Daniel Dan Liu

All Publications

• Purification and characterization of human neural stem and progenitor cells. Cell Liu, D. D., He, J. Q., Sinha, R., Eastman, A. E., Toland, A. M., Morri, M., Neff, N. F., Vogel, H., Uchida, N., Weissman, I. L. 2023; 186 (6): 1179

  Abstract

  The human brain undergoes rapid development at mid-gestation from a pool of neural stem and progenitor cells (NSPCs) that give rise to the neurons, oligodendrocytes, and astrocytes of the mature brain. Functional study of these cell types has been hampered by a lack of precise purification methods. We describe a method for prospectively isolating ten distinct NSPC types from the developing human brain using cell-surface markers. CD24-THY1-/lo cells were enriched for radial glia, which robustly engrafted and differentiated into all three neural lineages in the mouse brain. THY1hi cells marked unipotent oligodendrocyte precursors committed to an oligodendroglial fate, and CD24+THY1-/lo cells marked committed excitatory and inhibitory neuronal lineages. Notably, we identify and functionally characterize a transcriptomically distinct THY1hiEGFRhiPDGFRA- bipotent glial progenitor cell (GPC), which is lineage-restricted to astrocytes and oligodendrocytes, but not to neurons. Our study provides a framework for the functional study of distinct cell types in human neurodevelopment.

  View details for DOI 10.1016/j.cell.2023.02.017

  View details for PubMedID 36931245

• Prospective isolation of neural stem and progenitor cells from the developing human brain. STAR protocols Liu, D. D., He, J. Q., Uchida, N., Weissman, I. L., Sinha, R. 2023; 4 (4): 102674

  Abstract

  Prospective isolation of defined cell types is critical for the functional study of stem cells, especially in primary human tissues. Here, we present a protocol for purifying 10 transcriptomically and functionally distinct neural stem and progenitor cell types from the developing human brain using fluorescence-activated cell sorting. We describe steps for tissue dissociation, staining, and cell sorting as well as downstream functional experiments for measuring clonogenicity, differentiation, and engraftment potential of purified populations. For complete details on the use and execution of this protocol, please refer to Liu etal. (2023).1.

  View details for DOI 10.1016/j.xpro.2023.102674

  View details for PubMedID 37897731

• LCOR mediates interferon-independent tumor immunogenicity and responsiveness to immune-checkpoint blockade in triple-negative breast cancer. Nature cancer Perez-Nunez, I., Rozalen, C., Palomeque, J. A., Sangrador, I., Dalmau, M., Comerma, L., Hernandez-Prat, A., Casadevall, D., Menendez, S., Liu, D. D., Shen, M., Berenguer, J., Ruiz, I. R., Pena, R., Montanes, J. C., Alba, M. M., Bonnin, S., Ponomarenko, J., Gomis, R. R., Cejalvo, J. M., Servitja, S., Marzese, D. M., Morey, L., Voorwerk, L., Arribas, J., Bermejo, B., Kok, M., Pusztai, L., Kang, Y., Albanell, J., Celia-Terrassa, T. 2022

  Abstract

  Ligand-dependent corepressor (LCOR) mediates normal and malignant breast stem cell differentiation. Cancer stem cells (CSCs) generate phenotypic heterogeneity and drive therapy resistance, yet their role in immunotherapy is poorly understood. Here we show that immune-checkpoint blockade (ICB) therapy selects for LCORlow CSCs with reduced antigen processing/presentation machinery (APM) driving immune escape and ICB resistance in triple-negative breast cancer (TNBC). We unveil an unexpected function of LCOR as a master transcriptional activator of APM genes binding to IFN-stimulated response elements (ISREs) in an IFN signaling-independent manner. Through genetic modification of LCOR expression, we demonstrate its central role in modulation of tumor immunogenicity and ICB responsiveness. In TNBC, LCOR associates with ICB clinical response. Importantly, extracellular vesicle (EV) Lcor-messenger RNA therapy in combination with anti-PD-L1 overcame resistance and eradicated breast cancer metastasis in preclinical models. Collectively, these data support LCOR as a promising target for enhancement of ICB efficacy in TNBC, by boosting of tumor APM independently of IFN.

