VCOM Louisiana Research Day Program

Pharmacology

Hassan Y. Ebrahim 1 , PhD; Khaldoun S. Abdelwahed 1 , PhD; Abu Bakar Siddique 1 , PhD; Mohammed H. Qusa 1 , PhD; Ethar A. Mudhish 1 ; Ashkan H. Rad 1 ; Mourad Zerfaoui 3 , PhD; Zakaria Y. Abd Elmageed 2 , PhD; and Khalid A. El Sayed 1 , PhD 1 School of Basic Pharmaceutical and Toxicological Sciences, College of Pharmacy, University of Louisiana Monroe, Monroe, Louisiana; 2 Department of Pharmacology, Edward Via College of Osteopathic Medicine-Louisiana, Monroe, Louisiana; 3 Department of Surgery, School of Medicine, Tulane University, Louisiana 54 PSEUROTIN A MODULATES METASTATIC CASTRATION-RESISTANT PROSTATE CANCER RECURRENCE BY PCSK9/LDLR AXIS MODULATION

Context: Castration-resistant prostate cancer (CRPC) phenotype involves aggressive prostate cancer (PC) progression profile despite androgen deprivation therapy (ADT) and castrate levels of testosterone. CRPC lacks effective curative options. CRPC cells can biosynthesize androgens de novo by utilizing cholesterol, facilitating survival, progression, and motility. The low-density lipoprotein (LDL) is the main lipoprotein transporter of cholesterol in blood, which interacts with the low-density lipoprotein receptor (LDLR) facilitating the hepatocellular uptake of cholesterol, and therefore lowering circulating LDL-C. The level of LDLR is tightly regulated by proprotein convertase subtilisin/ kexin type 9 (PCSK9) through protein-protein interaction (PPI) mechanism, which ultimately induces receptor endocytosis and lysosomal degradation. PCSK9 emerged as a novel player in many human carcinomas, including the prostate. The fungal-derived spirocyclic alkaloid pseurotin A (PS) recently reported by our group as a novel lead suppressor for breast and prostate cancers progression and recurrence. Objective and/or Hypothesis: The current study provides a comprehensive validation of the role of PCSK9 in PC recurrence as well as the lead validation of PS as locoregional and distant recurrences suppressive entity by

targeting the PCSK9-axis in the most aggressive metastatic CRPC. Methods: Cell proliferation was assessed using the MTT-based cell viability assay. Cell migration was evaluated by the wound scratch assay, while colonization was gauged by the colony formation assay. The CRPC CWR-R1ca cells were transfected by small hairpin RNA for PCSK9 knockdown. The lentivirus-mediated transduction was implemented to tag CWR R1ca cells with luciferase (CWR-R1ca-Luc) for in vivo tumor bioluminescence tracing. Tissue microarray (TMA) was utilized to assess the expression level of PCSK9 in 64 human cases. High-fat diet (HFD, 11% total fat) was used to assess the role of high fat/cholesterol diet in CRPC progression and metastasis in a nude mouse model. Primary tumor growth and recurrence in vehicle control and PS-treated mice were monitored by IVIS bioluminescence imaging. Serum cholesterol, LDL-C and PCSK9 were tested to evaluate the pharmacodynamics effect of PS. Results: PS showed modest effect on CWR R1ca cell proliferation, while significantly suppressing motility and colonization. A similar pattern was also seen in PCSK9-KD CWR R1ca cells. TMA revealed that PCSK9 has a

higher protein expression in the cytoplasm of PC tissues compared to the normal tissues. The HFD induced the progression and metastasis by double-fold in a nude mouse model. Simultaneously, 10 mg/kg daily oral PS dosing significantly suppressed CWR-R1ca-Luc tumor Locoregional and distant recurrences, which was associated with reduction in serum cholesterol, LDL-C and PCSK9 levels. Conclusion(s): The current study proved the relevance of PCSK9 in PC pathogenesis. The study validated PS as a novel lead for the control of mCRPC recurrence through targeting PCSK9/ LDLR axis. PS is a novel entity amenable for further development as adjuvant therapy for the control of the aggressive mCRPC recurrences.

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