When breaks get hot: inflammatory signaling in BRCA1/2-mutant cancers
Authors: Marcel A.T.M. van Vugt, Eileen E. Parkes
Journal: Trends in cancer (2022)

Inflammatory signaling and genomic instability are hallmarks of cancer.Cancer-associated DNA repair defects, including defects in homologous recombination repair, lead to cytoplasmic DNA.Genomic instability fuels inflammatory signaling, triggering both tumor-suppressive as well as tumor-promoting traits.Through largely unknown mechanisms, tumor cells rewire inflammatory signaling to prevent immune clearance.Unique tumor microenvironment features of genomically unstable tumors may promote immunotherapy resistanceTherapeutic targeting of immune-suppressive mechanisms in genomically instable cancer may potentiate immune checkpoint inhibition. Genomic instability and inflammation are intricately connected hallmark features of cancer. DNA repair defects due to BRCA1/2 mutation instigate immune signaling through the cGAS/STING pathway. The subsequent inflammatory signaling provides both tumor-suppressive as well as tumor-promoting traits. To prevent clearance by the immune system, genomically instable cancer cells need to adapt to escape immune surveillance. Currently, it is unclear how genomically unstable cancers, including BRCA1/2-mutant tumors, are rewired to escape immune clearance. Here, we summarize the mechanisms by which genomic instability triggers inflammatory signaling and describe adaptive mechanisms by which cancer cells can 'fly under the radar' of the immune system. Additionally, we discuss how therapeutic activation of the immune system may improve treatment of genomically instable cancers. Genomic instability and inflammation are intricately connected hallmark features of cancer. DNA repair defects due to BRCA1/2 mutation instigate immune signaling through the cGAS/STING pathway. The subsequent inflammatory signaling provides both tumor-suppressive as well as tumor-promoting traits. To prevent clearance by the immune system, genomically instable cancer cells need to adapt to escape immune surveillance. Currently, it is unclear how genomically unstable cancers, including BRCA1/2-mutant tumors, are rewired to escape immune clearance. Here, we summarize the mechanisms by which genomic instability triggers inflammatory signaling and describe adaptive mechanisms by which cancer cells can 'fly under the radar' of the immune system. Additionally, we discuss how therapeutic activation of the immune system may improve treatment of genomically instable cancers. Genomic instability (see Glossary) is a trait of tumor cells that is observed in the majority of cancers. However, the level of genomic instability ranges significantly between tumors. High levels of genome instability are associated with hereditary or somatic mutations in DNA repair genes and oncogene-induced replication stress [1.Negrini S. et al.Genomic instability an evolving hallmark of cancer.Nat. Rev. Mol. Cell Biol. 2010; 11: 220-228Crossref PubMed Scopus (1327) Google Scholar]. In particular, defective repair of double-stranded DNA breaks (DSBs) and DNA crosslinks due to mutations in homologous recombination (HR) genes [e.g., Breast cancer 1, early onset (BRCA1) and Breast cancer 2, early onset (BRCA2)] or Fanconi anemia (FA) genes yields tumors with extensive genomic instability (Figure 1A ). These tumors are characterized by focal genomic deletions and amplifications as well as complex genomic rearrangements [2.Liu X. et al.Somatic loss of BRCA1 and p53 in mice induces mammary tumors with features of human BRCA1-mutated basal-like breast cancer.Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 12111-12116Crossref PubMed Scopus (327) Google Scholar,3.Davies H. et al.HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.Nat. Med. 2017; 23: 517-525Crossref PubMed Scopus (391) Google Scholar]. Notably, genomic instability drives intratumor heterogeneity and enables rapid acquisition of the genomic aberrations that drive therapy failure [4.Gerlinger M. Swanton C. How Darwinian models inform therapeutic failure initiated by clonal heterogeneity in cancer medicine.Br. J. Cancer. 2010; 103: 1139-1143Crossref PubMed Scopus (304) Google Scholar]. BRCA1 and BRCA2 are key regulators of DNA maintenance through HR [5.Wyman C. et al.Homologous recombination-mediated double-strand break repair.DNA Repair. 2004; 3: 827-833Crossref PubMed Scopus (0) Google Scholar]. BRCA1 functions in the initiation of HR by controlling DNA-end resection at DNA breaks, whereas BRCA2 functions downstream in HR, in loading the RAD51 recombinase to facilitate the actual recombination process (Figure 1B) [6.Chen C.-C. et al.Homology-directed repair and the role of BRCA1, BRCA2, and related proteins in genome integrity and cancer.Annu. Rev. Cancer Biol. 2018; 2: 313-336Crossref PubMed Google Scholar]. Besides these canonical roles, BRCA1 and BRCA2 function in DNA crosslink repair as part of the FA complex [7.Bunting S.F. et al.BRCA1 functions independently of homologous recombination in DNA interstrand crosslink repair.Mol. Cell. 2012; 46: 125-135Abstract Full Text Full Text PDF PubMed Scopus (201) Google Scholar] and were shown to protect nascent DNA at stalled replication forks from nucleolytic degradation (Figure 1B) [8.Schlacher K. et al.Double-strand break repair-independent role for BRCA2 in blocking stalled replication fork degradation by MRE11.Cell. 2011; 145: 529-542Abstract Full Text Full Text PDF PubMed Scopus (720) Google Scholar,9.Schlacher K. et al.A distinct replication fork protection pathway connects Fanconi anemia tumor suppressors to RAD51-BRCA1/2.Cancer Cell. 2012; 22: 106-116Abstract Full Text Full Text PDF PubMed Scopus (548) Google Scholar]. The relevance of BRCA1 and BRCA2 for genome maintenance and cellular viability became evident from genetic studies in mice. Specifically, loss of Brca1 or Brca2 leads to accumulation of DNA lesions, a consequent cell cycle arrest, and early embryonal death (Figure 1C) [10.Sharan S.K. et al.Embryonic lethality and radiation hypersensitivity mediated by Rad51 in mice lacking Brca2.Nature. 1997; 386: 804-810Crossref PubMed Google Scholar, 11.Hakem R. et al.The tumor suppressor gene Brca1 is required for embryonic cellular proliferation in the mouse.Cell. 1996; 85: 1009-1023Abstract Full Text Full Text PDF PubMed Scopus (563) Google Scholar, 12.Tutt A. et al.Absence of Brca2 causes genome instability by chromosome breakage and loss associated with centrosome amplification.Curr. Biol. 1999; 9: 1107-1110Abstract Full Text Full Text PDF PubMed Scopus (254) Google Scholar, 13.Hakem R. et al.Developmental studies of Brca1 and Brca2 knock-out mice.J. Mammary Gland Biol. Neoplasia. 1998; 3: 431-445Crossref PubMed Google Scholar, 14.Liu C.Y. et al.Inactivation of the mouse Brca1 gene leads to failure in the morphogenesis of the egg cylinder in early postimplantation development.Genes Dev. 1996; 10: 1835-1843Crossref PubMed Google Scholar, 15.Gowen L.C. et al.Brca1 deficiency results in early embryonic lethality characterized by neuroepithelial abnormalities.Nat. Genet. 