Stesura Seveso Archivio Italiano di Urologia e Andrologia 2020; 92, 4330 ORIGINAL PAPER No conflict of interest declared. DOI: 10.4081/aiua.2020.4.330 Perineural invasion in prostate needle biopsy: Prognostic value on radical prostatectomy and active surveillance Nuno Ramos, Alexandre Macedo, João Rosa, Miguel Carvalho Urology Department, Garcia de Orta Hospital, Almada, Portugal. tive treatment options such as radical prostatectomy (RP), external beam radiotherapy and brachytherapy to con- servative management strategies including active surveil- lance (AS) (2-5). Despite the use of adequate therapy in localized PCa, approximately 18% of patients will even- tually experience biochemical recurrence (BCR) (1, 6). Pathological stage, preoperative prostate-specific antigen (PSA) levels and Gleason score (GS) are widely used as risk factors for BCR (7). In other hand, AS has been increasingly adopted to prevent overtreatment in men with low-risk prostate cancer (8). This strategy pretends to identify patients with clinically indolent tumors and avoid or delay definitive treatment, without compromis- ing survival (9). Although the concept of AS is well established, there is no consensus regarding the optimal characteristics of patients who should be managed by this strategy. Therefore, there is a growing interest in the identification of new clinicopathological features in prostate needle biopsy specimens to improve the evalu- ation of the likelihood of BCR, as improving the selec- tion of patients to AS (2, 4, 10). In this setting, perineur- al invasion (PNI) has been increasingly recognized as prognostic marker (11). PNI is a histopathologic finding representing the infiltration of cancer cells in, through and/or around nerves and is present in 7%-43% of prostate needle biopsies with PCa (4, 12, 13). PNI has been implicated in PCa cell proliferation and extrapro- static spread, and the presence of PNI has been shown to be associated with adverse oncological outcomes (14- 16). Despite of PNI being a potential determinant in PCa behaviour, the association between PNI and PCa pro- gression is still a subject of debate. Due to the uncer- tainty regarding the role of PNI, we performed a study to evaluate the association of PNI in prostate needle biopsy with adverse pathological findings on RP specimens, as well the impact on BCR. We also pretend to evaluate the role of PNI in patients’ selection for AS (11). MATERIALS AND METHODS We performed a single center retrospective cohort study, on male patients, who underwent RP due to clinically localised PCa, from January 2010 to December 2016. All patients underwent a 12-core biopsy prior to RP and presence or absence of PNI was assessed. PNI was defined as the histopathologic finding of circumferential Purpose: The aim of this study was to evalu- ate the clinical impact of perineural inva- sion (PNI) in prostate biopsy in patients submitted to radical prostatectomy and on active surveillance (AS). Materials and methods: We performed a single center, retro- spective, cohort study on patients diagnosed with clinically localized prostate cancer and submitted to radical prostatecto- my between January 2010 and December 2016. We evaluated clinical and anatomopathological characteristics from the biop- sy and radical prostatectomy specimen and correlated with biochemical recurrence (BCR) using a survival analysis. We also evaluated the impact of PNI in patients with criteria for active surveillance. Results: The cohort analyzed consists of 107 patients, with a mean age of 63.1 years and a mean PSA prior to biopsy of 7.8 ng/ml. In prostate biopsy, 66.4% of the patients had a Gleason score of 6, 30.9% had a Gleason score of 7, and 2.7% had a Gleason score of 8 or higher, with PNI being detected in 57 (53.3%) of the patients. Regarding the anatomopathological characteristics of the surgical specimen, invasion of the semi- nal vesicles was observed in 6.5%, lymph nodes involvement in 9.3% and positive surgical margins in 27.1% of the cases. During follow-up, BCR was recorded in 24.3% of cases. Clinicopathological