DNA damage and repair (DDR) gene mutation profiles in driver gene wild-type advanced non-small cell lung cancer and the predictive role of response to platinum-based chemotherapy

Introduction

The treatment of non-small cell lung cancer (NSCLC) has evolved significantly in recent years, with advances in targeted therapy, anti-angiogenesis therapy, and programmed death-1 (PD-1)/programmed death ligand 1 (PD-L1) immunotherapy (1). However, for patients diagnosed with advanced NSCLC who lack driver mutations and have negative or low PD-L1 expression, the preferred first-line treatment option remains chemotherapy or a combination of chemotherapy and PD-1/PD-L1 immunotherapy (2). In addition, chemotherapy serves as a primary therapeutic approach for patients who have an unclear mechanism of drug resistance to targeted therapy; patients with increased susceptibility to tumor immunotoxicity and associated side effects or resistance to PD-1/PD-L1 inhibitors or as the main preoperative neoadjuvant therapeutic regimen for NSCLC (3,4). Therefore, platinum-based chemodublets remain the cornerstone for the treatment of NSCLC (5,6). However, it is unfortunate that the prognosis for patients with metastatic NSCLC who undergo first-line treatment with platinum-based chemodoublets remains poor (7). Fundamentally, the efficacy of platinum-based doublets in advanced NSCLC has reached a bottleneck. ECOG1594, a randomized phase III trial evaluating the superiority of three platinum-based chemotherapy regimens (cisplatin and gemcitabine, cisplatin and docetaxel) in NSCLC, demonstrated a response rate of 19% with a median survival of 7.9 months [95% confidence interval (CI): 7.3–8.3 months] in all eligible participants (8).

In recent years, there has been increasing focus on the connection between the DNA damage and repair (DDR) system and response to chemotherapy (9). Platinum-based agents cause various forms of DNA damage, including the formation of intra- and inter-strand crosslinks, single nucleotide damage, and strand breaks in cancer cells (10,11). The DDR system can repair DNA damaged by platinum by eliminating DNA adducts and restoring chromosome integrity (12,13). Theoretically, defects in DDR gene hinder repair of the cancer cell genome and cause enormous replication stress with stalled replication forks, enhancing the curative effects of chemotherapy.

There was increasing evidence that DDR gene predict the effectiveness of platinum-based therapy in various solid tumors. Tumors deficient in homologous recombination (HR), which represents a critical DDR signaling pathway, are more susceptible to platinum-based therapy in advanced breast cancer (14). A DDR-targeted panel including BRCA2, BRCA1, ATM, PALB2, FANCA, and CDK12 gene was used to predict response to platinum-based chemotherapy in prostate cancer (15). In NSCLC, the association between DDR mutation and the efficacy of platinum-based chemotherapy has been previously reported using single nucleotide polymorphism genotyping (SNP) by monitoring a limited number of DDR-related gene (16). Recently, deficiency of the 74-DDR gene set in 122 lung cancers (86 NSCLC and 36 small cell lung cancer patients), identified by whole genome sequencing, failed to demonstrate an association with treatment response and outcome (17). Therefore, the predict role of DDR-related gene for platinum-based chemotherapy in NSCLC is still controversial.

Direct inhibition of the DDR signaling pathway was achieved by targeting poly (ADP-ribose) polymerase (PARP) in ovarian cancer, breast cancer and prostate cancer (18). Deficiency of the HR pathway predicted the response to the PARP inhibitor through a mechanism termed synthetic lethal. Nevertheless, new DDR targets, including ATM, ATR, CHK1, CHK2, and DNA-PK, have been investigated in stage I clinical trials in solid tumors. Therefore, it is important to understand the overall picture of DDR gene status in NSCLC before conducting clinical trials targeting these targets or developing combination therapy.

In this study, we used a gene-specific next-generation sequencing (NGS) strategy to determine the status of 47 DDR-related gene involved in seven DDR signaling pathways in a cohort of NSCLC patients without actionable tumor driver gene. We examined the influence of the somatic DDR mutation on the effectiveness of cisplatin-based chemotherapy and also analyzed which DDR gene or signaling pathways play a crucial role in determining the response. We present this article in accordance with the STROBE reporting checklist (available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-24-972/rc).

