Insights into the DNA damage response and tumor drug resistance

Overview of the DNA harm response (DDR) in tumor cells

DDR is a extremely coordinated signaling community that repairs DNA harm attributable to intrinsic mobile processes and extrinsic insults, thereby stopping genome instability. Relying on the kind of harm, distinct DNA harm restore and DNA harm tolerance (DDT) pathways are concerned and coordinately regulated. For the reason that Nineteen Eighties, an more and more clear relationship between DDR and carcinogenesis has been demonstrated via in depth proof. DDR-associated genes are steadily mutated in tumor cells, thus resulting in genetic hypermutagenesis and consequently fueling carcinogenesis. On this context, inactivating mutations in DDR genes, such because the homologous recombination (HR) genes breast most cancers sort 1/2 susceptibility gene BRCA1/2; mismatch restore genes MutS homolog 2 (MSH2) or MutL homolog 1 (MLH1); and DNA translesion synthesis (TLS) polymerase genes, predispose people to the event of a variety of tumor sorts¹.

Given the important roles of the DDR in cell survival and proliferation, chemotherapeutic medicine have been developed and broadly used to induce DNA harm in quickly proliferating tumor cells, thereby inducing replication stress and cell dying. Double-strand breaks (DSBs) and inter-strand or intra-strand crosslinks are typical varieties of dangerous DNA harm induced by chemotherapeutic medicine, corresponding to etoposide, cisplatin, and mitomycin (Table 1). DSB is among the many most poisonous types of DNA harm and is repaired predominantly via the error-free HR and error-prone non-homologous end-joining (NHEJ) pathways. Generally, platinum-induced intra-strand crosslinks are particularly bypassed by DNA polymerase eta (Polη), a classical issue within the TLS pathway, thus reducing the anti-tumorigenic impact of platinum-based medicine². Sadly, DDR defects in tumor cells not solely confer sensitivity to DNA-damaging brokers but additionally trigger further genetic mutations, thus selling acquired chemoresistance and facilitating tumor recurrence. Due to this fact, DDR pathways have each advantages and downsides in most cancers prevention and chemotherapy. This angle is aimed toward summarizing current research on the molecular mechanisms underlying DDR in chemoresistance, which have offered novel and promising biomarkers and methods for enhancing the efficacy of chemotherapy.

Mechanisms of chemoresistance to DNA-damaging brokers

DNA-damaging brokers exert antitumor results by inducing DNA harm. The mechanisms underlying DDR-related tumor chemoresistance recognized to this point could be categorized into 3 most important teams: genetic mutation mechanisms, together with genetic reversion of DDR defects, allelic variation in DDR genes and DDR pathway rewiring; post-translational modification (PTM) alteration mechanisms; and epigenetic transforming mechanisms (Figure 1).

