Yang Zong

Basal epithelial stem cells are efficient targets for prostate cancer initiation

Prevailing theories suggest that luminal cells are the origin of prostate cancer because it is histologically defined by basal cell loss and malignant luminal cell expansion. We introduced a series of genetic alterations into prospectively identified populations of murine basal/stem and luminal cells in an in vivo prostate regeneration assay. Stromal induction of FGF signaling, increased expression of the ETS family transcription factor ERG1, and constitutive activation of PI3K signaling were evaluated. Combination of activated PI3K signaling and heightened androgen receptor signaling, which is associated with disease progression to androgen independence, was also performed. Even though luminal cells fail to respond, basal/stem cells demonstrate efficient capacity for cancer initiation and can produce luminal-like disease characteristic of human prostate cancer in multiple models. This finding provides evidence in support of basal epithelial stem cells as one target cell for prostate cancer initiation and demonstrates the propensity of primitive cells for tumorigenesis.

ETS family transcription factors collaborate with alternative signaling pathways to induce carcinoma from adult murine prostate cells

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.

Stromal epigenetic dysregulation is sufficient to initiate mouse prostate cancer via paracrine Wnt signaling

Significance Increasing evidence supports the notion that an aberrant microenvironment facilitates cancer development. The mechanisms underlying the phenotypic changes and the tumor-promoting function of cancer-associated stroma remain largely unknown. Overexpression of the chromatin remodeling protein Hmga2 in the stroma leads to the formation of multifocal prostatic precancerous lesions in the neighboring naïve epithelium in a paracrine Wnt-dependent manner. The potent growth-promoting effects of Hmga2-modified stroma collaborate significantly with enhanced stromal AR signaling to drive the development of prostate adenocarcinoma. Our findings provide compelling evidence for the critical contribution of epigenetic changes in stromal cells to cancer initiation and progression. Carcinomas most often result from the stepwise acquisition of genetic alterations within the epithelial compartment. The surrounding stroma can also play an important role in cancer initiation and progression. Given the rare frequencies of genetic events identified in cancer-associated stroma, it is likely that epigenetic changes in the tumor microenvironment could contribute to its tumor-promoting activity. We use Hmga2 (High-mobility group AT-hook 2) an epigenetic regulator, to modify prostate stromal cells, and demonstrate that perturbation of the microenvironment by stromal-specific overexpression of this chromatin remodeling protein alone is sufficient to induce dramatic hyperplasia and multifocal prostatic intraepithelial neoplasia lesions from adjacent naïve epithelial cells. Importantly, we find that this effect is predominantly mediated by increased Wnt/β-catenin signaling. Enhancement of Hmga2-induced paracrine signaling by overexpression of androgen receptor in the stroma drives frank murine prostate adenocarcinoma in the adjacent epithelial tissues. Our findings provide compelling evidence for the critical contribution of epigenetic changes in stromal cells to multifocal tumorigenesis.

Adaptation or selection—mechanisms of castration-resistant prostate cancer

Men with advanced prostate cancer are typically treated with hormonal therapy, which leads to tumour shrinkage. However, tumours relapse and develop into the lethal form of the disease, termed castration-resistant prostate cancer (CRPC). Two distinct, but not mutually exclusive, models have been proposed in the literature to describe the onset of CRPC: adaptation and selection. Although some studies indicate that tumour cells acquire new alterations that enable them to survive in the castrated state (adaptation), other research points to the outgrowth of rare, pre-existing cells capable of surviving hormonal therapy (selection). Targeting the cells that survive hormonal therapy—by either adaptation or selection—is necessary to prevent the development of CRPC. Current research is focused on not only understanding the cellular mechanisms of CRPC, but also defining critical pathways that can be targeted with combinatorial therapies in castration-resistant cancer cells.

Cell-autonomous activation of the PI3-kinase pathway initiates endometrial cancer from adult uterine epithelium

Epithelial-specific activation of the PI3-kinase pathway is the most common genetic alteration in type I endometrial cancer. In the majority of these tumors, PTEN expression is lost in the epithelium but maintained in tumor stroma. Currently reported PTEN knockout mouse models initiate type I endometrial cancer concomitant with loss of PTEN in both uterine epithelium and stroma. Consequently, the biologic outcome of selectively activating the PI3-kinase pathway in the endometrial epithelium remains unknown. To address this question, we established a malleable in vivo endometrial regeneration system from dissociated murine uterine epithelium and stroma. Regenerated endometrial glands responded to pharmacologic variations in hormonal milieu similar to the native endometrium. Cell-autonomous activation of the PI3-kinase pathway via biallelic loss of PTEN or activation of AKT in adult uterine epithelia in this model was sufficient to initiate endometrial carcinoma. AKT-initiated tumors were serially transplantable, demonstrating permanent genetic changes in uterine epithelia. Immunohistochemistry confirmed loss of PTEN or activation of AKT in regenerated hyperplastic glands that were surrounded by wild-type stroma. We demonstrate that cell-autonomous activation of the PI3-kinase pathway is sufficient for the initiation of endometrial carcinoma in naive adult uterine epithelia. This in vivo model provides an ideal platform for testing the response of endometrial carcinoma to targeted therapy against this common genetic alteration.

