DNA damage response in breast cancer and its significant role in guiding novel precise therapies

Abstract

DNA damage response (DDR) deficiency has been one of the emerging targets in treating breast cancer in recent years. On the one hand, DDR coordinates cell cycle and signal transduction, whose dysfunction may lead to cell apoptosis, genomic instability, and tumor development. Conversely, DDR deficiency is an intrinsic feature of tumors that underlies their response to treatments that inflict DNA damage. In this review, we systematically explore various mechanisms of DDR, the rationale and research advances in DDR-targeted drugs in breast cancer, and discuss the challenges in its clinical applications. Notably, poly (ADP-ribose) polymerase (PARP) inhibitors have demonstrated favorable efficacy and safety in breast cancer with high homogenous recombination deficiency (HRD) status in a series of clinical trials. Moreover, several studies on novel DDR-related molecules are actively exploring to target tumors that become resistant to PARP inhibition. Before further clinical application of new regimens or drugs, novel and standardized biomarkers are needed to develop for accurately characterizing the benefit population and predicting efficacy. Despite the promising efficacy of DDR-related treatments, challenges of off-target toxicity and drug resistance need to be addressed. Strategies to overcome drug resistance await further exploration on DDR mechanisms, and combined targeted drugs or immunotherapy will hopefully provide more precise or combined strategies and expand potential responsive populations.

Keywords: Breast cancer; DNA damage response; Homogenous recombination deficiency; Poly (ADP-ribose) polymerase (PARP) inhibitor.

Fig. 1

Summary of (A) history of studying DNA damage response (HR, MMR, and other DDR mechanisms), targeted therapies, and their clinical trials in breast cancer (B) total publication papers of DDR and DDR-related clinical trials in in breast cancer. A DDR in breast cancer has drawn much attention in targeted treatment since 1980 when PARP has been discovered in SSB repairing. The timeline in the upper part has summarized the history of DDR discovery, important mechanisms HR and MMR in breast cancer, representative clinical trials, and the evolution of indications of DDR-related therapies. B The line chart in the bottom has demonstrated publications and clinical trials on DDR in breast cancer since 1980. In 2007, olaparib was the first DDR-related drug under clinical trials, thus the total published papers increased. Clinical trials have increased since breast cancer had the first indications of PARP inhibitors approved by FDA in 2017. DDR, DNA damage response; SSB, DNA single-strand break; HRD, homogenous recombination deficiency; MMR, mismatch repair; MSI-H, high microsatellite instability; dMMR, deficient mismatch repair; PD-1, programmed cell death-1 protein; PD-L1, programmed cell death-1 ligand protein; RCT, randomized clinical trial; BC, breast cancer; TNBC, triple-negative breast cancer; MSI, microsatellite instability; CRC, colorectal cancer; PARP, poly-ADP-ribose polymerase; CHEK1, checkpoint kinase 1; ATR, ataxia telangiectasia and Rad3-related; PARG, poly (ADP-ribose) glycohydrolase