Determine 1

DDR-related mechanisms of chemoresistance to DNA-damaging brokers. Resistance to DNA-damaging brokers may end up from restoration of DDR exercise after genetic mutation of DDR genes, PTM alterations in DDR proteins, and epigenetic transforming. ① Genetic reversion of DDR deficiency. Therapy with DNA-damaging brokers could cause secondary mutations resulting in partial or full restoration of DDR protein exercise, as proven for BRCA2. ② Allelic variation in DDR genes has differential results on the chemotherapy response. C124R and C129E are 2 frequent missense mutations in PTEN that lower PTEN stability or confer achieve of operate, respectively. ③ Pathway rewiring attributable to genetic mutation of DDR genes shifts the steadiness amongst DDR pathways, corresponding to 53BP1-mediated NHEJ and BRCA1-mediated HR. ④ PTMs modulate DDR sign transduction, as proven for ATM/CHK2-catalyzed phosphorylation after DSB induction. ⑤ PTMs regulate DDR protein stability. C1QBP stabilizes MRE11/RAD50 and protects it from degradation, and inhibits MRE11-dependent nonspecific resection within the absence of stress situations. Within the case of DSBs, ATM phosphorylates MRE11; subsequently, C1QBP dissociation from MRE11/RAD50 promotes MRE11/RAD50/NBS1 (MRN) complicated meeting and DSB finish resection. ⑥ PTMs management the part separation of DDR elements. LLPS of FUS is inhibited by its methylation and phosphorylation. ⑦ Chromatin rest surrounding DNA harm websites is answerable for environment friendly DDR protein accumulation and the promotion of chemoresistance. After DSB formation, histone PARylation triggers nucleosomal disassembly, and chromatin remodelers corresponding to ALC1, CHD4, and MORC2 orchestrate additional alterations in chromatin construction. ⑧ Histone PTMs have an effect on DDR issue accumulation and DDR pathway alternative. For instance, H2AK15 monoubiquitination and H4K20 dimethylation promote 53BP1-mediated NHEJ and antagonize BRCA1-mediated HR at DSBs. ⑨ DNA epigenetic modifications modulate DDR protein expression, as proven for the promoter methylation or acetylation of BRCA1. 53BP1, p53 binding protein 1; ALC1, amplified in liver most cancers 1; ATM, ataxia telangiectasia mutated; BARD1, BRCA1 related RING area 1; BRCA1, breast most cancers sort 1 susceptibility protein; BRCA2, breast most cancers sort 2 susceptibility protein; C1QBP, complement C1q binding protein; CHD4, chromodomain helicase DNA binding protein 4; CHK2, checkpoint kinase 2; DSB, double-strand break; FUS, fused in sarcoma; H2AK15, histone H2A lysine 15; H4K20, histone 4 lysine 20; HR, homologous recombination; KAP1, KRAB-associated protein 1; MORC2, microrchidia CW-type zinc finger 2; MRE11, meiotic recombination 11; NBS1, Nijmengen breakage syndrome 1; NHEJ, non-homologous end-joining; PARP1/2, poly(ADP-ribose) polymerase 1/2; PTEN, phosphatase and tensin homolog; RIF1, replication timing regulatory issue 1. Determine was created in Phrase Processing System (WPS) and Figdraw.

Genetic mutation mechanisms of chemoresistance

Throughout chemotherapy, tumor cells survive by buying genetic aberrations. As a result of the features of DDR proteins could be both restored or abolished by mutations in DDR genes, genetic mutations have differential results on remedy resistance. Genetic reversion of DDR defects, potential penalties of other splicing, frameshift mutations, or germline mutation correction, have been recognized in DDR-deficient tumors with acquired chemotherapeutic resistance. The very best-known instance is poly(ADP-ribose) polymerase (PARP) inhibitor (PARPi) medicine, which selectively kill HR-deficient tumor cells by inducing DSBs. PARPi remedy has proven nice promise within the scientific remedy of BRCA-mutant ovarian and breast tumors. Nonetheless, some BRCA-deficient tumors exhibit PARPi resistance pushed by genetic restoration of BRCA protein operate and reversal of HR exercise³. In distinction, some mutations in DDR genes would possibly partially or utterly abolish the operate of DDR proteins. Consequently, the existence of allelic variation in DDR genes has differential results on remedy outcomes. The one-nucleotide polymorphism (SNP) rs4971059 enhances the expression of tripartite motif 46 (TRIM46), which in flip acts as an E3 ligase concentrating on histone deacetylase 1 (HDAC1) for degradation, thus resulting in transcriptional upregulation of a panel of DDR genes endowing chemoresistance in breast most cancers⁴.

Furthermore, DDR pathway rewiring in tumors contributes to the event of resistance to DNA-damaging brokers. HR is crucial for error-free S/G2 part DSB restore, whereas defects in HR result in PARPi hypersensitivity. The S/G2 phase-specific binding of BRCA1 to DSBs promotes DNA finish resection and thereby prevents NHEJ. Nonetheless, in some intrinsic HR-deficient tumors, suppression of the NHEJ pathway alleviates PARPi sensitivity, due to the shift from the NHEJ to the HR pathway. The lack of a number of important elements that inhibit DNA finish resection, corresponding to p53 binding protein 1 (53BP1), the Shieldin complicated, and REV7, promotes HR restore and results in resistance in tumor cells with BRCA1 deficiency⁵.