Role of autonomous androgen receptor signaling in prostate cancer initiation is dichotomous and depends on the oncogenic signal

The steroid hormone signaling axis is thought to play a central role in initiation and progression of many hormonally regulated epithelial tumors. It is unclear whether all cancer-initiating signals depend on an intact hormone receptor signaling machinery. To ascertain whether cell autonomous androgen receptor (AR) is essential for initiation of prostate intraepithelial neoplasia (PIN), the response of AR-null prostate epithelia to paracrine and cell autonomous oncogenic signals was assessed in vivo by using the prostate regeneration model system. Epithelial-specific loss of AR blocked paracrine FGF10-induced PIN, whereas the add back of exogenous AR restored this response. In contrast, PIN initiated by cell-autonomous, chronic-activated AKT developed independent of epithelial AR signaling. Our findings demonstrate a selective role for AR in the initiation of PIN, dependent on the signaling pathways driving tumor formation. Insights into the role of hormone receptor signaling in the initiation of epithelial tumors may help define this axis as a target for chemoprevention of carcinomas.

Interleukin-6 and oncostatin-M synergize with the PI3K/AKT pathway to promote aggressive prostate malignancy in mouse and human tissues

Chronic inflammation has been proposed as an etiological and progression factor in prostate cancer. In this study, we used a dissociated prostate tissue recombination system to interrogate the role of interleukin 6 (IL6) and the related cytokine oncostatin-M (OSM) in the initiation and progression of prostate cancer. We identified that prostatic intraepithelial neoplasia (PIN) lesions induced by PTEN loss of function (PTENLOF) progress to invasive adenocarcinoma following paracrine expression of either cytokine. Increased expression of OSM was also able to drive progression of benign human epithelium when combined with constitutively activated AKT. Malignant progression in the mouse was associated with invasion into the surrounding mesenchyme and increased activation of STAT3 in PTENLOF grafts expressing IL6 or OSM. Collectively, our work indicates that pro-inflammatory cytokines such as IL6 or OSM could activate pathways associated with prostate cancer progression and synergize with cell-autonomous oncogenic events to promote aggressive malignancy. Implications: Increased expression of IL6 or OSM synergizes with loss of PTEN to promote invasive prostate cancer. Visual Overview: http://mcr.aacrjournals.org/content/early/2013/09/02/1541-7786.MCR-13-0238/F1.large.jpg. Mol Cancer Res; 11(10); 1159–65. ©2013 AACR.

Functional screen identifies kinases driving prostate cancer visceral and bone metastasis

Significance Therapies are urgently needed to treat metastatic prostate cancer. Mutationally activated and wild-type kinases such as BCR-ABL and BTK are effective therapeutic targets in multiple cancers. Genetically altered kinases are rare in prostate cancer. Wild-type kinases may be implicated in prostate cancer progression, but their therapeutic potential in metastatic prostate cancer remains unknown. Using phosphoproteomics and gene expression datasets, we selected 125 wild-type kinases implicated in human prostate cancer metastasis to screen for metastatic ability in vivo. The RAF family, MERTK, and NTRK2 drove prostate cancer bone and visceral metastasis and were highly expressed in human metastatic prostate cancer tissues. These studies reveal that wild-type kinases can drive metastasis and that the RAF family, MERTK, and NTRK2 may represent important therapeutic targets. Mutationally activated kinases play an important role in the progression and metastasis of many cancers. Despite numerous oncogenic alterations implicated in metastatic prostate cancer, mutations of kinases are rare. Several lines of evidence suggest that nonmutated kinases and their pathways are involved in prostate cancer progression, but few kinases have been mechanistically linked to metastasis. Using a mass spectrometry-based phosphoproteomics dataset in concert with gene expression analysis, we selected over 100 kinases potentially implicated in human metastatic prostate cancer for functional evaluation. A primary in vivo screen based on overexpression of candidate kinases in murine prostate cells identified 20 wild-type kinases that promote metastasis. We queried these 20 kinases in a secondary in vivo screen using human prostate cells. Strikingly, all three RAF family members, MERTK, and NTRK2 drove the formation of bone and visceral metastasis confirmed by positron-emission tomography combined with computed tomography imaging and histology. Immunohistochemistry of tissue microarrays indicated that these kinases are highly expressed in human metastatic castration-resistant prostate cancer tissues. Our functional studies reveal the strong capability of select wild-type protein kinases to drive critical steps of the metastatic cascade, and implicate these kinases in possible therapeutic intervention.

A Two-Step Toward Personalized Therapies for Prostate Cancer

Therapeutic targeting of two proteins that drive tumorigenesis suggests personalized medicine is a possibility for prostate cancer. Identifying the dominant genetic alterations that drive tumorigenesis is essential for developing targeted cancer therapies. Recent work has demonstrated that prostate tumors can be stratified by dominant genetic alterations, such as chromosomal rearrangements involving ETS (Erythroblastosis virus E26 transformation-specific) family transcription factors or overexpression of SPINK1, a gene that encodes a secreted serine protease inhibitor. In this issue of Science Translational Medicine, Ateeq et al. provide evidence to support a rationale for targeting the SPINK1 protein in the SPINK1+/ETS– subset of prostate tumors and also describe a potential interaction of SPINK1 with epidermal growth factor receptor that could be an additional target for therapeutic intervention.

Dissociated prostate regeneration under the renal capsule

Tissue recombination models are useful for studying cancer initiation, progression, and metastasis. They also provide an in vivo environment in which to investigate the functional role of stem cells in tissue repair. In this protocol, we describe in detail the dissociated prostate regeneration assay. Dissociated adult murine prostate cells are combined with embryonic urogenital sinus mesenchymal cells and implanted under the renal capsule. Morphological tissue structures with appropriate epithelial-stroma interactions are reconstituted in the grafts.