1996; 12: 191-194Crossref PubMed Scopus (380) Google Scholar]. These findings were in apparent contrast with the observed tumor predisposition of BRCA mutation carriers and the full loss of BRCA1 or BRCA2 in the ensuing tumors (Figure 1A) [16.Elledge S.J. Amon A. The BRCA1 suppressor hypothesis: an explanation for the tissue-specific tumor development in BRCA1 patients.Cancer Cell. 2002; 1: 129-132Abstract Full Text Full Text PDF PubMed Scopus (81) Google Scholar]. This BRCA paradox was partly explained when Trp53 was conditionally inactivated along with Brca1/2, which allows cells to survive with damaged DNA and ultimately promotes tumor onset [2.Liu X. et al.Somatic loss of BRCA1 and p53 in mice induces mammary tumors with features of human BRCA1-mutated basal-like breast cancer.Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 12111-12116Crossref PubMed Scopus (327) Google Scholar,17.Jonkers J. et al.Synergistic tumor suppressor activity of BRCA2 and p53 in a conditional mouse model for breast cancer.Nat. Genet. 2001; 29: 418-425Crossref PubMed Scopus (764) Google Scholar,18.Xu X. et al.Conditional mutation of Brca 1 in mammary epithelial cells results in blunted ductal morphogenesis and tumour formation.Nat. Genet. 1999; 22: 37-43Crossref PubMed Scopus (0) Google Scholar]. The resulting tumors showed basal-like characteristics, including recurrent genomic features such as MYC amplification and RB1 loss [2.Liu X. et al.Somatic loss of BRCA1 and p53 in mice induces mammary tumors with features of human BRCA1-mutated basal-like breast cancer.Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 12111-12116Crossref PubMed Scopus (327) Google Scholar]. Moreover, these tumors displayed the complex genomic rearrangements that resemble those of human BRCA1/2-mutant cancers [2.Liu X. et al.Somatic loss of BRCA1 and p53 in mice induces mammary tumors with features of human BRCA1-mutated basal-like breast cancer.Proc. Natl. Acad. Sci. U. S. A. 2007; 104: 12111-12116Crossref PubMed Scopus (327) Google Scholar,19.Curtis C. et al.The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups.Nature. 2012; 486: 346-352Crossref PubMed Scopus (3261) Google Scholar, 20.Cancer Genome Atlas Network Comprehensive molecular portraits of human breast tumours.Nature. 2012; 490: 61-70Crossref PubMed Scopus (7479) Google Scholar, 21.Holstege H. et al.Cross-species comparison of aCGH data from mouse and human BRCA1- and BRCA2-mutated breast cancers.BMC Cancer. 2010; 10: 455Crossref PubMed Scopus (29) Google Scholar, 22.Hollern D.P. et al.A mouse model featuring tissue-specific deletion of p53 and Brca1 gives rise to mammary tumors with genomic and transcriptomic similarities to human basal-like breast cancer.Breast Cancer Res. Treat. 2019; 174: 143-155Crossref PubMed Scopus (6) Google Scholar]. Loss-of-function mutations in BRCA1/2, as well as mutations in other HR genes, lead to a profound defect in genome maintenance [23.Lord C.J. Ashworth A. BRCAness revisited.Nat. Rev. Cancer. 2016; 16: 110-120Crossref PubMed Scopus (623) Google Scholar], generically termed BRCAness. Defective HR yields aberrant genomes that are enriched for several mutational signatures, including base substitution signatures (SBS3, SBS8), indel signatures (ID6, ID8), and rearrangement signatures (RS1, RS3, and RS5) [3.Davies H. et al.HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.Nat. Med. 2017; 23: 517-525Crossref PubMed Scopus (391) Google Scholar,24.Alexandrov L.B. et al.The repertoire of mutational signatures in human cancer.Nature. 2020; 578: 94-101Crossref PubMed Scopus (613) Google Scholar, 25.Hwang T. et al.Defining the mutation signatures of DNA polymerase θ in cancer genomes.NAR Cancer. 2020; 2zcaa017Crossref PubMed Google Scholar, 26.Nguyen L. et al.Pan-cancer landscape of homologous recombination deficiency.Nat. Commun. 