Prostate Cancer Patient-derived Xenograft Models Research Articles

Patient-Derived Xenograft Models for Translational Prostate Cancer Research and Drug Development.

Patient-derived xenografts (PDXs) are a valuable preclinical research platform generated through transplantation of a patient's resected tumor into an immunodeficient or humanized mouse. PDXs serve as a high-fidelity avatar for both precision medicine and therapeutic testing against the cancer patient's disease state. While PDXs show mixed response to initial establishment, those that successfully engraft and can be sustained with serial passaging form a useful tool for basic and translational prostate cancer (PCa) research. While genetically engineered mouse (GEM) models and human cancer cell lines, and their xenografts, each play beneficial roles in discovery science and initial drug screening, PDX tumors are emerging as the gold standard approach for therapeutic proof-of-concept prior to entering clinical trial. PDXs are a powerful platform, with PCa PDXs shown to represent the original patient tumor cell population and architecture, histopathology, genomic and transcriptomic landscape, and heterogeneity. Furthermore, PDX response to anticancer drugs in mice has been closely correlated to the original patient's susceptibility to these treatments in the clinic. Several PDXs have been established and have undergone critical in-depth characterization at the cellular and molecular level across multiple PCa tumor subtypes representing both primary and metastatic patient tumors and their inherent levels of androgen responsiveness and/or treatment resistance, including androgen-sensitive, castration resistant, and neuroendocrine PCa. Multiple PDX networks and repositories have been generated for the collaborative and shared use of these vital translational cancer tools. Here we describe the creation of a PDX maintenance colony from an established well-characterized PDX, best practice for PDX maintenance in mice, and their subsequent application in preclinical drug testing. This chapter aims to serve as a go to resource for the preparation and adoption of PCa PDX models in the research laboratory and for their use as a valuable preclinical platform for translational research and therapeutic agent development.

Introduction of Androgen Receptor Targeting shRNA Inhibits Tumor Growth in Patient-Derived Prostate Cancer Xenografts.

Patient-derived xenograft (PDX) models have been established as important preclinical cancer models, overcoming some of the limitations associated with the use of cancer cell lines. The utility of prostate cancer PDX models has been limited by an inability to genetically manipulate them in vivo and difficulties sustaining PDX-derived cancer cells in culture. Viable, short-term propagation of PDX models would allow in vitro transfection with traceable reporters or manipulation of gene expression relevant to different studies within the prostate cancer field. Here, we report an organoid culture system that supports the growth of prostate cancer PDX cells in vitro and permits genetic manipulation, substantially increasing the scope to use PDXs to study the pathobiology of prostate cancer and define potential therapeutic targets. We have established a short-term PDX-derived in vitro cell culture system which enables genetic manipulation of prostate cancer PDXs LuCaP35 and BM18. Genetically manipulated cells could be re-established as viable xenografts when re-implanted subcutaneously in immunocompromised mice and were able to be serially passaged. Tumor growth of the androgen-dependent LuCaP35 PDX was significantly inhibited following depletion of the androgen receptor (AR) in vivo. Taken together, this system provides a method to generate novel preclinical models to assess the impact of controlled genetic perturbations and allows for targeting specific genes of interest in the complex biological setting of solid tumors.

Identification of the α2 chain of interleukin-13 receptor as a potential biomarker for predicting castration resistance of prostate cancer using patient-derived xenograft models.

Several treatment strategies use upfront chemotherapy or androgen receptor axis-targeting therapies for metastatic prostate cancer. However, there are no useful biomarkers for selecting appropriate patients who urgently require these treatments. Novel patient-derived xenograft (PDX) castration-sensitive and -resistant models were established and gene expression patterns were comprehensively compared. The function of a gene highly expressed in the castration-resistant models was evaluated by its overexpression in LNCaP prostate cancer cells. Protein expression in the tumors and serum of patients was examined by immunohistochemistry and ELISA, and correlations with castration resistance were analyzed. Expression of the α2 chain of interleukin-13 receptor (IL13Rα2) was higher in castration-resistant PDX tumors. LNCaP cells overexpressing IL13Rα2 acquired castration resistance in vitro and in vivo. In tissue samples, IL13Rα2 expression levels were significantly associated with castration-resistant progression (p < 0.05). In serum samples, IL13Rα2 levels could be measured in 5 of 28 (18%) castration-resistant prostate cancer patients. IL13Rα2 was highly expressed in castration-resistant prostate cancer PDX models and was associated with the castration resistance of prostate cancer cells. It might be a potential tissue and serum biomarker for predicting castration resistance in prostate cancer patients.

