Tumor irradiation induced immunogenic response: the impact of DNA damage induction and misrepair

Abstract

Abstract

Focal tumor irradiation, a cornerstone of cancer therapy, has been increasingly recognized for its capacity to provoke systemic immunogenic responses that extend beyond localized tumor control. Recent advances highlight DNA damage, especially DNA double-strand breaks (DSBs), as a central mediator linking radiotherapy to anti-tumor immune activation. Importantly, DNA misrepair, prevalent in cancer cells with deficient or dysregulated repair machinery, serves as a double-edged sword: while fostering tumor adaptation and genomic instability, it also fuels immune recognition through the accumulation of neoantigens, extracellular DNA release, immunogenic cell death, and the modulation of immune-related cytokines and chemokines. This review critically synthesizes the latest clinical and preclinical insights into the dynamic interplay between DNA damage, repair fidelity, and the immunogenic consequences of tumor irradiation. By focusing on the impact of DSB induction and misrepair processes, we underscore the emerging therapeutic opportunities of modulating DNA repair pathways during radiotherapy to potentiate anti-tumor immunity, particularly in synergy with immune checkpoint blockade. This article provides a comprehensive perspective on the molecular underpinnings and translational potential of harnessing irradiation-induced immunogenicity, offering a roadmap for future therapeutic strategies in radiation oncology and cancer immunotherapy.

Clinical trial number

Not applicable

Background

Radiotherapy remains a foundational modality in cancer care, with over half of all cancer patients receiving it as part of their treatment [1]. Although its principal efficacy has long been attributed to the direct cytotoxic effects of ionizing radiation (IR) on tumor cell DNA, clinical observations of the “abscopal effect”, where distant, non-irradiated tumors regress following localized irradiation, highlight the broader, systemic potential of radiotherapy through immune activation [2].

Preclinical and clinical studies have established that the abscopal effect is dependent on an intact and responsive immune system [2, 3]. This has driven efforts to leverage radiotherapy not only for local control but also as a catalyst for systemic anti-tumor immunity. Notably, combining radiotherapy with immune checkpoint inhibitors (ICIs) has been shown to boost abscopal responses, resulting in improved outcomes in cancers like non-small cell lung cancer [4–7]. Thus, IR-induced immunogenic effects are increasingly recognized as a major contributor to the overall therapeutic benefit of radiotherapy [8].

Tumor development involves a dynamic struggle with host immune surveillance [9]. While emerging tumor cells are initially targeted and cleared by the immune system, cancers acquire mechanisms for immune escape, facilitating an immunosuppressive tumor microenvironment (TME) through upregulation of inhibitory checkpoints, cytokines, downregulation of class I major histocompatibility complex (MHC-I) molecules, and recruitment of suppressive immune cells. These adaptations allow unchecked tumor growth. Importantly, IR can partially overcome these immune evasion strategies by enhancing tumor immunogenicity through DNA damage-driven processes [10].

Despite this, most cancers possess impaired DNA repair machinery, leading to frequent misrepair and persistent genomic instability after radiation. This instability not only promotes tumor adaptation but can also produce immunostimulatory signals that enhance anti-tumor immune responses [5]. However, radiotherapy alone rarely produces sustained immune-mediated tumor control, prompting the development of combination strategies that more effectively harness IR-induced immunogenicity [3, 10].

In this review, we examine radiation-induced tumor immunogenicity, focusing on the role of DNA damage and defective repair mechanisms. We discuss advances in targeting DNA repair to augment anti-tumor immunity and consider the prospects of using radiotherapy to prime therapeutic cancer vaccines. By synthesizing emerging data and mechanistic insights, this review aims to inform about strategies that harness radiation-induced immunogenicity to support long-term disease control in cancer patients.

