Telomeric allelic imbalance indicates defective DNA repair and sensitivity to DNA-damaging agents

Abstract

DNA repair competency is one determinant of sensitivity to certain chemotherapy drugs, such as cisplatin. Cancer cells with intact DNA repair can avoid the accumulation of genome damage during growth and also can repair platinum-induced DNA damage. We sought genomic signatures indicative of defective DNA repair in cell lines and tumors and correlated these signatures to platinum sensitivity. The number of subchromosomal regions with allelic imbalance extending to the telomere (N(tAI)) predicted cisplatin sensitivity in vitro and pathologic response to preoperative cisplatin treatment in patients with triple-negative breast cancer (TNBC). In serous ovarian cancer treated with platinum-based chemotherapy, higher levels of N(tAI) forecast a better initial response. We found an inverse relationship between BRCA1 expression and N(tAI) in sporadic TNBC and serous ovarian cancers without BRCA1 or BRCA2 mutation. Thus, accumulation of telomeric allelic imbalance is a marker of platinum sensitivity and suggests impaired DNA repair.

Significance: Mutations in BRCA genes cause defects in DNA repair that predict sensitivity to DNA damaging agents, including platinum; however, some patients without BRCA mutations also benefit from these agents. NtAI, a genomic measure of unfaithfully repaired DNA, may identify cancer patients likely to benefit from treatments targeting defective DNA repair.

Figure 1. Chromosomal aberrations and cisplatin sensitivity in vitro

The relationship between N_tAI and cisplatin sensitivity was analyzed in breast cancer cell lines. A and B: 10 cell lines were included in this study, 1: CAMA-1, 2: HCC1954, 3:MDA-MB-231, 4: MDA-MB-361, 5: HCC1187, 6:BT-549, 7: HCC1143, 8: MDA-MB-468, 9: BT-20, 10: T47D. A. IC₅₀ values for each of the 10 cell lines. A proliferation assay was used to assess viability after 48 hours of cisplatin exposure, and IC₅₀ was determined from the dose response curves. B. Relationship between N_tAI and cisplatin sensitivity. Breast cancer subtype is indicated as follows: ER− HER2−, red; HER2+, green, ER+ HER2−, blue. C. Relationship between N_tAI and cisplatin sensitivity as determined by GI₅₀ in breast cancer cell lines from Heiser et al. (18). Reported transcriptional subtype is indicated as follows: basal, red; claudin-low, pink; ERBB2Amp, green; luminal, blue. See supplemental methods for cell line identifiers.

Figure 2. N_tAI and cisplatin response in breast cancer

In two clinical trials, TNBC patients were given preoperative cisplatin (Cisplatin-1, Fig. 2 A-B) or cisplatin and bevacizumab (Cisplatin-2, Fig. 2 C-D). Cisplatin resistant tumors are indicated in black, cisplatin sensitive tumors are indicated in red. Tumors with germline mutations in BRCA1/2 are indicated with triangles. A and C. Box plots showing N_tAI distribution in cisplatin resistant and sensitive tumors. B and D. Receiver operating characteristic curves showing the ability of N_tAI to predict for sensitivity to cisplatin.

Figure 3. N_tAI and cisplatin response in serous ovarian cancer

Box plots showing N_tAI distribution in platinum sensitive and resistant tumors in cancers without BRCA1 or BRCA2 mutations (wtBRCA) and for cancers with germline or somatic mutation in BRCA1 (mBRCA1) or in BRCA2 (mBRCA2). Red indicate sensitive samples, triangles indicate samples with germline or somatic mutations in BRCA1 or BRCA2. Significant differences between resistant wtBRCA and sensitive groups are indicated. In addition, significant differences were found between sensitive wtBRCA and sensitive mBRCA2 (P = 0.047), and sensitive wtBRCA and sensitive mBRCA1 (P = 0.014).

Figure 4. Enrichment of common CNVs in tAI chromosomal breakpoints from TNBC

Association of tAI breakpoints with common CNV loci based on computational simulations that compared the expected number of breakpoints containing CNVs with the observed number in total cases in Cisplatin-1 (A) and Cisplatin-2 (B).

Figure 5. Association between BRCA1 transcript level and cisplatin sensitivity, BRCA1 promoter methylation, and N_tAI

Red indicates tumors sensitive to cisplatin. Tumors with a germline mutation in BRCA1 or BRCA2 are excluded in A. and B., but included in C., represented as triangles. In B. and C., BRCA1 transcript levels measured by qPCR were scaled and combined by centering the values, and dividing by the standard deviation within each trial. A. BRCA1 transcripts in resistant and sensitive tumors in the Cisplatin-2 cohort. B. BRCA1 expression in tumors by methylation status of the BRCA1 promoter region in the combined Cisplatin-1 and Cisplatin-2 cohorts. C. Relationship of BRCA1 transcript level and N_tAI in the combined Cisplatin-1 and Cisplatin-2 cohorts.

Figure 6. A model relating DNA repair to accumulation of telomeric AI and response to platinum agents

A. In DNA repair-competent cells, DNA breaks are repaired using error-free homologous recombination employing the identical sister chromatid as a template, resulting in no AI. B. and C. Compromised DNA repair favors the use of error-prone repair pathways, resulting in chromosome rearrangements and aberrant radial chromosome formation. After mitotic division, daughter cells will have imbalance in the parental contribution of telomeric segments of chromosomes (telomeric AI). B. Non-homologous end joining is one error-prone mechanism that joins a broken chromatid of one chromosome (dark blue) to the chromatid of another, usually nonhomologous, chromosome (white). Mitotic segregation results in cells with telomeric AI due to mono-allelic change in DNA copy number of the affected telomeric region. C. Mitotic recombination may result in rearrangements between homologous chromosomes (dark blue and light blue). Mitotic segregation results in cells with AI due to copy neutral loss of heterozygosity (LOH). D. The same compromise in DNA repair that causes telomeric AI may also result in the inability of the tumor cell to repair drug-induced DNA damage, leading to tumor sensitivity to drugs such as platinum salts.