Selected Articles from This Issue

Highlights| May 04 2022 Selected Articles from This Issue Author & Article Information Online Issn: 1557-3125 Print Issn: 1541-7786 ©2022 American Association for Cancer Research2022American Association for Cancer Research Mol Cancer Res (2022) 20 (5): 671. https://doi.org/10.1158/1541-7786.MCR-20-5-HI Related Content A commentary has been published: Modeling Androgen Deprivation Therapy–Induced Prostate Cancer Dormancy and Its Clinical Implications A commentary has been published: Targeted Rejuvenation of Exhausted Chimeric Antigen Receptor T Cells Regresses Refractory Solid Tumors A commentary has been published: Inactivation of p21-Activated Kinase 2 (Pak2) Inhibits the Development of Nf2-Deficient Tumors by Restricting Downstream Hedgehog and Wnt Signaling View more A commentary has been published: MOB3A Bypasses BRAF and RAS Oncogene-Induced Senescence by Engaging the Hippo Pathway View less Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Tools Icon Tools Get Permissions Cite Icon Cite Search Site Article Versions Icon Versions Version of Record May 4 2022 Citation Selected Articles from This Issue. Mol Cancer Res 1 May 2022; 20 (5): 671. https://doi.org/10.1158/1541-7786.MCR-20-5-HI Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest Search Advanced Search Androgen-deprivation therapy (ADT) is a frontline therapy for prostate cancer patients. However, many patients experience post-ADT disease recurrence owing to ADT-induced tumor cell dormancy and subsequent dormant tumor cell outgrowth. Elucidating mechanisms underlying ADT-induced tumor cell dormancy could facilitate improved prognostic and therapeutic options for prostate cancer patients, but dormant tumor cells are difficult to study given they are rare and difficult to isolate. To address these challenges, Dong and colleagues used prostate cancer patient-derived xenograft models to derive and study ADT-induced tumor cell dormancy. Immunohistochemical analysis revealed two subtypes of tumor dormancy: tumor mass dormancy and cellular dormancy. Transcriptomic analysis of samples from each subtype defined a corresponding signature of differentially expressed genes—termed the pre-disposed gene signature (PGS)—between the subtypes. Training and validation of the PGS using clinical samples demonstrated the ability of the PGS to predict ADT responsiveness, suggesting molecular characteristics underlying tumor dormancy subtypes discovered in this... You do not currently have access to this content.

Circulating Tumour Cells Indicate the Presence of Residual Disease Post-Castration in Prostate Cancer Patient-Derived Xenograft Models

Castrate-resistant prostate cancer (CRPC) is the lethal form of prostate cancer. Epithelial mesenchymal plasticity (EMP) has been associated with disease progression to CRPC, and prostate cancer therapies targeting the androgen signalling axis, including androgen deprivation therapy (ADT), promote EMP. We explored effects of castration on EMP in the tumours and circulating tumour cells (CTCs) of patient-derived xenograft (PDX)-bearing castrated mice using human-specific RT-qPCR assays and immunocytochemistry. Expression of prostate epithelial cell marker KLK3 was below detection in most tumours from castrated mice (62%, 23/37 mice), consistent with its known up-regulation by androgens. Endpoint tumour size after castration varied significantly in a PDX model-specific pattern; while most tumours were castration-sensitive (BM18, LuCaP70), the majority of LuCaP105 tumours continued to grow following castration. By contrast, LuCaP96 PDX showed a mixed response to castration. CTCs were detected in 33% of LuCaP105, 43% of BM18, 47% of LuCaP70, and 54% of LuCaP96 castrated mice using RPL32 mRNA measurement in plasma. When present, CTC numbers estimated using human RPL32 expression ranged from 1 to 458 CTCs per ml blood, similar to our previous observations in non-castrated mice. In contrast to their non-castrated counterparts, there was no relationship between tumour size and CTC burden in castrated mice. Unsupervised hierarchical clustering of the gene expression profiles of CTCs collected from castrated and non-castrated mice revealed distinct CTC sub-groups within the pooled population that were classified as having mesenchymal, epithelial, or EMP hybrid gene expression profiles. The epithelial signature was only found in CTCs from non-castrated mice. Hybrid and mesenchymal signatures were detected in CTCs from both castrated and non-castrated mice, with an emphasis towards mesenchymal phenotypes in castrated mice. Post-castration serum PSA levels were either below detection or very low for all the CTC positive samples highlighting the potential usefulness of CTCs for disease monitoring after androgen ablation therapy. In summary, our study of castration effects on prostate cancer PDX CTCs showed that CTCs were often detected in the castrate setting, even in mice with no palpable tumours, and demonstrated the superior ability of CTCs to reveal residual disease over the conventional clinical biomarker serum PSA.

