCLC and these findings take significant steps towards understanding the molecular events that enable VM. These results support therapeutic co-targeting of both NE and non-NE cells to curtail SCLC progression and to improve SCLC patient outcomes in the future.

    View details for DOI 10.1016/j.jtho.2023.07.012

    View details for PubMedID 37455012

  • Proteomics analysis of urine and catheter-associated biofilms in spinal cord injury patients. American journal of clinical and experimental urology Garcia-Marques, F. J., Zakrasek, E., Bermudez, A., Polasko, A. L., Liu, S., Stoyanova, T., Brooks, J. D., Lavelle, J., Pitteri, S. J. 2023; 11 (3): 206-219

    Abstract

    After spinal cord injury (SCI), use chronic urinary catheters for bladder management is common, making these patients especially vulnerable to catheter-associated complications. Chronic catheterization is associated with bacterial colonization and frequent catheter-associated urinary tract infections (CAUTI). One determinant of infection success and treatment resistance is production of catheter-associated biofilms, composed of microorganisms and host- and microbial-derived components. To better understand the biofilm microenvironment, we performed proteomics analysis of catheter-associated biofilms and paired urine samples from four people with SCI with chronic indwelling urinary catheters. We developed a novel method for the removal of adhered cellular components on catheters that contained both human and microbial homologous proteins. Proteins from seven microbial species were identified including: Escherichia coli, Klebsiella species (spp), Enterococcus spp, Proteus mirabilis, Pseudomonas spp, Staphylococcus spp, and Candida spp. Peptides identified from catheter biofilms were assigned to 4,820 unique proteins, with 61% of proteins assigned to the biofilm-associated microorganisms, while the remainder were human-derived. Contrastingly, in urine, only 51% were assigned to biofilm-associated microorganisms and 4,554 proteins were identified as a human-derived. Of the proteins assigned to microorganisms in the biofilm and paired urine, Enterococcus, Candida spp, and P. mirabilis had greater associations with the biofilm phase, whereas E. coli and Klebsiella had greater associations with the urine phase, thus demonstrating a significant difference between the urine and adhered microbial communities. The microbial proteins that differed significantly between the biofilm and paired urine samples mapped to pathways associated with amino acid synthesis, likely related to adaptation to high urea concentrations in the urine, and growth and protein synthesis in bacteria in the biofilm. Human proteins demonstrated enrichment for immune response in the catheter-associated biofilm. Proteomic analysis of catheter-associated biofilms and paired urine samples has the potential to provide detailed information on host and bacterial responses to chronic indwelling urinary catheters and could be useful for understanding complications of chronic indwelling catheters including CAUTIs, urinary stones, and catheter blockages.

    View details for PubMedID 37441441

    View details for PubMedCentralID PMC10333135

  • Siglec-7/9 are novel immune checkpoints for prostate cancer Wen, R., Stark, J. C., Marti, G., Garcia-Marques, F., Zhao, H., Nolley, R., Bertozzi, C. R., Pitteri, S. J., Brooks, J. D. AMER ASSOC IMMUNOLOGISTS. 2023
  • Original Proteomics analysis of urine and catheter-associated biofilms in spinal cord injury patients AMERICAN JOURNAL OF CLINICAL AND EXPERIMENTAL UROLOGY Garcia-Marques, F. J., Zakrasek, E., Bermudez, A., Polasko, A. L., Liu, S., Stoyanova, T., Brooks, J., Lavelle, J., Pitteri, S. J. 2023; 11 (3): 206-219
  • Integrated transcriptome-proteome analyses of human stem cells reveal source-dependent differences in their regenerative signature. Stem cell reports Ganguly, A., Swaminathan, G., Garcia-Marques, F., Regmi, S., Yarani, R., Primavera, R., Chetty, S., Bermudez, A., Pitteri, S. J., Thakor, A. S. 2022

    Abstract

    Mesenchymal stem cells (MSCs) are gaining increasing prominence as an effective regenerative cellular therapy. However, ensuring consistent and reliable effects across clinical populations has proved to be challenging. In part, this can be attributed to heterogeneity in the intrinsic molecular and regenerative signature of MSCs, which is dependent on their source of origin. The present work uses integrated omics-based profiling, at different functional levels, to compare the anti-inflammatory, immunomodulatory, and angiogenic properties between MSCs from neonatal (umbilical cord MSC [UC-MSC]) and adult (adipose tissue MSC [AD-MSC], and bone marrow MSC [BM-MSC]) sources. Using multi-parametric analyses, we identified that UC-MSCs promote a more robust host innate immune response; in contrast, adult-MSCs appear to facilitate remodeling of the extracellular matrix (ECM) with stronger activation of angiogenic cascades. These data should help facilitate the standardization of source-specific MSCs, such that their regenerative signatures can be confidently used to target specific disease processes.