DNA damage induction and repair mechanisms related to tumor irradiation

IR induces DNA damage through both direct energy deposition on DNA molecules and indirect effects via the generation of reactive oxygen species (ROS) from water radiolysis [11]. These processes lead to a broad spectrum of DNA lesions, including base modifications, single-strand breaks (SSBs), and double-strand breaks (DSBs). For instance, exposure to 1 Gy of X-rays produces approximately 1,000 single-strand breaks (SSBs), 40 double-strand breaks (DSBs), 30 DNA-DNA crosslinks, and more than 2,000 base lesions in a single mammalian cell [11]. The cumulative DNA insult inflicted during a typical radiotherapy course results in massive genotoxic stress in tumor cells.

Recognition and repair of IR-induced DNA damage are governed by a highly regulated DNA damage response (DDR), coordinated by key kinases such as ATM (ataxia-telangiectasia mutated), ATR (ATM and Rad3-related), and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) [12, 13]. DDR activation triggers cell cycle checkpoints, DNA repair, and in some cases cell death. DNA lesions are addressed by distinct repair pathways: base excision repair (BER) for small base lesions, nucleotide excision repair (NER) for bulky distortions, mismatch repair (MMR) for replication errors, interstrand crosslink (ICL) repair for covalent linkage between DNA strands [14–16], etc. (Fig. 1).

Fig. 1.

DNA repair pathways for IR-induced DNA damages. Base Excision Repair (BER), Mismatch Repair (MMR), Single-Strand Break Repair (SSBR), Homologous Recombination (HR), Non-Homologous End Joining (NHEJ), Alternative End-Joining (Alt-EJ) and Fanconi Anemia (FA) Pathway

Of all DNA damage types, DSBs are the most lethal and pivotal to the efficacy and biological consequences of radiotherapy. DSBs are primarily repaired by non-homologous end joining (NHEJ) and homologous recombination (HR) [17]. NHEJ is the predominant and rapid pathway, especially active in G1 phase, joining DNA ends with minimal processing but is prone to small insertions or deletions. HR, restricted to S/G2 phases, uses a homologous DNA template for error-free repair, requiring proteins such as BRCA1, BRCA2, and RAD51. Deficiency in HR (HRD) is a frequent driver of tumorigenesis, and influences sensitivity to DDR-targeting therapies, such as poly(ADP-ribose) polymerase inhibitors (PARPi) [18].

A third pathway, alternative end-joining (Alt-EJ) or microhomology-mediated end Joining (MMEJ), becomes prominent when NHEJ or HR is compromised [19–21]. MMEJ utilizes short microhomologous sequences near DSBs and involves factors such as poly(ADP-ribose) polymerase 1 (PARP1), polymerase theta (POLQ), and DNA ligase 1 (LIG1) or 3 (LIG3). This pathway is intrinsically error-prone, often causing deletions, insertions, and chromosomal translocations, contributing to genomic instability. MMEJ is usually upregulated in cancer cells, especially for those with defective DDR [22, 23].

Despite the efficiency of DDR, extensive DNA damage from radiotherapy can overwhelm cellular repair systems, increasing the likelihood of misrepair. Misrepair generates genomic alterations, including mutations and chromosomal aberrations, leading to mitotic catastrophe and reproductive cell death [20, 24]. Misrepair is an essential option for maintenance of genome integrity and prevention of cell death. Notably, in cancer cells, misrepair-driven genomic instability often provides a survival advantage, enhancing tumor progression and resistance to therapy [25]. Tumors with DDR gene mutations or overwhelmed repair systems show elevated misrepair rates, enhancing genomic instability and tumor evolution [24, 26, 27].

DNA damage-induced tumor immunogenicity

DNA damage, a threat to genomic stability and integrity, contributes to tumor immunogenicity by driving genetic mutations and genomic alterations. During radiotherapy, IR generates extensive DNA damage, which can increase the tumor mutational burden (TMB), defined as the total number of mutations per megabase of DNA in the cancer genome. High TMB enhances immunogenicity and predicts better responses to ICIs across cancer types [5]. Meta-analyses confirm TMB as a robust biomarker for ICI efficacy irrespective of tumor origin [28], and retrospective studies further associate high TMB with improved overall survival in ICI-treated patients [29]. Radiotherapy not only increases TMB but also alters the mutational landscape, particularly by inducing small and large deletions, as observed in gliomas [30].