Altered glucuronidation deregulates androgen dependent response profiles and signifies castration resistance in prostate cancer.

Glucuronidation controls androgen levels in the prostate and the dysregulation of enzymes in this pathway is associated with castration resistant prostate cancer. UDP-glucose dehydrogenase (UGDH) produces UDP-glucuronate, the essential precursor for glucuronidation, and its expression is elevated in prostate cancer. We compared protein and metabolite levels relevant to the glucuronidation pathway in five prostate cancer patient-derived xenograft models paired with their isogenic counterparts that were selected in vivo for castration resistant (CR) recurrence. All pairs showed changes in UGDH and associated enzymes and metabolites that were consistent with those we found in an isogenic androgen dependent (AD) and CR LNCaP prostate cancer model. Ectopic overexpression of UGDH in LNCaP AD cells blunted androgen-dependent gene expression, increased proteoglycan synthesis, significantly increased cell growth compared to controls, and eliminated dose responsive growth suppression with enzalutamide treatment. In contrast, the knockdown of UGDH diminished proteoglycans, suppressed androgen dependent growth irrespective of androgens, and restored androgen sensitivity in CR cells. Importantly, the knockdown of UGDH in both LNCaP AD and CR cells dramatically sensitized these cells to enzalutamide. These results support a role for UGDH in androgen responsiveness and a target for therapeutic strategies in advanced prostate cancer.

Diversity of Epithelial-Mesenchymal Phenotypes in Circulating Tumour Cells from Prostate Cancer Patient-Derived Xenograft Models.

Simple SummarySpread of prostate cancer to other parts of the body is responsible for the majority of deaths. Tumour cell epithelial mesenchymal plasticity (EMP) increases their metastatic potential and facilitates their survival in the blood as circulating tumour cells (CTCs). The aim of this study was to molecularly characterise CTCs in a panel of prostate cancer patient-derived xenografts using genes associated with epithelial and mesenchymal phenotypes, and to compare the EMP status of CTCs with their matched primary tumours. The study highlights high heterogeneity in CTC enumeration and EMP gene expression between tumour-bearing mice and within individual blood samples, and therefore caution should be taken when interpreting pooled CTC analyses. Critically, tumour cells were present in the epithelial-mesenchymal hybrid state in the circulation. The study also demonstrates that there is high variation in CTC size, which would introduce sample bias to size-based CTC isolation techniques.Metastasis is the leading cause of cancer-related deaths worldwide. The epithelial-mesenchymal plasticity (EMP) status of primary tumours has relevance to metastatic potential and therapy resistance. Circulating tumour cells (CTCs) provide a window into the metastatic process, and molecular characterisation of CTCs in comparison to their primary tumours could lead to a better understanding of the mechanisms involved in the metastatic cascade. In this study, paired blood and tumour samples were collected from four prostate cancer patient-derived xenograft (PDX) models (BM18, LuCaP70, LuCaP96, LuCaP105) and assessed using an EMP-focused, 42 gene human-specific, nested quantitative RT-PCR assay. CTC burden varied amongst the various xenograft models with LuCaP96 having the highest number of CTCs per mouse (mean: 704; median: 31) followed by BM18 (mean: 101; median: 21), LuCaP70 (mean: 73; median: 16) and LuCaP105 (mean: 57; median: 6). A significant relationship was observed between tumour size and CTC number (p = 0.0058). Decreased levels of kallikrein-related peptidase 3 (KLK3) mRNA (which encodes prostate-specific antigen; PSA) were observed in CTC samples from all four models compared to their primary tumours. Both epithelial- and mesenchymal-associated genes were commonly expressed at higher levels in CTCs compared to the bulk primary tumour, although some common EMT-associated genes (CDH1, VIM, EGFR, EPCAM) remained unchanged. Immunofluorescence co-staining for pan-cytokeratin (KRT) and vimentin (VIM) indicated variable proportions of CTCs across the full EMP axis, even in the same model. EMP hybrids predominated in the BM18 and LuCaP96 models, but were not detected in the LuCaP105 model, and variable numbers of KRT+ and human VIM+ cells were observed in each model. SERPINE1, which encodes plasminogen activator inhibitor-1 (PAI-1), was enriched at the RNA level in CTCs compared to primary tumours and was the most commonly expressed mesenchymal gene in the CTCs. Co-staining for SERPINE1 and KRT revealed SERPINE1+ cells in 7/11 samples, six of which had SERPINE+KRT+ CTCs. Cell size variation was observed in CTCs. The majority of samples (8/11) contained larger CTCs ranging from 15.3 to 37.8 µm, whilst smaller cells (10.7 ± 4.1 µm, similar in size to peripheral blood mononuclear cells (PBMCs)) were identified in 6 of 11 samples. CTC clusters were also identified in 9/11 samples, containing 2–100 CTCs per cluster. Where CTC heterogeneity was observed in the clusters, epithelial-like cells (KRT+VIM−) were located on the periphery of the cluster, forming a layer around hybrid (KRT+VIM+) or mesenchymal-like (KRT−VIM+) cells. The CTC heterogeneity observed in these models emphasises the complexity in CTC isolation and classification and supports the increasingly recognised importance of the epithelial-mesenchymal hybrid state in cancer progression and metastasis.

