Targeting Prolyl Peptidases in Tamoxifen Resistant Breast Cancer

 描述（由申请人提供）乳腺癌是最常见的女性癌症，约70-75%的病例表达雌激素受体α（ERα）。三苯氧胺（Tamoxifen，TAM）是一种雌激素受体拮抗剂，是雌激素受体α（ERα）阳性的绝经前乳腺癌的标准内分泌治疗药物。不幸的是，几乎所有晚期肿瘤都会对内分泌治疗产生耐药性。脯氨酰羧肽酶（PRCP）及其家族成员脯氨酰内肽酶（PREP）是脯氨酰肽酶家族的成员。这些酶切割神经肽G蛋白偶联受体（GPCR）激动剂以调节GPCR信号传导。PRCP是在基因筛选中鉴定的，用于促进MCF 7细胞中TAM抗性的因子。初步数据显示，PRCP高表达与乳腺癌患者的预后不良相关。PRCP过表达增加了AKT-mTOR活化，促进了TAM抗性，并诱导了MCF 7肿瘤异种移植物的自发转移。我们鉴定了一种有效的PRCP和PREP抑制剂，称为Y-ox。PRCP/PREP消耗或Y-ox抑制使IRS-1不稳定并抑制AKT-mTOR。由于PRCP和PREP切割肽GPCR激动剂，我们测试了PRCP/PREP是否以GPCR依赖性方式调节IRS-1和AKT-mTOR通路。我们的数据支持PRCP/PREP增加GPCR依赖的CaMK 2（钙/钙调蛋白激活激酶2）激活，激活的CaMK 2然后通过抑制AMPK稳定IRS-1的模型。最后，我们显示Y-ox使IRS-1不稳定并抑制雷帕霉素处理的细胞中AKT的反馈激活，并且与雷帕霉素组合增加了TAM抗性细胞的杀伤。基于这些结果，我们假设1）PRCP/PREP以GPCR和CaMK 2依赖的方式维持IRS-1和AKT/mTOR通路，导致TAM抗性和转移，2）原发性乳腺肿瘤中的PRCP、PREP和/或CaMK 2表达将与TAM抗性和不良预后相关，3）PRCP/PREP和mTOR的组合抑制将减少PI 3 K/AKT的反馈激活，从而改善内分泌抗性和转移性乳腺肿瘤的治疗。

 DESCRIPTION (provided by applicant) Breast cancer is the most common female cancer and approximately 70-75% of cases express estrogen receptor alpha (ERα). Tamoxifen (TAM) is an estrogen receptor antagonist and the standard endocrine therapy for premenopausal women with ERα positive breast cancer. Unfortunately, resistance to endocrine therapy develops in almost all advanced tumors. Prolylcarboxypeptidase (PRCP) and its family member prolylendopeptidase (PREP) are members of the prolyl- peptidase family. These enzymes cleave neuropeptide G-protein coupled receptor (GPCR) agonists to regulate GPCR signaling. PRCP was identified in a genetic screen for factors that promote TAM resistance in MCF7 cells. Preliminary data show high PRCP expression is correlated with worse prognosis in breast cancer patients. PRCP over-expression increased AKT-mTOR activation, promoted TAM resistance, and induced spontaneous metastasis in MCF7 tumor xenografts. We identified a potent inhibitor of PRCP and PREP termed Y-ox. PRCP/PREP depletion or inhibition by Y-ox destabilized IRS-1 and inhibited AKT-mTOR. Because PRCP and PREP cleave peptide GPCR agonists, we tested if PRCP/PREP regulate IRS-1 and the AKT-mTOR pathway in a GPCR-dependent manner. Our data support a model in which PRCP/PREP increase GPCR-dependent activation of CaMK2 (calcium/calmodulin activated kinase 2), and activated CaMK2 then stabilizes IRS-1 by inhibiting AMPK. Finally, we showed Y-ox destabilized IRS-1 and inhibited feedback activation of AKT in rapamycin treated cells and, in combination with rapamycin increased killing of TAM- resistant cells. Based on these results, we hypothesize 1) PRCP/PREP maintain IRS-1 and the AKT/mTOR pathway in a GPCR and CaMK2-dependent manner, leading to TAM resistance and metastasis, 2) PRCP, PREP, and/or CaMK2 expression in primary breast tumors will correlate with TAM resistance and poor prognosis, 3) combined inhibition of PRCP/PREP and mTOR will reduce feedback activation of the PI3K/AKT and consequently improve treatment of endocrine resistant and metastatic breast tumors.