  View details for DOI 10.1038/s43018-022-00339-4

  View details for PubMedID 35301507

• A Clinical PET Imaging Tracer ([18F]DASA-23) to Monitor Pyruvate Kinase M2 Induced Glycolytic Reprogramming in Glioblastoma. Clinical cancer research : an official journal of the American Association for Cancer Research Beinat, C., Patel, C. B., Haywood, T., Murty, S., Naya, L., Castillo, J. B., Reyes, S. T., Phillips, M., Buccino, P., Shen, B., Park, J. H., Koran, M. E., Alam, I. S., James, M. L., Holley, D., Halbert, K., Gandhi, H., He, J. Q., Granucci, M., Johnson, E., Liu, D. D., Uchida, N., Sinha, R., Chu, P., Born, D. E., Warnock, G. I., Weissman, I., Hayden Gephart, M., Khalighi, M. M., Massoud, T. F., Iagaru, A., Davidzon, G., Thomas, R., Nagpal, S., Recht, L. D., Gambhir, S. S. 2021

  Abstract

  PURPOSE: Pyruvate kinase M2 (PKM2) catalyzes the final step in glycolysis, a key process of cancer metabolism. PKM2 is preferentially expressed by glioblastoma (GBM) cells with minimal expression in healthy brain. We describe the development, validation, and translation of a novel positron emission tomography (PET) tracer to study PKM2 in GBM. We evaluated 1-((2-fluoro-6-[18F]fluorophenyl)sulfonyl)-4-((4-methoxyphenyl)sulfonyl)piperazine ([18F]DASA-23) in cell culture, mouse models of GBM, healthy human volunteers, and GBM patients.EXPERIMENTAL DESIGN: [18F]DASA-23 was synthesized with a molar activity of 100.47 {plus minus} 29.58 GBq/mol and radiochemical purity >95%. We performed initial testing of [18F]DASA-23 in GBM cell culture and human GBM xenografts implanted orthotopically into mice. Next we produced [18F]DASA-23 under FDA oversight, and evaluated it in healthy volunteers, and a pilot cohort of glioma patients.RESULTS: In mouse imaging studies, [18F]DASA-23 clearly delineated the U87 GBM from surrounding healthy brain tissue and had a tumor-to-brain ratio (TBR) of 3.6 {plus minus} 0.5. In human volunteers, [18F]DASA-23 crossed the intact blood-brain barrier and was rapidly cleared. In GBM patients, [18F]DASA-23 successfully outlined tumors visible on contrast-enhanced magnetic resonance imaging (MRI). The uptake of [18F]DASA-23 was markedly elevated in GBMs compared to normal brain, and it identified a metabolic non-responder within 1-week of treatment initiation.CONCLUSIONS: We developed and translated [18F]DASA-23 as a new tracer that demonstrated the visualization of aberrantly expressed PKM2 for the first time in human subjects. These results warrant further clinical evaluation of [18F]DASA-23 to assess its utility for imaging therapy-induced normalization of aberrant cancer metabolism.

  View details for DOI 10.1158/1078-0432.CCR-21-0544

  View details for PubMedID 34475101

• Pulsed focused ultrasound enhances the therapeutic effect of mesenchymal stromal cell-derived extracellular vesicles in acute kidney injury. Stem cell research & therapy Ullah, M., Liu, D. D., Rai, S., Razavi, M., Concepcion, W., Thakor, A. S. 2020; 11 (1): 398