Beyond mutational effects, IR-induced DNA damage influences tumor-immune interactions by modulating immune ligand expression. For example, a positive correlation exists between γH2AX (DSB marker) and programmed death-ligand 1 (PD-L1) expression in lung squamous cell carcinoma, indicating that DSBs drive PD-L1 upregulation [31]. Consistently, the ATM/ATR/CHK1 kinases, which are activated in response to DSBs, regulate PD-L1 expression [32]. As expected, IR can upregulate PD-L1 expression in cancer cells, potentially contributing to radioresistance [33, 34]. High PD-L1 expression, nevertheless, predicts improved outcomes with ICI therapy, and combinations of radiation, anti-CTLA4 and anti-PD-L1 have shown promoted response and immunity in a metastatic melanoma study [35].

In parallel, DNA damage boosts the expression of immunostimulatory ligands, such as NKG2D and DNAM-1, recognized by NK and T cells and instrumental in anti-tumor immunity [36, 37]. Severe DNA damage also triggers the release of damage-associated molecular patterns (DAMPs), including HMGB1, heat shock proteins, ATP, calreticulin (CALR), and others, which activate innate immunity and drive adaptive immune responses [38]. Elevated DAMP levels, such as nuclear-to-cytoplasmic HMGB1 translocation or CALR upregulation, correlate with favorable therapeutic outcomes in rectal, head and neck, and lung cancers after radiotherapy [39–41].

The process of immunogenic cell death (ICD), characterized by CALR exposure, HMGB1 release, and ATP secretion, is frequently induced by IR and has been linked to superior immune responses and clinical benefits in various cancers [42–45]. Additionally, radiotherapy-induced DNA damage promotes cytosolic DNA release, especially double-stranded DNA (dsDNA), from DSBs or micronuclei formation. Such DNA is sensed by the cyclic GMP-AMP (cGAMP) synthase (cGAS), which triggers the cGAS-STING pathway, stimulating production of type I interferons (e.g., IFNβ) and interferon-stimulated genes (ISGs), and enhancing dendritic cell-mediated antigen presentation [46–52]. Notably, defects in classical DSB repair (e.g., DNA-PKcs or BRCA1 loss) enhance cytosolic DNA accumulation and further potentiate cGAS-STING signal transduction, thereby increasing the tumor immunogenicity [53–56].

Immunogenic effects of DNA damage misrepair in irradiated cancer cells

Misrepair of IR-induced DNA damage promotes profound immunogenic effects by increasing genomic instability, generating neoantigens, and releasing immunostimulatory signals (Fig. 2) [57, 58]. Aberrant DDR mechanisms, frequently observed in cancer, underlie these misrepair events [59]. Radiotherapy further enhances misrepair-driven immunity by overwhelming repair pathways with extensive DNA damage. For example, tumors with MMR deficiencies display high TMB and neoantigen loads, which not only confer increased effects to ICIs [60, 61], but also are associated with improved responses to irradiation [62].

Fig. 2.

Immunogenic effects of DNA damage misrepair in irradiated cancer cells. IR elicits immunogenic cell death (ICD) through stimulation of damage-associated molecular patterns (DAMPs) (A); IR modulates the immune synapse by inducing expression of immune checkpoint molecules on tumor cell surfaces (B); IR enhances tumor cell antigenicity via generation of tumor neoantigens (C); Activation of cytosolic immunity pathways enhances tumor radiosensitivity (D). ATP: adenosine triphosphate; CALR: calreticulin; DC: dendritic cell; ER: endoplasmic reticulum; HMGB1:high mobility group box 1; ISG: interferon-stimulated genes; MHC: major histocompatibility complex; M6P: mannose 6-phosphate; NK: natural killer; p:phosphorylated; PD1:programmed death 1; PDL1:programmed death ligand 1; TCR: T cell receptor; P2RX7:purinergic receptor P2X 7; LRP1:low density lipoprotein related protein 1; FAS: factor-related apoptosis; FASL: factor-related apoptosis ligand; IGF2R: insulin like growth factor 2 receptor; TRAIL: tumor-necrosis factor related apoptosis-inducing ligand; TRAILR: TRAIL-receptor; CXCL10:C-X-C motif chemokine ligand 10; CXCR3:C-X-C motif chemokine receptor 3; IFNB1:interferon beta 1; IFNAR1:interferon beta receptor 1; CCL5:C-C motif chemokine ligand 5