Alterations in PTM mechanisms in chemoresistance

Past genetic mutations in DDR genes, PTM alterations are thought-about important in controlling chemotherapy outcomes by modulating the transduction of DDR alerts, the steadiness of DDR proteins, and the dynamic part separation of DDR elements. PTM entails dynamic and enzyme-mediated processes that covalently add practical teams at particular residues of substrate proteins. DDR proteins bear a number of varieties of PTMs, together with phosphorylation, ubiquitination, methylation, glycosylation, and lactylation. Specifically, ataxia telangiectasia mutated (ATM), ataxia telangiectasia mutated and Rad3-related (ATR), and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) are 3 principal kinases triggered by DNA harm at early levels of DDR, their activation is answerable for the phosphorylation of downstream DDR effectors and the magnitude of the DDR signaling cascade. Extra not too long ago, lactate-induced lactylation of DDR proteins, corresponding to meiotic recombination 11 (MRE11) and Nijmengen breakage syndrome 1 (NBS1), have been proven to advertise HR and contribute to scientific resistance to cisplatin or PARPi^6,7. These findings counsel that tumor metabolites can modulate DDR and chemoresistance. To restore DSBs, quite a few DDR elements, notably 53BP1 and fused in sarcoma (FUS), focus at harm websites and type droplet-like membraneless buildings through liquid-liquid part separation (LLPS)⁸. PTMs have been proposed to manage LLPS of DDR elements, thereby contributing to the DDR course of by altering protein conformation and protein–protein interactions. For instance, arginine methylation, along with phosphorylation, has been discovered to interrupt FUS LLPS formation^9,10.

So far, in depth crosstalk amongst PTM sorts has been described in DDR. Dysregulated expression of the enzymes answerable for DDR protein PTMs has been noticed in quite a lot of tumors and related to poor outcomes of remedy with DNA-damaging brokers. For instance, the methyltransferase SYMD3—which features in HR restore and genome stability via ATM methylation, thereby selling the DDR signaling cascade—is dysregulated and contributes to chemoresistance in colorectal most cancers, hepatocellular most cancers, and breast most cancers¹¹. In response to cisplatin remedy, proliferating cell nuclear antigen (PCNA), a homotrimeric sliding clamp within the replisome, is mono-ubiquitinated by the E3 ubiquitin ligase RAD18 and subsequently initiates the TLS pathway. Past its position in TLS, RAD18 coordinates different DDR pathways, together with HR and Fanconi anemia. Nonetheless, the expression of RAD18 is altered in varied tumors. Suppression of RAD18 decreases PCNA mono-ubiquitination, and will increase tumor sensitivity to cisplatin, etoposide, and camptothecin. Collectively, DDR protein-related PTM alterations are potential drivers of acquired most cancers drug resistance.

