Establishing novel, representative prostate cancer cell lines from fresh prostatectomy tissues via an optimized protocol

Background: The existing panel of prostate cancer (PCa) cell lines, often established from metastatic sites or after extensive passaging, frequently fails to fully recapitulate the genetic heterogeneity and tumor microenvironment dependence of primary prostate tumors. Additionally, there is a crucial need for diverse, population-specific cell lines to investigate the biological factors underlying disparities in patient outcomes and therapeutic response. To address this critical resource gap, we optimized and validated a highly efficient protocol for the de novo establishment of patient-derived, primary PCa cell lines from fresh surgical specimens. Methods: Patients were consented in accordance with VCU IRB protocol #20210623, and their self-reported race was recorded as either White (W) or African American (AA). Fresh tissues were obtained as punched biopsies from 36 radical prostatectomy specimens. Each specimen yielded 1-3 biopsy samples. H&E slides were graded by a licensed pathologist. Optimized from Maitland et al. 2016 protocol, tissue samples digested enzymatically were separated into distinct populations through differential centrifugation, and propagated in optimal media for epithelial and fibroblastic cell lines. The cellular origin of patient-matched lines were confirmed using cytochemistry and further characterized through proliferation assays, Western blot, and short tandem repeat (STR) profiling. Results: The optimized protocol, carried out in the latter quarter of the trial, demonstrated high efficacy with an 86% success rate (13 out of 15 samples), bringing the total to 30 novel, matched PCa epithelial and fibroblast cell lines successfully established from 18 patients. Among the 30 samples, 18 were benign (11 W and 7 AA) and 10 were cancerous (6 W and 4 AA). Additionally, 3 patient prostatic specimens yielded matched cell lines from both the tumor and the contralateral non-malignant tissue. Critically, these cell lines retained the expression of androgen receptor (AR) and prostate-specific antigen (PSA). Furthermore, the lines exhibited heterogeneous growth patterns reflecting the diversity of the source tumors. Conclusion: We have established a robust, highly effective protocol for generating patient-derived, primary PCa cell lines directly from prostatectomy tissues. These novel, characterized models significantly enrich the translational research toolkit, providing invaluable resources for several critical applications, including biomarker identification, understanding tumor initiation and progression, resistance mechanism studies, drug discovery and validation, and, most importantly, for studying population-specific differences to guide the development of truly equitable and personalized treatment strategies.