Repair of DSBs is central to the immunogenic effects of radiotherapy. Inhibition of key DDR kinases involved in DSB repair, such as ATM or ATR, leads to the accumulation of cytosolic DNA and activation of the cGAS-STING pathway, thereby amplifying anti-tumor immune responses following irradiation [63, 64]. Moreover, substantial evidence indicates that abnormalities in major DSB repair pathways, including NHEJ, HR, and alt-EJ, are linked to enhanced tumor immunogenicity, particularly in the context of IR treatment.

NHEJ

NHEJ has an error-prone characteristic in DSB repair when compared with HR. NHEJ-mediated misrepair can generate chromosomal rearrangements and aberrant fragments, which missegregate during cell division and form micronuclei. The rupture of these micronuclei releases genomic dsDNA into the cytoplasm, activating the cGAS-STING pathway and eliciting immunogenic responses [65, 66]. Because NHEJ is primarily used to maintain genomic stability, its deficiency exacerbates DSB repair defects and increases immunogenicity in cancer cells. For example, PRKDC (DNA-PKcs) nonsense mutations in non-small cell lung cancer (NSCLC) are associated with high TMB and better responses to ICIs [67]. Consistent findings show that inhibition of DNA-PKcs enhances neoantigen diversity, upregulates MHC-I, and boosts innate immunity in melanoma and small cell lung cancer (SCLC) [68, 69]. In pancreatic cancer models, combined DNA-PK inhibition and IR enhanced anti-tumor immunity via cytosolic RNA sensor RIG-I and RNA polymerase III pathways, independent of cGAS-STING [70]. In addition, depletion of ARID1A, an epigenetic regulator required for both HR and NHEJ, promotes micronuclei formation and increases immune cell infiltration after irradiation [71].

HR

Tumors with HRD demonstrate increased rates of misrepair, resulting in higher TMB and neoantigen loads compared to HR-proficient tumors [72]. HRD is characterized by unique mutational patterns, such as specific base substitutions and indels with microhomology, which contribute to a distinctive immunogenic profile [73]. Additionally, HRD is associated with three specific chromosomal alterations: telomeric allelic imbalance, loss of heterozygosity, and large-scale state transitions, which together comprise the HRD score and predict responses to DDR-targeted therapy [72]. Tumors with high HRD scores tend to have active immune signaling pathways and are more responsive to ICIs [74]. IR further enhances genomic instability and ISG expression in HRD tumors, particularly those with BRCA1 or BRCA2 mutations [75]. Notably, BRCA2 mutations may have a stronger impact on both adaptive and innate immunity than BRCA1 mutations, indicating complex HRD-dependent immunogenic mechanisms [76].

Alt-EJ

Alt-EJ is a highly mutagenic pathway for DSB repair, which frequently induces chromosomal translocations, deletions, and indels with microhomology at the junction sites, fostering neoantigen generation [19, 20, 77–80]. TCGA analyses reveal that alt-EJ gene expression correlates with mutation signature ID6, and indel-rich profiles associate closely with tumor-specific neoantigen production [23, 81, 82]. High alt-EJ activity, especially in tumors with low TGFβ signaling, is linked to improved ICI response [83]. Key alt-EJ proteins, such as PARP1 and POLQ, have additional immunogenic roles: PARP1 suppresses PD-L1 transcription and is associated with TMB, immune infiltration, and ICI responsiveness [84, 85], and is required for IR-induced NF-κB activation [86]. Elevated POLQ expression in bladder cancer is related to both chromosomal instability and an active tumor microenvironment [87]. Radiotherapy amplifies alt-EJ-dependent misrepair, especially in contexts of deficient TGFβ signaling; loss of POLQ abrogates these effects, further supporting alt-EJ’s importance in immunomodulation [88]. Notably, high alt-EJ activity with low TGFβ signaling is associated with favorable prognosis in radiotherapy-treated patients [23].

