Abstract 870: The role of the intermediate prolactin receptor in breast cancer oncogenesis

Abstract

Abstract Epidemiological, cellular, and genetic analyses indicate the hormone prolactin (PRL) and its cognate receptor in humans (hPRLr) are significantly involved in breast cancer pathogenesis. Recent evidence demonstrated that a truncated mouse PRLr (mPRLrT) is oncogenic when expressed alongside its canonical long form counterpart (mPRLrL) [Cell Reports 17, 249-260]. The mPRLrT shares significant sequence homology with a naturally-occurring and widely-expressed hPRLr splice variant, the intermediate hPRLr (hPRLrI) isoform. Given this similarity, we hypothesized hPRLrI may also induce transformation, when expressed alongside wild-type long hPRLr (hPRLrL). Like the mPRLrT, hPRLrI co-expression with hPRLrL in the immortalized but not transformed human breast cell line MCF10A resulted in a significant increase in proliferation, migration, viability, and anchorage-independent growth. These results were not observed following overexpression of either isoform alone, demonstrating that hPRLrL and hPRLrI co-expression is necessary to induce transformation of normal mammary epithelia. To further characterize this transformation, we established MCF10A xenografts using female NOD scid gamma (NSG) mice. Following intraductal injection, we observed rapid tumor growth in the hPRLrL/I co-expression cohort, significantly over that of the cohorts harboring either isoform alone, validating our in vitro findings in vivo. To determine mechanisms of transformation, we examined both differential protein stability and altered signaling events. In analyzing receptor degradation, a cycloheximide assay revealed hPRLrL stability is increased when heterodimerized with hPRLrI. hPRLrL turnover has been reported to be impaired in human breast cancer, indicating this phenomenon may be involved in the observed hPRLrI-mediated transformation. In regards to differential signaling, we examined the Jak2/Stat5a pathway. Jak2 is a promiscuous kinase whose significant oncogenic actions are well-characterized, while Stat5a is a transcription factor whose activities are critical in attenuating the actions of Jak2. Following PRL stimulation, it was observed that hPRLrL/I co-expression induced approximately two-fold greater Jak2-Y1007/1008 phosphorylation (pJak2) compared to that induced by hPRLrL expression alone. Further, it was observed that hPRLrL/I co-expression induced ten-fold less Stat5a-Y694 phosphorylation (pY-Stat5a) than hPRLrL expression alone. These data indicate unchecked pJak2 activity may also be a contributing mechanism in the observed transformation. Overall, these results demonstrate that hPRLrI, alongside hPRLrL, is sufficient for transformation of normal breast tissue. Citation Format: Jacqueline M. Grible, Charles V. Clevenger. The role of the intermediate prolactin receptor in breast cancer oncogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 870.