Prostate cancer induced in primary human prostate basal cells recapitulates disease initiation and progression in immunodeficient mice.Luminal cells are believed to be the cells-of-origin for human prostate cancer because the disease is characterized by luminal cell expansion and absence of basal cells. Yet functional studies addressing the origin of human prostate cancer have not previously been reported due to a lack of relevant in vivo human models. Here we show that basal cells, from primary benign human prostate tissue, can initiate prostate cancer in immunodeficient mice. The cooperative effects of AKT, ERG, and androgen receptor (AR) in basal cells recapitulated the histological and molecular features of human prostate cancer with loss of basal cells and expansion of luminal cells expressing prostate-specific antigen (PSA) and alpha-methylacylCoA racemase (AMACR). Our results demonstrate that histological characterization of cancers does not necessarily correlate with the cellular origins of the disease.Prostate cancer research has been hindered by an absence of model systems in which the disease is initiated from primary human prostate epithelial cells, precluding investigation of transforming alterations and cells-of-origin. Commonly used human prostate cancer cell lines and xenografts were derived from metastatic lesions. Murine prostate cancer models prohibit testing of species-specific therapies such as monoclonal antibodies against human proteins (1). An ideal model system would be human cell-derived and present as a multifocal disease to accurately represent the heterogeneity of prostate malignancy (2). The

The epithelium of the adult prostate contains 3 distinct cell types: basal, luminal, and neuroendocrine. Tissue-regenerative activity has been identified predominantly from the basal cells, isolated by expression of CD49f and stem cell antigen-1 (Sca-1). An important question for the field is whether all basal cells have stem cell characteristics. Prostate-specific microarray databases were interrogated to find candidate surface antigens that could subfractionate the basal cell population. Tumor-associated calcium signal transducer 2 (TACSTD2/Trop2/M1S1/GA733-1) was identified because it was enriched after castration, in prostate sphere cells and in the basal fraction. In the murine prostate, Trop2 shows progenitor characteristics such as localization to the region of the gland proximal to the urethra and enrichment for sphere-forming and colony-forming cells. Trop2 subfractionates the basal cells into 2 populations, both of which express characteristic basal cell markers by quantitative PCR. However, only the basal cells expressing high levels of Trop2 were able to efficiently form spheres in vitro. In the human prostate, where Sca-1 is not expressed, sphere-forming progenitor cells were also isolated based on high expression of Trop2 and CD49f. Trop2-expressing murine basal cells could regenerate prostatic tubules in vivo, whereas the remaining basal cells had minimal activity. Evidence was found for basal, luminal, and neuroendocrine cells in prostatic tubules regenerated from Trop2 hi basal cells. In summary, functionally distinct populations of cells exist within the prostate basal compartment and an epithelial progenitor can give rise to neuroendocrine cells in vivo.neuroendocrine ͉ progenitor ͉ sphere assay

ABSTRACT/SUMMARY The successful isolation and cultivation of prostate stem cells will allow us to study their unique biological properties and their application in therapeutic approaches. Here we provide step-by-step procedures on the basis of previous work in our laboratory for: the harvesting of primary prostate cells from adolescent male mice by a modified enzymatic procedure; the isolation of an enriched population of prostate stem cells through cell sorting; the cultivation of prostate stem cells in vitro; and characterization of these cells and their stem-like activity, including in vivo tubule regeneration. Normally it will take approximately 8 hours to harvest prostate cells, isolate the stem cell enriched population, and set up the in vitro sphere assay. It will take up to 8 weeks to analyze the unique properties of the stem cells, including their regenerative capacity in vivo.

Chromosomal rearrangements involving erythroblast transformation specific (ETS) family transcription factors were recently defined as the most common genetic alterations in human prostate cancer. Despite their prevalence, it is unclear what quantitative role they play in either initiation or progression of the disease. Using a lentiviral transduction and dissociated cell prostate regeneration approach, we find that acutely increased expression of ETS proteins in adult murine prostate epithelial cells is sufficient to induce the formation of epithelial hyperplasia and focal prostatic intraepithelial neoplasia (PIN) lesions, but not progression to carcinoma. However, combined expression of ERG with additional genetic alternations associated with human prostate cancer can lead to aggressive disease. Although ERG overexpression does not cooperate with loss of the tumor suppressor p53, it does collaborate with alterations in PI3K signaling, such as Pten knockdown or AKT up-regulation, to produce a well-differentiated adenocarcinoma. Most striking is our finding that overexpression of androgen receptor (AR) does not give rise to any hyperplastic lesions, but when combined with high levels of ERG, it promotes the development of a more poorly differentiated, invasive adenocarcinoma. These findings suggest that in human prostate cancer, the most potent function of ETS gene fusions may be to synergize with alternative genetic events and provide different pathways for carcinoma production and invasive behavior. Our results provide direct evidence for selective cooperating events in ERG-induced prostate tumorigenesis and offer a rational basis for combined therapeutic interventions against multiple oncogenic pathways in prostate cancer.AKT ͉ androgen receptor ͉ ERG ͉ prostate cancer ͉ PTEN H uman cancer is a multigene disorder in which several somatic mutations in cancer predisposition genes are required in a stepwise manner (1). The first genetic alteration usually confers a selective growth advantage and subsequently results in clonal expansion to initiate the neoplastic process. But the cells must accumulate several other rate-limiting mutations over a long period to become malignant (2). For prostate cancer (PCa), various oncogenes and tumor suppressor genes, including NKX3.1, PTEN, p27, and p53, and androgen receptor (AR) signaling have been implicated in the disease progression (3). However, the key initiating steps in prostate carcinogenesis remain uncertain, and most genetic alterations that can collaborate to drive tumor progression have yet to be elucidated.Chromosomal translocations play critical roles in hematological malignancies (4). Several lines of evidence in epithelial cancers of distinct tissue origins suggest that acquired chromosomal rearrangements could also be the common genetic events in many, if not all, epithelial cancers (5). Recent studies showed that 50-80% of patients with PCa were found to harbor fusion transcripts between an androgen-regulated gene, TMPRSS2, and members of the erythroblast tr...

The relationship between the cells that initiate cancer and the cancer stem-like cells that propagate tumors has been poorly defined. In a human prostate tissue transformation model, basal cells expressing the oncogenes Myc and myristoylated AKT can initiate heterogeneous tumors. Tumors contain features of acinartype adenocarcinoma with elevated eIF4E-driven protein translation and squamous cell carcinoma marked by activated betacatenin. Lentiviral integration site analysis revealed that alternative histological phenotypes can be clonally derived from a common cell of origin. In advanced disease, adenocarcinoma can be propagated by self-renewing tumor cells with an androgen receptor-low immature luminal phenotype in the absence of basal-like cells. These data indicate that advanced prostate adenocarcinoma initiated in basal cells can be maintained by luminal-like tumor-propagating cells. Determining the cells that maintain human prostate adenocarcinoma and the signaling pathways characterizing these tumor-propagating cells is critical for developing effective therapeutic strategies against this population.