In vivo quantitative phosphoproteomic profiling identifies novel regulators of castration-resistant prostate cancer growth

Abstract

Prostate cancer remains a leading cause of cancer-related mortality worldwide owing to our inability to treat effectively castration-resistant tumors. To understand the signaling mechanisms sustaining castration-resistant growth, we implemented a mass spectrometry-based quantitative proteomic approach and use it to compare protein phosphorylation in orthotopic xenograft tumors grown in either intact or castrated mice. This investigation identified changes in phosphorylation of signaling proteins such as MEK, LYN, PRAS40, YAP1 and PAK2, indicating the concomitant activation of several oncogenic pathways in castration-resistant tumors, a notion that was confirmed by tumor transcriptome analysis. Further analysis demonstrated that the activation of mTORC1, PAK2 and the increased levels of YAP1 in castration-resistant tumors can be explained by the loss of androgen inhibitory actions. The analysis of clinical samples demonstrated elevated levels of PAK2 and YAP1 in castration-resistant tumors, whereas knockdown experiments in androgen-independent cells demonstrated that both YAP1 and PAK2 regulate cell colony formation and cell invasion activity. PAK2 also influenced cell proliferation and mitotic timing. Interestingly, these phenotypic changes occur in the absence of obvious alterations in the activity of AKT, MAPK or mTORC1 pathways, suggesting that PAK2 and YAP1 may represent novel targets for the treatment of castration-resistant prostate cancer. Pharmacologic inhibitors of PAK2 (PF-3758309) and YAP1 (Verteporfin) were able to inhibit the growth of androgen-independent PC3 xenografts. This work demonstrates the power of applying high-resolution mass spectrometry in the proteomic profiling of tumors grown in vivo for the identification of novel and clinically relevant regulatory proteins.

YAP1 and AR interactions contribute to the switch from androgen-dependent to castration-resistant growth in prostate cancer

“…Besides, Jiang et al . 44 identified additional phosphorylation sites (S163/164 and T63) on YAP1 in the lysate of LNCaP xenografts through a quantitative proteomic approach; however, that study did not show a change in phospho-YAP1–S127 levels. One possible explanation for it was that the MST1 kinase activity might be inactivated in LNCaP xenografts, possibly due to the expression of the constitutively active AKT1, which is known to inactivate MST1 via phosphorylation 59 .…”

“…Activation of YAP1 due to the gene amplification or the loss-of-function of MST1/2 or LATS1/2 is linked to the aetiology of many cancers including lung 35 , colon 36 , ovarian 37 , head and neck 38 , liver 39,40 , meningioma 41 , thyroid 42 and stomach 43 . In addition, emerging evidence suggests that YAP1 may also play a critical role in the pathobiology of PC 44 . However, the role and mechanism of YAP1 in PC remains to be explored.…”

Verteporfin, a suppressor of YAP–TEAD complex, presents promising antitumor properties on ovarian cancer

“…Lately, VP has shown promising anticancer effect in various malignancies such as breast cancer,13,14 prostate carcinoma,15 pancreatic ductal adenocarcinoma,16 and esophageal cancer,17 especially when combined with standard chemotherapy.…”

Expansion of BCR/ABL1+ cells requires PAK2 but not PAK1

“…Cell cycle progression is mildly affected upon loss of PAK2, which is in line with a reduced mitotic timing (Jiang et al , 2015). However, this defect did not translate into differences in growth curve kinetics, suggesting compensation by PAK1.…”

“…Human prostatic adenocarcinoma cells and skin epidermal cells that are subjected to epidermal growth factor‐induced transformation show reduced colony formation upon loss of PAK2 (Li et al , 2011; Jiang et al , 2015). …”