Molecular features of prostate cancer after neoadjuvant therapy in the phase 3 CALGB 90203 trial

Abstract

Background: The phase 3 CALGB 90203 (Alliance) trial evaluated neoadjuvant chemohormonal therapy for high-risk localized prostate cancer before radical prostatectomy. We dissected the molecular features of post-treated tumors with long-term clinical outcomes to explore mechanisms of response and resistance to chemohormonal therapy.

Methods: We evaluated 471 radical prostatectomy tumors, including 294 samples from 166 patients treated with 6 cycles of docetaxel plus androgen deprivation therapy before radical prostatectomy and 177 samples from 97 patients in the control arm (radical prostatectomy alone). Targeted DNA sequencing and RNA expression of tumor foci and adjacent noncancer regions were analyzed in conjunction with pathologic changes and clinical outcomes.

Results: Tumor fraction estimated from DNA sequencing was significantly lower in post-treated tumor tissues after chemohormonal therapy compared with controls. Higher tumor fraction after chemohormonal therapy was associated with aggressive pathologic features and poor outcomes, including prostate-specific antigen-progression-free survival. SPOP alterations were infrequently detected after chemohormonal therapy, while TP53 alterations were enriched and associated with shorter overall survival. Residual tumor fraction after chemohormonal therapy was linked to higher expression of androgen receptor-regulated genes, cell cycle genes, and neuroendocrine genes, suggesting persistent populations of active prostate cancer cells. Supervised clustering of post-treated high-tumor-fraction tissues identified a group of patients with elevated cell cycle-related gene expression and poor clinical outcomes.

Conclusions: Distinct recurrent prostate cancer genomic and transcriptomic features are observed after exposure to docetaxel and androgen deprivation therapy. Tumor fraction assessed by DNA sequencing quantifies pathologic response and could be a useful trial endpoint or prognostic biomarker. TP53 alterations and high cell cycle transcriptomic activity are linked to aggressive residual disease, despite potent chemohormonal therapy.

Figure 1.

Relationship between sequencing-based tumor fraction and pathologic features or clinical outcomes. A) Study design. B) Box plot showing the sequencing-based tumor fraction (also known as tumor purity) in samples from arm A vs arm B. At the bottom of the plot, horizontal blue or orange filled rectangles illustrate the proportion of samples in each arm that had evidence of residual tumor DNA by sequencing. C) Association between sequencing-based tumor fraction and pathologic tumor cellularity (from assessment of an hematoxylin-eosin slide) in each arm. No samples from arm B had <20% tumor cellularity on hematoxylin-eosin slides by pathology assessment. D) Association between sequencing-based tumor fraction and pathologic features in samples from arm A (for results from arm B samples, see Supplementary Figure 7, available online). E, F) Kaplan-Meier survival analysis for PSA progression-free survival and event-free survival in patients with and without sequencing-based tumor fractions detected in arm A. For patients with multiple samples, the sample with the highest sequencing-based tumor fraction was used. G) Multivariate analysis for patients in arm A. Twenty-five patients were excluded from this model for lack of annotation for intraductal carcinoma or unevaluable pathologic tumor cellularity (from sole use of core sections). P values were estimated using a Fisher exact test or Mann-Whitney U test (B, D) or univariate Cox proportional hazards regression analysis (E, F). CHT = chemohormone therapy; CI = confidence interval; HR = hazard ratio; ISUP = International Society of Urological Pathology; mRNA = messenger RNA; NR = not reported; PSA = prostate-specific antigen; pts = patients; RP = radical prostatectomy; REF = reference group.

Figure 2.