Methods

Patients and study design

In a prospective study, 182 consecutive treatment-naïve patients with advanced driver gene wild-type NSCLC were enrolled between November 2016 and September 2021, attending the Department of Respiratory Medicine, Xinqiao Hospital (Chongqing, China), Fuling Center Hospital (Chongqing, China) and Third Affiliated Hospital of Chongqing Medical University (Chongqing, China). The study included patients who received at least four cycles of first-line treatment with platinum-based doublets. We excluded patients with common actionable driver mutations (EGFR, ALK, and ROS-1); patients with an Eastern Cooperative Oncology Group performance status (ECOG PS) score of ≥2; patients receive radiotherapy or surgery; patients treated with PD-1/PD-L1 inhibitors, antiangiogenic agents, or other antineoplastic treatments. We collected geographic and clinical data on age, gender, Tumor, Node, and Metastasis (TNM) stage, histological types, smoking status, chemotherapy regimens, and treatment outcomes. Treatment response was evaluated according to the Response Evaluation Criteria in Solid Tumors version 1.1 (RECIST 1.1) (19). The primary endpoint was objective response rate (ORR) [complete response (CR) + partial response (PR)]. The secondary endpoints were progression-free survival (PFS, time from initiation of platinum-based chemotherapy to first documentation of disease progression or death from any cause), overall survival (OS, time from first treatment to death), and time to response (TTR, time from the start of treatment to the first objective tumor response observed in patients achieving a CR or PR), duration of disease control (DDC, time from the first objective tumor response of CR, PR or stable disease (SD) to the first documentation of the course of the disease or a cause of death). This study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). This study was registered with Chinese Clinical Trials (Registration No. ChiCTR1800015470) and approved by the Ethics Committee of the Second Affiliated Hospital Medical of Army Medical University (No. 2018-022-01). Written informed consent was obtained from all individual participants.

DDR gene panel design

The selection of the DDR gene was based on a comprehensive review of the current scientific literature and biological relevance to DNA damage response and repair pathways. Our primary goal was to focus on gene that are well-established in their roles within the DDR mechanisms and have been implicated in the prediction of response to platinum-based chemotherapy in various cancer types, including NSCLC.

The DDR gene panel targets 47 gene involved in seven important DNA repair processes. Base excision repair (BER) (MUTYH, PARP1, POLD1, POLE), nucleotide excision repair (NER) (CUL3, ERCC1, ERCC2, ERCC3, ERCC4, ERCC5), mismatch repair (MMR) (MLH1, MSH2, MSH6, PMS1, PMS2), Fanconi anemia (FA) signaling pathway (BLM, BRIP1, FANCA, FANCC, FANCE, FANCF, FANCG, FANCL, PALB2, FANCD2, RAD52), HR (BARD1, BRCA1, BRCA2, CDK12, MRE11A, NBN, RAD50, RAD51, RAD51B, RAD51C, RAD51D, RAD54L, RECQL4), non-homologous end joining (NHEJ) (AURKA, BAP1, PRKDC) and cell-cycle checkpoint (CP) (ATM, ATR, CHEK1, CHEK2, MDC1).

Tumor DNA and plasma circulating tumor DNA (ctDNA) sequencing

In our study, we focused on sequencing the coding regions of the 47 DDR gene because these regions are most directly involved in the functional outcomes of gene mutations, thereby increasing the sensitivity and specificity of our mutation detection.