  Abstract

  BACKGROUND: Acute kidney injury (AKI) is characterized by rapid failure of renal function and has no curative therapies. Mesenchymal stromal cell (MSC)-derived extracellular vesicles (EVs) are known to carry therapeutic factors, which have shown promise in regenerative medicine applications, including AKI. However, there remains an unmet need to optimize their therapeutic effect. One potential avenue of optimization lies in pulsed focused ultrasound (pFUS), where tissues-of-interest are treated with sound waves. pFUS has been shown to enhance MSC therapy via increased cell homing, but its effects on cell-free EV therapy remain largely unexplored.METHODS: We combine pFUS pretreatment of the kidney with MSC-derived EV therapy in a mouse model of cisplatin-induced AKI.RESULTS: EVs significantly improved kidney function, reduced injury markers, mediated increased proliferation, and reduced inflammation and apoptosis. While pFUS did not enhance EV homing to the kidney, the combined treatment resulted in a superior therapeutic effect compared to either treatment alone. We identified several molecular mechanisms underlying this synergistic therapeutic effect, including upregulation of proliferative signaling (MAPK/ERK, PI3K/Akt) and regenerative pathways (eNOS, SIRT3), as well as suppression of inflammation.CONCLUSION: Taken together, pFUS may be a strategy for enhancing the therapeutic efficacy of MSC-derived EV treatment for the treatment of AKI.

  View details for DOI 10.1186/s13287-020-01922-1

  View details for PubMedID 32928310

• The role of ultrasound in enhancing mesenchymal stromal cell-based therapies. Stem cells translational medicine Liu, D. D., Ullah, M., Concepcion, W., Dahl, J. J., Thakor, A. S. 2020

  Abstract

  Mesenchymal stromal cells (MSCs) have been a popular platform for cell-based therapy in regenerative medicine due to their propensity to home to damaged tissue and act as a repository of regenerative molecules that can promote tissue repair and exert immunomodulatory effects. Accordingly, a great deal of research has gone into optimizing MSC homing and increasing their secretion of therapeutic molecules. A variety of methods have been used to these ends, but one emerging technique gaining significant interest is the use of ultrasound. Sound waves exert mechanical pressure on cells, activating mechano-transduction pathways and altering gene expression. Ultrasound has been applied both to cultured MSCs to modulate self-renewal and differentiation, and to tissues-of-interest to make them a more attractive target for MSC homing. Here, we review the various applications of ultrasound to MSC-based therapies, including low-intensity pulsed ultrasound, pulsed focused ultrasound, and extracorporeal shockwave therapy, as well as the use of adjunctive therapies such as microbubbles. At a molecular level, it seems that ultrasound transiently generates a local gradient of cytokines, growth factors, and adhesion molecules that facilitate MSC homing. However, the molecular mechanisms underlying these methods are far from fully elucidated and may differ depending on the ultrasound parameters. We thus put forth minimal criteria for ultrasound parameter reporting, in order to ensure reproducibility of studies in the field. A deeper understanding of these mechanisms will enhance our ability to optimize this promising therapy to assist MSC-based approaches in regenerative medicine.

  View details for DOI 10.1002/sctm.19-0391

  View details for PubMedID 32157802

• Mesenchymal Stromal Cell Homing: Mechanisms and Strategies for Improvement. iScience Ullah, M. n., Liu, D. D., Thakor, A. S. 2019; 15: 421–38

  Abstract

  Mesenchymal stromal cells (MSCs) have been widely investigated for their therapeutic potential in regenerative medicine, owing to their ability to home damaged tissue and serve as a reservoir of growth factors and regenerative molecules. As such, clinical applications of MSCs are reliant on these cells successfully migrating to the desired tissue following their administration. Unfortunately, MSC homing is inefficient, with only a small percentage of cells reaching the target tissue following systemic administration. This attrition represents a major bottleneck in realizing the full therapeutic potential of MSC-based therapies. Accordingly, a variety of strategies have been employed in the hope of improving this process. Here, we review the molecular mechanisms underlying MSC homing, based on a multistep model involving (1) initial tethering by selectins, (2) activation by cytokines, (3) arrest by integrins, (4) diapedesis or transmigration using matrix remodelers, and (5) extravascular migration toward chemokine gradients. We then review the various strategies that have been investigated for improving MSC homing, including genetic modification, cell surface engineering, in vitro priming of MSCs, and in particular, ultrasound techniques, which have recently gained significant interest. Contextualizing these strategies within the multistep homing model emphasizes that our ability to optimize this process hinges on our understanding of its molecular mechanisms. Moving forward, it is only with a combined effort of basic biology and translational work that the potential of MSC-based therapies can be realized.