2020; 11: 5584Crossref PubMed Scopus (54) Google Scholar]. These genomic alterations can be explained by the usage of alternative, nonconservative DNA repair mechanisms to repair DNA DSBs (Figure 1C) [3.Davies H. et al.HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.Nat. Med. 2017; 23: 517-525Crossref PubMed Scopus (391) Google Scholar,27.Stok C. et al.Shaping the BRCAness mutational landscape by alternative double-strand break repair, replication stress and mitotic aberrancies.Nucleic Acids Res. 2021; 49: 4239-4257Crossref PubMed Scopus (3) Google Scholar]. In particular, the usage of non-homologous end-joining, polymerase Theta (POLQ)-mediated end-joining (also referred to as alternative end-joining), or single-strand annealing leads to deletions and translocations that are characteristic of HR-deficient cancers [3.Davies H. et al.HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures.Nat. Med. 2017; 23: 517-525Crossref PubMed Scopus (391) Google Scholar,24.Alexandrov L.B. et al.The repertoire of mutational signatures in human cancer.Nature. 2020; 578: 94-101Crossref PubMed Scopus (613) Google Scholar,25.Hwang T. et al.Defining the mutation signatures of DNA polymerase θ in cancer genomes.NAR Cancer. 2020; 2zcaa017Crossref PubMed Google Scholar,28.Nik-Zainal S. et al.Landscape of somatic mutations in 560 breast cancer whole-genome sequences.Nature. 2016; 534: 47-54Crossref PubMed Scopus (1083) Google Scholar,29.Póti Á. et al.Correlation of homologous recombination deficiency induced mutational signatures with sensitivity to PARP inhibitors and cytotoxic agents.Genome Biol. 2019; 20: 240Crossref PubMed Scopus (7) Google Scholar]. In addition to the effects of defective DSB repair, genome integrity in BRCA1/2-mutant cancers is affected by defective protection of stalled replication forks [8.Schlacher K. et al.Double-strand break repair-independent role for BRCA2 in blocking stalled replication fork degradation by MRE11.Cell. 2011; 145: 529-542Abstract Full Text Full Text PDF PubMed Scopus (720) Google Scholar]. This defect is independent of the HR defect [30.Daza-Martin M. et al.Isomerization of BRCA1–BARD1 promotes replication fork protection.Nature. 2019; 571: 521-527Crossref PubMed Scopus (41) Google Scholar] and leads to fork collapse and persistent DNA breaks. Specific to BRCA1-mutant cells, re-replication of tracts of DNA adjacent to stalled forks leads to tandem duplications [31.Willis N.A. et al.Mechanism of tandem duplication formation in BRCA1-mutant cells.Nature. 2017; 551: 590-595Crossref PubMed Scopus (57) Google Scholar], which are a recurrent feature of BRCA1-mutant cancers [28.Nik-Zainal S. et al.Landscape of somatic mutations in 560 breast cancer whole-genome sequences.Nature. 2016; 534: 47-54Crossref PubMed Scopus (1083) Google Scholar,32.Menghi F. et al.The tandem duplicator phenotype is a prevalent genome-wide cancer configuration driven by distinct gene mutations.Cancer Cell. 2018; 34: 197-210Abstract Full Text Full Text PDF PubMed Scopus (68) Google Scholar,33.Menghi F. Liu E.T. Reply to Watkins et al.: Whole-genome sequencing-based identification of diverse tandem duplicator phenotypes in human cancers.Proc. Natl. Acad. Sci. U. S. A. 2016; 113: E5259-E5260Crossref PubMed Scopus (0) Google Scholar]. The effects of BRCA1/2 mutation on genome integrity will likely extend beyond the effects of impaired HR and fork protection, as BRCA1 and BRCA2 (and multiple other HR genes) have been described to perform multiple other functions in tumor suppression, including roles for BRCA1 in regulating gene expression and chromatin remodeling [34.Savage K.I. et al.Identification of a BRCA1-mRNA splicing complex required for efficient DNA repair and maintenance of genomic stability.Mol. Cell. 2014; 54: 445-459Abstract Full Text Full Text PDF PubMed Scopus (110) Google Scholar, 35.Filipponi D. et al.Wip1 controls global heterochromatin silencing via ATM/BRCA1-dependent DNA methylation.Cancer Cell. 2013; 24: 528-541Abstract Full Text Full Text PDF PubMed Scopus (45) Google Scholar, 36.Zhu Q. et al.BRCA1 tumour suppression occurs via heterochromatin-mediated silencing.Nature. 