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Figure S1: Additional analyses of NP and NPp53 GEM models Figure S2: Tolerance and efficacy of abiraterone in mouse prostate Figure S3: Additional analyses of tumor volumes in GEM models before and after treatment Figure S4: Analyses of abiraterone treatment of cell lines from NP and NPp53 tumors Figure S5: Examples of alternative histopathologies of NPp53 CRPC Figure S6: Additional cross-species computational analyses Figure S7: Additional analyses of expression of SOX11 and other NE genes Table S1: Description of human datasets used in this study Table S2: Summary of preclinical phenotypic analyses Table S3: Summary of histopathology of individual mouse tumors Table S4: List of master regulators Table S5: Quantitation of synaptophysin-positive cells in mouse CRPC Table S6: List of antibodies used in this study Table S7: List of primers and shRNA used in this study Detailed experimental procedures

Differentially expressed genes for mouse CRPC tumors

PDF file, 3314K, Representative IHC analysis of ALDH1A1 expression in large tissues prostate sections.

PDF file, 46K, Analysis of the expression of ALDH specific isoforms in BPH and PCa surgical specimens.

PDF file, 39K, Characteristics of PCa patients.

PDF file, 1824K, Differential expression and localization of ALDH1A1 and ALDH7A1 proteins in BPH.

PDF file, 41K.

Purpose: High aldehyde dehydrogenase (ALDH) has been suggested to selectively mark cells with high tumorigenic potential in established prostate cancer cell lines. However, the existence of cells with high ALDH activity (ALDHbright) in primary prostate cancer specimens has not been shown so far. We investigated the presence, phenotype, and clinical significance of ALDHbright populations in clinical prostate cancer specimens.Experimental Design: We used ALDEFLUOR technology and fluorescence-activated cell-sorting (FACS) staining to identify and characterize ALDHbright populations in cells freshly isolated from clinical prostate cancer specimens. Expression of genes encoding ALDH-specific isoforms was evaluated by quantitative real-time PCR in normal prostate, benign prostatic hyperplasia (BPH), and prostate cancer tissues. ALDH1A1-specific expression and prognostic significance were assessed by staining two tissue microarrays that included more than 500 samples of BPH, prostatic intraepithelial neoplasia (PIN), and multistage prostate cancer.Results: ALDHbright cells were detectable in freshly excised prostate cancer specimens (n = 39) and were mainly included within the EpCAM(+) and Trop2(+) cell populations. Although several ALDH isoforms were expressed to high extents in prostate cancer, only ALDH1A1 gene expression significantly correlated with ALDH activity (P < 0.01) and was increased in cancers with high Gleason scores (P = 0.03). Most importantly, ALDH1A1 protein was expressed significantly more frequently and at higher levels in advanced-stage than in low-stage prostate cancer and BPH. Notably, ALDH1A1 positivity was associated with poor survival (P = 0.02) in hormone-naïve patients.Conclusions: Our data indicate that ALDH contributes to the identification of subsets of prostate cancer cells of potentially high clinical relevance. Clin Cancer Res; 19(19); 5361–71. ©2013 AACR.

Item does not contain fulltext Patient-derived xenografts (PDXs) are generated by engrafting human tumours into mice. Serially transplantable PDXs are used to study tumour biology and test therapeutics, linking the laboratory to the clinic. Although few prostate cancer PDXs are available in large repositories, over 330 prostate cancer PDXs have been established, spanning broad clinical stages, genotypes and phenotypes. Nevertheless, more PDXs are needed to reflect patient diversity, and to study new treatments and emerging mechanisms of resistance. We can maximize the use of PDXs by exchanging models and datasets, and by depositing PDXs into biorepositories, but we must address the impediments to accessing PDXs, such as institutional, ethical and legal agreements. Through collaboration, researchers will gain greater access to PDXs representing diverse features of prostate cancer. 01 juni 2023