Synthetic Lethal Metabolic Targeting of Androgen-Deprived Prostate Cancer Cells with Metformin.

The initiation of androgen-deprivation therapy (ADT) induces susceptibilities in prostate cancer cells that make them vulnerable to synergistic treatment and enhanced cell death. Senescence results in cell-cycle arrest, but cells remain viable. In this study, we investigated the mechanisms by which prostate cancer cells undergo senescence in response to ADT, and determined whether an FDA-approved antidiabetic drug metformin has a synergistic effect with ADT in prostate cancer both in vitro and in vivo Our results show that longer term exposure to ADT induced senescence associated with p16INK4a and/or p27kip2 induction. The activation of PI3K/AKT and inactivation of AMPK in senescent cells resulted in mTORC1 activation. In addition, the antiapoptotic protein XIAP expression was increased in response to ADT. The addition of metformin following ADT induced apoptosis, attenuated mTOR activation, reduced senescent cell number in vitro, and inhibited tumor growth in prostate cancer patient-derived xenograft models. This study suggests that combining ADT and metformin may be a feasible therapeutic approach to remove persistent prostate cancer cells after ADT.

BRN4 Is a Novel Driver of Neuroendocrine Differentiation in Castration-Resistant Prostate Cancer and Is Selectively Released in Extracellular Vesicles with BRN2.

Neuroendocrine prostate cancer (NEPC), an aggressive variant of castration-resistant prostate cancer (CRPC), often emerges after androgen receptor-targeted therapies such as enzalutamide or de novo, via trans-differentiation process of neuroendocrine differentiation. The mechanistic basis of neuroendocrine differentiation is poorly understood, contributing to lack of effective predictive biomarkers and late disease recognition. The purpose of this study was to examine the role of novel proneural Pit-Oct-Unc-domain transcription factors (TF) in NEPC and examine their potential as noninvasive predictive biomarkers.Experimental Design: Prostate cancer patient-derived xenograft models, clinical samples, and cellular neuroendocrine differentiation models were employed to determine the expression of TFs BRN1 and BRN4. BRN4 levels were modulated in prostate cancer cell lines followed by functional assays. Furthermore, extracellular vesicles (EV) were isolated from patient samples and cell culture models, characterized by nanoparticle tracking analyses, Western blotting, and real-time PCR. We identify for the first time that: (i) BRN4 is amplified and overexpressed in NEPC clinical samples and that BRN4 overexpression drives neuroendocrine differentiation via its interplay with BRN2, a TF that was previously implicated in NEPC; (ii) BRN4 and BRN2 mRNA are actively released in prostate cancer EVs upon neuroendocrine differentiation induction; and (iii) enzalutamide treatment augments release of BRN4 and BRN2 in prostate cancer EVs, promoting neuroendocrine differentiation induction. Our study identifies a novel TF that drives NEPC and suggests that as adaptive mechanism to enzalutamide treatment, prostate cancer cells express and secrete BRN4 and BRN2 in EVs that drive oncogenic reprogramming of prostate cancer cells to NEPC. Importantly, EV-associated BRN4 and BRN2 are potential novel noninvasive biomarkers to predict neuroendocrine differentiation in CRPC.