Patients suspected of having lung cancer underwent bronchoscopy or percutaneous lung biopsy from November 2016 to September 2021. After the pathological diagnosis of NSCLC, ten 10-µm formalin-fixed, paraffin-embedded (FFPE) sections from 43 NSCLC patients (tumor cell ≥30%) or 8–10 mL blood samples placed in cell-free DNA tubes from 74 NSCLC patients were collected were sent to the core facility of Tongshu Biotechnology Co., Ltd (Shanghai, China) for tumor DNA or plasma ctDNA extraction and subsequent sequencing. Germline mutations can be identified primarily through paired white blood cell (WBC) sequencing. According to the manufacturer’s protocol, germline DNA was extracted from WBC using the HiPure Blood DNA Mini Kit (Qiagen, Guangzhou, China; Cat# D3111-03), and tumor tissue DNA was extracted using the Nucleic Acid Extraction Kit (Changzhou Tongshu, Cat# FD-50/FD-250). For ctDNA, DNA was isolated from plasma samples using the HiPure Circulated DNA Midi Kit (Magen, Cat# IVD3182) according to the product instructions. All DNA concentrations were verified using Qubit 4.0 (Invitrogen) using the Qubit™ dsDNA HS Assay Kit (Invitrogen, Cat# Q32854). For all previously extracted DNA (ctDNA, gDNA and tDNA), we use customized Shanghai Tongshu Multiple Panels (Shanghai Tongshu Biotechnology Co., Ltd., Shanghai, China) and TruePrep DNA Library Prep Kit V2 for Illumina (#TD501, V azyme, Nanjing, China) for target detection and library construction. Paired-end sequencing was performed on the NovaSeq 6000 system (Illumina, CA, USA) with coverage depths of 2,000× for tumor DNA or 10,000× for ctDNA. After obtaining the sequencing data, BWA (version: 0.7.12) was used to align the sequencing to the human genome (hg19), then Sambamba (version: 0.7.1) was used to match the repeated sequences in the alignment results filter, and the Genome Analysis Toolkit (GATK) was performed to ensure recalibration of the base quality score. After comparison, the single and paired sample modes of Mutect2 software were used to detect somatic mutations. Annovar and VEP software (version 100.2) was used to annotate the population mutation status of each site in the ExAC, genomAD, and ESP databases in the mutation analysis results, the location of each mutation site in the gene, and the impact on function. Somatic SNV/indel calling requires at least 20 reads covering the mutated region and 5 reads supporting the variant allele. In addition, the sequencing depth must be ≥20× and the reads supporting variants at the same location in the control sample should be <5. Variations with a minor allele frequency (MAF) >1% in databases such as ExAC, gnomAD and esp6500 were screened as common germline variants (20,21) and removed, the remaining mutations were used for subsequent analysis. We classified deleterious mutations in the structure and function of a protein using Polyphen2 (http://genetics.bwh.harvard.edu/pph2/), MutationTaster (https://www.mutationtaster.org/), and MutationAssessor (http://mutationassessor.org/r3/) databases. In addition, we identified hotspot mutation information using the COSMIC database (https://cancer.sanger.ac.uk/cosmic) and annotated the pathogenicity of ClinVar (www.ncbi.nlm.nih.gov/clinvar). The mutant information of each sample was extracted using the Maftools R package, and the mutant frequency of each gene was calculated using ComparMaf.

Outcomes

Patients were defined as DDR-mutant (DDRmut) if they had at least one deleterious somatic alteration in a set of gene listed above. Tumors or plasma ctDNA that had no mutation or only variants of unknown significance in these gene were classified as DDR wild type (DDRwt). An abnormality in the DDR signaling pathway was defined as a mutation in one of the gene involved in the corresponding DDR signaling pathways.

Statistical analysis

The characteristics of DDR mutants and DDRwt probands were compared using the unpaired t-test or Wilcoxon signed-rank test for continuous variables and the χ² or Fisher exact test for categorical variables. Multiple comparisons were made using the Kruskal-Wallis test. The association between DDR mutation and treatment response (ORR) was assessed using χ² or Fisher’s exact test. A P value of less than 0.05 was considered significant. Survival analyzes were performed using the Kaplan-Meier method and the log-rank test. The Cox proportional hazards model was used to estimate crude or adjusted hazard ratios and 95% CIs adjusted for age, sex, smoking, histotypes, stage, metastasis, ECOG PS score and chemotherapy regimens. Statistical analysis was performed using SPSS (v22.0; SPSS Inc., Chicago, IL, USA) and R (version 4.2.0, R Foundation for Statistical Computing) software.

Results

Characteristics of the cohort

A total of 117 enrolled treatment-naïve driver gene wild-type NSCLC patients receiving first-line platinum-based chemotherapy underwent genomic profiling of 47 DDR gene, and 101 NSCLC patients received qualified DDR profiling results from either tumor DNA (36 cases) or serum ctDNA (65 cases) (Figure 1). In this cohort, 43 of 101 patients (42.6%) were older than 60 years. There were 87 male patients (86.1%) and 14 female patients (13.9%). The pathological type included 42 (41.6%) cases of adenocarcinoma, 58 (57.4%) cases of squamous cell carcinoma, and 1 case of large cell carcinoma (0.99%); Most patients were current or former smokers (82/101, 81.2%) and 19 (18.8%) patients were never smokers; 97 (96.0%) patients had distant metastases. All patients received at least 4 cycles of first-line platinum-based chemotherapy. The demographic, clinicopathological characteristics and treatment regimens were comparable between the DDRmut and DDRwt patients (Table 1).