DDR Inhibition promotes IR-induced tumor immunogenicity

DDR inhibitors enhance tumor cell death when combined with radiotherapy [89]. By exacerbating DNA damage, these agents accelerate micronuclei formation, increase TMB, promote genomic instability, facilitate neoantigen generation, and activate immunostimulatory pathways [90–95]. Consequently, DDR inhibition markedly augments tumor immunogenicity in irradiated cancers. In recent years, agents targeting DDR components, particularly those involved in DSB repair, have attracted considerable interest in translational research. Several DDR inhibitors are currently being evaluated in clinical trials as part of combination regimens with radiotherapy and ICIs (Table 1) [53, 96].

Table 1.

Clinical trials of combined radiotherapy, DDR inhibitors, and ICIs

Trial ID	Phase	Tumor type	RT type	DDRi		ICI
NCT04550104	Phase 1	NSCLC	RT	Ceralasertib AZD1390 Olaparib Saruparib	ATRi ATMi PARPi	Durvalumab
NCT04068194	Phase 1/2	Cholangiocarcinoma Gallbladder Carcinoma	HFRT	Peposertib	DNA-PKi	Avelumab
NCT04711824	Phase 1/2	Breast cancer	SRS	Olaparib	PARPi	Durvalumab
NCT05568550	Phase 2	Prostate Cancer	RT	Olaparib	PARPi	Pembrolizumab
NCT05379972	Phase 2	Gastric Cancer GastroEsophageal Cancer	SBRT	Olaparib	PARPi	Pembrolizumab
NCT06074692	Phase 2	Sarcoma	SBRT	Fluzoparib	PARPi	Camrelizumab
NCT05568550	Phase 2	Prostate Cancer	RT	Olaparib	PARPi	Pembrolizumab
NCT05366166	Phase 2	HNSCC	IMRT	Olaparib	PARPi	Pembrolizumab
NCT04380636	Phase 3	NSCLC	RT	Olaparib	PARPi	Pembrolizumab
NCT04624204	Phase 3	LS-SCLC	RT	Olaparib	PARPi	Pembrolizumab
NCT05411094	Phase 1	Pancreatic Cancer	RT	Olaparib	PARPi	Durvalumab
NCT04728230	Phase1/2	ES-SCLC	RT	Olaparib	PARPi	Durvalumab
NCT06197581	Not Applicable	Breast Cancer	RT	Olaparib	PARPi	Pembrolizumab
NCT04683679	Phase 2	Breast Cancer	SBRT	Olaparib	PARPi	Pembrolizumab
NCT04837209	Phase 2	Breast Cancer	RT	Niraparib	PARPi	Dostarlimab
NCT04681469	Phase 3	Prostate Cancer	RT	Olaparib	PARPi	Pembrolizumab
NCT03834519	Phase 3	Prostate Cancer	EBRT	Olaparib	PARPi	Pembrolizumab
NCT03307785	Phase 1	NSCLC	RT	Niraparib	PARPi	Dostarlimab

Note: Full access to the research studies description is available on ClinicalTrials.gov

Abbreviations: NSCLC, non-small cell lung cancer; HNSCC, head and neck squamous cell carcinoma; LS-SCLC, limited-stage small-cell lung cancer; ES-SCLC, extensive-stage small-cell lung cancer; RT, radiotherapy; HFRT, hypofractionated radiotherapy; SBS, stereotactic radiosurgery; SBRT, stereotactic body radiation therapy; IMRT, intensity modulated radiotherapy; EBRT, external-beam radiation therapy