2011; 477: 179-184Crossref PubMed Scopus (310) Google Scholar]. Recently, inflammatory signaling was identified as a consequence of genomic instability, including in BRCA1/2-mutant cancers. Tumor-cell intrinsic inflammatory signaling can activate immune cells in the tumor microenvironment (TME) and has been linked to responses to immune checkpoint inhibitors. Although important features to respond to cancer immunotherapy are clearly present in genomically instable cancers, patients with such cancers paradoxically only minimally benefit from immune checkpoint inhibitors. Apparently, genomically instable cancers somehow evade immune eradication. To expand benefit of cancer immunotherapy to patients with genomically instable cancers, it is crucial to understand how these cancers have adapted to inflammatory signaling and to investigate if and how these adaptive mechanisms can be therapeutically targeted. We review the mechanisms that drive inflammatory signaling in response to genomic instability caused by BRCA1/2 inactivation. We also discuss alterations in these tumors to evade immune clearance and options to improve treatment of these cancers. In line with their DNA repair defects and consequent tumor mutational burden (TMB), BRCA1/2 mutation status in high-grade serous ovarian cancer associates with a predicted higher neoantigen load and improved overall survival [37.Strickland K.C. et al.Association and prognostic significance of BRCA1/2-mutation status with neoantigen load, number of tumor-infiltrating lymphocytes and expression of PD-1/PD-L1 in high grade serous ovarian cancer.Oncotarget. 2016; 7: 13587-13598Crossref PubMed Scopus (345) Google Scholar]. In this study, neoantigen load correlates with HR deficiency, even in non-BRCA mutations. Similarly, BRCA1/2 mutation in breast cancer associates with increased TMB and increased expression of the PD1/PDL-1 immune checkpoint components, albeit only in BRCA1-mutant cancers [38.Wen W.X. Leong C.-O. Association of BRCA1- and BRCA2-deficiency with mutation burden, expression of PD-L1/PD-1, immune infiltrates, and T cell-inflamed signature in breast cancer.PLoS One. 2019; 14e0215381Crossref PubMed Scopus (7) Google Scholar]. Even so, this increased TMB does not reach the scale of that observed in tumors classically responsive to immune checkpoint blockade: microsatellite unstable colorectal cancer, melanoma, and non-small cell lung cancer [39.Lawrence M.S. et al.Mutational heterogeneity in cancer and the search for new cancer-associated genes.Nature. 2013; 499: 214-218Crossref PubMed Scopus (3303) Google Scholar]. It is likely that the increased immunogenicity of BRCA1/2-mutant cancers cannot solely be attributed to a moderately elevated TMB, as HR mutations are predictors of response to immune checkpoint inhibitors, independent of TMB [40.Hsiehchen D. et al.DNA repair gene mutations as predictors of immune checkpoint inhibitor response beyond tumor mutation burden.Cell Reports Med. 2020; 1100034Abstract Full Text Full Text PDF PubMed Google Scholar]. Beyond mutational burden, inflammatory signaling appears to strongly associate with immunogenicity, likely in part because it is a direct read-out of immune cell activation. BRCA1/2 mutation in ovarian cancer associates with expression of immune-related genes, including type II interferon gamma (IFNγ) and TNFR [37.Strickland K.C. et al.Association and prognostic significance of BRCA1/2-mutation status with neoantigen load, number of tumor-infiltrating lymphocytes and expression of PD-1/PD-L1 in high grade serous ovarian cancer.Oncotarget. 