Figure 1 Patients screening and disposition flow diagram. ALK, anaplastic lymphoma kinase; ctDNA, circulating tumor DNA; DDR, DNA damage and repair; EGFR, epidermal growth factor receptor; FFPE, formalin-fixed paraffin-embedded; NSCLC, non-small cell lung cancer; PD-1, programmed death-1; ROS-1, ROS proto-oncogene 1.

DDR mutation signatures in NSCLC

A total of 335 somatic mutations were detected in 47 DDR gene from 101 NSCLC patients, including 258 missense_mutations, 18 frame_shift_del, 7 in_frame_del, 23 nonsense_mutation, 9 frameshift_ins, 2 translation_start_site, 6 in_frame_ins, 11 splice_site, 1 nonstop_mutation. 32 samples showed more than one type of mutation in the same gene and were designated as multi-hit. DDR deficiency was observed in 67 (66.33%, 67/101) patients. The 10 mutated DDR gene were BARD1 (17.82%), POLE (16.83%), PRKDC (14.85%), BRCA2 (13.86%), ATM (11.88%), RECQL4 (9.9%), CDK12 (8.91%), ERCC3 (8.91%), FANCE (8.91%), ATR (7.92%) (Figure 2A). missense_mutations (95%) was the most common mutation type in BARD1 and the most common mutation type p.V507M was identified in 60% of samples. BRCA2 gene showed multi-hit in 5 patients, with 1 patient having 4 mutation forms, including 1 site with nonsense_mutation, 2 sites with in_frame_ins, and 1 site with missense_mutation. By integrating a single mutated DDR gene into the DDR signaling pathway, changes were observed in the MMR, NER, HR, FA, BER, NHEJ, and CP signaling pathways (Figure 2B). The HR pathway mutation was detected most frequently (41.58%), followed by the FA pathway (35.64%). The mutation in the NHEJ pathway was relatively rare, accounting for 14.85%.

Figure 2 Landscapes of DDR gene mutations in NSCLC. (A) Oncoplot depicting the top 30 most frequently mutated DDR-related genes. The oncoprint (bottom panel) shows details of the types of alterations in DDR genes, as well as clinical characteristics including gender, pathological type, smoking and sample type. (B) The proportion of mutated samples in the 7 DDR-related pathways. (C-F) The correlation of DDR variant number with pathological type (C), TNM staging (D), smoking history (E) and age (F). (G) Co-mutation analysis. The co-mutation included co-occurring (green) and mutual exclusion (brown), “*”, P<0.05; “•”, P<0.1. BER, base excision repair; CP, cell-cycle checkpoint; DDR, DNA damage and repair; FA, Fanconi anemia pathway; HR, homologous recombination; LUAD, lung adenocarcinoma; LUSC, lung squamous cell carcinoma; MMR, mismatch repair; NER, nucleotide excision repair; NHEJ, non-homologous end joining; NSCLC, non-small cell lung cancer; TMB, tumor mutation burden; TNM, Tumor, Node, and Metastasis.

In lung adenocarcinoma, ATM (17%) was the most frequently mutated DDR gene, followed by BARD1 (14%), ERCC3 (12%), POLE (12%), and ATR (10%) (Figure S1A). In lung squamous cell carcinoma, BARD1, POLE and PRKDC showed a mutation frequency of more than 20%. BRCA2, CHEK2, BRIP1, CDK12, and RECCQL4 were detected in >10% of lung squamous cell carcinomas (Figure S1B). The proportion of ATM and ATR mutations was lower in squamous cell carcinoma than in adenocarcinoma, and ERCC3 mutation was not frequently detected. Overall, the DDR mutation burden in squamous cell carcinomas was comparable to that in adenocarcinomas (Figure 2C).

We also examined the distribution of DDR mutation burden in patients with different clinical features. No significant difference in stage (P=0.30, Figure 2D) or smoking (P=0.67, Figure 2E) was observed between DDRmut and DDRwt patients. A trend toward increased DDR mutation burden was noted with increasing age, but there was no statistical significance (P=0.09, Figure 2F). DDR comutation was observed in 76% (51/67) of NSCLC patients with DDR alteration. The average number of mutations was 4,657±4,343/person. DDR gene comutations occur in both the same and different DDR pathways. BARD1 was observed to commute with ERCQL4, BRCA2, RAD50 in the same HR pathway and with MSH6 in the MMR pathway, PALB2 in the FA pathway, ERCC5 in the NER pathway, ATR, ATM in the CP pathway and POLE in the BER pathway. path commutated (Figure 2G). Compared to co-mutation, mutually exclusive mutation in DDR gene was not frequently observed.