    View details for DOI 10.1016/j.stemcr.2022.11.006

    View details for PubMedID 36493779

  • Siglec-7/9-sialic acid interactions inhibit T cell immune response in prostate cancer Wen, R. M., Stark, J., Garcia-Marques, F., Nolley, H., Bertozzi, C. R., Pitteri, S. J., Brooks, J. D. AMER ASSOC CANCER RESEARCH. 2022
  • Interplay between the Chd4/NuRD Complex and the Transcription Factor Znf219 Controls Cardiac Cell Identity INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES El Abdellaoui-Soussi, F., Yunes-Leites, P. S., Lopez-Maderuelo, D., Garcia-Marques, F., Vazquez, J., Redondo, J., Gomez-del Arco, P. 2022; 23 (17)

    Abstract

    The sarcomere regulates striated muscle contraction. This structure is composed of several myofibril proteins, isoforms of which are encoded by genes specific to either the heart or skeletal muscle. The chromatin remodeler complex Chd4/NuRD regulates the transcriptional expression of these specific sarcomeric programs by repressing genes of the skeletal muscle sarcomere in the heart. Aberrant expression of skeletal muscle genes induced by the loss of Chd4 in the heart leads to sudden death due to defects in cardiomyocyte contraction that progress to arrhythmia and fibrosis. Identifying the transcription factors (TFs) that recruit Chd4/NuRD to repress skeletal muscle genes in the myocardium will provide important information for understanding numerous cardiac pathologies and, ultimately, pinpointing new therapeutic targets for arrhythmias and cardiomyopathies. Here, we sought to find Chd4 interactors and their function in cardiac homeostasis. We therefore describe a physical interaction between Chd4 and the TF Znf219 in cardiac tissue. Znf219 represses the skeletal-muscle sarcomeric program in cardiomyocytes in vitro and in vivo, similarly to Chd4. Aberrant expression of skeletal-muscle sarcomere proteins in mouse hearts with knocked down Znf219 translates into arrhythmias, accompanied by an increase in PR interval. These data strongly suggest that the physical and genetic interaction of Znf219 and Chd4 in the mammalian heart regulates cardiomyocyte identity and myocardial contraction.

    View details for DOI 10.3390/ijms23179565

    View details for Web of Science ID 000851178500001

    View details for PubMedID 36076959

    View details for PubMedCentralID PMC9455175

  • Inhibition of triple negative breast cancer metastasis via Enolase-1 modulation Tailor, D., Dheeraj, A., Garcia-Marques, F., Pandrala, M., Bermudez, A., Pitteri, S., Malhotra, S. V. AMER ASSOC CANCER RESEARCH. 2022
  • SU086, an inhibitor of HSP90, impairs glycolysis and represents a treatment strategy for advanced prostate cancer. Cell reports. Medicine Rice, M. A., Kumar, V., Tailor, D., Garcia-Marques, F. J., Hsu, E., Liu, S., Bermudez, A., Kanchustambham, V., Shankar, V., Inde, Z., Alabi, B. R., Muruganantham, A., Shen, M., Pandrala, M., Nolley, R., Aslan, M., Ghoochani, A., Agarwal, A., Buckup, M., Kumar, M., Going, C. C., Peehl, D. M., Dixon, S. J., Zare, R. N., Brooks, J. D., Pitteri, S. J., Malhotra, S. V., Stoyanova, T. 2022; 3 (2): 100502

    Abstract

    Among men, prostate cancer is the second leading cause of cancer-associated mortality, with advanced disease remaining a major clinical challenge. We describe a small molecule, SU086, as a therapeutic strategy for advanced prostate cancer. We demonstrate that SU086 inhibits the growth of prostate cancer cells invitro, cell-line and patient-derived xenografts invivo, and exvivo prostate cancer patient specimens. Furthermore, SU086 in combination with standard of care second-generation anti-androgen therapies displays increased impairment of prostate cancer cell and tumor growth invitro and invivo. Cellular thermal shift assay reveals that SU086 binds to heat shock protein 90 (HSP90) and leads to a decrease in HSP90 levels. Proteomic profiling demonstrates that SU086 binds to and decreases HSP90. Metabolomic profiling reveals that SU086 leads to perturbation of glycolysis. Our study identifies SU086 as a treatment for advanced prostate cancer as a single agent or when combined with second-generation anti-androgens.

    View details for DOI 10.1016/j.xcrm.2021.100502

    View details for PubMedID 35243415

  • Protein signatures to distinguish aggressive from indolent prostate cancer. The Prostate Garcia-Marques, F., Liu, S., Totten, S. M., Bermudez, A., Tanimoto, C., Hsu, E. C., Nolley, R., Hembree, A., Stoyanova, T., Brooks, J. D., Pitteri, S. J. 2022

    Abstract

    Distinguishing men with aggressive from indolent prostate cancer is critical to decisions in the management of clinically localized prostate cancer. Molecular signatures of aggressive disease could help men overcome this major clinical challenge by reducing unnecessary treatment and allowing more appropriate treatment of aggressive disease.We performed a mass spectrometry-based proteomic analysis of normal and malignant prostate tissues from 22 men who underwent surgery for prostate cancer. Prostate cancer samples included Grade Groups (3-5), with 8 patients experiencing recurrence and 14 without evidence of recurrence with a mean of 6.8 years of follow-up. To better understand the biological pathways underlying prostate cancer aggressiveness, we performed a systems biology analysis and gene enrichment analysis. Proteins that distinguished recurrent from nonrecurrent cancer were chosen for validation by immunohistochemical analysis on tissue microarrays containing samples from a larger cohort of patients with recurrent and nonrecurrent prostate cancer.In all, 24,037 unique peptides (false discovery rate < 1%) corresponding to 3,313 distinct proteins were identified with absolute abundance ranges spanning seven orders of magnitude. Of these proteins, 115 showed significantly (p < 0.01) different levels in tissues from recurrent versus nonrecurrent cancers. Analysis of all differentially expressed proteins in recurrent and nonrecurrent cases identified several protein networks, most prominently one in which approximately 24% of the proteins in the network were regulated by the YY1 transcription factor (adjusted p < 0.001). Strong immunohistochemical staining levels of three differentially expressed proteins, POSTN, CALR, and CTSD, on a tissue microarray validated their association with shorter patient survival.The protein signatures identified could improve understanding of the molecular drivers of aggressive prostate cancer and be used as candidate prognostic biomarkers.