ATM inhibitors

ATM, a key sensor of DSBs, orchestrates the DDR and safeguards genomic stability [12]. ATM inhibition has been shown to enhance tumor immunogenicity: it promotes the release of both chromosomal and mitochondrial DNA into the cytosol, leading to robust activation of the cGAS-STING pathway and increased lymphocyte infiltration in the TME [97]. Further, ATM inhibition alone or combined with radiotherapy has been reported to enhance tumor immunogenicity by activating type I interferon signaling in a TANK-binding kinase 1 (TBK1)-dependent manner, increasing PD-L1 expression, and sensitizing pancreatic cancer to ICI therapy in vivo [91]. ATM inhibition also disrupts pro-survival pathways in cancer, counteracting IL-6-mediated resistance in glioblastoma, and inhibiting survival pathways such as NF-κB and TIGAR, thereby sensitizing tumors to radiotherapy and radiomimetics [98, 99].

ATR inhibitors

ATR, activated in response to replication stress and fork collapse, facilitates end resection for HR repair of replication fork-derived DSBs [12]. Pharmacological inhibition of ATR increases genomic instability and radiosensitizes cancer cells [92]. Beyond promoting direct tumor cell kill, ATR inhibition in combination with IR induces pronounced immunogenic effects across cancer types [92, 93, 100, 101]. Dillon et al. demonstrated that ATR inhibition potentiates the immunogenic effects of radiotherapy by increasing antigen presentation, immune cell infiltration, and cytokine modulation, driven by augmented interferon signaling and nucleic acid sensing pathway activation [92]. Vendetti et al. further showed that ATR inhibition prevents IR-induced PD-L1 upregulation, increases CD8⁺ T cell activity, and generates durable immunologic memory [101].

Mechanistic studies suggest that cGAS-STING, along with other nucleic acid-sensing pathways (e.g., MAVS-dependent RNA sensing), plays a crucial role in the immunogenic effects of ATR inhibition in irradiated tumors [64]. In hepatocellular carcinoma, ATR inhibition combined with radiotherapy synergistically improved anti-PD-L1 responses via cGAS-STING activation [93]. In colorectal cancer, ATR inhibitors in combination with IR increased cGAS-STING signaling, CD8⁺ T cell infiltration, cytokine production, and NFκB-P65 activation [102]. In HNSCC, ATR inhibition augmented radiation-induced inflammation, boosted NK cell activity, and improved responses to ICI therapy [103]. ATR inhibition has also been implicated in the induction of ICD in irradiated cancer cells [104].

DNA-PK inhibitors

DNA-PK, a key component of the NHEJ pathway, plays a critical role in addressing IR-induced DSBs [105]. DNA-PK inhibition significantly increases genome instability, sensitizes tumors to radiotherapy, and enhances anti-tumor immune responses. For instance, DNA-PK inhibition suppresses PD-1 expression in T cells, increases granzyme B expression in NK cells, and augments type I interferon signaling in irradiated tumor models, leading to durable tumor control via immunologic memory [106]. DNA-PK inhibition promotes micronuclei formation by disrupting chromosomal alignment and segregation, inducing cGAS-STING activation in irradiated cancer cells, resulting in enhanced CD8⁺ T cell infiltration and heightened efficacy of dual-targeting ICI therapy [94]. In pancreatic cancer, co-targeting DNA-PK and IR increased cytosolic dsDNA and stimulated anti-tumor immunity via RNA polymerase III–dependent pathways [70]. In HNSCC models, DNA-PK inhibitors combined with IR and PD-1 blockade synergistically increased cGAS-STING signaling and CD8⁺ T cell infiltration, leading to superior tumor control [107].

