Functional screen identifies kinases driving prostate cancer visceral and bone metastasis

Abstract

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.kinases | metastasis | prostate cancer | bone metastasis M etastatic prostate cancer is responsible for the deaths of ∼30,000 men in the United States each year (1, 2). Ninety percent of patients develop bone metastases, and other major sites of metastases include lymph nodes, liver, adrenal glands, and lung (3). First-line treatments for metastatic disease are androgen deprivation therapies that block androgen synthesis or signaling through the androgen receptor (AR) (2). Inevitably, metastatic prostate cancer becomes resistant to androgen blockade. Second-line treatments such as chemotherapy (docetaxel, cabazitaxel) and radiation only extend survival 2-4 mo (4, 5).Identifying new therapeutic targets for metastatic prostate cancer has proven difficult. Exome and whole-genome sequencing of human metastatic prostate cancer tissues have found frequent mutations and/or chromosomal aberrations in numerous genes, including AR, TP53, PTEN, BRCA2, and MYC (6-11). The precise functional contribution of these genes to prostate cancer metastasis remains unknown. Genomic and phosphoproteomic analyses have also revealed that metastatic prostate cancer is molecularly heterogeneous, which has complicated the search for common therapeutic targets (12). Few murine models of prostate cancer develop metastases. Mice having prostate-specific homozygous deletions in SMAD4 and PTEN or expression of mutant KRAS develop metastases in visceral organs but rarely in bone (13-15).Targeting genetically altered constitutively active protein kinases such as BCR-ABL in chronic myelogenous leukemia and BRAF V600E in melanoma has led to dramat...

Driven to metastasize: Kinases as potential therapeutic targets in prostate cancer

“…Coincidentally, there are also no US Food and Drug Administrationapproved kinase inhibitors for the treatment of this disease. In PNAS, Faltermeier et al (4) investigated the landscape of kinases in prostate cancer and performed functional screens to identify candidate kinases that drive metastatic progression in preclinical models of disease.…”

“…Although there are over 500 kinases comprising the human kinome, Faltermeier et al (4) identify 125 kinases of potential relevance in metastatic prostate cancer, from a combination of analyses of phosphoproteomic and genomic/transcriptomic datasets coupled with a literature search. Using an in vivo lung colonization screen based upon overexpression of candidate kinases in prostate tumor-derived, PTENnull murine Cap8 cells, the authors investigated the ability of their candidate kinases, screening five at a time, to promote metastases in a mouse model.…”

“…Using an in vivo lung colonization screen based upon overexpression of candidate kinases in prostate tumor-derived, PTENnull murine Cap8 cells, the authors investigated the ability of their candidate kinases, screening five at a time, to promote metastases in a mouse model. Having identified 20 kinases that promoted metastases in this initial screen, Faltermeier et al (4) then conducted a second screen in which they overexpressed these 20 kinases in RWPE-1 immortalized normal prostate epithelial cells, and then performed tail vein injections in NOS scid gamma mice. This second screen demonstrated that five kinases, all three rapidly accelerated fibrosarcoma (RAF) family members (ARAF, BRAF, and CRAF), as well as C-Mer proto-oncogene tyrosine kinase (MERTK) and neurotrophic tyrosine kinase receptor, type 2 (NTRK2), drove bone and visceral metastases, as assessed with PET/computed tomography imaging and histology.…”

“…Kinase inhibitors as a therapeutic strategy for mCRPC. Using preclinical models of prostate cancer, kinases that potentially drive metastatic progression may be identified through in vivo screening assays for metastases, as described by Faltermeier et al (4). Upon identification of biomarkers of response, patients could be selected for treatment with a specific kinase inhibitor to optimize the potential for disease response.…”

“…Given the subtle, yet important, differences in the activity of ARAF, BRAF and CRAF, a more detailed understanding of RAF kinases in prostate cancer may be required to define the mechanism by which RAF family members promote metastases and to define the role of RAF inhibitors in prostate cancer. As pointed out by Faltermeier et al (4), given that RAF inhibitors may paradoxically activate WT BRAF in certain contexts (15), inhibition of downstream RAF effectors, such as mitogen-activated protein kinase kinase 1 (MEK), may ultimately represent a more efficacious strategy.…”