    View details for DOI 10.1002/pros.24307

    View details for PubMedID 35098564

  • Engineered Cell-Derived Vesicles Displaying Targeting Peptide and Functionalized with Nanocarriers for Therapeutic microRNA Delivery to Triple-Negative Breast Cancer in Mice. Advanced healthcare materials Bose, R. J., Kumar, U. S., Garcia-Marques, F., Zeng, Y., Habte, F., McCarthy, J. R., Pitteri, S., Massoud, T. F., Paulmurugan, R. 2021: e2101387

    Abstract

    Polymeric nanocarriers (PNCs) can be used to deliver therapeutic microRNAs (miRNAs) to solid cancers. However, the ability of these nanocarriers to specifically target tumors remains a challenge. Alternatively, extracellular vesicles (EVs) derived from tumor cells show homotypic affinity to parent cells, but loading sufficient amounts of miRNAs into EVs is difficult. Here, we investigate whether uPAR-targeted delivery of nanococktails containing PNCs loaded with therapeutic antimiRNAs, and coated with uPA engineered extracellular vesicles (uPA-eEVs) can elicit synergistic antitumor responses. The uPA-eEVs coating on PNCs increases natural tumor targeting affinities, thereby enhancing the antitumor activity of antimiRNA nanococktails. The systemic administration of uPA-eEV-PNCs nanococktail showed a robust tumor tropism, which significantly enhanced the combinational antitumor effects of antimiRNA-21 and antimiRNA-10b, and led to significant tumor regression and extension of progression free survival for syngeneic 4T1 tumor-bearing mice. In addition, the uPA-eEV-PNCs-antimiRNAs nanococktail plus low dose doxorubicin resulted in a synergistic antitumor effect as evidenced by inhibition of tumor growth, reduction of lung metastases, and extension of survival of 4T1 tumor-bearing mice. Our targeted combinational nanococktail strategy could be readily translated to the clinical setting by using autologous cancer cells that have flexibility for ex vivo expansion and genetic engineering. This article is protected by copyright. All rights reserved.

    View details for DOI 10.1002/adhm.202101387

    View details for PubMedID 34879180

  • Multi-omics analysis of spatially distinct stromal cells reveals tumor-induced O-glycosylation of the CDK4-pRB axis in fibroblasts at the invasive tumor edge. Cancer research Bouchard, G., Garcia-Marques, F. J., Karacosta, L. G., Zhang, W., Bermudez, A., Riley, N. M., Varma, S., Mehl, L. C., Benson, J. A., Shrager, J. B., Bertozzi, C. R., Pitteri, S. J., Giaccia, A. J., Plevritis, S. K. 2021

    Abstract

    The invasive leading edge represents a potential gateway for tumor metastasis. The role of fibroblasts from the tumor edge in promoting cancer invasion and metastasis has not been comprehensively elucidated. We hypothesize that crosstalk between tumor and stromal cells within the tumor microenvironment (TME) results in activation of key biological pathways depending on their position in the tumor (edge vs core). Here we highlight phenotypic differences between tumor-adjacent-fibroblasts (TAF) from the invasive edge and tumor core fibroblasts (TCF) from the tumor core, established from human lung adenocarcinomas. A multi-omics approach that includes genomics, proteomics, and O-glycoproteomics was used to characterize crosstalk between TAFs and cancer cells. These analyses showed that O-glycosylation, an essential post-translational modification resulting from sugar metabolism, alters key biological pathways including the cyclin-dependent kinase 4 and phosphorylated retinoblastoma protein (CDK4-pRB) axis in the stroma and indirectly modulates pro-invasive features of cancer cells. In summary, the O-glycoproteome represents a new consideration for important biological processes involved in tumor-stroma crosstalk and a potential avenue to improve the anti-cancer efficacy of CDK4 inhibitors.

    View details for DOI 10.1158/0008-5472.CAN-21-1705

    View details for PubMedID 34853070

  • Identifying a novel glycolytic inhibitor for treatment of aggressive prostate cancer. Stoyanova, T., Rice, M. A., Kumar, V., Tailor, D., Garcia-Marques, F., Bermudez, A., Kanchustambham, V., Shankar, V., Inde, Z., Pandrala, M., Nolley, R., Ghoochani, A., Liu, S., Aslan, M., Agarwal, A., Buckup, M., Hsu, E., Going, C. C., Peehl, D. M., Dixon, S. J., Zare, R. N., Brooks, J. D., Pitteri, S. J., Malhotra, S. V., Stoyanova, T. AMER ASSOC CANCER RESEARCH. 2021
  • Y box binding protein 1 inhibition as a targeted therapy for ovarian cancer. Cell chemical biology Tailor, D., Resendez, A., Garcia-Marques, F. J., Pandrala, M., Going, C. C., Bermudez, A., Kumar, V., Rafat, M., Nambiar, D. K., Honkala, A., Le, Q., Sledge, G. W., Graves, E., Pitteri, S. J., Malhotra, S. V. 2021

    Abstract

    Y box binding protein 1 (YB-1) is a multifunctional protein associated with tumor progression and the emergence of treatment resistance (TR). Here, we report an azopodophyllotoxin small molecule, SU056, that potently inhibits tumor growth and progression via YB-1 inhibition. This YB-1 inhibitor inhibits cell proliferation, resistance to apoptosis in ovarian cancer (OC) cells, and arrests in the G1 phase. Inhibitor treatment leads to enrichment of proteins associated with apoptosis and RNA degradation pathways while downregulating spliceosome pathway. Invivo, SU056 independently restrains OC progression and exerts a synergistic effect with paclitaxel to further reduce disease progression with no observable liver toxicity. Moreover, invitro mechanistic studies showed delayed disease progression via inhibition of drug efflux and multidrug resistance 1, and significantly lower neurotoxicity as compared with etoposide. These data suggest that YB-1 inhibition may be an effective strategy to reduce OC progression, antagonize TR, and decrease patient mortality.