  View details for DOI 10.1016/j.isci.2019.05.004

  View details for PubMedID 31121468

• Trends in the characteristics of human functional genomic data on the gene expression omnibus, 2001-2017. Laboratory investigation; a journal of technical methods and pathology Liu, D. D., Zhang, L. 2019; 99 (1): 118-127

  Abstract

  The gene expression omnibus (GEO) is the world's largest public repository of functional genomic data. Despite its broad use in secondary genomic analyses, the temporal trends in the characteristics of genomic data on GEO, including experimental procedures, geographic origin, funder(s), and related disease, have not been examined. We identified 75,376 Series deposited to the GEO during 2001-2017 and built a database of all human genomic data (39,076 Series, 51.8% of all Series). Using the associated publications, we obtained funding information and identified the related disease area. Of the Series with classified disease areas, the two most common were cancer (n = 12,688, 32.5%) and immunologic diseases (n = 2,393, 6.1%), while the percentages of all other disease areas were below 5%, including neurological diseases (n = 1733, 4.4%), infectious diseases (n = 1225, 3.1%), diabetes (n = 828, 2.1%), and cardiovascular diseases (n = 299, 0.8%). In recent years, there has been a significant increase in the use of high-throughput sequencing (HTS), protein array and multiple-platform technologies, as well as in the proportion of North American deposits. Compared to those from other regions, North American deposits appeared to lead the shift from array-based to HTS technologies (odds ratio [OR], 95% confidence intervals [CI] = 3.39, 3.23-3.55, P = 9.40E-323), and were less likely to focus on a major disease area (OR = 0.64, 95% CI: 0.61-0.67, P = 5.02E-107), suggesting a greater emphasis on basic science in North America. Furthermore, the Series utilizing HTS were less likely to be disease-classified compared to other technologies (OR = 0.39, 95% CI: 0.37-0.41, P = 1.00E-322), suggesting a preferential use or adoption of HTS in basic science settings. Finally, funding from the NHGRI, NCI, NIEHS, and NCCR resulted in a higher number of GEO Series per grant than other NIH institutes, demonstrating different preferences on genomic studies among awardees of NIH institutes. Our findings demonstrate geographic, technological, and funding disparities in the trends of GEO deposit characteristics.

  View details for DOI 10.1038/s41374-018-0125-5

  View details for PubMedID 30206311

• Hysteresis control of epithelial-mesenchymal transition dynamics conveys a distinct program with enhanced metastatic ability NATURE COMMUNICATIONS Celia-Terrassa, T., Bastian, C., Liu, D., Ell, B., Aiello, N. M., Wei, Y., Zamalloa, J., Blanco, A. M., Hang, X., Kunisky, D., Li, W., Williams, E. D., Rabitz, H., Kang, Y. 2018; 9: 5005

  Abstract

  Epithelial-mesenchymal transition (EMT) have been extensively characterized in development and cancer, and its dynamics have been modeled as a non-linear process. However, less is known about how such dynamics may affect its biological impact. Here, we use mathematical modeling and experimental analysis of the TGF-β-induced EMT to reveal a non-linear hysteretic response of E-cadherin repression tightly controlled by the strength of the miR-200s/ZEBs negative feedback loop. Hysteretic EMT conveys memory state, ensures rapid and robust cellular response and enables EMT to persist long after withdrawal of stimuli. Importantly, while both hysteretic and non-hysteretic EMT confer similar morphological changes and invasive potential of cancer cells, only hysteretic EMT enhances lung metastatic colonization efficiency. Cells that undergo hysteretic EMT differentially express subsets of stem cell and extracellular matrix related genes with significant clinical prognosis value. These findings illustrate distinct biological impact of EMT depending on the dynamics of the transition.