features were stratified according to the presence or absence of PNI, with statistical significance in relation to the Gleason Score (p = 0.001), pathologic T stage (p = 0.001), D’Amico risk (p = 0.002) and upstaging of the Gleason score (p = 0.045). The survival analysis revealed a relationship between PNI and BCR (hazard ratio = 2.98; 95% CI: 1.36-6.58; p = 0.007). Regarding the men potentially eligi- ble for AS, the presence of PNI on the biopsy presented a sig- nificant relation with Gleason upgrade (p = 0.004) and extraprostatic extension (p = 0.017). Conclusions: The presence of PNI in prostate biopsy is related to adverse anatomopathological factors, being a potential pre- dictor of BCR and have a possible role in the selection of patients for AS. KEY WORDS: Prostate cancer; Active Surveillance; Prostatectomy. Submitted 22 July 2020; Accepted 3 September 2020 INTRODUCTION Prostate cancer (PCa) is the second most commonly diag- nosed cancer in men and the third cause of cancer relat- ed death (1, 2). The best treatment strategies in clinical- ly localized PCa remains unclear and varies from defini- Summary 331Archivio Italiano di Urologia e Andrologia 2020; 92, 4 Perineural invasion in prostate needle biopsy or longitudinal tracking of PCa cells along a nerve, with- in the perineural space. PNI was not always reported on pathology reports and we excluded patients with unknown PNI status to minimize the potential for mis- classification bias. Clinical and the biopsy parameters were evaluated, including age, PSA level prior to surgery, prostate volume, number of positive cores, total percent of core involvement and GS. We also assessed histopathologic finding on RP specimens: GS, margin positivity, stage, seminal vesicle involvement and lym- phatic invasion. The primary objective of this study was to report the association between PNI in prostate needle biopsy and adverse pathological findings on RP specimen, specifical- ly, the presence of extraprostatic extension, surgical Gleason upgrading, positive surgical margin, and lymph node involvement. Gleason upgrading was defined as pathologic GS higher than the GS in the prostate biopsy. A sub-analysis was focused on evaluating the role of PNI on BCR following RP, defined as two successive postoper- ative PSA values of 0.2 ng/mL or greater. A secondary objective of this study, was to evaluate the impact of PNI in prostate needle biopsy on the selection of patients for AS. The AS cohort was based on the patients who per- formed RP but could be potentially selected for AS, defined according to the Epstein criteria (clinical stage ≤ T2a, PSA density < 0.15 ng/mL, PSA < 10 ng/mL, biopsy Gleason score ≤ 6, ≤ 2 positive biopsy cores, and ≤ 50% cancer involvement in any biopsy core). A descriptive analysis on the study population was performed. Categorical data were compared using Pearson's chi- squared test and continuous variables with Student t-test. The biochemical recurrence-free survival was calculated through the Kaplan-Meier analysis. To estimate the prog- nostic value of PNI we used Cox proportional hazard regression. A two-sided p value < 0.05 was considered as statistically significant. Statistical analysis was performed using SPSS®, version 23.0 (SPSS Inc., Chicago, IL, USA). RESULTS A total of 107 patients were included in the study, of whom 57 (53.3%) had PNI of the biopsy specimens. The demographic and the clinical characteristics of the patients are shown in Table 1. Patient age ranged from 48 to 73 years (mean, 63.1 years; standard deviation (SD), 5.3 years). Preoperative serum PSA levels ranged from 1.51 to 21.9 ng/mL (mean, 7.8 ng/mL; SD, 3.9 ng/mL) and the clinical T stage was T1c in 48 (44.9%) of the patients, T2a in 28 (26.2%), T2b in 20 (18.7%) and T2c in 11 (10.3%). According to D’Amico risk classification 65 (60,7%) patients were low risk, 29 (27,1%) interme- diate risk and 13 (12,2%) high risk. From the prostate biopsy specimens, 71 (66.4%) had a Gleason score of 6, 33 (30.9%) had a Gleason score of 7, and 3 (2.7%) had a Gleason score of 8 or higher. The