    View details for DOI 10.1016/j.chembiol.2021.02.014

    View details for PubMedID 33713600

  • Discovery of CASP8 as a potential biomarker for high-risk prostate cancer through a high-multiplex immunoassay. Scientific reports Liu, S. n., Garcia-Marques, F. n., Zhang, C. A., Lee, J. J., Nolley, R. n., Shen, M. n., Hsu, E. C., Aslan, M. n., Koul, K. n., Pitteri, S. J., Brooks, J. D., Stoyanova, T. n. 2021; 11 (1): 7612

    Abstract

    Prostate cancer remains the most common non-cutaneous malignancy among men in the United States. To discover potential serum-based biomarkers for high-risk prostate cancer, we performed a high-multiplex immunoassay utilizing patient-matched pre-operative and post-operative serum samples from ten men with high-grade and high-volume prostate cancer. Our study identified six (CASP8, MSLN, FGFBP1, ICOSLG, TIE2 and S100A4) out of 174 proteins that were significantly decreased after radical prostatectomy. High levels of CASP8 were detected in pre-operative serum samples when compared to post-operative serum samples and serum samples from patients with benign prostate hyperplasia (BPH). By immunohistochemistry, CASP8 protein was expressed at higher levels in prostate cancer tissues compared to non-cancerous and BPH tissues. Likewise, CASP8 mRNA expression was significantly upregulated in prostate cancer when compared to benign prostate tissues in four independent clinical datasets. In addition, mRNA levels of CASP8 were higher in patients with recurrent prostate cancer when compared to patients with non-recurrent prostate cancer and high expression of CASP8 was associated with worse disease-free survival and overall survival in renal cancer. Together, our results suggest that CASP8 may potentially serve as a biomarker for high-risk prostate cancer and possibly renal cancer.

    View details for DOI 10.1038/s41598-021-87155-5

    View details for PubMedID 33828176

  • MCM2-7 complex is a novel druggable target for neuroendocrine prostate cancer. Scientific reports Hsu, E. C., Shen, M., Aslan, M., Liu, S., Kumar, M., Garcia-Marques, F., Nguyen, H. M., Nolley, R., Pitteri, S. J., Corey, E., Brooks, J. D., Stoyanova, T. 2021; 11 (1): 13305

    Abstract

    Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer that rarely develops de novo in primary tumors and is commonly acquired during the development of treatment resistance. NEPC is characterized by gain of neuroendocrine markers and loss of androgen receptor (AR), making it resistant to current therapeutic strategies targeting the AR signaling axis. Here, we report that MCM2, MCM3, MCM4, and MCM6 (MCM2/3/4/6) are elevated in human NEPC and high levels of MCM2/3/4/6 are associated with liver metastasis and poor survival in prostate cancer patients. MCM2/3/4/6 are four out of six proteins that form a core DNA helicase (MCM2-7) responsible for unwinding DNA forks during DNA replication. Inhibition of MCM2-7 by treatment with ciprofloxacin inhibits NEPC cell proliferation and migration in vitro, significantly delays NEPC tumor xenograft growth, and partially reverses the neuroendocrine phenotype in vivo. Our study reveals the clinical relevance of MCM2/3/4/6 proteins in NEPC and suggests that inhibition of MCM2-7 may represent a new therapeutic strategy for NEPC.

    View details for DOI 10.1038/s41598-021-92552-x

    View details for PubMedID 34172788

  • Oncogene-mediated metabolic gene signature predicts breast cancer outcome. NPJ breast cancer Aslan, M., Hsu, E. C., Garcia-Marques, F. J., Bermudez, A., Liu, S., Shen, M., West, M., Zhang, C. A., Rice, M. A., Brooks, J. D., West, R., Pitteri, S. J., Győrffy, B., Stoyanova, T. 2021; 7 (1): 141

    Abstract

    Breast cancer remains the second most lethal cancer among women in the United States and triple-negative breast cancer is the most aggressive subtype with limited treatment options. Trop2, a cell membrane glycoprotein, is overexpressed in almost all epithelial cancers. In this study, we demonstrate that Trop2 is overexpressed in triple-negative breast cancer (TNBC), and downregulation of Trop2 delays TNBC cell and tumor growth supporting the oncogenic role of Trop2 in breast cancer. Through proteomic profiling, we discovered a metabolic signature comprised of TALDO1, GPI, LDHA, SHMT2, and ADK proteins that were downregulated in Trop2-depleted breast cancer tumors. The identified oncogene-mediated metabolic gene signature is significantly upregulated in TNBC patients across multiple RNA-expression clinical datasets. Our study further reveals that the metabolic gene signature reliably predicts poor survival of breast cancer patients with early stages of the disease. Taken together, our study identified a new five-gene metabolic signature as an accurate predictor of breast cancer outcome.