Establishing a cryopreservation protocol for patient-derived xenografts of prostate cancer.

Serially transplantable patient-derived xenografts (PDXs) are invaluable preclinical models for studying tumor biology and evaluating therapeutic agents. As these models are challenging to establish from prostate cancer specimens, the ability to preserve them through cryopreservation has several advantages for ongoing research. Despite this, there is still uncertainty about the ability to cryopreserve PDXs of prostate cancer. This study compared three different cryopreservation protocols to identify a method that can be used to reproducibly cryopreserve a diverse cohort of prostate cancer PDX models. One serially transplantable prostate cancer PDX from the Melbourne Urological Research Alliance cohort was used to compare three cryopreservation protocols: slow freezing in fetal calf serum (FCS) with 10% dimethyl sulfoxide (DMSO), FCS with 10% DMSO supplemented with the Rho-associated kinase (ROCK) inhibitor Y-27632 and vitrification. The efficiency of the slow freezing protocols was then assessed in 17 additional prostate cancer PDXs. Following cryopreservation, PDXs were re-established in host mice that were either intact and supplemented with testosterone or castrated. Graft take rate, tumor growth, histological features, and transcriptome profiles before and after cryopreservation were compared. Slow freezing maintained the viability and histological features of prostate cancer PDXs, and the addition of a ROCK inhibitor increased their growth following cryopreservation. Using the slow freezing method, we re-established 100% of PDXs grown in either testosterone-supplemented or castrated host mice. Importantly, the long-term tumor growth rate and transcriptome profile were maintained following cryopreservation. This study has identified a protocol to reliably cryopreserve and re-establish a diverse cohort of serially transplantable PDXs of prostate cancer. This study has the potential to significantly improve the practicality of maintaining PDX models. Cryopreservation may also increase the accessibility of these important resources and provide new opportunities for preclinical studies on a broader spectrum of prostate tumors.

Application of Prostate Cancer Models for Preclinical Study: Advantages and Limitations of Cell Lines, Patient-Derived Xenografts, and Three-Dimensional Culture of Patient-Derived Cells.

Various preclinical models have been developed to clarify the pathophysiology of prostate cancer (PCa). Traditional PCa cell lines from clinical metastatic lesions, as exemplified by DU-145, PC-3, and LNCaP cells, are useful tools to define mechanisms underlying tumorigenesis and drug resistance. Cell line-based experiments, however, have limitations for preclinical studies because those cells are basically adapted to 2-dimensional monolayer culture conditions, in which the majority of primary PCa cells cannot survive. Recent tissue engineering enables generation of PCa patient-derived xenografts (PDXs) from both primary and metastatic lesions. Compared with fresh PCa tissue transplantation in athymic mice, co-injection of PCa tissues with extracellular matrix in highly immunodeficient mice has remarkably improved the success rate of PDX generation. PDX models have advantages to appropriately recapitulate the molecular diversity, cellular heterogeneity, and histology of original patient tumors. In contrast to PDX models, patient-derived organoid and spheroid PCa models in 3-dimensional culture are more feasible tools for in vitro studies for retaining the characteristics of patient tumors. In this article, we review PCa preclinical model cell lines and their sublines, PDXs, and patient-derived organoid and spheroid models. These PCa models will be applied to the development of new strategies for cancer precision medicine.

Abstract A004: MEDI3726 (ADCT-401), a novel antibody-drug conjugate targeting PSMA, has potent in vivo antitumor activity in prostate cancer patient-derived xenograft models

