Identification Of Prognosis And Enzalutamide Resistance-related Genes In Prostate Cancer Patients Based On Single-cell RNA Sequencing Analysis

Research Background and Objective:Prostate cancer is a malignancy of the male reproductive system and has the second-highest incidence among cancers in men.In Asian countries,3.8 per 100,000 people die from prostate cancer each year,which puts severe health and economic burdens on society.Male death from prostate cancer usually has a close relationship with metastasis and castration resistance.Although survival has improved with therapeutic strategies,the heterogeneous biological processes of prostate cancer in different patients lead to diverse clinical outcomes.Increasing evidence suggests that the presence of minor cell subpopulations in prostate cancer(PCa)could drive drug resistance.This study aimed to identify and annotate novel cell subpopulations based on the single-cell RNA sequencing,performed on Enzalutamide-sensitive and-resistant PCa cells.Material and Method:C4-2 cells were treated with gradually improved Enzalutamide dose,and after a long time culturing,the C4-2R cells were obtained,which was resistant to enzalutamide.Then,single-cell RNA sequencing was performed on Enzalutamide-sensitive and-resistant PCa cells to identify and annotate novel cell subpopulations.Then,Based on public datasets,we validated the prognostic values of critical candidates representing critical clusters identified by the single-cell RNA sequencing analyses.Firstly,the Kaplan-Meier and log-rank analyses were used to identify recurrence-free survival(RFS)-related genes;subsequently,the Least Absolute Shrinkage and Selection Operator(LASSO)Cox regression analyses were used to establish the predicting signatures in each cohort.The stability and significance of each signature were determined by Kaplan-Meier and log-rank,and receiver operative characteristics(ROC)analyses.All the statistical analyses were performed on R software(R Core Team,Miami,FL,USA).Results:By clustering 448 differentially expressed genes,six subclones were identified,showing a previously inappreciable cellular heterogeneity level related to Enzalutamide resistance.Notably,cluster 5 displayed Interferon-regulatory features.In silico analyses demonstrated that marker-gene-cluster was linked to the RFS of PCa,serving as an independent indicator of RFS prognosis.The nomogram receiver operating characteristic(ROC)curve synthesizing the signatures with clinicopathological features proved the signature added clinical values to the current staging system.Besides,the study verified the function of three critical markers,whose functions were consistent with their validated protein expressions.Conclusion:The study provides a better understanding of how PCa cells develop heterogeneously to Enzalutamide treatment.The study also revealed the interferon-regulating properties of cell subgroup 5,and it is also an important factor that may lead to enzalutamide resistance in prostate cancer cells.