    View details for DOI 10.1038/s41523-021-00341-6

    View details for PubMedID 34711841

  • Plectin is a regulator of prostate cancer growth and metastasis. Oncogene Buckup, M., Rice, M. A., Hsu, E., Garcia-Marques, F., Liu, S., Aslan, M., Bermudez, A., Huang, J., Pitteri, S. J., Stoyanova, T. 2020

    Abstract

    Prostate cancer is responsible for over 30,000 US deaths annually, attributed largely to incurable metastatic disease. Here, we demonstrate that high levels of plectin are associated with localized and metastatic human prostate cancer when compared to benign prostate tissues. Knock-down of plectin inhibits prostate cancer cell growth and colony formation in vitro, and growth of prostate cancer xenografts in vivo. Plectin knock-down further impairs aggressive and invasive cellular behavior assessed by migration, invasion, and wound healing in vitro. Consistently, plectin knock-down cells have impaired metastatic colonization to distant sites including liver, lung, kidney, bone, and genitourinary system. Plectin knock-down inhibited number of metastases per organ, as well as decreased overall metastatic burden. To gain insights into the role of plectin in prostate cancer growth and metastasis, we performed proteomic analysis of prostate cancer plectin knock-down xenograft tissues. Gene set enrichment analysis shows an increase in levels of proteins involved with extracellular matrix and laminin interactions, and a decrease in levels of proteins regulating amino acid metabolism, cytoskeletal proteins, and cellular response to stress. Collectively these findings demonstrate that plectin is an important regulator of prostate cancer cell growth and metastasis.

    View details for DOI 10.1038/s41388-020-01557-9

    View details for PubMedID 33219316

  • A Network of Macrophages Supports Mitochondrial Homeostasis in the Heart CELL Nicolas-Avila, J. A., Lechuga-Vieco, A. V., Esteban-Martinez, L., Sanchez-Diaz, M., Diaz-Garcia, E., Santiago, D. J., Rubio-Ponce, A., Li, J., Balachander, A., Quintana, J. A., Martinez-de-Mena, R., Castejon-Vega, B., Pun-Garcia, A., Traves, P. G., Bonzon-Kulichenko, E., Garcia-Marques, F., Cusso, L., A-Gonzalez, N., Gonzalez-Guerra, A., Roche-Molina, M., Martin-Salamanca, S., Crainiciuc, G., Guzman, G., Larrazabal, J., Herrero-Galan, E., Alegre-Cebollada, J., Lemke, G., Rothlin, C., Jesus Jimenez-Borreguero, L., Reyes, G., Castrillo, A., Desco, M., Munoz-Canoves, P., Ibanez, B., Torres, M., Ng, L., Priori, S. G., Bueno, H., Vazquez, J., Cordero, M. D., Bernal, J. A., Enriquez, J. A., Hidalgo, A. 2020; 183 (1): 94-+

    Abstract

    Cardiomyocytes are subjected to the intense mechanical stress and metabolic demands of the beating heart. It is unclear whether these cells, which are long-lived and rarely renew, manage to preserve homeostasis on their own. While analyzing macrophages lodged within the healthy myocardium, we discovered that they actively took up material, including mitochondria, derived from cardiomyocytes. Cardiomyocytes ejected dysfunctional mitochondria and other cargo in dedicated membranous particles reminiscent of neural exophers, through a process driven by the cardiomyocyte's autophagy machinery that was enhanced during cardiac stress. Depletion of cardiac macrophages or deficiency in the phagocytic receptor Mertk resulted in defective elimination of mitochondria from the myocardial tissue, activation of the inflammasome, impaired autophagy, accumulation of anomalous mitochondria in cardiomyocytes, metabolic alterations, and ventricular dysfunction. Thus, we identify an immune-parenchymal pair in the murine heart that enables transfer of unfit material to preserve metabolic stability and organ function. VIDEO ABSTRACT.

    View details for DOI 10.1016/j.cell.2020.08.031

    View details for Web of Science ID 000576772900010

    View details for PubMedID 32937105

  • Functional role of respiratory supercomplexes in mice: SCAF1 relevance and segmentation of the Q(pool) SCIENCE ADVANCES Calvo, E., Cogliati, S., Hernansanz-Agustin, P., Loureiro-Lopez, M., Guaras, A., Casuso, R. A., Garcia-Marques, F., Acin-Perez, R., Marti-Mateos, Y., Silla-Castro, J. C., Carro-Alvarellos, M., Huertas, J. R., Vazquez, J., Enriquez, J. A. 2020; 6 (26): eaba7509

    Abstract

    Mitochondrial respiratory complexes assemble into supercomplexes (SC). Q-respirasome (III2 + IV) requires the supercomplex assembly factor (SCAF1) protein. The role of this factor in the N-respirasome (I + III2 + IV) and the physiological role of SCs are controversial. Here, we study C57BL/6J mice harboring nonfunctional SCAF1, the full knockout for SCAF1, or the wild-type version of the protein and found that exercise performance is SCAF1 dependent. By combining quantitative data-independent proteomics, 2D Blue native gel electrophoresis, and functional analysis of enriched respirasome fractions, we show that SCAF1 confers structural attachment between III2 and IV within the N-respirasome, increases NADH-dependent respiration, and reduces reactive oxygen species (ROS). Furthermore, the expression of AOX in cells and mice confirms that CI-CIII superassembly segments the CoQ in two pools and modulates CI-NADH oxidative capacity.