Fig. 2

Summary of DNA damage response mechanisms and targeted therapies in breast cancer. BER is the primary pathway to repair SBSs. PARP binds to damaged SBSs to recruit DNA repair effectors. First, aberrant bases are identified and cleaved by DNA glycosylation enzymes. Cleavage of damaged N-glycosidic bond of the base results in an AP site creation and AP spots can be recognized by AP endonucleases. APE cleaves to the AP site to generate 3′-OH and 5′-dR termini. Intrinsic dRP lyase activity of DNA Pol β cleaves dRP residue to produce 5′-phosphates. DSBs are caused by UV light and aromatic chemicals and repaired by NHEJ or HR. In NHEJ, 53BP1 and RIF1 localize to DSBs, resulting in BRCA1 recruitment inhibition, blocking DNA excision, and promoting NHEJ repair pathway. The essential components of NHEJ are heterodimers consisting of Ku70/Ku80 and catalytic subunits of DNA-PKcs, identifying DSB and activating downstream signaling factors, including XRCC4, XLF, and DNA ligase IV. HR is initiated by MRN binds to DSB terminal, and MRN recruits and activates ATM and CHEK2 and phosphorylates downstream substrates. CtIP binds to BRCA1 and nucleases EXO1, further interacts with MRN and excises DSB ends together. Excised DNA ends are coated with hyperphosphorylated single-stranded RPA to form a nucleoprotein filament. γH2AX is activated and phosphorylated by apical kinases (ATM and ATR). Diffusion of γH2AX along chromosomes recruits and accumulates additional DDR proteins, including 53BP1 and BRCA1. Under PALB2 localization, BRCA2 binds to BRCA1, promotes recombinant enzyme RAD51 loading on ssDNA, and finally RAD51 prevents secondary structure formation and shields DNA ends from degradation. NER removes bulky adducts and crosslinks lesions, especially those induced by UV light. NER employs TC-NER and GG-NER, both utilizing XPC and RAD23 to excise the damage. DNA-binding proteins XPA and RPA enhance translocation and damage-verification activity of TFIIH and stabilize and orient endonucleases XPF/ERCC1 and XPG. Replication factors involved include PCNA and RFC, DNA polymerases δ, ε, and κ, and ligases I and III. MMR corrects base match errors. First, MutS (MSH2/MSH6) determines base pair error. MutS/MutL complex binds to MutH (MLH1/PMS2). MutH is then activated to cleave the unmethylated strands. Exonuclease removes the nascent strand from the cut point to error base-pair portion. Other DDR pathways including DNA damage tolerance, interstrand crosslink repair and direct damage reversal are not demonstrated. Among proteins involved in DDR, PARP, PARG, and XRCC1 in BER, DNA-PK in NHEJ, and ATR, ATM, and RAD51 in HR can be targeted in DDR-related therapies. Drugs under RCT and in preclinical stage are summarized below (searching results on https://clinicaltrials.gov by 31st August 2024). BER, base excision repair; SSBs, DNA single-strand breaks; AP, apyrimidine; dRP, deoxyribose phosphate; DSBs, DNA double-strand breaks; UV, ultraviolet; NHEJ, non-homogenous end joint; HR, homogenous recombination; 53BP1, p53-binding protein; DNA-PKcs, DNA-dependent protein kinases; XLF, XRCC4-like factor; MRN, MRE11-RAD50-NBS1; ATM, ataxic telangiectasia mutation; CHEK, checkpoint kinase; CtIP, CtBP-interacting protein; EXO1, exonuclease 1; TC-NER, transcription-coupled NER; GG-NER, global genome NER; XRCC, X-ray repair cross-complementing protein; TFIIH, transcription factor II H; PCNA; proliferating cell nuclear antigen; MMR, mismatch repair; PARP, poly-ADP-ribose polymerase; PARG, poly (ADP-ribose) glycohydrolase; PNKP, polynucleotide kinase 3'-phosphatase; APTX: aprataxin; pol, polymerase; ATRIP, ATR-interacting protein; TOPBP1: topoisomerase 2-binding protein 1; CDK: cyclin-dependent kinase; MDC1, mediator of DNA damage checkpoint protein 1; CIP2A, cellular inhibitor of PP2A; CK, casein kinase; ATR, ataxia telangiectasia and Rad3-related; RPA, Replication protein A; Me, dimethylation; P, phosphorylation; Ub, ubiquitylation; RCT, randomized clinical trial

Fig. 3

Mechanisms of cell cycle-dependent DNA damage repair activation. DNA damage checkpoints would be activated to by the presence of DNA damage, leading to cell cycle arrest to allow for DNA repair. ATM-CHEK2-p53 axis and ATR-CHEK1-WEE1 axis will be activated in response to DSBs and SSBs exposure, respectively. First, the MRN complex senses DSBs and activates ATM at break sites to orchestrate DDR signaling. During G1, ATM promotes activation of RNF168, which ubiquitylates histone H2A. This modification, together with the histone H4 Lys20 dimethylation mark results in recruitment of 53BP1 and ATM-mediated phosphorylation of 53BP1, which promotes its interaction with RIF1, REV7 and SHLD. ATM kinase activity also controls p53 stability, which triggers G1 arrest in response to DNA damage. During S phase, the BRCA1-BARD1 complex is recruited to DSBs and counteracts 53BP1 via recognition of unmethylated H4 Lys20 and H2A Lys15 ubiquitylation to promote RAD51 to load onto resected DNA ends. RPA-coated single-stranded DNA also recruits ATR via interaction with ATRIP. ETAA1 binding to RPA-coated single-stranded DNA or TOPBP1 binding to double-stranded DNA and single-stranded DNA junctions activates ATR, phosphorylating CHEK1 to promote CDC25A degradation, thus preventing activation of CDK1 and CDK2. Kinases WEE1 and PKMYT1 mediate inhibitory phosphorylation of CDK1 and CDK2, which prevents cells progressing into mitosis. In mitosis, ATM-dependent H2AX phosphorylation recruits MDC1, which binds TOPBP1 and CIP2A via its CK2-mediated phosphorylation. Aurora-A and PLK1 are implicated in mitotic entry partially through phosphorylation on WEE1 and PKMYT1 that result in their degradation. Among proteins involved in cell cycle-dependent DNA damage repair activation, ATM in G1 phase, CHEK1/2 in G1, G2 and S phase, ATR in S phase, and WEE1 and PKMYT1 in S and G2 phase can be targeted in DDR-related therapies. Drugs under RCT and in the preclinical stage are summarized in the column below (searching results on https://clinicaltrials.gov by 31st August 2024)