mean number of cores involved by tumor was 4.83 (SD, 1.5), and the mean percent of core involvement was 25.8% (SD, 20%). Pathologic findings at RP are summarized in Table 2. Regarding the RP specimens, 25 (23.4%) showed extraprostatic extension and 29 (27.1%) had positive margins. The seminal vesicles were invaded in 7 (6.5%) cases and lymph nodes were involved in 10 (9.3%). Surgical GS upgrading was observed in 26 (24.2%) patients. The mean follow-up time was 71 months (SD 25.3 months) and, in this period, 26 (24.3%) men expe- rienced BCR. Clinical and anatomopathological characteristics from the biopsy and RP specimen were stratified according to the presence or absence of PNI. Significant differences were found, patients with PNI presented higher GS on biopsy and RP specimen (p = 0.036 and p = 0.001), Table 1. Demographic, clinical and prostate biopsy characterization. PNI absence PNI presence Total p (n = 50, 46.7%) (n = 57, 53.3%) (n = 107) Age, mean (years) 62.4 (SD 5.2) 63.7 (SD 5.4) 63.1 (SD 5.38) 0.189 PSA level (ng/ml) 7.64 (SD 3.6) 7.94 (SD 4.5) 7.8 (SD 3.9) 0.53 Prostate volume(mean, gr) 46.5 (SD 17.5) 45.1 (SD 14.8) 45.8 (SD 16.1) 0.46 GS biopsy 0.036 6 39 (78%) 32 (56.1%) 71 (66.4%) 7 10 (20%) 23 (40.3%) 33 (30.9%) ≥ 8 1 (2%) 2 (3.6%) 3 (2.7%) Number of positive cores (mean) 4.56 (SD 2.36) 5.07 (SD 2.77) 4.83 (SD 2.5) 0.31 Percent of core involvement (mean) 19.48% (SD 17%) 31.3%(SD 20.95%) 25.8% (SD 20%) 0.04 Clinical stage 0.019 T1c 23 (46%) 25 (43.9%) 48 (44.9%) T2a 20 (40%) 8 (14%) 28 (26.2%) T2b 6 (12%) 14 (24.6%) 20 (18.7%) T2c 1 (2%) 10 (17.6%) 11 (10.3%) Age, PSA level prior surgery, clinical stage and anatomopathological characteristics from biopsy stratified in two groups: patients with PNI absence and patients with PNI presence in the biopsy. Categorical data were compared using Pearson's chi-squared test and continuous variables with Student t-test. Table 2. Clinical and histopathologic finding on RP specimens’ characterization. PNI absence PNI presence Total p (n = 50, 46.7%) (n = 57, 53.3%) (n = 107) D’Amico risk 0.002 Low 38 (76%) 27 (47.4%) 65 (60.7%) Intermediate 10 (20%) 19 (33.3%) 29 (27.1%) High 2 (4%) 11 (19.3%) 13 (12.2%) Gleason score 0.001 6 34 (68%) 20 (35.1%) 71 (66.4%) 7 16 (32%) 34 (59.6%) 33 (30.9%) ≥ 8 - 3 (5.3%) 3 (2.7%) Pathological stage 0.001 pT2a 8 (16%) 7 (12.3%) 15 (14%) pT2b 5 (10%) 1 (1.8%) 6 (5.6%) pT2c 33 (66%) 27 (47.4%) 60 (56.1%) ≥ pT3 4 (8%) 22 (38.6%) 26 (24.3%) GS upgrading 8 (16%) 18 (31.6%) 26 (24.2%) 0.065 Lymphatic invasion 3 (6%) 7 (12.3%) 10 (9.3%) 0.27 SV invasion 1 (2%) 6 (10.5%) 7 (6.5%) 0.11 Extraprostatic extension 4 (8%) 21 (36.8%) 25 (23.4%) 0.001 Margin positivity 8 (16%) 17 (29.8%) 25 (23.4%) 0.09 BCR 9 (18%) 22 (38.6%) 31 (29%) 0.032 Clinical and histopathologic finding on RP specimens stratified in two groups: patients with PNI absence and patients with PNI presence. Categorical data were compared using Pearson's chi-squared test and continuous variables with Student t-test. SV-seminal vesicle. Archivio Italiano di Urologia e Andrologia 2020; 92, 4 N. Ramos, A. Macedo, J. Rosa, M. Carvalho 332 higher percent of core involvement (p = 0.04), higher clinical and pathologic T stage (p = 0.019 and p = 0.001), higher D’Amico risk (p = 0.002) , extrapro- static extension (p = 0.001), and higher BCR (p = 0.032) comparing with patients without PNI. On other hand, no significant differences were identified with respect to age (p = 0.189), preoperative PSA (p = 0.53), number of positive cores (p = 0.31), GS upgrading (p = 0.065), surgi- cal margin involvement (p = 0.09), lymph node invasion (p = 0.27) and seminal vesicles involvement (p = 0.11). The Kaplan-Meier curve (Figure 1) revealed a poorer recurrence-free survival in patients with PNI (Log rank test p = 0.04). On uni- variate Cox analysis (Table 3), PNI was asso- ciated with BCR (HR: 2.98, 95% CI: 1.36- 6.58, p = 0.007), as GS on RP specimen (HR: 3.01, 95% CI: 1.34-6.76, p = 0.008), surgical margin positivity (HR: 3.86, 95% CI: 1.90- 7.94, p = 0.0001) and presence of extrapro- static extension (HR: 4.02, 95% CI: 