Serum PSA levels fluctuations in patients with non–muscle-invasive bladder cancer (NMIBC) undergoing BCG therapy: a prospective study

Abstract

Background

Intravesical bacillus Calmette–Guérin (BCG) therapy is the standard adjuvant treatment for intermediate-high risk non–muscle-invasive bladder cancer (NMIBC). Transient elevations in prostate-specific antigen (PSA) following BCG instillation can prompt unnecessary prostate biopsies and increase patient anxiety. This study investigated temporal changes in total PSA levels (TPSA) and Free-to-Total PSA ratio (f/tPSA) in NMIBC patients receiving intravesical BCG therapy.

Methods

We prospectively enrolled 25 male patients with NMIBC and baseline TPSA < 4 ng/mL. Serum TPSA and free PSA were measured at three time points: prior to BCG initiation as baseline, immediately after the induction course, and three months post-treatment. Patients with prior prostate cancer, inflammatory prostate conditions, or use of medications affecting PSA were excluded. Data were analyzed using Friedman test to compare the three time points, followed by post-hoc pairwise comparisons using Wilcoxon signed-rank tests (SPSS v27).

Results

Twenty patients completed the study (mean age 64.0 ± 8.4 years), Sixteen patients (80%) exhibited a significant elevation of TPSA immediately following induction therapy. In 14 of these patients (> 80%), PSA levels spontaneously returned to near-baseline values within three months. Mean TPSA rose from 1.17 ± 1.24 ng/mL at baseline to 2.02 ± 1.48 ng/mL following intravesical BCG therapy course, the declined to 1.27 ± 1.24 ng/mL at three months post-treatment. The mean f/tPSA exhibited a steadier trend, declining from 0.30 ± 0.16 at baseline to 0.26 ± 0.12 ng/mL post-treatment, and a slight increasing to 0.28 ± 0.16 ng/mL at three months. Statistical analysis confirmed that the post-treatment elevations in TPSA and was significant (p < 0.001). No patient developed clinical or histopathological evidence of prostate cancer during follow-up.

Conclusion

Transient PSA elevation is a common but reversible phenomenon after intravesical BCG instillation, reflecting local inflammatory and immune responses rather than malignant transformation. In the absence of additional clinical findings, a strategy of “watchful waiting” with repeat PSA testing at three months may prevent unnecessary prostate biopsies, reduce patient anxiety, and optimize healthcare resource utilization.

Background

Bladder cancer is among the most common malignancies of the urinary tract, representing a public health concern globally and in Iran. According to GLOBOCAN 2020, blader cancer ranks as the tenth most prevalent cancer globally, with over 573,000 new cases and 213,000 deaths annually [1–3]. In Iran, it is one of the top four malignancies in men, with an age-standardized incidence rate (ASR) of approximately 8.5 per 100,000 population [4, 5]. The majority of cases (70–75%) present as non–muscle-invasive bladder cancer (NMIBC), a subtype associated with favorable long-term survival yet characterized by high recurrence and variable risk of progression. These clinical features necessitate close monitoring and effective adjuvant treatments to improve patient outcomes [6].

Intravesical Bacillus Calmette–Guérin (BCG) therapy remains an important adjuvant treatment for patients with intermediate- and high-risk NMIBC following transurethral resection of bladder tumor (TURBT) [7], reducing the risk of recurrence and potentially the risk of progression as well [8, 9]. Despite its proven efficacy, BCG therapy is associated with a spectrum of local and systemic adverse effects, including such as fever, hematuria, epididymitis, granulomatous cystitis, and granulomatous prostatitis, and prostatic abscess [10, 11]. Among these, BCG-induced granulomas have been widely documented in prostate biopsy specimens, cytological aspirates, and prostatectomy specimens [12, 13].

