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The DNA mismatch repair (MMR) proteins recognize and repair DNA base pair mismatches and insertions/deletions of DNA that have occurred during DNA replication. Additionally, they are involved in regulation of the DNA damage response, including cell cycle checkpoints and apoptosis. Therefore, regulation of these proteins is essential for maintaining genomic integrity. It has been recognized that post-translational modifications, such as phosphorylation, ubiquitination, and acetylation, are being used as an important means to regulate the functions and stability of MMR proteins. Here, we report that a histone acetyltransferase CREB binding protein (CBP) interacts with and acetylates MLH1, a component of the MutLα complex (MLH1–PMS2). Moreover, CBP stabilizes MLH1 by preventing it from degradation via the ubiquitin–proteasome degradation pathway. Consistently, acetylation induced by a pan-histone deacetylase inhibitor, Trichostatin A, promotes the assembly between the MutSα (MSH2–MSH6) and MutLα complexes. Furthermore, overexpression of CBP enhances MMR activities in cells. Overall, our results suggest a novel role of CBP in prolonging MLH1 stability and enhancing MutSα–MutLα complex formation, leading to increased cellular MMR activity.

Additional Characterization of PTEN K128R

Legends for all 4 supplemental figures

Resistance to cancer chemotherapies leads to deadly consequences, yet current research focuses only on the roles of somatically acquired mutations in this resistance. The mutational status of the germline is also likely to play a role in the way cells respond to chemotherapy. The carrier status for the POLB rs3136797 germline mutation encoding P242R DNA polymerase beta (Pol β) is associated with poor prognosis for lung cancer, specifically in response to treatment with cisplatin. Here, it is revealed that the P242R mutation is sufficient to promote resistance to cisplatin in human cells and in mouse xenografts. Mechanistically, P242R Pol β acts as a translesion polymerase and prefers to insert the correct nucleotide opposite cisplatin intrastrand cross-links, leading to the activation of the nucleotide excision repair (NER) pathway, removal of crosslinks, and resistance to cisplatin. In contrast, wild-type (WT) Pol β preferentially inserts the incorrect nucleotide initiating mismatch repair and cell death. Importantly, in a mouse xenograft model, tumors derived from lung cancer cells expressing WT Pol β displayed a slower rate of growth when treated with cisplatin, whereas tumors expressing P242R Pol β had no response to cisplatin. Pol β is critical for mediating crosstalk in response to cisplatin. The current data strongly suggest that the status of Pol β influences cellular responses to crosslinking agents and that Pol β is a promising biomarker to predict responses to specific chemotherapies. Finally, these results highlight that the genetic status of the germline is a critical factor in the response to cancer treatment.Implications: Pol β has prognostic biomarker potential in the treatment of cancer with cisplatin and perhaps other intrastrand crosslinking agents. Mol Cancer Res; 15(3); 269–80. ©2017 AACR.

The development of therapeutic agents that specifically target cancer cells while sparing healthy tissue could be used to enhance the efficacy of cancer therapy without increasing its toxicity. Specific targeting of cancer cells can be achieved through the use of pH-low insertion peptides (pHLIP), which take advantage of the acidity of the tumor microenvironment to deliver cargoes selectively to tumor cells. We developed a pHLIP–peptide nucleic acid (PNA) conjugate as an antisense reagent to reduce expression of the otherwise undruggable DNA double-strand break repair factor, KU80, and thereby radiosensitize tumor cells. Increased antisense activity of the pHLIP–PNA conjugate was achieved by partial mini-PEG sidechain substitution of the PNA at the gamma position, designated pHLIP-αKu80(γ). We evaluated selective effects of pHLIP-αKu80(γ) in cancer cells in acidic culture conditions as well as in two subcutaneous mouse tumor models. Fluorescently labeled pHLIP-αKu80(γ) delivers specifically to acidic cancer cells and accumulates preferentially in tumors when injected i.v. in mice. Furthermore, pHLIP-αKu80(γ) selectively reduced KU80 expression in cells under acidic conditions and in tumors in vivo. When pHLIP-αKu80(γ) was administered to mice prior to local tumor irradiation, tumor growth was substantially reduced compared with radiation treatment alone. Furthermore, there was no evidence of acute toxicity associated with pHLIP-αKu80(γ) administration to the mice. These results establish pHLIP-αKu80(γ) as a tumor-selective radiosensitizing agent.Implications:This study describes a novel agent, pHLIP-αKu80(γ), which combines PNA antisense and pHLIP technologies to selectively reduce the expression of the DNA repair factor KU80 in tumors and confer tumor-selective radiosensitization.