Epigenetic transforming mechanisms of chemoresistance

Epigenetic transforming mechanisms are answerable for DDR dysregulation and therefore drug resistance. Throughout DDR, chromatin undergoes transient rest, which in flip allows DDR proteins to entry broken DNA; after DDR completion, chromatin condensation happens. Histones are important in mediating chromatin conformation and DDR protein accumulation. In response to DNA-damaging brokers, the dynamic conformational shift of chromatin rest depends strongly on nucleosomal disassembly triggered by the PARylation of core histones. After histone PARylation, chromatin remodelers corresponding to amplified in liver most cancers 1 (ALC1) and chromodomain helicase DNA binding protein 4 (CHD4) are recruited by PARP1 and PARylated histones. These remodelers additional orchestrate alterations in chromatin construction, that are answerable for environment friendly DDR protein accumulation and promotion of chemoresistance. The expression of chromatin remodelers governs the efficacy of DNA-damaging brokers. For instance, ALC1 deficiency decreases chromatin accessibility and promotes apurinic/apyrimidinic endodeoxyribonuclease 1 (APE1)-dependent DSB formation and replication fork stalling, thereby leading to PARPi hypersensitivity in BRCA-mutant cells¹². A current examine has revealed that dynamic SUMOylation of microrchidia CW-type zinc finger 2 (MORC2), an ATPase-dependent chromatin transforming enzyme, contributes to correct chromatin transforming and DDR. Inhibition of MORC2 SUMOylation enhances the sensitivity of breast most cancers cells to chemotherapeutic medicine¹³. Past chromatin transforming, histone PTMs surrounding DNA harm websites immediately have an effect on DDR issue accumulation and DDR pathway alternative. For instance, the standing of histone H2A lysine 15 (H2AK15) ubiquitination and histone 4 lysine 20 (H4K20) methylation determines whether or not 53BP1-mediated NHEJ or BRCA1-mediated HR happens at DSBs¹⁴. Diacetylated H2A lysine 5 and lysine 9 (H2AK5acK9ac) round DSBs additionally antagonizes NHEJ restore by selling BRD4-Ku80 LLPS. The mitotic deacetylase complicated (MiDAC) removes di-acetyls from H2AK5K9 and suppresses BRD4-KU condensates, and due to this fact is a probable potential regulator in tumor chemotherapy¹⁵. Furthermore, DDR exercise is altered by modulation of the expression of DDR proteins, corresponding to BRCA1, through promoter epigenetic modifications, together with DNA methylation and acetylation, which steadily happen in tumors. Therefore, concentrating on epigenetic transforming together with DNA-damaging brokers would possibly reverse tumor cell resistance.

Overcoming chemoresistance through the DDR pathway

Given the important roles of DDR in intrinsic and purchased drug resistance throughout chemotherapy, efforts have been made to handle chemoresistance by rising the toxicity of DNA-damaging medicine throughout chemotherapy. In precept, past efforts to boost drug uptake, intrinsic drug resistance in tumors may very well be prevented with exact drugs regimens emphasizing the exact matching of particular DDR-deficient sufferers to focused therapies. To maximise the efficacy of chemotherapy and overcome hostile results in regular cells, creating biomarkers of DDR pathways that predict the response to DNA-damaging brokers is important. Hereditary mutations or copy quantity aberrations of DDR genes are referred to as potential predictive biomarkers for choosing optimum therapeutics. For instance, HR deficiency (HRD) phenotype detection is used for affected person stratification in scientific PARPi remedy. Past the two most typical indicators of HR exercise, BRCA1 and BRCA2, a computational software, signature multivariate evaluation (SigMA), has been demonstrated to precisely detect HRD candidates¹⁶. Furthermore, quite a few DDR regulators, together with proteins and noncoding RNAs, have been recognized as potential practical biomarkers and druggable targets. For instance, the lengthy noncoding RNA CTD-2256P15.2-derived micropeptide PACMP, which performs a twin position within the upkeep of CtBP-interacting protein (CtIP) abundance and poly(ADP-ribosyl)ation stimulation, causes resistance to a number of DNA-damaging brokers, together with epirubicin, camptothecin, and PARPi¹⁷. Just lately, vestigial-like 3 (VGLL3), a vital transcription cofactor, has been proven to advertise HR restore, whose inhibition sensitizes tumor cells to etoposide remedy and additional inhibits tumor development¹⁸. Consequently, on the idea of quite a few classical and potential DDR biomarkers, higher therapeutic outcomes could be achieved via personalised drug remedy and novel drug improvement.