  View details for DOI 10.1038/s41467-018-07538-7

  View details for Web of Science ID 000451310400010

  View details for PubMedID 30479345

  View details for PubMedCentralID PMC6258667

• Ets2 anchors the prometastatic function of mutant p53 in osteosarcoma. Genes & development Liu, D. D., Kang, Y. 2017; 31 (18): 1823-1824

  Abstract

  Mutations in the tumor suppressor p53 occur in a majority of human cancers. Some gain-of-function (GOF) p53 mutations endow tumor cells with increased metastatic ability, although our understanding of the underlying mechanism remains incomplete. In this issue of Genes & Development, Pourebrahim and colleagues (pp. 1847-1857) develop a new mouse model of osteosarcoma in which a GOF mutant p53 allele is expressed specifically in osteoblasts, while the tumor microenvironment remains wild type for p53, allowing for the study of cell-autonomous functions. In this model, the role of GOF mutant p53 in promoting lung metastasis is shown to be critically dependent on the transcription factor Ets2 and is accompanied by the elevated expression of a cluster of small nucleolar RNAs (snoRNAs).

  View details for DOI 10.1101/gad.307439.117

  View details for PubMedID 29051386

  View details for PubMedCentralID PMC5695082

• Identification of Nidogen 1 as a lung metastasis protein through secretome analysis. Genes & development Alečković, M., Wei, Y., LeRoy, G., Sidoli, S., Liu, D. D., Garcia, B. A., Kang, Y. 2017; 31 (14): 1439-1455

  Abstract

  Secreted proteins play crucial roles in mediating tumor-stroma interactions during metastasis of cancer to different target organs. To comprehensively profile secreted proteins involved in lung metastasis, we applied quantitative mass spectrometry-based proteomics and identified 392 breast cancer-derived and 302 melanoma-derived proteins secreted from highly lung metastatic cells. The cancer-specific lung metastasis secretome signatures (LMSSs) displayed significant prognostic value in multiple cancer clinical data sets. Moreover, we observed a significant overlap of enriched pathways between the LMSSs of breast cancer and melanoma despite an overall small overlap of specific proteins, suggesting that common biological processes are executed by different proteins to enable the two cancer types to metastasize to the lung. Among the novel candidate lung metastasis proteins, Nidogen 1 (NID1) was confirmed to promote lung metastasis of breast cancer and melanoma, and its expression is correlated with poor clinical outcomes. In vitro functional analysis further revealed multiple prometastatic functions of NID1, including enhancing cancer cell migration and invasion, promoting adhesion to the endothelium and disrupting its integrity, and improving vascular tube formation capacity. As a secreted prometastatic protein, NID1 may be developed as a new biomarker for disease progression and therapeutic target in breast cancer and melanoma.

  View details for DOI 10.1101/gad.301937.117

  View details for PubMedID 28827399

  View details for PubMedCentralID PMC5588926

• Normal and cancerous mammary stem cells evade interferon-induced constraint through the miR-199a-LCOR axis NATURE CELL BIOLOGY Celia-Terrassa, T., Liu, D. D., Choudhury, A., Hang, X., Wei, Y., Zamalloa, J., Alfaro-Aco, R., Chakrabarti, R., Jiang, Y., Koh, B., Smith, H. A., DeCoste, C., Li, J., Shao, Z., Kang, Y. 2017; 19 (6): 711-+

  Abstract

  Tumour-initiating cells, or cancer stem cells (CSCs), possess stem-cell-like properties observed in normal adult tissue stem cells. Normal and cancerous stem cells may therefore share regulatory mechanisms for maintaining self-renewing capacity and resisting differentiation elicited by cell-intrinsic or microenvironmental cues. Here, we show that miR-199a promotes stem cell properties in mammary stem cells and breast CSCs by directly repressing nuclear receptor corepressor LCOR, which primes interferon (IFN) responses. Elevated miR-199a expression in stem-cell-enriched populations protects normal and malignant stem-like cells from differentiation and senescence induced by IFNs that are produced by epithelial and immune cells in the mammary gland. Importantly, the miR-199a-LCOR-IFN axis is activated in poorly differentiated ER- breast tumours, functionally promotes tumour initiation and metastasis, and is associated with poor clinical outcome. Our study therefore reveals a common mechanism shared by normal and malignant stem cells to protect them from suppressive immune cytokine signalling.

  View details for DOI 10.1038/ncb3533

  View details for Web of Science ID 000402525200018

  View details for PubMedID 28530657

  View details for PubMedCentralID PMC5481166