Clinically, BCG-induced prostatitis can closely resemble prostate cancer in multiple dimensions —laboratory, radiological, and clinical, thereby complicating management. On digital rectal examination, the prostate may appear nodular and firm, and ultrasonography may reveal suspicious abnormalities consistent with underlying pathological changes. Elevated serum prostate-specific antigen (PSA) levels may further confound the clinical picture, often raising suspicion for concurrent malignancy [13, 14]. Elevated serum PSA may be found in patients treated with BCG, even with no clinical evidence of granulomatous prostatitis [14]. Importantly, these PSA elevations are generally transient and self-limiting; however, they may be misinterpreted as early indicators of prostate cancer, prompting unnecessary invasive procedures such as prostate biopsy. Such interventions not only expose patients to procedural risks but also contribute to psychological stress and increased healthcare costs [11, 13, 14].

Despite global research, evidence from Middle Eastern populations remains infrequent. In particular, no national instruction in Iran currently provides recommendations on how clinicians should interpret PSA fluctuations in patients undergoing BCG therapy. This gap may contribute to inconsistent clinical decision-making and unwarranted diagnostic interventions across institutions.

In light of these challenges, the present study was designed to evaluate short-term changes in total PSA levels (TPSA) and Free-to-Total PSA ratio (f/tPSA) in Iranian patients with NMIBC undergoing intravesical BCG therapy. Our research intends to clarify the clinical significance of PSA fluctuations in this population, inform clinical decision-making, and potentially reduce the frequency of unnecessary prostate biopsies. Ultimately, this study seeks to fill an important knowledge gap and contribute to the optimization of patient care both within Iran and in broader international contexts.

Methods

Study design and setting

This prospective cohort study was conducted between August 2023 and August 2025 at three tertiary referral hospitals affiliated with Mashhad University of Medical Sciences, namely Imam Reza, Qaem, and Omid hospitals (Mashhad, Iran). The study protocol was approved by the Institutional Review Board and Ethics Committee (IR.MUMS.MEDICAL.REC.1402.228).

Participants

Eligible participants were adults (≥ 18 years) diagnosed with non–muscle-invasive bladder cancer (NMIBC) who had normal baseline prostate-specific antigen (PSA) levels (< 4 ng/mL) and were candidates for intravesical Bacillus Calmette–Guérin (BCG) therapy. Written informed consent was obtained from all patients prior to enrollment. Exclusion criteria included prior or concurrent prostate cancer, elevated baseline PSA, muscle-invasive bladder cancer, tuberculosis, immunodeficiency, or inability/unwillingness to comply with follow-up. Participants were withdrawn upon withdrawal of consent, occurrence of severe BCG-related clinical or laboratory adverse events, or incomplete treatment or missing data.

Intravesical BCG instillation protocol

All eligible patients diagnosed with NMIBC underwent intravesical Bacillus Calmette–Guérin (BCG) immunotherapy following complete transurethral resection of bladder tumor (TURBT). Treatment initiation was scheduled after mucosal healing and absence of urinary tract infection, typically within 2–4 weeks post-TURBT. The induction regimen consisted of six weekly intravesical instillations, each administered with a standard dose of BCG (PastoCys^®, Pasteur Institute of Iran). Each vial contains 120 mg of Mycobacterium bovis Pasteur strain 1173P2, which was reconstituted with sterile normal saline to a final volume of 50 ml [15]. Prior to each instillation, patients were instructed to completely empty their bladder. Instillations were administered under aseptic conditions via a transurethral Foley catheter, after which the catheter was removed. Patients were asked to retain the BCG solution in their bladder for approximately two hours, before voiding naturally.

Serum PSA levels were measured at baseline (prior to BCG instillation), immediately after completion of BCG induction, and three months post-treatment. Digital rectal examination (DRE) was also performed to evaluate potential prostate changes.

Data collection

Demographic and clinical data (age, occupation, smoking history, comorbidities, medications) were collected using a structured checklist. Serum total and free PSA were measured at three time points: baseline (prior to BCG therapy), immediately after completion of BCG induction, and three months post-treatment. Patients with incomplete BCG courses or missing follow-up data were excluded from final analysis. All data were managed confidentially in accordance with ethical guidelines.