Abstract Prostate specific membrane antigen (PSMA), a type II membrane glycoprotein, is highly expressed in nearly all prostate cancers, with the highest expression in metastatic castration-resistant prostate cancer (mCRPC). The prevalence, increased surface expression in prostate tumors, and constitutive internalization make PSMA an attractive target for an antibody-drug conjugate (ADC) approach in anticancer treatment of patients with mCRPC. MEDI3726 (previously known as ADCT-401) is an ADC consisting of an engineered version of anti-PSMA antibody J591, site-specifically conjugated with DNA cross-linking pyrrolobenzodiazepine (PBD) dimer SG3249 for targeting prostatic cancer cells. Using prostate cancer cell line models, we have previously shown that MEDI3726 specifically binds to the extracellular domain of PSMA and, once internalized, releases the PBD dimer to cross link DNA and achieve potent in vitro and in vivo cytotoxicity. Here we investigated the in vivo activity of MEDI3726 in a series of LuCaP prostate cancer patient-derived xenograft (PDX) models. The selected LuCaP models had varying PSMA expression and heterogeneous genetic and phenotypic backgrounds. In agreement with the earlier cell line xenograft data, dose-dependent antitumor activity was observed in PSMA-positive PDX models with durable tumor regressions in models with high PSMA expression. In the PSMA-negative LuCaP 35CR PDX model, MEDI3726 did not have significant antitumor activity, thus highlighting target-mediated in vivo activity. Increased phosphorylation of histone H2AX was observed in xenografts dosed with MEDI3726, confirming DNA damage induced by interstrand cross-linking PBD dimer as the mechanism of antitumor activity of MEDI3726. In summary, MEDI3726 demonstrated potent and specific in vivo antitumor activity, concurrent with DNA damage, in clinically relevant prostate cancer PDX models. MEDI3726 is being evaluated in phase 1 clinical trial as an anticancer treatment in patients with metastatic castrate-resistant prostate cancer (NCT02991911). Citation Format: Song Cho, Francesca Zammarchi, Noel R. Monks, Kapil Vashisht, Ravinder Tammali, Kevin Schifferli, Patrick Strout, Wanda King, Karma Dacosta, Ryan Fleming, David G. Williams, Karin Havenith, Mary Jane Masson Hinrichs, Simon Chivers, Nazzareno Dimasi, Phil W. Howard, John A. Hartley, Steve Coats, Ronald Herbst, Patrick H. van Berkel, David A. Tice. MEDI3726 (ADCT-401), a novel antibody-drug conjugate targeting PSMA, has potent in vivo antitumor activity in prostate cancer patient-derived xenograft models [abstract]. In: Proceedings of the AACR Special Conference: Prostate Cancer: Advances in Basic, Translational, and Clinical Research; 2017 Dec 2-5; Orlando, Florida. Philadelphia (PA): AACR; Cancer Res 2018;78(16 Suppl):Abstract nr A004.

Movember GAP1 PDX project: An international collection of serially transplantable prostate cancer patient-derived xenograft (PDX) models.

While it has been challenging to establish prostate cancer patient-derived xenografts (PDXs), with a take rate of 10-40% and long latency time, multiple groups throughout the world have developed methods for the successful establishment of serially transplantable human prostate cancer PDXs using a variety of immune deficient mice. In 2014, the Movember Foundation launched a Global Action Plan 1 (GAP1) project to support an international collaborative prostate cancer PDX program involving eleven groups. Between these Movember consortium members, a total of 98 authenticated human prostate cancer PDXs were available for characterization. Eighty three of these were derived directly from patient material, and 15 were derived as variants of patient-derived material via serial passage in androgen deprived hosts. A major goal of the Movember GAP1 PDX project was to provide the prostate cancer research community with a summary of both the basic characteristics of the 98 available authenticated serially transplantable human prostate cancer PDX models and the appropriate contact information for collaborations. Herein, we report a summary of these PDX models. PDX models were established in immunocompromised mice via subcutaneous or subrenal-capsule implantation. Dual-label species (ie, human vs mouse) specific centromere and telomere Fluorescence In Situ Hybridization (FISH) and immuno-histochemical (IHC) staining of tissue microarrays (TMAs) containing replicates of the PDX models were used for characterization of expression of a number of phenotypic markers important for prostate cancer including AR (assessed by IHC and FISH), Ki67, vimentin, RB1, P-Akt, chromogranin A (CgA), p53, ERG, PTEN, PSMA, and epithelial cytokeratins. Within this series of PDX models, the full spectrum of clinical disease stages is represented, including androgen-sensitive and castration-resistant primary and metastatic prostate adenocarcinomas as well as prostate carcinomas with neuroendocrine differentiation. The annotated clinical characteristics of these PDXs were correlated with their marker expression profile. Our results demonstrate the clinical relevance of this series of PDXs as a platform for both basic science studies and therapeutic discovery/drug development. The present report provides the prostate cancer community with a summary of the basic characteristics and a contact information for collaborations using these models.