1.98-8.15 p = 0.0001). However, the prognostic role of PNI disappeared in multivariate analysis when adjusted for other predictive factors. In the cohort, a total of 29 men submitted to RP were potentially eligible for AS, of whom 13 (44.8%) had biopsy PNI. The characteris- tics of this subgroup are listed in Table 4. A significant relation between PNI and GS upgrade (p = 0.004) and extraprostatic extension (p = 0.017) was found in the AS group. DISCUSSION The evaluation of pathological features that may predict oncologic outcomes are important for counselling patients and therapy selection, as treatment options in PCa differ according to defined risk groups (17). PNI was identified as a possible significant marker of adverse pathologic findings in localized PCa, however, the onco- logical significance and prognostic value is still contro- versial, with inconsistent results among studies (13). The incidence of PNI in biopsy specimens in the present study is similar to that reported by Ravery et al. (53,3% VS 47%) (18, 19). Although, a wide range of incidences have been reported in the literature due to variation in pathologic definitions and interpretation of PNI (16). There are many challenges in looking for PNI and a neg- ative result may either indicate that there no nerves iden- tified in the biopsy or that nerves were present without invasion (2). On our study, PNI presence on the biopsy cores was associated with adverse parameters in RP specimens, including higher GS, higher pathologic T staging and extraprostatic extension. Such associations were also reported in a recent meta-analysis were PNI was a signif- icant marker to predict high stage disease (4). Despite hypothesized that spread along intraprostatic nerves may facilitating extraprostatic tumoral extension, there are controversial results in studies investigating the correlation between PNI and BCR. Jeon et al. and Kang et al. showed that PNI is associated with adverse patholog- ic findings and is an independent predictor for BCR in PCa patients who undergo RP (6, 20). The same results were observed by Yu et al. and Wong et al. in patients who undergo external beam radiotherapy (21, 22). On the Figure 1. Kaplan-Meier curve and Log rank test. Biochemical recurrence-free survival in patients with or without PNI. Table 3. Association between disease characteristics and biochemical recurrence-free survival. Univariate Multivariate HR (95% IC) P value HR (95% IC) P value GS in RP specimen < 7 1 ≥ 7 3.01 (1.34 -6.76) 0.008 1.65 (1.04-2.60) 0.032 Extraprostatic extension absence 1 presence 4.02 (1.98-8.15) 0.0001 1.26 (0.48-3.27) 0.64 Surgical margin negative 1 positive 3.89 (1.90-7.94) 0.0001 3.51 (1.45-8.52) 0.005 PNI absence 1 presence 2.98 (1.36-6.58) 0.007 1.64 (0.7-3.86) 0.25 Univariate and multivariate Cox models. CI: confidence interval; HR: hazard ratio; GS: Gleason score. Table 4. Clinical and histopathologic finding on active surveillance patients. PNI absence PNI presence Total p (n = 16, 55.2%) (n = 13, 44.8%) (n = 29) Age, mean (years) 62.5 (SD 4.9) 63.3 (SD 6.7) 62.86 (52-73) 0.362 PSA level (ng/ml) 7.31 (SD 4.9) 7 (SD 4.1) 7.1 (SD 4.2) 0.41 Prostate volume (mean, gr) 56.9 (SD 24.5) 48.7 (SD 18.4) 53.3 (SD 22) 0.31 GS upgrading 1 (6.3%) 7 (53.8%) 8 (27.6%) 0.004 extraprostatic extension - 4 (30.8%) 4 (13.8%) 0.017 CSub-group of patients potentially selected to active surveillance. Clinical and histopathologic finding stratified in two groups: patients with PNI absence and patients with PNI presence in the prostate biopsy. 