    View details for DOI 10.1126/sciadv.aba7509

    View details for Web of Science ID 000543504100029

    View details for PubMedID 32637615

    View details for PubMedCentralID PMC7314541

  • Complement C5 Protein as a Marker of Subclinical Atherosclerosis JOURNAL OF THE AMERICAN COLLEGE OF CARDIOLOGY Martinez-Lopez, D., Roldan-Montero, R., Garcia-Marques, F., Nunez, E., Jorge, I., Camafeita, E., Minguez, P., Rodriguez de Cordoba, S., Lopez-Melgar, B., Lara-Pezzi, E., Fernandez-Ortiz, A., Ibanez, B., Manuel Valdivielso, J., Fuster, V., Michel, J., Miguel Blanco-Colio, L., Vazquez, J., Luis Martin-Ventura, J. 2020; 75 (16): 1926–41

    Abstract

    The mechanisms underlying early atherosclerotic plaque formation are not completely understood. Moreover, plasma biomarkers of subclinical atherosclerosis are lacking.The purpose of this study was to analyze the temporal and topologically resolved protein changes taking place in human aortas with early atherosclerosis to find new potential diagnostic and/or therapeutic targets.The protein composition of healthy aortas (media layer) or with early atheroma (fatty streak and fibrolipidic, media and intima layers) was analyzed by deep quantitative multiplexed proteomics. Further analysis was performed by Western blot, immunohistochemistry, real-time polymerase chain reaction, and enzyme-linked immunosorbent assay. Plasma levels of complement C5 were analyzed in relation to the presence of generalized (>2 plaques) or incipient (0 to 2 plaques) subclinical atherosclerosis in 2 independent clinical cohorts (PESA [Progression of Early Subclinical Atherosclerosis] [n = 360] and NEFRONA [National Observatory of Atherosclerosis in Nephrology] [n = 394]).Proteins involved in lipid transport, complement system, immunoglobulin superfamily, and hemostasis are increased in early plaques. Components from the complement activation pathway were predominantly increased in the intima of fibrolipidic plaques. Among them, increased C5 protein levels were further confirmed by Western blot, enzyme-linked immunosorbent assay and immunohistochemistry, and associated with in situ complement activation. Plasma C5 was significantly increased in individuals with generalized subclinical atherosclerosis in both PESA and NEFRONA cohorts, independently of risk factors. Moreover, in the PESA study, C5 plasma levels positively correlated with global plaque volume and coronary calcification.Activation of the complement system is a major alteration in early atherosclerotic plaques and is reflected by increased C5 plasma levels, which have promising value as a novel circulating biomarker of subclinical atherosclerosis.

    View details for Web of Science ID 000531872400010

    View details for PubMedID 32327104

  • Trop2 is a driver of metastatic prostate cancer with neuroendocrine phenotype via PARP1. Proceedings of the National Academy of Sciences of the United States of America Hsu, E. C., Rice, M. A., Bermudez, A. n., Marques, F. J., Aslan, M. n., Liu, S. n., Ghoochani, A. n., Zhang, C. A., Chen, Y. S., Zlitni, A. n., Kumar, S. n., Nolley, R. n., Habte, F. n., Shen, M. n., Koul, K. n., Peehl, D. M., Zoubeidi, A. n., Gambhir, S. S., Kunder, C. A., Pitteri, S. J., Brooks, J. D., Stoyanova, T. n. 2020

    Abstract

    Resistance to androgen deprivation therapy, or castration-resistant prostate cancer (CRPC), is often accompanied by metastasis and is currently the ultimate cause of prostate cancer-associated deaths in men. Recently, secondary hormonal therapies have led to an increase of neuroendocrine prostate cancer (NEPC), a highly aggressive variant of CRPC. Here, we identify that high levels of cell surface receptor Trop2 are predictive of recurrence of localized prostate cancer. Moreover, Trop2 is significantly elevated in CRPC and NEPC, drives prostate cancer growth, and induces neuroendocrine phenotype. Overexpression of Trop2 induces tumor growth and metastasis while loss of Trop2 suppresses these abilities in vivo. Trop2-driven NEPC displays a significant up-regulation of PARP1, and PARP inhibitors significantly delay tumor growth and metastatic colonization and reverse neuroendocrine features in Trop2-driven NEPC. Our findings establish Trop2 as a driver and therapeutic target for metastatic prostate cancer with neuroendocrine phenotype and suggest that high Trop2 levels could identify cancers that are sensitive to Trop2-targeting therapies and PARP1 inhibition.

    View details for DOI 10.1073/pnas.1905384117

    View details for PubMedID 31932422

  • Discovery of PTN as a serum-based biomarker of pro-metastatic prostate cancer. British journal of cancer Liu, S. n., Shen, M. n., Hsu, E. C., Zhang, C. A., Garcia-Marques, F. n., Nolley, R. n., Koul, K. n., Rice, M. A., Aslan, M. n., Pitteri, S. J., Massie, C. n., George, A. n., Brooks, J. D., Gnanapragasam, V. J., Stoyanova, T. n. 2020

    Abstract

    Distinguishing clinically significant from indolent prostate cancer (PC) is a major clinical challenge. We utilised targeted protein biomarker discovery approach to identify biomarkers specific for pro-metastatic PC. Serum samples from the cancer-free group; Cambridge Prognostic Group 1 (CPG1, low risk); CPG5 (high risk) and metastatic disease were analysed using Olink Proteomics panels. Tissue validation was performed by immunohistochemistry in a radical prostatectomy cohort (n = 234). We discovered that nine proteins (pleiotrophin (PTN), MK, PVRL4, EPHA2, TFPI-2, hK11, SYND1, ANGPT2, and hK14) were elevated in metastatic PC patients when compared to other groups. PTN levels were increased in serum from men with CPG5 compared to benign and CPG1. High tissue PTN level was an independent predictor of biochemical recurrence and metastatic progression in low- and intermediate-grade disease. These findings suggest that PTN may represent a novel biomarker for the presence of poor prognosis local disease with the potential to metastasise warranting further investigation.