Generation of Prostate Cancer Patient Derived Xenograft Models from Circulating Tumor Cells

Patient derived xenograft (PDX) models are gaining popularity in cancer research and are used for preclinical drug evaluation, biomarker identification, biologic studies, and personalized medicine strategies. Circulating tumor cells (CTC) play a critical role in tumor metastasis and have been isolated from patients with several tumor types. Recently, CTCs have been used to generate PDX experimental models of breast and prostate cancer. This manuscript details the method for the generation of prostate cancer PDX models from CTCs developed by our group. Advantages of this method over conventional PDX models include independence from surgical sample collection and generating experimental models at various disease stages. Density gradient centrifugation followed by red blood cell lysis and flow cytometry depletion of CD45 positive mononuclear cells is used to enrich CTCs from peripheral blood samples collected from patients with metastatic disease. The CTCs are then injected into immunocompromised mice; subsequently generated xenografts can be used for functional studies or harvested for molecular characterization. The primary limitation of this method is the negative selection method used for CTC enrichment. Despite this limitation, the generation of PDX models from CTCs provides a novel experimental model to be applied to prostate cancer research.

Generation of Prostate Cancer Patient Derived Xenograft Models from Circulating Tumor Cells.

Patient derived xenograft (PDX) models are gaining popularity in cancer research and are used for preclinical drug evaluation, biomarker identification, biologic studies, and personalized medicine strategies. Circulating tumor cells (CTC) play a critical role in tumor metastasis and have been isolated from patients with several tumor types. Recently, CTCs have been used to generate PDX experimental models of breast and prostate cancer. This manuscript details the method for the generation of prostate cancer PDX models from CTCs developed by our group. Advantages of this method over conventional PDX models include independence from surgical sample collection and generating experimental models at various disease stages. Density gradient centrifugation followed by red blood cell lysis and flow cytometry depletion of CD45 positive mononuclear cells is used to enrich CTCs from peripheral blood samples collected from patients with metastatic disease. The CTCs are then injected into immunocompromised mice; subsequently generated xenografts can be used for functional studies or harvested for molecular characterization. The primary limitation of this method is the negative selection method used for CTC enrichment. Despite this limitation, the generation of PDX models from CTCs provides a novel experimental model to be applied to prostate cancer research.

Abstract A43: Novel prostate cancer patient-derived xenograft models of bone metastatic castrate-resistant prostate cancer

Abstract Prostate cancer metastasis to bone occurs in 50-90% of men with advanced disease for which there is no cure. Bone metastasis leads to debilitating fractures and severe bone pain. It is associated with disease progression, therapy resistance, poor prognosis, and rapid decline. Androgen ablation therapy is standard of care for advanced prostate cancer, however, the role of androgens in bone metastatic prostate cancer is not understood. The effects of anti-androgens as seen on bone scans can also be inconsistent with the biochemical PSA response. There are few pre-clinical models to understand the interaction between the bone microenvironment and prostate cancer. It is essential to understand the unique interaction of prostate cancer with the bone environment and to develop novel therapies that target these pathways. Here we report the development of novel patient-derived intra-femoral xenograft models of prostate bone metastatic cancer. METHODS: Surgical prostate cancer bone metastasis specimens were transplanted by direct injection into the femurs of Rag2-/-γc-/- mice or sub-cutaneously into the right flank. Patient-derived xenograft (PDX) tumors that grew out were analyzed for prostate cancer biomarker expression using quantitative RT-PCR and immunohistochemistry. Bone lesion formation was measured using micro-computed tomography (μCT). RESULTS: Prostate cancer surgical bone metastasis specimens have been collected from which we have established new serially transplantable, prostate cancer bone metastasis xenograft models – PCSD1, PCSD4 and PCSD5. PCSD1 (Prostate Cancer San Diego 1) was molecularly characterized as advanced, luminal epithelial-type prostate cancer. PCSD1 intra-femoral xenografts formed mixed osteoblastic/osteolytic lesions that closely mimicked those of the patient. Treatment with the anti-androgen, bicalutamide, did not inhibit intra-femoral PCSD1 xenograft growth although there was a decrease in PSA as seen in some patients treated with anti-androgen who had discordant PSA and bone scan tests. CONCLUSION: PCSD1, PCSD4 and PCSD5 are new patient-derived prostate cancer bone metastasis-derived xenograft models. PCSD1 xenograft model closely recapitulates the mixed osteolytic/osteoblastic bone metastatic lesions seen in patients, and we are using it to develop novel therapies for inhibiting prostate cancer growth in the bone-niche. Citation Format: Christina Jamieson, Christina Wu, Amy Strasner, Jason R. Woo, Michelle Muldong, Young B. Jeong, Michael A. Liss, Omer Raheem, Tomonori Yamaguchi, Heather Leu, Deborah Marshall, Sheldon Morris, Nicholas A. Cacalano, Koichi Masuda, Catriona H.M. Jamieson, Anna A. Kulidjian, Christopher J. Kane. Novel prostate cancer patient-derived xenograft models of bone metastatic castrate-resistant prostate cancer. [abstract]. In: Proceedings of the AACR Special Conference: The Translational Impact of Model Organisms in Cancer; Nov 5-8, 2013; San Diego, CA. Philadelphia (PA): AACR; Mol Cancer Res 2014;12(11 Suppl):Abstract nr A43.