ORR of DDRmt versus DDRwt NSCLC to platinum-based chemotherapy

Best responses to platinum-based chemotherapy were assessed in the 101 enrolled NSCLC patients after 4–6 cycles of standard treatment. Overall, 43 patients (42.6%) achieved ORR, including 1 patient (1.0%) with CR. The disease control rate (DCR) was 81.2% and the median PFS (mPFS) was 5.0 months (Table 2). In DDRmut patients, the ORR was 52.2% compared to 23.5% in DDRwt NSCLC (ORs 3.5, 95% CI: 1.8 to 6.9, P<0.001, Figure 3A). The depth of efficacy response of the target lesion is shown in the waterfall plot of Figure 3B. For individual gene, RAD50 (87.5% vs. 38.7%, P=0.01, Figure 3C) and BARD1 (61.1% vs. 38.6%, P=0.11, Figure 3D) mutations were found to be associated with increased proportion of PR+CR. For the DDR pathways, we found that mutations in the HR pathway (61.9% vs. 28.8%, P=0.001, Figure 3E) were associated with a better response.

Figure 3 Response to platinum-based chemotherapy in patients with DDRmut and DDRwt disease. (A) Histogram of ORR between DDRmut and DDRwt NSCLC. (B) Waterfall plot showing the best change of target lesion’s diameter from baseline for patients with DDRmut and DDRwt disease. (C-E) Histogram of ORR between NSCLC patients with or without RAD50 (C), BARD1 (D), or HR pathway (E) alterations. CR, complete response; DDR, DNA damage and repair; DDRmut, DDR-mutant; DDRwt, DDR-wild type; HR, homologous recombination; ORR, objective response rate; PD, progressive disease; PR, partial response; SD, stable disease.

Predictive role of DDR in prognosis of NSCLC treated with platinum-based chemotherapy

Univariate analyzes of the COX risk regression model showed that only DDR gene mutation status was an influencing factor on PFS (hazard ratio =3.102; 95% CI: 1.981 to 4.857; P<0.001, Table S1) and OS (hazard ratio =2.217; 95% CI: 1.229–3.997; P=0.008, Table S2). mPFS from initiation of platinum-based therapy was significantly longer in the DDRmut group than in the DDRwt group (6.30 vs. 3.30 months, P<0.001, Figure 4A). Similarly, the DDRmut group demonstrated improved median OS (mOS) compared to the DDRwt group (16.80 vs. 9.40 months, P=0.007, Figure 4B). The mean DDC (mDDC) in the DDRmut group was significantly longer than in the DDRwt group (4.30 vs. 1.40 months, P<0.001, Figure 4C). TTR was comparable between the DDRmut and DDRwt groups (1.90 vs. 1.90 months, P=0.83, Figure 4D). Forest plot of multi-factor COX regression model for PFS and OS showed the relationship between outcomes and various covariates, including DDR status, gender, smoking history, tumor histology type, TNM stage, treatment regimens, ECOG score, and lymph node metastasis. This analysis showed that the correlation between platinum-based chemotherapy efficacy PFS (hazard ratio =3.166; 95% CI: 1.947 to 5.149; P<0.001, Figure 4E)/OS (hazard ratio =2.431; 95% CI: 1.286 to 4.595; P=0.006, Figure 4F) and gene 47 DDR status remains significant.

Figure 4 Outcome of NSCLC with DDRmut and DDRwt disease. (A-D) PFS (A), OS (B), DDC (C) and TTR (D) on platinum-based chemotherapy for DDRmut and DDRwt patients. (E,F) Forest plot of NSCLC patients PFS (E) and OS (F) from multivariate Cox regression. CI, confidence interval; DDC, duration of disease control; DDR, DNA damage and repair; DDRmut, DDR-mutant; DDRwt, DDR-wild type; ECOG, eastern cooperative oncology group; NSCLC, non-small cell lung cancer; OS, overall survival; PFS, progression-free survival; TNM, Tumor, Node, and Metastasis; TTR, time to response.