2016; 7: 13587-13598Crossref PubMed Scopus (345) Google Scholar]. BRCA1 mutation is associated with the immunoreactive subtype of high-grade serous ovarian cancers, which are characterized by high levels of tumor-infiltrating immune cells [41.George J. et al.Nonequivalent gene expression and copy number alterations in high-grade serous ovarian cancers with BRCA1 and BRCA2 mutations.Clin. Cancer Res. 2013; 19: 3474-3484Crossref PubMed Scopus (64) Google Scholar]. Also, BRCA1/2 status is associated with increased abundance of CD3+ and CD4+ tumor-infiltrating lymphocytes [42.McAlpine J.N. et al.BRCA1 and BRCA2 mutations correlate with TP53 abnormalities and presence of immune cell infiltrates in ovarian high-grade serous carcinoma.Mod. Pathol. 2012; 25: 740-750Crossref PubMed Scopus (118) Google Scholar] and increased expression of the immune checkpoint molecules PD1/PDL1 [37.Strickland K.C. et al.Association and prognostic significance of BRCA1/2-mutation status with neoantigen load, number of tumor-infiltrating lymphocytes and expression of PD-1/PD-L1 in high grade serous ovarian cancer.Oncotarget. 2016; 7: 13587-13598Crossref PubMed Scopus (345) Google Scholar,43.Wieser V. et al.BRCA1/2 and TP53 mutation status associates with PD-1 and PD-L1 expression in ovarian cancer.Oncotarget. 2018; 9: 17501-17511Crossref PubMed Scopus (52) Google Scholar]. In a separate study only focusing on BRCA1 status, increased levels of DNA damage in BRCA1-mutant ovarian cancers were associated with elevated stimulator of interferon genes (STING) levels, increased STAT1 signaling, and increased T cell infiltrate [44.Bruand M. et al.Cell-autonomous inflammation of BRCA1-deficient ovarian cancers drives both tumor-intrinsic immunoreactivity and immune resistance via STING.Cell Rep. 2021; 36109412Abstract Full Text Full Text PDF PubMed Scopus (5) Google Scholar]. Moreover, the tumor-associated inflammation in BRCA1-mutant tumors is a favorable prognostic feature [45.van Verschuer V.M.T. et al.Tumor-associated inflammation as a potential prognostic tool in BRCA1/2-associated breast cancer.Hum. Pathol. 2015; 46: 182-190Crossref PubMed Google Scholar,46.de Boo L. et al.Tumour-infiltrating lymphocytes (TILs) and BRCA-like status in stage III breast cancer patients randomised to adjuvant intensified platinum-based chemotherapy versus conventional chemotherapy.Eur. J. Cancer. 2020; 127: 240-250Abstract Full Text Full Text PDF PubMed Scopus (9) Google Scholar]. Analysis of gene expression in patient samples with BRCA1/2 or FA mutations yielded a gene set signature, which was highly enriched for genes involved in immune signaling [47.Mulligan J.M. et al.Identification and validation of an anthracycline/cyclophosphamide-based chemotherapy response assay in breast cancer.J. Natl. Cancer Inst. 2014; 106: djt335Crossref PubMed Scopus (74) Google Scholar]. Lymphocyte infiltration only partially explains the expression of immune-related genes, pointing towards cell intrinsic inflammatory signaling [47.Mulligan J.M. et al.Identification and validation of an anthracycline/cyclophosphamide-based chemotherapy response assay in breast cancer.J. Natl. Cancer Inst. 2014; 106: djt335Crossref PubMed Scopus (74) Google Scholar]. Subsequent studies have revealed that inactivation of BRCA1, BRCA2, or FANCD2 results in tumor-cell intrinsic inflammatory signaling, involving secretion of proinflammatory cytokines CXCL10, CCL5, and TNF-α and attraction of immune cells [48.Parkes E.E. et al.Activation of STING-dependent innate immune signaling by S-phase-specific DNA damage in breast cancer.J. Natl. Cancer Inst. 2017; 109djw199Crossref PubMed Scopus (197) Google Scholar,49.Heijink A.M. et al.BRCA2 deficiency instigates cGAS-mediated inflammatory signaling and confers sensitivity to tumor necrosis factor-alpha-mediated cytotoxicity.Nat. Commun. 