Fig. 4

The impact of DNA damage on immune tumor microenvironment. Immune tumor microenvironment can adapt to DNA damage in various manners. The STING pathway can be activated by increased TMB, the upstream cytosolic DNA sensor cGAS or ATM and TRAF6 after perceiving DNA damaging agents or DDR alterations, leading to immune activation. This activation makes a conformation change of STING, which leads to an endoplasmic reticulum to perinuclear endosome shuttling. The activated STING stimulates IRF3, which promotes the production of type I IFNs. Consequently, NK cells are inactivated, while T cells and dendrite cells are activated. STING also activates NK-κB by ATM-TRAF6, induces IL, TGF-β, and other cytokine production, further recruits Tregs and M2-like macrophages. DDR deficiencies also improve tumor recognition through generating neoantigens. Additionally, DNA damage signaling and DDR deficiencies role as important regulators in upregulating PD-L1 expression. ATM-TRAF6 activates NF-kB and induces tumor cell upregulation of PD-L1 that may elicit immune escape. Besides this mechanism, IFN I itself (secreted upon STING activation) is the main factor inducing transcription and expression of PD-L1. DDR, DNA damage response; STING, stimulator of interferon genes; TMB, tumor mutation burden; cGAS, cyclic GMPeAMP synthase; IRF3, interferon regulatory factor 3; NK, natural kill; NK-κB, transcription factor nuclear factor κB; TGF-β, transforming growth factor (TGF)-β; IFN, interferon; Treg, regulatory T cells; PD-L1, programmed cell death-ligand 1

Fig. 5

Summary of clinical trials with HRD stratification in breast cancer. Since 2007, the studies on HRD evaluation in breast cancer have been conducted systematically; more clinical trials with HRD stratification have been published in recent years. The x-axis represented the publication year of the study. The y-axis represented the percentage of breast cancer with HRD features in all breast cancer population. Each spot was a study with a size matched its population. Different colors showed different breast cancer population (pink, breast cancer (without subtype information); red, TNBC; orange, HER2-negative breast cancer; blue, breast cancer with HR-related genes mutations). HR-related genes refer to BRCA1/2 and BRCAness gene (RAD51, CDK12, PALB2, ATM, CHEK2, etc.) mutations. The prevalence of HRD differs among different breast cancer subgroups. TNBC tends to demonstrate higher HRD proportion (all > 30%), thus its spots were concentrated on the upper part of the graph; vice versa for HER2-negative breast cancer in the lower part with lower HRD frequency. HRD, homogenous recombination deficiency; BC, breast cancer; TNBC, triple-negative breast cancer

Fig. 6

Mechanism of PARP inhibitor resistance and strategies to overcome. The efficacy of PARP inhibitors in patients with breast cancer is limited by the development of mechanisms of resistance in most patients. Many mechanisms of resistance have been characterized, including upregulation of drug efflux transporters, restoration of homologous recombination activity, and mitigation of the replication stress via replication fork stabilization. Novel strategies are under investigation to overcome resistance to PARP inhibitors, such as new drug combinations and targeting acquired deficiency. Ongoing research efforts are also uncovering novel biomarkers to identify additional patients who may benefit from these agents. HR, homologous recombination; ctDNA indicates circulating tumor DNA; PD-1, programmed cell death-1 protein; PD-L1, programmed cell death-1 ligand protein; PARP, poly (ADP-ribose) polymerase; PARG, poly (ADP-ribose) glycohydrolase; PROTAC, PROteolysis-TArgeting chimera