The outcome of platinum-based chemotherapy with a single DDR gene or specific DDR pathway alteration

We examined the difference in PFS (Figure S2), DDC (Figure S3) and OS (Figure 5) of patients with specific DDR pathway mutations in DDRwt participants. Consistently, there was a better prognosis as measured by PFS, DDC, or OS in NSCLC with at least one DDR pathway abnormality. However, when we compare the difference of PFS, DDC and OS of patients with or without a specific DDR pathway, the significant predictive effect was not seen (data not shown). This may be due to the complementary role of other DDR pathway alterations in patients without a specific DDR pathway mutation.

Figure 5 OS of NSCLC patients with alteration in NHEJ pathway (A), NER pathway (B), MMR pathway (C), HR pathway (D), FA pathway (E), CP pathway (F), and BER pathway (G) compared with DDRwt group. BER, base excision repair; CP, cell-cycle checkpoint; DDR, DNA damage and repair; FA, Fanconi anemia pathway; HR, homologous recombination; MMR, mismatch repair; NHEJ, non-homologous end joining; NER, nucleotide excision repair; NSCLC, non-small cell lung cancer; OS, overall survival.

For a single DDR gene, mutation of BRAD1 (P=0.002, Figure 6A), POLE (P<0.001, Figure 6B), PRKDC (P<0.001, Figure 6C), BRCA2 (P=0.002, Figure 6D), or ATM (P=0.01, Figure 6E) was associated with better PFS. However, only mutation of a single PRKDC gene (P=0.02, Figure 6F) was associated with better OS in NSCLC treated with platinum-based chemotherapy.

Figure 6 PFS of NSCLC patients with alteration in BRAD1 (A), POLE (B), PRKDC (C), BRCA2 (D), and ATM (E) compared with DDRwt group. OS of NSCLC patients with alteration in PRKDC (F) compared with DDRwt group. DDR, DNA damage and repair; DDRwt, DDR-wild type; NSCLC, non-small cell lung cancer; OS, overall survival; PFS, progression-free survival.

Discussion

Our study provides a comprehensive analysis of the prevalence and predictive value of DDR gene mutations in patients with advanced NSCLC treated with platinum-based chemotherapy. The DDR mutation rate in the cohort was 66.33%. The HR signaling pathway gene BARD1 was the most frequently detected DDR gene in NSCLC. HR pathway mutation was detected in 41.58% of enrolled participants. Lung adenocarcinoma and squamous cell carcinoma of the lung showed different mutation patterns. Our results show that DDR gene mutations are significantly associated with improved treatment outcomes, including ORR, PFS, and OS.

Germline phenotyping of DDR gene has been reported in NSCLC cancer (22). Now more attention is being paid to the somatic mutation of DDR gene, which may be associated with sensitivity to anticancer drugs, synthetic lethality targets, or even the tumor immune microenvironment (17). Owen et al. reported somatic mutations of 9 DDR gene (ATM, BLM, BRCA2, ERCC2, FANCA, FANCM, MSH6, PRKDC and RAD50) in a cohort of NSCLC patients enrolled in a phase II trial of carboplatin and nab-paclitaxel participants (NCT00729612) (23), 84% (21/25) of NSCLC patients had reported pathogenic DDR gene mutations. However, the form and frequency of the mutation have not been documented. Dai et al. examined the mutational landscape of 74 DDR gene in 122 advanced lung cancers (including NSCLC and SCLC) and showed that 54.9% (67/122) of patients had deleterious alterations of DDR gene, including POLQ, BRCA2, ATM, ATR, PARP4, POLD1, BARD1 were most frequently observed. In our study, DDR alteration was observed in 66.33% of NSCLC. The most frequently observed mutations of BARD1, ATM, ATR et al. were also reported in the study by Dai et al. (17).