333Archivio Italiano di Urologia e Andrologia 2020; 92, 4 Perineural invasion in prostate needle biopsy contrary, Reeves et al. and Freedland et al. reported that PNI is not correlated with BCR in PCa after RP (23, 24). In our cohort, we found an association between PNI and BCR on univariate but not on multivariate analysis. This questions whether PNI is an independent prognostic fac- tor or just a risk factor for BCR, since significance is lost when PNI presence is controlled for other biopsy param- eters, such as Gleason score and extraprostatic extension (25). One possible explanation for these findings is that PNI may only be an important prognostic factor in a spe- cific sub-group of patients. D’Amico et al. found that biopsy PNI showed statistical significance on multivari- ate analysis only in the low risk group and Quinn et al. reported that biopsy PNI was a significant prognostic fac- tor on multivariate analysis of patients with PSA values more than 10 ng/ml (26, 27). In the same setting, rather than evaluated the presence or absence of PNI, quantification could have a better pre- dictive value. Maru et al. found that PNI diameter > 0.25 mm was an independent prognostic indicator for bio- chemical recurrence on multivariate analysis (28). Moreover, Sun et al. demonstrated that multifocal PNI, rather than unifocal PNI, is correlate to shorter biochem- ical recurrence-free survival in patients with PCa (10). AS has been widely accepted as an observational strategy, in the last decade, in response to the over-treatment of men with low-risk PCa (16). The selection of patients is based on pathologic findings on needle core biopsy. At present, biopsy PNI is not included in the established criteria for AS selection, thus, whether PNI is potentially associated with worse prognosis and preclude a conservative management is not known. In our analysis, the presence of PNI in patients who met criteria for AS has been associated with adverse pathologic findings at prostatectomy, including GS upstaging and extraprostatic extension. These finding sug- gest a potential role for biopsy PNI in identifying men at risk for progression on AS. This evidence is corroborated by a retrospective review of the REduction by Dutasteride of clinical progression Events in Expectant Management (REE- DEM) study, with 302 men on AS, who describe that PNI is an independent predictor of clinical progression (73% after 2 years) (29). Similarly, in a cohort of 165 men on AS, Cohn et al. reported that biopsy PNI remained a significant predictor for AS failure after adjustment biopsy parameters such as tumour length (30). While these data suggest that patients with biopsy PNI on initial biopsy may not be good candidates for AS, future study is required to assess prog- nostic value of this pathologic finding. Several limitations should be acknowledged in this study. First, the small number of cases, similar to other series published, limits the statistical power of the conclusions. Second, the study design, as a single-institution retro- spective analysis with risk of unmeasured bias, does not allow to generalize the conclusions. Third, prostatectomy specimens did not undergo a centralized review, so there may be variability in the reporting of PNI. Ideally, a mul- ticenter randomized prospective study with a larger sam- ple would answer many questions raised in our study. Fourth, we only report PNI as a binary variable because quantitative measures of PNI were not included in pathology reports. Finally, a short follow-up with a mean of 71 months can undervalue the BCR. CONCLUSIONS In conclusion, despite the limitations listed above, this study recognizes the clinicopathological importance and potential prognostic value of PNI in PCa. The presence of PNI on prostate biopsy cores is an important predictive of aggressive disease in patients submitted to RP with clinically localized PCa and an indicator of BCR in uni- variate analysis. Additionally, among men who met crite- ria for AS, biopsy PNI is associate with GS upgrade and extraprostatic extension and could have a role in the selection of patients for AS. 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The prognostic significance of perineural invasion and race in men considering active surveil- lance. BJU Int. 2014; 114:75-80. Correspondence Nuno Ramos, MD (Corresponding Author) nuno.ramos@hgo.min-saude.pt nunoandre33@gmail.com Alexandre Macedo, MD alex.m.macedo89@gmail.com João Rosa, MD jpmrosa@yahoo.com Miguel Carvalho, MD uro.miguelcarvalho@gmail.com Urology Department, Garcia de Orta Hospital Av. Torrado da Silva, 2801-951, Almada (Portugal)