Common genomic alterations present in residual cancers after chemohormone therapy. A) Driver genomic landscape of residual high-risk, localized prostate cancer after treatment with neoadjuvant chemohormonal therapy before radical prostatectomy (arm A) vs prostate cancers not exposed to therapy (arm B). Only patients who had molecular residual disease are shown (ie, detected sequencing-derived tumor fraction: 91 in arm A and 82 in arm B). The highest sequencing-derived tumor fraction sample from each patient is represented if multiple same-patient tumor samples were available. Patients were sorted by the presence or absence of alterations in TP53 or SPOP and sequencing-derived tumor fraction (high to low). Fourteen genes associated with prostate cancer were provided (see Supplementary Figure 13 for additional genes, available online). Per-arm sequencing-derived tumor fraction estimates in SPOP-mutant and TP53-mutant tumors were compared using the Mann-Whitney U test. B, C, D) Kaplan-Meier survival analysis of PSA progression-free survival, event-free survival, and overall survival in patients according to TP53 alteration status (patients without residual tumor [ie, undetected sequencing-derived tumor fraction] who are unevaluable for TP53 alteration status are represented by the gray line). Statistical significance was measured using multivariate Cox proportional hazards regression analysis. Seq-TF = sequencing-based tumor fraction; CI = confidence interval; HR = hazard ratio; PSA = prostate-specific antigen; NR = not reached; REF = reference group.

Figure 3.

Transcriptomic changes in prostate cancer samples after chemohormonal therapy. A) Differentially expressed genes between arm A and arm B. The volcano plot shows the fold-change (x-axis) vs the significance (y-axis) of 155 core transcripts. Differentially expressed genes are established at change >2 and FDR < 0.05. Red dots indicate the genes that were significantly upregulated in arm A. Blue dots indicate the genes that were significantly downregulated in arm A. Black dots represent the remaining genes in the integrated panel that were not significantly changed after treatment therapy. B) Differences in the expression of AR, AR-variant 7 (AR-V7), FOLH1 (PSMA), androgen receptor–targeted genes (KLK2, KLK3, TMPRSS2), and a subset of neuroendocrine and plasticity genes (CD56, CHGB [chromogranin B], ENO2 [NSE], NEUROD1, SPDEF, SST [somatostatin]) in arm A vs arm B. C) Heatmap of gene expression levels (z-score) showing hierarchically clustered genes (rows; 155 core transcripts) and samples in arm A (columns), with dendrograms based on all core transcripts. TP53 and SPOP alteration status is provided for each sample, along with tumor fraction from targeted DNA sequencing. The dendrogram for samples is shown on top of the heat map. Values are measured by euclidean distance, with a complete linkage clustering algorithm. FC = fold change; FDR = false discovery rate; mRNA = messenger RNA; Seq-TF = sequencing-based tumor fraction.

Figure 4.

Cluster 1 and cluster 2 gene expression subgroups based on sequencing-based tumor fraction. A) Hierarchical clustering heatmap analysis of differentially expressed genes between sequencing-based tumor fraction–detected (n = 77) vs not detected (n = 67) within arm A revealed 2 clusters (1 and 2) (P = .03857). Red in the heatmap denotes upregulation, while blue denotes downregulation. The horizontal axis refers to the samples in arm A, and the vertical axis denotes the differentially expressed genes based on sequencing-based tumor fraction status. On top of the heatmap are the same annotations as Figure 3, C. The dendrogram values are measured by euclidean distance, with a complete linkage clustering algorithm. B) Box plot shows the sequencing-based tumor fraction in samples from arm A, cluster1; arm A, cluster2; and arm B. At the bottom of the plot, horizontal blue, orange, or green filled rectangles illustrate the proportion of samples in each group that had evidence of residual tumor DNA by sequencing. C) Association between clusters and clinical outcomes in patients with high-risk, localized prostate cancer following exposure to chemohormonal therapy. D, E) Kaplan-Meier survival analysis for prostate-specific antigen progression-free survival and event-free survival in patients between 2 clusters, identified based on sequencing-based tumor fraction status in arm A. P values were estimated using a Pearson χ² test with Yates continuity correction, Fisher exact test, Mann-Whitney U test (A, B, C), or univariate Cox proportional hazards regression analysis (D, E). CI = confidence interval; HR = hazard ratio; Seq-TF = sequencing-based tumor fraction.