Synthesis And Biological Evaluation Of 3-(4-fluorophenyl)-1H-pyrazole Derivatives As AR Antagonists

Prostate cancer (PCa) is one of the most commonly diagnosed cancers, and the second and third leading cause of cancer death among men in the United States and Europe, respectively. It is estimated that there will be approximately 180,890 new cases diagnosed, and 26,120 deaths from PCa in 2016 in United States. PCa is, apart from hereditary cases, a desease of old age with a peak incidence in men of 70 years old.The standard care for patients with metastatic PCa is androgen deprivation therapy (ADT) by targeting the androgen receptor (AR) signaling pathway. However, after initial reponse to ADT, tumor cells ultimately become castrated-resistant resulting in progression of the disease despite castrate level of serum androgen. This stage of desease is termed as castration-resistant PCa (CRPC). The chief formal criteria for CRPC are serial increases in PSA level and/or radiographic progression of metastatic lesions in patients with testosterone level<50ng/dL or<1.7nmol/L. Other clinical manifestations of CRPC include painful bone or lynph node metastases, and cachexia. Death typically occurs within 2-4 years of CRPC. The reactivation of AR is a crucial motivation to the survival of CRPC. Therefore AR is still considered as an important therapy target for CRPC.AR is a ligand dependent transcription factor, expressed within the luminal layer and in the stoma of prostate epithelium, which controls the balance between cell differentiation and proliferation. In nomal prostate epithelium, the primary function of AR is to induce expression of genes required to promote termimal differentiation, suppress proliferation and promote secretion. While in PCa, the function of AR is induce expression of genes required for cell survival and proliferation.AR Antagonists, also known as antiandrogen, prevent androgens from carrying out their biological activity by directly binding AR ligand binding domain (LBD) and inducing repressive activity. Initial AR antagonists are chemicals with steroidal skeleton to ensure binding to the receptor. However, severe darawbacks such as hepatotoxicity, interference with libido and potency, cardiovascular side effects and low efficacy have limited their clinical use. Non-steroidal antiandrogens (NSAAs) were first introduced in 1989 in clinical practice as treatment for advanced and metastatic PCa. The non-steroidal structure avoids the typical constraints associated with the previous steroidal antiandrogens. The NSAAs used in clinical includes the first generation flutamide (Flut), bicalutamide (Bic) and nilutamide (Nil) and the second generation enzalutamide (Enz) and ARN-509. Although antiandrogen treatment usually exhibit favorable responses, AR gene mutation in LBD, altering ligand specificity and/or function, finally lead to the emergence of agonistic activity of NSAAs and the failure of the treatment. There is no available therapy for CRPC currently. Therefore, it is urgent to develop novel AR antagonists.In this work, a series of derivatives of compound T3, a potent AR antagonist discovered in virtual screening performed in our lab previously, were designed by modifying the linker between the two aromatic rings and prepared using fluoroacetophenone as a starting material by aldo-ester condensation reaction, cycliztion reation, methylation reaction, hydrolysis reaction, rearrangement reaction, addition reaction, reduction reaction, halogenation reaction and nucleophilic substitution reaction. The stuctures were identified by 1H-nuclear magnetic resonance (’H-NMR),13C-nuclear magnetic resonance (13C-NMR). The ability of inhibiting PSA expression and AR transcription, and anti-proliferative activity towards two cell lines, LNCaP cell which is androgen dependent prostate cancer cell line and PC-3 which is androgen independent prostate cancer cell line, were determined by enzyme linked immunosorbent assay (ELISA), luciferase reporter gene assay and MTT assay repectively.Results indicated that most of the derivatives showed promising anti-proliferative activity and inhibitory effects on PSA expression compared to lead compound T3, and some of the compounds selectively inhibit cell growth of androgen dependent cell LNCaP while have no affect on the proliferation of androgen independent cell PC-3. Among them, the most potent of them are compound 13d and 13e, whose cell growth inhibitory effect towards LNCaP (IC50) were 27μM and 17μM, and inhibitory effects on PSA expression at the concentration of 5μM were 39% and 46% respectively, better than that of lead compound T3. Further luciferase reporter gene assay showed that both of the two compounds, especially 13e, exhibited good antagonistic activity towards AR, compared to that of Enzalutamide while have no agonistic activity. Compounds 13d and 13e can be used as lead compounds for further optimization.