SAT635 Role Of The Human Prolactin Receptor I-tail In The Pathogenesis Of Breast Cancer

Abstract

Abstract Disclosure: C.V. Clevenger: None. S. Shen: None. Evidence from epidemiological, cellular, and genetic analyses indicate that the hormone prolactin (PRL) and its receptor (hPRLr) in humans are significantly involved in breast cancer pathogenesis. The intermediate form of hPRLr (hPRLrI) induces significant increases in proliferation and anchorage-independent growth of normal mammary epithelia in vitro when co-expressed with the long form hPRLr (hPRLrL). However, the exact mechanism of how the hPRLrI transforms mammary epithelium is not well understood. The hPRLrI is produced by alternative splicing that induces a frameshift, resulting in a premature stop codon and a novel 13 amino acid tail ("I-Tail") gain. The ubiquitin-like protein neural precursor cell expressed developmentally down-regulated protein 8 (NEDD8) was found to be associated with the I-tail by yeast two-hybrid analysis. Treatment with a selective NEDD8 inhibitor resulted in the altered stability of the hPRLrL and the death of breast cancer cells. Given this, we hypothesized that the hPRLrI I-tail contributes to mammary transformation. hPRLrL, hPRLrI, hPRLrIΔ13 (I-tail removed mutant due to loss of 13 C-terminal amino acid resides) were delivered to MCF10AT cells by lentivirus transduction and subsequently analyzed. Anchorage-independent cell growth was determined by soft agar assay, cell proliferation was determined with a Incucyte analysis, and cell motility was examined by wound healing assay. In addition, signaling pathway and differential gene expression alterations mediated by the I-tail were determined by RNA-seq and signaling analysis. Successful expression of the hPRLrL+hPRLrIΔ13 in the MCF10AT transfectant resulted in significant reductions or alterations of anchorage-independent growth, cell proliferation, ERK1/2 activation, and PRL-induced different gene expression. These findings demonstrate that the hPRLrI I-tail contributes to the transformation of normal mammary epithelia cells. Presentation: Saturday, June 17, 2023