PARP inhibitors

PARP1 is a crucial mediator of DDR, particularly in DSB repair through the alt-EJ pathway. PARP inhibitors are established therapeutics for HRD cancers [108]. PARP inhibitor can trap PARP1 protein onto DNA, generating complex DNA damages that require HR for restoration [109]. These genotoxic effects of PARP inhibitors increase tumor immunogenicity. Indeed, accumulated TMB and neoantigens, upregulated PD-L1 expression and enhanced interferon signaling have been observed in tumors treated with PARP inhibitor, which facilitate a better response to anti-PD-L1 immunotherapy [95].

The combination of PARP inhibition and radiotherapy amplifies these immunogenic effects. For example, in tumors harboring IDH1 mutations, PARP inhibition with IR remodels the immunosuppressive microenvironment by increasing DNA damage, cGAS-STING activation, and infiltration of CD8⁺ T cells [110]. In a NSCLC model, PARPi synergizes with radiotherapy to activate cGAS-STING signaling, promote CD8⁺ T lymphocyte infiltration, and suppress tumor growth [111]. Similarly, in SCLC, the combination of PARPi and IR induces an inflamed tumor microenvironment and enhances anti-PD-1 efficacy, partly through cGAS-STING activation and the induction of ICD [112]. In colorectal cancer, this combination induces cGAS-mediated ferroptosis [113], and enhances responsiveness to anti-PD-1 therapy [114]. Additionally, radiotherapy with PARPi triggered STING activation and reinforced both local and abscopal tumor control in hepatocellular carcinoma [115].

POLQ inhibitors

Cancer cells often overexpress POLQ to cope with DSBs by alt-EJ, making POLQ a compelling therapeutic target [21]. High POLQ expression is associated with an inflamed TME and improved immunotherapy responses in muscle-invasive bladder cancer [87]. POLQ inhibition may further expose the tumor immunogenicity. Notably, POLQ inhibition is synthetically lethal in HRD tumors and, when ablated, increases micronuclei formation, activates cGAS-STING signaling, and recruits activated CD8⁺ T cells, as demonstrated in BRCA2-deficient pancreatic cancer models [116]. The combination of POLQ inhibition with radiotherapy exacerbates DNA damage and further elevates cytosolic DNA and type I interferon expression [117].

While combining DDR inhibition with radiotherapy and immune checkpoint inhibitors shows promising effects, several challenges remain for clinical translation. Key issues include optimizing treatment sequencing, overcoming tumor heterogeneity and immune escape, and managing increased toxicity from combination strategies. Continued research into patient selection, biomarker-driven approaches, and careful toxicity monitoring will be vital to fully realize the therapeutic potential of DDR-RT-ICI regimens.

Conclusions

Radiotherapy stands out as a compelling partner for immunotherapy, offering both direct tumor cell kill and immunomodulatory effects through the induction of DNA damage [118, 119]. Among these DDR-related effects, DNA damage misrepair is particularly important, as it not only promotes tumor adaptation but also underpins enhanced immunogenicity. This review underscores how DNA misrepair increases tumor antigenicity by raising TMB, generating neoantigens, and activating innate immune pathways such as cGAS-STING. Understanding the interplay between genomic instability and immune response can reveal tumor vulnerabilities and inform novel treatment strategies.

However, considerable inter-tumor heterogeneity in DDR capacity and repair pathway preference, as well as differences in radiotherapy dose and fractionation, create variable immunogenic outcomes [120]. Tailoring radiotherapeutic approaches to accommodate tumor DNA repair deficiencies and immune responsiveness may significantly improve therapeutic outcomes [121].

Targeted inhibition of DDR pathways represents a transformative approach for amplifying the immunogenic effects of radiotherapy. By disrupting key repair pathways such as NHEJ, HR, and alt-EJ, DDR inhibitors augment DNA damage misrepair, driving heightened tumor genomic instability, enhanced neoantigen production, and robust pro-inflammatory immune signaling [24]. The integration of DDR inhibitors with radiotherapy and immunotherapy holds strong potential for durable, immune-mediated tumor control.

Looking ahead, precision oncology will benefit from further research clarifying the links between DNA misrepair and tumor immunogenicity, supporting the design of individualized, multimodal combination therapies. Such efforts may ultimately enable more effective and lasting anti-tumor immune responses across a broad range of cancers.