    View details for DOI 10.1038/s41416-020-01200-0

    View details for PubMedID 33288843

  • Genomic analysis of Vascular Invasion in Hepatocellular Carcinoma (HCC) Reveals Molecular Drivers and Predictive Biomarkers. Hepatology (Baltimore, Md.) Krishnan, M. S., Rajan Kd, A. n., Park, J. n., Arjunan, V. n., Garcia Marques, F. J., Bermudez, A. n., Girvan, O. A., Hoang, N. S., Yin, J. n., Nguyen, M. H., Kothary, N. n., Pitteri, S. n., Felsher, D. W., Dhanasekaran, R. n. 2020

    Abstract

    Vascular invasion is a critical risk factor for hepatocellular carcinoma (HCC) recurrence and poor survival. The molecular drivers of vascular invasion in HCC are largely unknown. Deciphering the molecular landscape of invasive HCC will help identify novel therapeutic targets and noninvasive biomarkers. To this end, we undertook this study to evaluate the genomic, transcriptomic, and proteomic profile of tumors with vascular invasion using the multi-platform cancer genome atlas (TCGA) data (n=373). In the TCGA liver hepatocellular carcinoma (LIHC) cohort, macrovascular invasion was present in 5% (n=17) of tumors and microvascular invasion in 25% (n=94) of tumors. Functional pathway analysis revealed that the MYC oncogene was a common upstream regulator of the mRNA, miRNA and proteomic changes in vascular invasion. We performed comparative proteomic analyses of invasive human HCC and MYC driven murine HCC and identified fibronectin to be proteomic biomarker of invasive HCC (mouse Fn1 p= 1.7 X 10-11 ; human FN1 p=1.5 X 10-4 ) conserved across the two species. Mechanistically, we show that FN1 promotes the migratory and invasive phenotype of HCC cancer cells. We demonstrate tissue overexpression of fibronectin in human HCC using a large independent cohort of human HCC tissue microarray (n=153; p<0.001). Lastly, we showed that plasma fibronectin levels were significantly elevated in patients with HCC (n=35, mean=307.7 μg/ml, SEM=35.9) when compared to cirrhosis (n=10, mean=41.8 μg/ml, SEM=13.3; p<0.0001). CONCLUSION: Our study evaluates the molecular landscape of tumors with vascular invasion, identifying distinct transcriptional, epigenetic and proteomic changes driven by the MYC oncogene. We show that MYC upregulates fibronectin expression which promotes HCC invasiveness. In addition, we identify fibronectin to be a promising non-invasive proteomic biomarker of vascular invasion in HCC.

    View details for DOI 10.1002/hep.31614

    View details for PubMedID 33140851

  • Reconstructed Apoptotic Bodies as Targeted "Nano Decoys" to Treat Intracellular Bacterial Infections within Macrophages and Cancer Cells. ACS nano Bose, R. J., Tharmalingam, N. n., Garcia Marques, F. J., Sukumar, U. K., Natarajan, A. n., Zeng, Y. n., Robinson, E. n., Bermudez, A. n., Chang, E. n., Habte, F. n., Pitteri, S. J., McCarthy, J. R., Gambhir, S. S., Massoud, T. F., Mylonakis, E. n., Paulmurugan, R. n. 2020

    Abstract

    Staphylococcus aureus (S. aureus) is a highly pathogenic facultative anaerobe that in some instances resides as an intracellular bacterium within macrophages and cancer cells. This pathogen can establish secondary infection foci, resulting in recurrent systemic infections that are difficult to treat using systemic antibiotics. Here, we use reconstructed apoptotic bodies (ReApoBds) derived from cancer cells as "nano decoys" to deliver vancomycin intracellularly to kill S. aureus by targeting inherent "eat me" signaling of ApoBds. We prepared ReApoBds from different cancer cells (SKBR3, MDA-MB-231, HepG2, U87-MG, and LN229) and used them for vancomycin delivery. Physicochemical characterization showed ReApoBds size ranges from 80 to 150 nm and vancomycin encapsulation efficiency of 60 ± 2.56%. We demonstrate that the loaded vancomycin was able to kill intracellular S. aureus efficiently in an in vitro model of S. aureus infected RAW-264.7 macrophage cells, and U87-MG (p53-wt) and LN229 (p53-mt) cancer cells, compared to free-vancomycin treatment (P < 0.001). The vancomycin loaded ReApoBds treatment in S. aureus infected macrophages showed a two-log-order higher CFU reduction than the free-vancomycin treatment group. In vivo studies revealed that ReApoBds can specifically target macrophages and cancer cells. Vancomycin loaded ReApoBds have the potential to kill intracellular S. aureus infection in vivo in macrophages and cancer cells.