Theoretically, tumor cells with dysfunction of DNA damage response and repair could be more sensitive to chemotherapy than with a lack of DNA repair after DNA damage caused by chemotherapy. A previous study showed that the expression of DDR-related gene correlates with the prognosis of platinum-based chemotherapy. ERCC1, a member of the NER pathway, has been proposed as a biomarker associated with survival from adjuvant cisplatin-based chemotherapy (24). Using RT-PCR, Leng et al. showed that patients with negative ERCC1 expression had significantly longer PFS (P=0.001) and OS (P=0.001) than patients with positive expression (25). However, a meta-analysis showed that high ERCC1 expression could be a favorable prognosis and predict drug resistance in NSCLC (26). NSCLC patients with negative expression of APE1, ERCC1 or TUBB3 benefited from first-line chemotherapy with platinum plus paclitaxel (27). However, these studies focused on a specific DDR gene or pathway and could not evaluate DDR-related gene as an integrated system. Evaluating DDR-related gene mutations with NGS may facilitate comprehensive assessment of treatment response in tumors receiving DNA-damaging therapies (i.e., chemotherapy) based on multiple DDR-related gene. Using RNA sequencing transcriptome data (RNA-seq) from 986 NSCLC patients from The Cancer Genome Atlas (TCGA) database, Li et al. have successfully established and validated a predictive model for the 6 DDR related prognostic genes (CDC25C, NEIL3, H2AFX, NBN, XRCC5 and RAD1) (28).

In our study, we observed that the ORR of DDRmut patients was significantly higher (52.2% vs. 23.5%, P<0.001) than that of DDRwt NSCLC. The mPFS from the start of platinum-based therapy was significantly longer in the DDRmut group than in the DDRwt group (6.3 vs. 3.3 months, P<0.001). mOS was also significantly longer in DDRmut NSCLC than in the DDRwt group (16.80 vs. 9.40 months, P=0.007). Our results showed that comprehensive assessment of 47 DDR gene covering seven DDR signaling pathways using NGS could predict the response and prognosis of first-line platinum-based chemotherapy in NSCLC. The Dai et al. generated combination of 76 DDR-related gene showed no significant correlations with ORR or DCR and could not predict prognosis in terms of PFS and OS. The discrepancy between our study and theirs may be due to several factors: first, the patients included in our study were NSCLC, while they also included small cell lung cancer (SCLC); second, the 47 DDR gene panel targeted in our study was selected based on previous publications with predictive roles in NSCLC and also functional significance in DDR signaling pathways. Third, the combination of other cancer treatments such as PD-1/PD-L1 inhibitors in their study may not truly reflect the predictive role of DDR gene for platinum-based chemotherapy.

In a retrospective study conducted on a large Chinese cohort, Xiao et al. found that DDR gene mutations are significantly associated with the efficacy of platinum-based chemotherapy and the combination of immunotherapy with platinum-based chemotherapy in advanced NSCLC patients (29). This result is consistent with our study, which also indicates that DDR gene mutations are associated with improved ORR and longer PFS and OS. Their research complements our findings and highlights the potential clinical application value of DDR gene mutations in the treatment of NSCLC. The study by Xiao et al. While providing valuable insights, it has several limitations, including its retrospective design and limitation to a single ethnic cohort. Future studies should validate these results in patient populations of different ethnicities and regions and investigate the specific mechanisms by which DDR gene mutations influence responses to immunotherapy.

The BARD1 gene, which encodes a BRCA1-associated RING domain protein, plays a critical role in the DNA damage response, particularly in the HR repair pathway. The V507M variant, located in the BRCT domain of BARD1, has been the subject of debate regarding its classification as benign or deleterious. In our study, we used the following approach to classify the BARD1 V507M variant: (I) literature search: we conducted a thorough search of the current literature, including databases such as ClinVar, to collect information about the reported associations of the variants with cancer susceptibility (30,31). (II) Clinical correlation: we hypothesize that the presence of the V507M variant correlates with clinical outcomes. including response to platinum-based chemotherapy and overall survival in our cohort of NSCLC patients. Our analysis revealed that the BARD1 V507M variant was associated with a better response to platinum-based chemotherapy in NSCLC patients. This finding is in contrast to its classification as “likely benign” in some databases. We hypothesize that the V507M variant may have a context-dependent effect, potentially exerting a deleterious role in the DNA damage response in the context of lung cancer.

The concept of tumor mutation burden (TMB) as a predictive biomarker has gained considerable attention, particularly in the context of immunotherapy. Lin et al. (32) and Song et al. (33) have shown that first-line chemotherapy is more effective in patients with low TMB. Recent studies have shown that patients with DDR gene mutations have higher TMB (32,34), but the relationship between TMB and chemotherapy efficacy in patients with DDR gene mutations has not been explored. Although we did not directly measure TMB in our study, we observed a high prevalence of DDR gene mutations in patients with advanced NSCLC and their association with improved response to platinum-based chemotherapy. This observation leads us to consider the possible interplay between TMB, DDR gene mutations, and chemotherapy outcomes. However, the association between TMB, DDR gene mutations and chemotherapy effectiveness is not clear and may vary depending on the specific DDR gene involved, the type of chemotherapy and the tumor microenvironment. Further studies are needed to investigate the complex interactions between TMB, DDR gene mutations, and response to chemotherapy.