Abstract 2030: Culturing patient-derived prostate cancer xenografts using a three-dimensional hydrogel system for drug testing and mechanistic studies

Abstract Advanced prostate cancer (PCa) typically metastasizes to bone and current treatments are mainly palliative. While a mechanistic understanding of PCa progression in bone is needed to develop new therapies, the complex phenotypes in PCa are poorly reflected by cell lines and cell line-based xenograft models, impeding progress in this endeavor. To overcome this limitation, a panel of PCa patient-derived xenograft (PDX) models was established at the University of Texas MD Anderson Cancer Center. While these PDX models better recapitulate the phenotypic signature of the original tumors as compared to cell lines, limitations exist, including the inability to manipulate these cells ex vivo before implantation in vivo and issues associated with the foreign animal physiology. Based on previous studies demonstrating the potential of using hyaluronan (HA)-based hydrogels for the 3D engineering of PCa cell line-derived ectotumors suitable for mechanistic studies, we evaluated the ability of these hydrogels to maintain viability of the PCa PDX tumors in vitro and optimize culture conditions. Using the MDA PCa 183 and 118b PDX tumors as proof-of-concept, tumors freshly harvested were digested and encapsulated within a HA-based hydrogel, comprised of thiol-modified HA crosslinked with poly(ethylene glycol)-diacrylate. Tumor cells remained as multicellular aggregates or ectotumors in 3D, and were viable even up to 2 weeks in culture. Furthermore, cells were positive for Ki-67, indicating that the hydrogel system is able to retain the proliferative capacity of the tumor cells. In probing for the androgen receptor (AR) expression, we found that while the hydrogel-encapsulated MDA PCa 183 cells stained positive for AR, it was not the case for the MDA PCa 118b, as would be expected in vivo, demonstrating that the hydrogel system is also able to preserve critical phenotypic characteristics of the original tumors. Lastly, to evaluate this novel system as a potential platform for rapid drug testing, we treated the 3D PDX ectotumors to docetaxel, and compared their response against a traditional bone metastatic PCa cell line, C4-2B. Interestingly, the 3D PDX ectotumors exhibited a significantly greater resistance to the drug than the C4-2B cells, reminiscent of chemoresistant tumors in vivo. We demonstrate for the first time that PCa PDX tumors are amenable to in vitro culture with the use of a 3D HA-based hydrogel system. The hydrogel supported the maintenance of multicellular ectotumors of PCa cells, which remained viable and phenotypically similar to the tumors of origin. Furthermore, a differential drug response to docetaxel was observed for both the 3D PCa PDX ectotumors tested as compared to the C4-2B cell line, the clinical relevance of which is currently being investigated. This model may have broad applicability for the culture of other bone metastatic cancers and enable mechanistic studies of disease biology. Citation Format: Eliza L.S. Fong, Mariane Martinez, Jun Yang, Leland M.K. Chung, Antonios G. Mikos, Nora M. Navone, Daniel A. Harrington, Mary C. Farach-Carson. Culturing patient-derived prostate cancer xenografts using a three-dimensional hydrogel system for drug testing and mechanistic studies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2030. doi:10.1158/1538-7445.AM2014-2030