    View details for DOI 10.1021/acsnano.0c00921

    View details for PubMedID 32347709

  • NOTCH Activation Promotes Valve Formation by Regulating the Endocardial Secretome. Molecular & cellular proteomics : MCP Torregrosa-Carrión, R., Luna-Zurita, L., García-Marqués, F., D'Amato, G., Piñeiro-Sabarís, R., Bonzón-Kulichenko, E., Vázquez, J., de la Pompa, J. L. 2019; 18 (9): 1782-1795

    Abstract

    The endocardium is a specialized endothelium that lines the inner surface of the heart. Functional studies in mice and zebrafish have established that the endocardium is a source of instructive signals for the development of cardiac structures, including the heart valves and chambers. Here, we characterized the NOTCH-dependent endocardial secretome by manipulating NOTCH activity in mouse embryonic endocardial cells (MEEC) followed by mass spectrometry-based proteomics. We profiled different sets of soluble factors whose secretion not only responds to NOTCH activation but also shows differential ligand specificity, suggesting that ligand-specific inputs may regulate the expression of secreted proteins involved in different cardiac development processes. NOTCH signaling activation correlates with a transforming growth factor-β2 (TGFβ2)-rich secretome and the delivery of paracrine signals involved in focal adhesion and extracellular matrix (ECM) deposition and remodeling. In contrast, NOTCH inhibition is accompanied by the up-regulation of specific semaphorins that may modulate cell migration. The secretome protein expression data showed a good correlation with gene profiling of RNA expression in embryonic endocardial cells. Additional characterization by in situ hybridization in mouse embryos revealed expression of various NOTCH candidate effector genes (Tgfβ2, Loxl2, Ptx3, Timp3, Fbln2, and Dcn) in heart valve endocardium and/or mesenchyme. Validating these results, mice with conditional Dll4 or Jag1 loss-of-function mutations showed gene expression alterations similar to those observed at the protein level in vitro. These results provide the first description of the NOTCH-dependent endocardial secretome and validate MEEC as a tool for assaying the endocardial secretome response to a variety of stimuli and the potential use of this system for drug screening.

    View details for DOI 10.1074/mcp.RA119.001492

    View details for PubMedID 33451535

  • APOA1 oxidation is associated to dysfunctional high-density lipoproteins in human abdominal aortic aneurysm EBIOMEDICINE Martinez-Lopez, D., Camafeita, E., Cedo, L., Roldan-Montero, R., Jorge, I., Garcia-Marques, F., Gomez-Serrano, M., Bonzon-Kulichenko, E., Blanco-Vaca, F., Miguel Blanco-Colio, L., Michel, J., Carles Escola-Gil, J., Vazquez, J., Luis Martin-Ventura, J. 2019; 43: 43–53

    Abstract

    High-density lipoproteins (HDL) are a complex mixture of lipids and proteins with vasculoprotective properties. However, HDL components could suffer post-translational modifications (PTMs) under pathological conditions, leading to dysfunctional HDL. We studied whether HDL are modified in abdominal aortic aneurysm (AAA) and the effect on HDL functionality.HDL were isolated by ultracentrifugation from AAA tissue (HDL-T) and from plasma of healthy volunteers and then incubated with AAA tissue-conditioned medium (HDL-AAA CM). PTMs from these particles were characterized using Comet-PTM. The ability of HDL-AAA CM for promoting cholesterol efflux was determined ex vivo and in vivo by using J774A.1 [3H]cholesterol-labeled mouse macrophages and after injecting [3H]cholesterol-labeled mouse macrophages and HDL into the peritoneal cavity of wild-type C57BL/6 mice, respectively. Trp50 and Trp108 oxidized forms of APOA1 in HDL incubated with conditioned-medium of activated neutrophils and in plasma of AAA patients and controls were measured by targeted parallel reaction monitoring.Oxidation was the most prevalent PTM in apolipoproteins, particularly in APOA1. Trp50 and Trp108 in APOA1 were the residues most clearly affected by oxidation in HDL-T and in HDL-AAA CM, when compared to their controls. In addition, cholesterol efflux was decreased in macrophages incubated with HDL-AAA CM in vitro and a decreased macrophage-to-serum reverse cholesterol transport was also observed in mice injected with HDL-AAA CM. Finally, both oxidized Trp50 and Trp108 forms of APOA1 were increased in HDL incubated with conditioned-medium of activated neutrophils and in plasma of AAA patients in relation to controls.Oxidative modifications of HDL present in AAA tissue and plasma were closely associated with the loss of vasculoprotective properties of HDL in AAA. FUND: MINECO, ISCiii-FEDER, CIBERDEM, CIBERCV and LA CAIXA.

    View details for DOI 10.1016/j.ebiom.2019.04.012

    View details for Web of Science ID 000470091600019

    View details for PubMedID 30982767

    View details for PubMedCentralID PMC6562066

  • SanXoT: a modular and versatile package for the quantitative analysis of high-throughput proteomics experiments BIOINFORMATICS Trevisan-Herraz, M., Bagwan, N., Garcia-Marques, F., Manuel Rodriguez, J., Jorge, I., Ezkurdia, I., Bonzon-Kulichenko, E., Vazquez, J. 2019; 35 (9): 1594–96

    Abstract

    Mass spectrometry-based proteomics has had a formidable development in recent years, increasing the amount of data handled and the complexity of the statistical resources needed. Here we present SanXoT, an open-source, standalone software package for the statistical analysis of high-throughput, quantitative proteomics experiments. SanXoT is based on our previously developed weighted spectrum, peptide and protein statistical model and has been specifically designed to be modular, scalable and user-configurable. SanXoT allows limitless workflows that adapt to most experimental setups, including quantitative protein analysis in multiple experiments, systems biology, quantification of post-translational modifications and comparison and merging of experimental data from technical or biological replicates.Download links for the SanXoT Software Package, source code and documentation are available at https://wikis.cnic.es/proteomica/index.php/SSP.jvazquez@cnic.es or ebonzon@cnic.es.Supplementary information is available at Bioinformatics online.

    View details for DOI 10.1093/bioinformatics/bty815

    View details for Web of Science ID 000469491000025

    View details for PubMedID 30252043

    View details for PubMedCentralID PMC6499250