Sample size calculation

Based on Beltrami et al. [11], which reported mean PSA levels of 1.58 ng/ml (IQR 0.79–2.39 ng/ml) at baseline, 3.45 ng/ml (IQR 1.49–5.54 ng/ml) at the end of induction (post-treatment), and 3.68 (IQR 1.64–5.36 ng/ml) at 3 months post-BCG, a minimum sample size of 25 (with 25% dropout adjustment) was estimated for 90% power, α = 0.05, assuming a mean paired difference of 1.9 ± 1.81 ng/mL.

Statistical analysis

Data were analyzed using SPSS version 27 (IBM Corp., Armonk, NY, USA). Continuous variables were expressed as mean ± standard deviation (SD) and median [interquartile range (IQR)], and categorical variables as frequencies and percentages. Normality of distribution was assessed using the Kolmogorov–Smirnov test. Given the non-parametric nature of the total PSA values, the Friedman test was used to compare the three time points, followed by post-hoc pairwise comparisons using Wilcoxon signed-rank tests. To evaluate temporal changes in the f/tPSA ratio across the three time points, a repeated-measures General Linear Model (GLM) was performed. The assumption of sphericity was tested using Mauchly’s test. Associations between age and total PSA levels at each time point were evaluated using Spearman’s rank correlation coefficient (ρ). Data were visualized with pandas, matplotlib, seaborn, and plotly libraries in Python 3.13.7. A two-tailed p-value < 0.05 was considered statistically significant.

Results

Demographic data

A total of 25 patients were initially recruited, of whom 20 completed follow-up and were included in the final analysis. The mean age was 64.0 ± 8.4 years (range 55–85). The majority of patients (80%) were aged between 50 and 69 years (40% each in the 50–59 and 60–69 year categories). Only a small proportion of patients were aged 70 years or older (20% in total). One-half of the participants reported a history of smoking, and 15% had history of type 2 diabetes mellitus.

Serum total PSA and Free-to-Total PSA ratio

Baseline total PSA (TPSA) averaged 1.17 ± 1.24 ng/mL, rising markedly to 2.02 ± 1.48 ng/mL immediately following BCG therapy, and subsequently returning to 1.27 ± 1.24 ng/mL at three months. The Free-to-Total PSA ratio (f/tPSA) exhibited a steadier pattern, declining from 0.30 ± 0.16 ng/mL at baseline to 0.26 ± 0.12 ng/mL post-treatment, before slight increasing to 0.28 ± 0.16 ng/mL at the three-month follow-up. (Table 1; Fig. 1).

Table 1.

Serum PSA levels and pairwise comparisons following intravesical BCG therapy

Parameter	Timepoint	Median [IQR] (ng/mL)	Mean ± SD (ng/mL)	Change	p-value
TPSA	Baseline (pre-BCG)	0.67 [0.21–1.99]	1.17 ± 1.24	—	—
	Immediately post-BCG	1.48 [0.90–3.18]	2.02 ± 1.48	↑ vs. Baseline	< 0.001 *
	Three months post-BCG	0.80 [0.28–2.00]	1.27 ± 1.24	↓ vs. Post-BCG	< 0.001 *
				↔ vs. Baseline	0.339
f/tPSA	Baseline (pre-BCG)	0.27 [0.19–0.38]	0.30 ± 0.16	—	—
	Immediately post-BCG	0.25 [0.21–0.31]	0.26 ± 0.12	↓ vs. Baseline	0.921
	Three months post-BCG	0.25 [0.17–0.39]	0.28 ± 0.16	↑ vs. Post-BCG	1.000
				↓ vs. Baseline	1.000

PSA Prostate specific antigen, TPSA Total PSA, f/tPSA Free-to-Total PSA ratioPairwise p-values: Wilcoxon Signed-Rank test for TPSA, repeated measures General Linear Model with Bonferroni adjustment for f/tPSA ratio

Directional symbols indicate changes over time: ↑ = increase, ↓ = decrease, and ↔ = no significant change

Statistically significant results (p < 0.05) are shown in bold and marked with an asterisk (*)

Fig. 1.