2019; 10: 100Crossref PubMed Scopus (42) Google Scholar]. Inflammatory signaling in these cells likely stems from the DNA lesions that arise due to their defective DNA maintenance, as similar effects are observed in response to DNA damaging agents [48.Parkes E.E. et al.Activation of STING-dependent innate immune signaling by S-phase-specific DNA damage in breast cancer.J. Natl. Cancer Inst. 2017; 109djw199Crossref PubMed Scopus (197) Google Scholar,50.Sistigu A. et al.Cancer cell–autonomous contribution of type I interferon signaling to the efficacy of chemotherapy.Nat. Med. 2014; 20: 1301-1309Crossref PubMed Scopus (576) Google Scholar,51.Legrier M.E. et al.Activation of IFN/STAT1 signalling predicts response to chemotherapy in oestrogen receptor-negative breast cancer.Br. J. Cancer. 2016; 114: 177-187Crossref PubMed Google Scholar]. These findings are in line with observations in genetically engineered mouse models. Combined tissue-specific loss of Trp53 and Brca1 leads to development of cancers with signatures associated with higher levels of Th2 cells, T-regulatory cells (Tregs), central memory cells, and exhausted T cells, as well as elevated expression of immune checkpoint genes, Pd1 and Ctla4 [22.Hollern D.P. et al.A mouse model featuring tissue-specific deletion of p53 and Brca1 gives rise to mammary tumors with genomic and transcriptomic similarities to human basal-like breast cancer.Breast Cancer Res. Treat. 2019; 174: 143-155Crossref PubMed Scopus (6) Google Scholar]. Whereas BRCA1/2 mutation and other HR deficiencies are all characterized by severe genomic instability, not all HR deficiencies are equal in terms of subsequent effects on the TME. For instance, gene expression analysis revealed that ovarian cancers with mutant or hypermethylated BRCA1 but not BRCA2-mutant cancers are associated with the immunoreactive subtype [41.George J. et al.Nonequivalent gene expression and copy number alterations in high-grade serous ovarian cancers with BRCA1 and BRCA2 mutations.Clin. Cancer Res. 2013; 19: 3474-3484Crossref PubMed Scopus (64) Google Scholar]. In a study of BRCA-mutant breast cancers, distinct features of the immune TME of BRCA1- versus BRCA2-mutant were identified, with increased immunosuppressive tumor-associated macrophages in Brca1-null mouse models and increased immune checkpoint expression in BRCA1-mutant breast cancer [52.Samstein R.M. et al.Mutations in BRCA1 and BRCA2 differentially affect the tumor microenvironment and response to checkpoint blockade immunotherapy.Nat. Cancer. 2020; 1: 1188-1203Crossref PubMed Scopus (34) Google Scholar]. A pan-cancer study of BRCA1- versus BRCA2-mutant cancers treated with immune checkpoint blockade suggests benefit in BRCA2-mutant disease only [53.Zhou Z. Li M. Evaluation of BRCA1 and BRCA2 as indicators of response to immune checkpoint inhibitors.JAMA Netw. Open. 2021; 4e217728Crossref PubMed Scopus (0) Google Scholar]. Whether similar patterns could be observed in response to other treatments such as chemotherapy is not known. Some studies have instead observed BRCA1/2-mutant cancers to be poorly immunogenic compared with BRCA1/2-wild type. Analysis of breast cancers in patients with a germline BRCA mutation or clonal loss of heterozygosity found these tumors to have a lower immune gene expression score compared with BRCA1/2-wild type or tumors with subclonal loss of heterozygosity, correlating with reduced immune infiltration on immunohistochemical analysis of germline BRCA1-mutant breast cancers [54.Kraya A.A. et al.Genomic signatures predict the immunogenicity of BRCA-deficient breast cancer.Clin. Cancer Res. 2019; 25: 