As mentioned above, acquired drug resistance ensuing from dysregulated DDR exercise poses a significant impediment to using DNA-damaging brokers in scientific settings. Mixture remedy methods that inhibit the response of key DDR proteins to chemotherapeutic brokers or goal particular DDR defects maintain promise for re-sensitizing tumors to chemotherapeutics. Due to this fact, the event of DDR inhibitors is a gorgeous technique for reversing drug resistance (Table 2). Given the first roles of ATM and ATR kinases in DDR signaling cascade activation, the inhibition of ATM or ATR, coupled with chemotherapy medicine, will increase antitumor efficacy¹⁹. Equally, inhibitors of downstream targets of those kinases, corresponding to checkpoint kinase 1 (CHK1), exacerbate the cytotoxicity of DNA-damaging brokers²⁰. The TLS pathway allows bypassing of cisplatin-induced intra-strand crosslinks and contributes to cisplatin resistance. Nonetheless, elevated TLS capability has been noticed in tumor cells, thus resulting in cell survival and chemoresistance. Compounds that restrict TLS exercise are anticipated to be potential chemosensitizers in tumor remedy. Past the small chemical molecules that inhibit the TLS polymerase REV1, the pure TLS inhibitor ganoboninketal C, derived from Ganoderma mushrooms, has not too long ago been reported to sensitize tumor cells to cisplatin remedy²¹. Furthermore, restored HR exercise in HR-deficient tumors could be eradicated by inhibitors concentrating on DDR proteins, together with RAD51 and ATR, thus additional enhancing the sensitivity of tumor cells to PARPi. Drug treatment-induced epigenetic adaptation contributes to the buildup of drug-resistant cells. Due to this fact, concentrating on epigenetic regulators is an efficient technique to reverse drug resistance. Inhibitors of chromatin remodelers, corresponding to HDACs, have synergistic killing results when utilized in mixture with PARPi remedy in triple-negative breast most cancers; this routine is of nice curiosity in testing the reversibility of PARPi resistance.

Abstract

Typically, most chemotherapy medicine are DNA-damaging brokers that set off distinct varieties of DNA harm and due to this fact cell dying. Nonetheless, the emergence of innate or acquired resistance happens in lots of tumors throughout remedy. Tumor drug resistance is a multifactorial phenomenon intently related to the DDR—a course of essential for genomic integrity. Three totally different however not mutually unique mechanisms involving genetic mutation, PTM alteration, and epigenetic transforming have been implicated within the dysregulation of DDR and poor chemoresponse in tumor cells. Inhibitors concentrating on DDR proteins, mixture therapies involving DNA damaging brokers, and novel DDR factor-targeting compounds are steadily used methods to enhance antitumor efficacy.

After publicity to DNA-damaging brokers, cells rewire organic actions corresponding to rRNA biosynthesis, protein translation, endomembrane morphological change, and power metabolism, thereby integrating them with the DDR and fine-tuning the systemic stress response. Specifically, whereas elevated ranges of 2-hydroxyglutarate (2HG), succinate, and fumarate suppress HR, the buildup of lactate, a byproduct of the Warburg impact, promotes DDR exercise in most cancers cells, thus demonstrating a robust synergy between oncometabolites and the DDR. Due to this fact, oncometabolite interventions could also be a promising strategy for enhancing chemotherapy outcomes. Just lately, an inhibitor of lactate dehydrogenase (LDHA), the glycolytic enzyme catalyzing pyruvate to lactate, has been discovered to alleviate the resistance of most cancers cells to DNA-damaging brokers. The innate immune response will also be activated by the DNA lesions attributable to DNA-damaging brokers, and the mix of DDR inhibitors with immune checkpoint blockade is one other promising technique for reversing chemoresistance. Given the excessive molecular heterogeneity amongst sufferers with most cancers, predictive biomarkers of DDR inhibitors ought to be rigorously validated in additional research and scientific trials. In conclusion, a deeper understanding of DDR regulation in tumor cells wouldn’t solely reveal how chemoresistance happens after DNA harm remedy but additionally assist within the improvement of methods to beat chemoresistance.

Writer contributions

Conceived and designed the evaluation: Xiaolu Ma and Caixia Guo.

Wrote the paper: Xiaolu Ma and Zina Cheng.

Revised the paper: Caixia Guo.