Abbreviations

IR

Ionizing radiation

ICIs

Immune checkpoint inhibitors

NSCLC

Non-small cell lung cancer

TME

Tumor microenvironment

MHC-I

Class I major histocompatibility complex

ROS

Oxygen species

SSBs

Single-strand breaks

ATM

Ataxia-telangiectasia mutated

ATR

ATM and Rad3 related

DNA-PKcs

DNA-dependent protein kinase catalytic subunit

DDR

DNA damage response

ICL

Interstrand crosslink

DSBs

Double-strand breaks

BER

Base excision repair

NER

Nucleotide excision repair

FA

Fanconi anemia

NHEJ

Non-homologous end Joining

HR

Homologous recombination

LIG4

Ligase 4

ssDNA

single-stranded DNA

HRD

HR deficiency

Alt-EJ

Alternative end-joining

MMEJ

Microhomology-mediated end joining

PARP1

Poly ADP-ribose polymerase-1

POLQ

Polymerase theta

LIG1

Ligase 1

TMB

Tumor mutational burden

PD-L1

Programmed death-ligand 1

NKG2D

Natural killer group 2D

DNAM-1

DNAX accessory molecule-1

DAMPs

Damage-associated molecular patterns

HMGB1

High-mobility group box 1

HSPs

Heat shock proteins

CALR

Calreticulin

dsDNA

Double-stranded DNA

HNSCC

Head and neck squamous cell carcinoma

ICD

Immunogenic cell death

cGAS

cyclic GMP-AMP synthase

cGAMP

cyclic GMP-AMP

ISGs

Interferon-stimulated genes

TCGA

The Cancer Genome Atlas

SCLC

Small cell lung cancer

HPV

Human papillomavirus

TBK1

TANK-binding kinase 1

IL-6

Interleukin-6

TIGAR

TP53-induced glycolysis and apoptosis regulator

MMR

Mismatch Repair

SSBR

Single-strand break repair

DAMPs

Damage-associated molecular patterns

ATP

Adenosine triphosphate

DC

Dendritic cell

ER

Endoplasmic reticulum

HMGB1

High mobility group box 1

M6P

Mannose 6-phosphate

NK

Natural killer

p

phosphorylated

PD1

Programmed death 1

TCR

T cell receptor

P2RX7

Purinergic receptor P2X 7

LRP1

Low density lipoprotein related protein 1

FAS

Factor-related apoptosis

FASL

Factor-related apoptosis ligand

IGF2R

Insulin like growth factor 2 receptor

TRAIL

Tumor-necrosis factor related apoptosis-inducing ligand

TRAILR

TRAIL-receptor

CXCL10

C-X-C motif chemokine ligand 10

CXCR3

C-X-C motif chemokine receptor 3

IFNB1

Interferon beta 1

IFNAR1

Interferon beta receptor 1

CCL5

C-C motif chemokine ligand 5

RT

Radiotherapy

HFRT

Hypofractionated radiotherapy

SBS

Stereotactic radiosurgery

SBRT

Stereotactic body radiation therapy

IMRT

Intensity modulated radiotherapy

EBRT

External-beam radiation therapy

Author contributions

Conceptualization, Q. L. and L.M.; investigation, Q.L., N.T. and X.S.; writing—original draft preparation, Q.L., N.T., and X.S.; writing—review and editing, Q.L., X.S., and L.M.; visualization, X.S. and Q.L.; supervision, Q.L. and L.M.; funding acquisition, Q. L. and L.M.

Funding

This study was supported by National Natural Science Foundation of China (#82373212 to Q.L.; #82203967 to L.M.); Guangdong Basic and Applied Basic Research Foundation (#2023A1515011945 to L.M.); Shenzhen Science and Technology Program (#JCYJ20240813142112017 to L.M.).

Data availability

No datasets were generated or analysed during the current study.

Declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.