4363-4374Crossref PubMed Scopus (33) Google Scholar]. These observations add granularity to the association of BRCA1/2 mutation and immune infiltration, indicating that detailed molecular analysis is needed to clearly understand this complex relationship. Moreover, intratumoral heterogeneity plays a role in enabling immune evasion in BRCA-mutant cancers. Using digital pathology to define morphological diversity in ovarian cancer, a significant association between low BRCA1 expression and increased diversification was noted [55.Heindl A. et al.Microenvironmental niche divergence shapes BRCA1-dysregulated ovarian cancer morphological plasticity.Nat. Commun. 2018; 9: 3917Crossref PubMed Scopus (23) Google Scholar]. Further morphological examination revealed a lack of tumor invasion by CD3+ T cells in morphologically diverse regions, with a suggestion of active exclusion of these cells from tumor nests by upregulation of the immune checkpoint galectin-3 [55.Heindl A. et al.Microenvironmental niche divergence shapes BRCA1-dysregulated ovarian cancer morphological plasticity.Nat. Commun. 2018; 9: 3917Crossref PubMed Scopus (23) Google Scholar]. The relationship between BRCA1/2 mutation and a favorable immune environment is far from straightforward, with genetic and spatial heterogeneity contributing to a spectrum of immunogenicity in genomically unstable cancers. BRCA1/2-mutant cancers almost invariably harbor inactivating TP53 mutations [19.Curtis C. et al.The genomic and transcriptomic architecture of 2,000 breast tumours reveals novel subgroups.Nature. 2012; 486: 346-352Crossref PubMed Scopus (3261) Google Scholar], therefore, these cells have incomplete cell cycle control and frequently transmit DNA damage into mitosis [56.Schoonen P.M. et al.Progression through mitosis promotes PARP inhibitor-induced cytotoxicity in homologous recombination-deficient cancer cells.Nat. Commun. 2017; 8: 15981Crossref PubMed Scopus (54) Google Scholar,57.Feng W. Jasin M. BRCA2 suppresses replication stress-induced mitotic and G1 abnormalities through homologous recombination.Nat. Commun. 2017; 8: 525Crossref PubMed Scopus (79) Google Scholar]. These mitotic DNA lesions are positive for the replication stress marker FANCD2 [56.Schoonen P.M. et al.Progression through mitosis promotes PARP inhibitor-induced cytotoxicity in homologous recombination-deficient cancer cells.Nat. Commun. 2017; 8: 15981Crossref PubMed Scopus (54) Google Scholar] and likely represent 'joint DNA molecules', either due to incomplete DNA replication or unresolved intermediates of repair attempts. Consequently, BRCA1/2 inactivation leads to elevated numbers of mitotic chromatin bridges and ultrafine DNA bridges [56.Schoonen P.M. et al.Progression through mitosis promotes PARP inhibitor-induced cytotoxicity in homologous recombination-deficient cancer cells.Nat. Commun. 2017; 8: 15981Crossref PubMed Scopus (54) Google Scholar,58.Laulier C. et al.The relative efficiency of homology-directed repair has distinct effects on proper anaphase chromosome separation.Nucleic Acids Res. 2011; 39: 5935-5944Crossref PubMed Scopus (38) Google Scholar] and results in 53BP1 nuclear bodies in the subsequent G1 cells [57.Feng W. Jasin M. BRCA2 suppresses replication stress-induced mitotic and G1 abnormalities through homologous recombination.Nat. Commun. 2017; 8: 525Crossref PubMed Scopus (79) Google Scholar]. Unresolved mitotic DNA damage frequently results in micronuclei [59.Löbrich M. Jeggo P.A. The impact of a negligent G2/M checkpoint on genomic instability and cancer induction.Nat. Rev. Cancer. 2007; 7: 861-869Crossref PubMed Scopus (431) Google Scholar,60.Lewis C.W. Golsteyn R.M. Cancer cells that survive checkpoint adaptation contain micronuclei that ha