The gray-zone prostate-specific antigen (PSA) range is accepted to be 4-10 ng/ml and is considered to vary according to age. We aimed to investigate whether fusion prostate biopsy (FPB), which has been reported to have relatively higher cancer detection rates, has an effect on gray-zone PSA cutoff value. Material and methods This retrospective study included patients that underwent standard prostate biopsy (SPB) or multiparametric magnetic resonance imaging (MpMRI)-guided FPB (SPB+ targeted biopsy). All the patients included in the study were detected with a Prostate Imaging Reporting and Data System (PI-RADS) ≥3 lesion on MpMRI (the FPB group only). The demographics, clinical characteristics, and histopathological diagnoses were recorded for each patient. Results A total of 1,628 patients comprising 1,208 patients in the SPB group and 420 patients in the FPB group were included in the study. The mean PSA level was 9.75±6.68 ng/ml in the FBP group and 10.46±6.46 ng/ml in the SPB group (p=0.053). Prostate cancer (PCa) detection rate was significantly higher in the FPB group as compared to the SPB group (42.4% vs. 36.4%). The PSA cutoff value for PCa was 9.75 ng/ml (sensitivity and specificity, 81%) in the SPB group and was 7.55 ng/ml (sensitivity and specificity, 81% and 84%, respectively) in the FPB group. In the FPB group, the cancer detection rate among the patients with a PSA level of 7.55-10.00 ng/ml was 56.1%. Conclusion The results indicated that the introduction of FPB into clinical practice, which has relatively higher cancer detection rates, has further lowered the upper limit for gray-zone PSA.