Boxplots showing temporal dynamics of PSA levels following intravesical BCG therapy. Left panel: Total PSA levels at baseline, immediately post-BCG, and three months post-BCG. Right panel: Free-to-Total PSA ratio (f/tPSA) at the same time points

Analysis of TPSA using the Friedman test demonstrated a significant time-dependent change across the three timepoints (p < 0.001). Post-hoc Wilcoxon Signed-Rank tests revealed significant temporal changes in total TPSA over the three time points. Pairwise comparisons demonstrated a significant increase in TPSA immediately following BCG therapy relative to baseline (p < 0.001), whereas no significant difference was observed between baseline and the three-month follow-up (p = 0.339). TPSA at three months was significantly lower than immediately post-treatment (p < 0.001) (Table 1).

The free-to-total PSA (f/tPSA) ratio was normally distributed across all time points. Analysis using a repeated measures General Linear Model (GLM) showed no statistically significant change in the f/tPSA ratio between the three measurement time points (p = 0.617). Overall, the f/tPSA ratio remained relatively stable throughout the treatment and follow-up period, indicating intravesical BCG therapy did not produce a measurable alteration in the ratio (Table 1).

There was no significant correlation between age and TPSA at any time point (Baseline: r = 0.010; Immediately post-BCG: r = − 0.035; Three months post-BCG: r = − 0.036; all p > 0.05). In contrast, strong positive correlations were observed among TPSA measurements at different time points. TPSA measured immediately after BCG therapy correlated significantly with both baseline TPSA (r = 0.771, p < 0.001) and TPSA at three months (r = 0.855, p < 0.001). Similarly, TPSA at three months showed a strong positive correlation with baseline TPSA (r = 0.787, p < 0.001) (Fig. 2).

Fig. 2.

Spearman’s correlation between age and total PSA levels at different time points. Statistical significance indicated as p < 0.05 (*). Positive correlations are shown in red, negative correlations in blue, with intensity proportional to the magnitude of correlation coefficient.)

Linear regression was conducted to identify predictors of TPSA at three months post-BCG. Baseline TPSA emerged as the sole significant predictor (B = 0.87, 95% CI: 0.63–1.11, p < 0.001). In contrast, neither age (B = − 0.099, p = 0.421) nor TPSA immediately post-BCG (B = 0.446, p = 0.077) contributed significantly to the model.

Discussion

The purpose of this study was to gain a better understanding of the temporal fluctuations in serum total PSA levels (TPSA) and Free-to-Total PSA ratio (f/tPSA) in patients with non-muscle-invasive bladder cancer (NMIBC) undergoing intravesical Bacillus Calmette–Guérin (BCG) therapy. While PSA is widely used as a biomarker for prostate malignancy, transient elevations following BCG instillation can pose diagnostic challenges, potentially leading to unnecessary prostate biopsies. The results of the present study support the hypothesis that BCG therapy induces a significant yet temporary inflammatory response in the prostate, reflected by a transient PSA elevation. The key findings indicate a significant increase in TPSA from a mean of 1.17 ± 1.24 ng/mL to 2.02 ± 1.48 ng/mL immediately following the induction course (p < 0.001). Crucially, this elevation was self-limiting, with TPSA levels returning to a baseline state (1.27 ± 1.24 ng/mL) at the three-month post-treatment mark, showing no significant difference from the initial measurement (p = 0.339). Taken together, our findings underscore that a PSA rise immediately post-BCG is predominantly a consequence of a localized inflammatory response rather than an indicator of malignancy, supporting a cautious “watchful waiting” clinical approach.

This pattern of results is consistent with the previous literature. The core observation of a temporary PSA increase following BCG instillation is a consistent theme across multiple studies. The present results are consistent with Leibovici et al.’s [14] work that deals with PSA elevation post-BCG, where they reported a significant increase in 75% of patients that returned to baseline within three months. Furthermore, our findings provide supporting evidence that the normalization window is typically around three months, as Serretta et al. [16] also found that PSA elevations above 4 ng/ml were rare and, when present, were no longer evident by the 90-day mark. They also observed PSA elevation in more than 70% of patients during intravesical therapy, independent of whether BCG or chemotherapy was used. Notably, they found the increase was greater in the BCG group (0.3 vs. 0.1 ng/ml). This led them to recommend monitoring and proposing biopsy only if the elevation persists beyond three months. Our study strengthens this consensus by demonstrating the same temporal pattern using repeated-measures analysis in a cohort with strictly normal baseline PSA.