There are several limitations in this study. The panel we designed covered most and important DDR gene in the literature. Some less important or unidentified DDR-related gene were not considered, which may result in selection bias. For example, the study design missed KRAS and TP53, two genes that are commonly mutated in NSCLC. The role of KRAS and TP53 gene in NSCLC cannot be ignored. As one of the most common oncogenes, KRAS mutation status has an important impact on patient treatment options and prognosis. The TP53 gene serves as a tumor suppressor gene and its mutations are closely related to the ability to repair DNA damage, tumor progression and sensitivity to chemotherapy. In NSCLC, the mutation status of TP53 is particularly important because it is directly related to the genome integrity and DDR function of the tumor. Because the DDR gene panel used in our study was established in late 2016 and early 2017 and the research literature on DDR gene was relatively limited at that time, these two key gene were not included. The exclusion of the KRAS and TP53 gene may limit the general applicability of our results. This limitation may have influenced our overall understanding of the relationship between DDR gene mutations and response to platinum-based chemotherapy. Despite the above limitations, our study provides evidence of the high prevalence of DDR gene mutations in NSCLC patients and suggests that DDR gene defects are favorable predictive biomarkers for predicting response to first-line platinum-based chemotherapy in patients with advanced NSCLC. However, to further improve our understanding of the genomic heterogeneity of NSCLC and optimize individual treatment options, future studies need to include KRAS and TP53 gene and examine their interactions with other DDR gene. Second, one of the critical considerations in our study involves the use of two distinct types of biological samples for NGS: FFPE tumor tissues and plasma ctDNA. The concordance between mutations detected in FFPE and ctDNA can vary, potentially affecting the observed association between DDR gene mutations and response to platinum-based chemotherapy. Thus, the potential biases of our study cannot be entirely eliminated. While our study provides valuable insights into the role of DDR gene mutations in NSCLC patients treated with platinum-based chemotherapy, the use of different sample types (FFPE and ctDNA) may influence the interpretation of our results. We suggest that future studies should aim to use a more homogeneous sample type or incorporate both types to validate findings. This will provide a more comprehensive understanding of the impact of DDR gene mutations on treatment response and prognosis in NSCLC patients. Third, the study is based on a genomic modification of DDR gene; the function of the corresponding mutated molecules has not been validated.

Conclusions

In conclusion, a high prevalence of DDR mutation was observed in advanced NSCLC with wild-type EGFR/ALK/ROS1. DDR deficiency, manifested by an alteration in the 47-gene DDR panel readout, is a favorable predictive biomarker for first-line platinum-based chemotherapy in patients with advanced NSCLC.

Acknowledgments

None.

Footnote

Reporting Checklist: The authors have completed the STROBE reporting checklist. Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-24-972/rc

Data Sharing Statement: Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-24-972/dss

Peer Review File: Available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-24-972/prf

Funding: This work was supported by the Key Project of Clinical Research Of Second Affiliated Hospital (Xinqiao Hospital) of the Army Medical University (No. 2016YLC02) and the Key Support Object Training Project of Army Medical University (No. 2019R025).

Conflicts of Interest: All authors have completed the ICMJE uniform disclosure form (available at https://tlcr.amegroups.com/article/view/10.21037/tlcr-24-972/coif). The authors have no conflicts of interest to declare.

Ethical Statement: The authors are accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. This study was conducted in accordance with the Declaration of Helsinki (as revised in 2013). This study was registered with Chinese Clinical Trials (Registration No. ChiCTR1800015470) and approved by the Ethics Committee of the Second Affiliated Hospital Medical of Army Medical University (No. 2018-022-01). Written informed consent was obtained from all individual participants.

Open Access Statement: This is an Open Access article distributed in accordance with the Creative Commons Attribution-NonCommercial-NoDerivs 4.0 International License (CC BY-NC-ND 4.0), which permits the non-commercial replication and distribution of the article with the strict proviso that no changes or edits are made and the original work is properly cited (including links to both the formal publication through the relevant DOI and the license). See: https://creativecommons.org/licenses/by-nc-nd/4.0/.

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