Introduction: Muscular Invasive Bladder Cancer (MIBC) is a highly heterogeneous disease. A consensus classification, based on gene expression profiling, has identified six MIBC subgroups, ranging from basal/squamous (Ba/Sq) to luminal papillary (LumP). This classification is a static representation of the disease and does not account for cellular plasticity, which may induce phenotype switch and cancer progression. Here, we aimed at identifying factors involved in cellular plasticity using bladder cancer patient-derived organoids (PDO) models. Methods: Bladder cancer RNA sequencing data from bulk and single cell experiments were downloaded from public repositories. Samples from both datasets were classified using the consensus molecular classification. Pathway activation scores and cell-cell interaction scores were calculated for each subtype. Differential gene expression analysis was performed with the R packages edgeR and Seurat. MIBC samples were collected from bladder cancer patients undergoing surgical treatment. The samples were mechanically and enzymatically digested to generate a cell suspension which was seeded in Matrigel and immersed in growth-factor supplemented medium. Phenotypical characterization of long-term PDO lines was performed through immunofluorescence (IF). The PDOs were classified as Luminal, Basal or Mixed, based on the number of cells expressing the luminal marker CK8 or the basal marker CK5. Three selected organoids lines were cultured in alginate, immersed in a serum-free chemically-defined medium. Results: Long-term PDO lines were successfully generated from 6 out of 13 MIBC samples. Organoids from the same line showed a consistent expression of basal and luminal markers. One line was classified as Basal, 2 lines as Mixed and 3 lines as Luminal. Exploring bulk RNAseq and scRNAseq datasets, we identified several genes whose expression was significantly altered in Ba/Sq and LumP MIBC subtypes. In particular, SHH and related BMP genes were significantly less expressed in Ba/Sq samples than in LumP. In addition, we observed an overexpression of BMP3 in LumP samples and of TGFβ2 in Ba/Sq samples. As preliminary test, we formulated a chemically-defined medium containing inhibitors of the proteins encoded by the identified genes (TGFβ inhibitor and Noggin); using this medium, we were able to maintain organoids from a Luminal, Basal and Mixed subtype in culture for up to two weeks. Interestingly, after two weeks of culture, IF analysis highlighted the presence of cells expressing luminal markers in the basal organoid line, suggesting culture-induced change of phenotype. Conclusions: Bladder cancer PDOs represent promising models to study MIBC plasticity. The integration of RNA sequencing data and the successful culture of PDOs in chemically-defined culture conditions may allow to identify candidate factors impacting the phenotype of bladder cancer cells. Citation Format: Michele Garioni, Luca Roma, Renaud Mével, Tatjana Vlajnic, Heike Pueschel, Dagmar Iber, Hans-Helge Seifert, Cyrill A. Rentsch, Lukas Bubendorf, Clémentine Le Magnen. Bladder cancer patient-derived organoids to decipher cellular plasticity and cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3073.

Introduction: An important clinical need remains to improve survival of men with advanced metastatic prostate cancer (PCa). A better understanding of the mechanisms underlying PCa progression and treatment resistance is a prerequisite to address this challenge, which is currently limited by the scarcity of experimental models. Here, we aimed at establishing and characterizing new patient-derived organoids xenograft (PDOX) models of advanced PCa. Materials & methods: We used two patient-derived organoid lines (PDOs), previously generated in our laboratory. P20-11 PDOs were derived from a lung metastasis obtained from a patient with hormone-naïve PCa. P20-23 PDOs were derived from a transurethral prostate resection obtained from a patient with metastatic castration-resistant PCa, previously treated with goserelin, docetaxel, and enzalutamide. PDOX were generated by subcutaneous injection of PDOs in NOD scid gamma (NSG) male mice. Organoids were derived from the PDOX tumors (PDOX-O). Matched patients’ tumor, PDOs, PDOX and PDOX-O samples were characterized using immunohistochemistry (IHC), immunofluorescence (IF), and whole exome sequencing (WES). Organoids were treated with different concentrations of drugs and cell viability was measured using CellTiter-Glo 3D, after a 5-day long treatment. Results: P20-11 PDOs developed tumors in 2 out of 6 mice, 10 months after injection. IHC analysis highlighted a loss of PTEN expression, overexpression of ERG and P53, as well as strong AR and NKX3.1 expression in the tumor, PDOs, PDOX and PDOX-O samples. WES analysis uncovered mutations in CTNNB1, PTEN and TP53 in all samples, and approximately 87% of shared non-synonymous mutations between the PDOX and the original patients’ tumor. Similar to the original PDOs, P20-11 PDOX-O displayed androgen sensitivity in vitro. P20-23 PDOs formed tumors in 3 out of 3 mice 8 months following injection. IHC and IF analyses highlighted a strong expression of CK8, PSMA, AR and NKX3.1, as well as a loss of PTEN expression in the tumor, PDOs, PDOX and PDOX-O samples. WES identified a pathogenic mutation in the PCa-associated gene ZMYM3 in all samples, associated with loss of protein expression. The activating AR point mutation L702H, previously linked to AR signaling inhibitors resistance, was detected in the tumor and in 2 out of 3 PDOX and their derived organoids. Overall, an average of 80% of non-synonymous mutations were shared between the patients’ tumor and PDOX. Finally, P20-23 PDOX-O did not respond to docetaxel or enzalutamide but exhibited sensitivity to the PI3K/AKT inhibitor ipatasertib. Conclusion: We have successfully generated two novel PDOX models, which highly resemble the original patients’ tumor and can be further cultured as organoids. These models are representative of relevant clinical and molecular subtypes of advanced PCa, providing further opportunities for translational studies. Citation Format: Raphaëlle Servant, Zoi Diamantopoulou, Michele Garioni, Luca Roma, Tatjana Vlajnic, Arnoud J. Templeton, Heike Pueschel, Salvatore Piscuoglio, Nicola Aceto, Helge Seifert, Cyrill A. Rentsch, Bubendorf Lukas, Clémentine Le Magnen. Establishment and characterization of two novel patient-derived organoid xenograft models of advanced prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3085.