Our findings are in line with those of Beltrami et al. [11], who prospectively evaluated 106 intermediate or high-risk NMIBC patients (77 cases treated with immunotherapy and 29 with chemotherapy). The median PSA in their immunotherapy group increased from 1.58 ng/mL at baseline to 3.45 ng/mL at the end of induction, peaked at 3.68 ng/mL at three months, and subsequently declined to 1.67 ng/mL at twelve months. By contrast, PSA levels in their intravesical chemotherapy group exhibited steadier trend (1.58, 1.90, and 1.68 ng/mL, respectively). This detailed kinetic profile supports our observation of a significant but temporary inflammatory response specific to BCG. The results of Ogreden et al. [17] also support our observations, showing a significant mean PSA increase from 1.7 ng/mL pre-treatment to 4.3 ng/mL post-induction BCG, though their single follow-up point limited the analysis of temporal dynamics. Collectively, these studies confirm the temporal nature of PSA elevations and the absence of clinical prostate malignancy during follow-up, emphasizing that PSA fluctuations are a benign, treatment-related phenomenon.

In our cohort of NMIBC patients with baseline PSA levels below 4 ng/mL, the f/tPSA ratio remained remarkably stable throughout the course of intravesical BCG therapy, despite transient elevations in TPSA. The median f/tPSA ratio was 0.27 at baseline, declined modestly to 0.25 immediately following BCG instillation, and remained at 0.25 three months post-treatment, with pairwise comparisons indicating no statistically significant changes over time. Importantly, 15–20% of patients exhibited an f/tPSA ratio below 0.15 at any time point, suggesting that the majority of patients remained within a range considered low-risk for prostate cancer. There are similarities between our results those of previous studies in men with PSA < 4 ng/mL, where an f/tPSA cutoff of 0.15 demonstrated 80% sensitivity and 81% specificity for prostate cancer detection, with a negative predictive value of 96% [18]. Although recommended cutoff values for f/tPSA can vary, with no universally accepted international standard, clinical practice generally applies a 25% threshold for men with PSA levels between 4.0 and 10.0 ng/mL, and a 15% threshold for men with lower PSA levels [18–21].

Despite the overall agreement on the PSA fluctuation phenomenon during BCG therapy, some variability exists among studies, prompting consideration of methodological and biological factors that might explain these inconsistencies. These discrepancies across studies may be explained by differences in study methodology, including variations in BCG strain, induction schedules, or patient characteristics. One interpretation of these findings is that while a general trend of normalization exists, a subset of patients may experience a more prolonged inflammatory response. A crucial dimension for interpreting PSA elevation is provided by Hong et al. [22], who introduced the variable of bladder tumor location. Their research demonstrated that the detection of prostate cancer after BCG therapy was highly associated with the tumor’s location, being significantly more likely when the tumor was distant from the bladder neck. Although our study did not stratify patients by tumor location, this finding should be considered in the context for interpreting PSA elevations.

In addition, other patient-specific characteristics, such as age and prostate volume, may further modulate PSA kinetics during and after BCG therapy. The absence of a significant correlation between age and PSA fluctuation in our cohort aligns with prior studies [11, 23], although some reports suggest a slightly delayed normalization in older patients [14], potentially due to larger prostate volumes or coexisting benign prostatic hyperplasia (BPH). Variability in PSA responses among patients may be explained by differences in immune response to BCG, tumor proximity to the bladder neck, prior urologic procedures, or comorbid prostatic conditions.

