Serial Molecular Profiling of Low-grade Prostate Cancer to Assess Tumor Upgrading: A Longitudinal Cohort Study

Abstract

Background: The potential for low-grade (grade group 1 [GG1]) prostate cancer (PCa) to progress to high-grade disease remains unclear.

Objective: To interrogate the molecular and biological features of low-grade PCa serially over time.

Design, setting, and participants: Nested longitudinal cohort study in an academic active surveillance (AS) program. Men were on AS for GG1 PCa from 2012 to 2017.

Intervention: Electronic tracking and resampling of PCa using magnetic resonance imaging/ultrasound fusion biopsy.

Outcome measurements and statistical analysis: ERG immunohistochemistry (IHC) and targeted DNA/RNA next-generation sequencing were performed on initial and repeat biopsies. Tumor clonality was assessed. Molecular data were compared between men who upgraded and those who did not upgrade to GG ≥ 2 cancer.

Results and limitations: Sixty-six men with median age 64 yr (interquartile range [IQR], 59-69) and prostate-specific antigen 4.9 ng/mL (IQR, 3.3-6.4) underwent repeat sampling of a tracked tumor focus (median interval, 11 mo; IQR, 6-13). IHC-based ERG fusion status was concordant at initial and repeat biopsies in 63 men (95% vs expected 50%, p < 0.001), and RNAseq-based fusion and isoform expression were concordant in nine of 13 (69%) ERG⁺ patients, supporting focal resampling. Among 15 men who upgraded with complete data at both time points, integrated DNA/RNAseq analysis provided evidence of shared clonality in at least five cases. Such cases could reflect initial undersampling, but also support the possibility of clonal temporal progression of low-grade cancer. Our assessment was limited by sample size and use of targeted sequencing.

Conclusions: Repeat molecular assessment of low-grade tumors suggests that clonal progression could be one mechanism of upgrading. These data underscore the importance of serial tumor assessment in men pursuing AS of low-grade PCa.

Patient summary: We performed targeted rebiopsy and molecular testing of low-grade tumors on active surveillance. Our findings highlight the importance of periodic biopsy as a component of monitoring for cancer upgrading during surveillance.

Keywords: Cancer progression; Gene fusions; Immunohistochemistry; Low-grade cancer; Next-generation sequencing; Prostate cancer; Tumor clonality.

Fig. 1 –

Potential clinical trajectories of grade group 1 (GG1) prostate cancer following diagnosis. In men on active surveillance for GG1 prostate cancer (yellow area), follow-up biopsies may reveal (A) stable GG1 disease, that is, no upgrading or (B) cancer upgrading (>GG1 disease; blue area). Cancer upgrading detected on follow-up biopsy could be explained by: (B1) clonal grade progression of the initially sampled cancer focus; (B2) initial undersampling of a high-grade component of the same cancer focus; (B3) an initially unsampled, anatomically distinct, high-grade cancer focus; and/or (B4) de novo development of high-grade cancer. Our findings support the occurrence of scenario B1 in at least some cases of upgrading. (C) Molecular analysis of GG1 cancer from initial biopsy was not predictive of cancer upgrading on follow-up biopsy. PCa = prostate cancer.

Fig. 2 –

Tracking low-grade prostate cancer with multiparametric magnetic resonance imaging/ultrasound (mpMRI/US) fusion biopsy platform. (A1) mpMRI (T2W MRI, axial cut shown) acquired prior to initial (t₀) prostate biopsy shows a PIRADS 5 region of interest (ROI; yellow enclosure) in the right posterior prostate peripheral zone. (A2) mpMRI/US fusion biopsy of the ROI and a systematic 12-core biopsy were performed. All biopsy sites were recorded and stored in a 3D reconstruction model to facilitate repeat sampling during surveillance (ROI—red, targeted [ROI] cores—yellow, and systematic cores—green). (A3) mpMRI prior to repeat biopsy (t₁, 11 mo later) shows the same ROI meeting PIRADS 4 criteria. (A4) All positive biopsy cores (ROI and systematic) were resampled, tracked, and stored in a 3D reconstruction model, as shown. (B1 and B2) H&E stain (10×) of ROI biopsy core and corresponding ERG IHC (10×) are shown, demonstrating GG1 cancer exhibiting positive ERG expression at t₀. (B3 and B4) At t₁, GG5 (Gleason score 4 + 5) cancer was sampled from the ROI with concordant positive ERG expression on IHC. The corresponding molecular data (from targeted DNA and RNA sequencing) at both time points are described in Supplementary Figure 2. Taken together, it is unclear whether upgrading in this case indicates undersampling due to intratumoral heterogeneity at initial biopsy or temporal clonal grade progression. Bx = biopsy; 3D = three dimensional; GG = grade group; H&E = hematoxylin and eosin; IHC = immunohistochemistry; PIRADS = Prostate Imaging Reporting and Data System; T2W = T2 weighted.

Fig. 3 –

Integrative targeted genomic and transcriptomic profiling of tracked prostate cancer foci. Each column represents a patient sample ordered by patient number and by time points (t₀ and t₁). Paired samples from the same patient are indicated with “A” and “B” to represent biopsy time points t₀ and t₁, respectively. The panel to the left includes patients with evaluable samples at both time points, while the panel to the right includes patients with data available at one time point only. Patients who progressed to higher-grade disease at time t₁ are denoted by an asterisk (*). Overall, 63 (95%) patients showed concordant ERG immunohistochemistry (IHC) status between paired t₀ and t₁ biopsies, supporting frequent sampling of the same clonal focus overtime. Among the 74 specimens passing NGS quality control, three (51A, 51B, and 60B) demonstrated heterogeneous staining on IHC, and thus could not be classified as concordant or discordant with the NGS-based ERG fusion assessment (positive vs negative). Among the remaining 71 samples, ERG fusion status by IHC and targeted RNA NGS were concordant in 69 (97%). All 11 samples with SPINK1 overexpression were ERG fusion negative. GG = grade group; ISUP = International Society of Urological Pathology; NGS = next-generation sequencing.