Defects in DNA Repair Genes Predict Response to Neoadjuvant Cisplatin-based Chemotherapy in Muscle-invasive Bladder Cancer

Abstract

Background: Cisplatin-based neoadjuvant chemotherapy (NAC) before cystectomy is the standard of care for muscle-invasive bladder cancer (MIBC), with 25-50% of patients expected to achieve a pathologic response. Validated biomarkers predictive of response are currently lacking.

Objective: To discover and validate biomarkers predictive of response to NAC for MIBC.

Design, setting, and participants: Pretreatment MIBC samples prospectively collected from patients treated in two separate clinical trials of cisplatin-based NAC provided the discovery and validation sets. DNA from pretreatment tumor tissue was sequenced for all coding exons of 287 cancer-related genes and was analyzed for base substitutions, indels, copy number alterations, and selected rearrangements in a Clinical Laboratory Improvements Amendments-certified laboratory.

Outcome measurements and statistical analysis: The mean number of variants and variant status for each gene were correlated with response. Variant data from the discovery cohort were used to create a classification tree to discriminate responders from nonresponders. The resulting decision rule was then tested in the independent validation set.

Results and limitations: Patients with a pathologic complete response had more alterations than those with residual tumor in both the discovery (p=0.024) and validation (p=0.018) sets. In the discovery set, alteration in one or more of the three DNA repair genes ATM, RB1, and FANCC predicted pathologic response (p<0.001; 87% sensitivity, 100% specificity) and better overall survival (p=0.007). This test remained predictive for pathologic response in the validation set (p=0.033), with a trend towards better overall survival (p=0.055). These results require further validation in additional sample sets.

Conclusions: Genomic alterations in the DNA repair-associated genes ATM, RB1, and FANCC predict response and clinical benefit after cisplatin-based chemotherapy for MIBC. The results suggest that defective DNA repair renders tumors sensitive to cisplatin.

Patient summary: Chemotherapy given before bladder removal (cystectomy) improves the chance of cure for some but not all patients with muscle-invasive bladder cancer. We found a set of genetic mutations that when present in tumor tissue predict benefit from neoadjuvant chemotherapy, suggesting that testing before chemotherapy may help in selecting patients for whom this approach is recommended.

Keywords: ATM; Biomarkers; Bladder cancer; Cisplatin resistance; Cisplatin sensitivity; DNA repair; FANCC; Neoadjuvant chemotherapy; RB1; Urothelial carcinoma.

Figure 1

Distribution of alteration in samples by alteration type and responder status for the (A) AMVAC and (B) DDGC data sets. For each panel, the top graph indicates the alteration counts per sample. Somatic mutations in all sequenced genes were taken into account. Samples were subdivided into nonresponders (left section of the graph) and responders (right section) and sorted by the total number of all alterations in descending order. For each panel, the right-hand graph provides alteration counts per gene. For each panel, the main field indicates the presence of the mutation in a given sample in a given gene. Only the most deleterious mutation in the indicated gene is shown for cases in which two or more mutations were identified in the same patient. The type of mutation is color-coded as shown by the legend. MIBC = muscle-invasive bladder cancer.

Figure 1

Distribution of alteration in samples by alteration type and responder status for the (A) AMVAC and (B) DDGC data sets. For each panel, the top graph indicates the alteration counts per sample. Somatic mutations in all sequenced genes were taken into account. Samples were subdivided into nonresponders (left section of the graph) and responders (right section) and sorted by the total number of all alterations in descending order. For each panel, the right-hand graph provides alteration counts per gene. For each panel, the main field indicates the presence of the mutation in a given sample in a given gene. Only the most deleterious mutation in the indicated gene is shown for cases in which two or more mutations were identified in the same patient. The type of mutation is color-coded as shown by the legend. MIBC = muscle-invasive bladder cancer.

Fig. 2

Progression-free survival (PFS) and overall survival (OS) by ATM/RB1/FANCC mutation status for the AMVAC discovery and DDGC validation sets. Alteration in any one of ATM/RB1/FANCC predicts better PFS (p = 0.0085) and OS (p = 0.007) in the AMVAC discovery set, with a trend towards significance for OS (p = 0.0545) in the DDGC validation set. wt = wild type; mut = mutation; PTs = patients.

Fig. 3

ATM and RB protein domains and structures annotated with alterations. (A) RB domains and variants. The positions of RB missense variants and truncations are mapped with respect to known domains. The domains of RB are denoted along with their sequence ranges (except Rb-C, which corresponds to residues 829–872). RbN A and RbN B denote the A and B N-terminal domains. Pocket A and Pocket B denote the two pocket domains of RB. Rb-C is the C-terminal conserved motif. Truncations are marked with arrows. Red triangles denote missense mutations in responders, while green triangles denote missense mutations in nonresponders. Mutations predicted to be deleterious are marked with a black border around the triangles; those predicted to be neutral do not have a border. Mutations found in the same patient are connected with thin red lines. TCGA mutations associated with bladder cancer are denoted below the protein domain diagram with blue triangles. (B) ATM domains and variants. The positions of ATM missense variants and truncations are mapped with respect to known domains. (C) Predicted structure of ATM. The FAT (light green), PI-3/PI-4 kinase (light blue), and FATc (orange helix in the background) domains in a predicted structure of ATM are shown in ribbon representation. The wild-type residues found where missense variants were determined in this study are shown with red spheres. The magenta spheres represent a PI-3 kinase inhibitor molecule and thus mark the active site of the kinase domain.(D) Structure of RB1. RB-C domain bound to transcription factor Dp-1 and transcription factor E2F1 (Protein Data Bank entry 2AZE). The RB-C domain is shown in orange, Dp-1 in green, and E2F1 in cyan. The wild-type residue for mutation S862G is shown in red spheres; it forms a side- chain/side-chain hydrogen bond with E864 of RB, shown in orange spheres. It forms backbone hydrogen bonds with C274 of Dp-1 (not shown).