One explanation for the present set of findings is that intravesical BCG instillation causes a localized, granulomatous prostatitis, leading to the transient release of PSA into the serum. This idea is further supported by results of the study by Akagi et al. [24], which provides compelling evidence by extending the clinical implications to evaluate the diagnostic yield of prostate biopsy in NMIBC patients who had previously undergone intravesical BCG. They found that prostate biopsies performed within one year of BCG treatment had a significantly lower cancer detection rate (13.9%) compared to a control group (78.5%), with all cancer diagnoses in their BCG cohort occurring more than one-year post-treatment. This strongly supports the “watchful waiting” approach and aligns perfectly with our observation, underscoring the importance of cautious interpretation of PSA increases in the early post-BCG period and recommendation to defer prostate biopsy unless PSA elevations persist beyond 12 months or are when accompanied by progressive suspicious findings. They also described that serum PSA levels rose significantly following BCG (from 3.2 to 5.8 ng/mL) but subsequently declined toward baseline (4.0 ng/mL) on follow-up, which reinforces the transient and non-malignant nature of PSA elevation following immunotherapy.

Taken together, these findings integrate into a coherent body of evidence emphasizing the benign, self-limited, and inflammation-driven nature of PSA elevation following BCG instillation.

Strengths and limitations

There are some limitations that must be acknowledged when interpreting our findings. First, the study’s single-center design, small sample size, and absence of a control group constrain statistical power and generalizability. Second, our study relied solely on biochemical markers (PSA) and did not include histological confirmation via prostate biopsy. Additionally, the lack of concurrent imaging (e.g., TRUS or MRI) and inflammatory biomarker assessments, combined with a follow-up limited to three months, restricts the ability to evaluate long-term PSA dynamics and the broader applicability of our findings. Finally, potential confounding variables were not systematically analyzed. As discussed, the precise location of the bladder tumor is a critical factor that was not recorded or controlled for in our analysis. Other factors, such as urinary tract infections, recent catheterizations or digital rectal examination, variations in prostate volume, individual patient inflammatory responses, could also influence PSA levels and were not accounted for.

Future research should aim to address these limitations by conducting larger, multi-center studies with extended follow-up periods. Incorporating tumor location as a key variable and correlating PSA changes with multiparametric MRI findings and, where clinically indicated, histopathology, would provide a more comprehensive understanding of this clinical issue and further refine patient management strategies.

Conclusions

The elevation in total PSA levels following intravesical BCG therapy is a significant yet transient phenomenon, typically resolving within a three-month period. This fluctuation primarily reflects an inflammatory response rather than underlying prostate malignancy, emphasizing the importance of interpreting PSA values within temporal context. Clinicians should postpone prostate biopsy in the absence of supporting clinical or imaging findings, opting instead for repeat PSA testing and close monitoring. These insights can guide evidence-based clinical decision-making, minimize unnecessary invasive interventions, and support the development of refined management protocols for patients undergoing BCG therapy.

Abbreviations

ASR

Age-standardized incidence rate

BCG

Bacillus calmette–guérin

DRE

Digital rectal examination

f/tPSA

Free-to-total prostate-specific antigen ratio

IQR

Interquartile range

NMIBC

Non–muscle-invasive bladder cancer

PSA

Prostate-specific antigen

SD

Standard deviation

SPSS

Statistical package for the social sciences

TURBT

Transurethral resection of bladder tumor

TPSA

Total prostate-specific antigen

Authors’ contributions

- [S.P.A] performed statistical analysis and generated figures, assisted in data interpretation, and contributed to manuscript writing.- [F.S] assisted in data interpretation, and contributed to manuscript writing.- [B.K] assisted in patient recruitment, and collected clinical and laboratory data.- [Z.S] conceptualized the study, supervised patient recruitment, and contributed to manuscript writing.- All authors read and approved the final manuscript.

Funding

This study was supported by “Mashhad University of Medical Sciences” (Grant No. 4020504”). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.

Data availability

The dataset generated and analyzed during the current study are available from the corresponding author on reasonable request.

Declarations

Ethics approval and consent to participate

This prospective cohort study was conducted in accordance with the Declaration of Helsinki and was approved by the Institutional Review Board and Ethics Committee of Mashhad University of Medical Sciences (IR.MUMS.MEDICAL.REC.1402.228). Written informed consent was obtained from all participants prior to enrollment.

Consent for publication

Not applicable. No individual patient data or identifiable images are included in this manuscript.

Competing interests

The authors declare no competing interests.