Hypoxia induces genomic instability through replication stress and dysregulation of vital DNA repair pathways. The Fanconi anemia (FA) proteins, FANCD2 and FANCI, are key members of a DNA repair pathway that responds to replicative stress, suggesting that they undergo regulation by hypoxic conditions. Here acute hypoxic stress activates the FA pathway via ubiquitination of FANCD2 and FANCI in an ATR-dependent manner. In addition, the presence of an intact FA pathway is required for preventing hypoxia-induced DNA damage measurable by the comet assay, limiting the accumulation of γH2AX (a marker of DNA damage or stalled replication), and protecting cells from hypoxia-induced apoptosis. Furthermore, prolonged hypoxia induces transcriptional repression of FANCD2 in a manner analogous to the hypoxic downregulation of BRCA1 and RAD51. Thus, hypoxia-induced FA pathway activation plays a key role in maintaining genome integrity and cell survival, while FA protein downregulation with prolonged hypoxia contributes to genomic instability.Implications: This work highlights the critical role of the FA pathway in response to hypoxic stress and identifies the pathway as a therapeutic target under hypoxic conditions. Mol Cancer Res; 12(7); 1016–28. ©2014 AACR.

Fig. S1: pHLIP-αKu80(γ) binds to the mouse Ku80 sequence. Fig. S2: Validation of tumor vs. stromal cell sorting in EMT6-GFP cells. Fig. S3: Partial suppression of Ku80 expression induces radiosensitivity in cultured cancer cells. Fig. S4: pHLIP-αKu80(γ) causes tumor radiosensitization. Fig. S5: pHLIP-αKu80(γ) causes in vitro radiosensitization. Fig. S6: pHLIP-αKu80(γ) does not induce electrolyte abnormalities. Fig. S7: pHLIP-αKu80(γ) does not cause bone marrow suppression.

TH-302 is a 2-nitroimidazole triggered hypoxia-activated prodrug (HAP) of bromo-isophosphoramide mustard currently undergoing clinical evaluation. Here, we describe broad-spectrum activity, hypoxia-selective activation, and mechanism of action of TH-302. The concentration and time dependence of TH-302 activation was examined as a function of oxygen concentration, with reference to the prototypic HAP tirapazamine, and showed superior oxygen inhibition of cytotoxicity and much improved dose potency relative to tirapazamine. Enhanced TH-302 cytotoxicity under hypoxia was observed across 32 human cancer cell lines. One-electron reductive enzyme dependence was confirmed using cells overexpressing human NADPH:cytochrome P450 oxidoreductase and radiolytic reduction established the single-electron stoichiometry of TH-302 fragmentation (activation). Examining downstream effects of TH-302 activity, we observed hypoxia-dependent induction of γH2AX phosphorylation, DNA cross-linking, and cell-cycle arrest. We used Chinese hamster ovary cell–based DNA repair mutant cell lines and established that lines deficient in homology-dependent repair, but not lines deficient in base excision, nucleotide excision, or nonhomologous end-joining repair, exhibited marked sensitivity to TH-302 under hypoxia. Consistent with this finding, enhanced sensitivity to TH-302 was also observed in lines deficient in BRCA1, BRCA2, and FANCA. Finally, we characterized TH-302 activity in the three-dimensional tumor spheroid and multicellular layer models. TH-302 showed much enhanced potency in H460 spheroids compared with H460 monolayer cells under normoxia. Multicellular layers composed of mixtures of parental HCT116 cells and HCT116 cells engineered to express an oxygen-insensitive bacterial nitroreductase showed that TH-302 exhibits a significant bystander effect. Mol Cancer Ther; 11(3); 740–51. ©2011 AACR.

Validation of DSB repair assays.

NHEJ activity is unchanged upon FOXO3a knockdown in AV16.

SFig 1: SAHA reduces H3K9 trimethylation in IDH1 mutant cells while increasing H3K9 and H4 acetylation, SFig 2: Characterization of U2OS EJDR cells after SAHA, metabolite and siRNA treatment, SFig 3: SAHA elevates DNA damage to a greater degree in IDH1 mutant cells, comparable to BRCA1 and RAD51 siRNA knockdown, SFig 4: SAHA represses RAD51 and increases γ-H2AX foci formation, SFig 5: Mutant IDH1 sensitivity to PARPi treatment, SFig 6: Synergy of SAHA combination with radiation, TMZ and PARPi, SFig 7: SAHA down-regulates HDR factor expression, elevates DNA damage and has greater effect alone and plus PARPi against IDH1 mutant HCT116 cells

S1. Dose-response curve of MCF7 cells. S2. Western blotting analysis of Pol beta in cell lines. S3. Pol beta and XPA interact in vitro. S4. The effect of E295K Pol beta expression on survival following cisplatin treatment. Table S1. Difference in viability of crosslinking agents from high-throughput screen.

PDF file - 1436KB, S1. FANCD2 is ubiquitinated and transcriptionally down-regulated upon treatment with DFX in MCF7 cells. S2. FANCD2 is ubiquitinated and transcriptionally down-regulated upon treatment with DFX in A549 cells. S3. FANCD2 and FANCI ubiquitination does not occur at less severe levels of hypoxia. S4. siRNA depletion of ATR, but not ATM, inhibits FANCD2 and FANCI ubiquitination induced by hypoxia and DFX. S5. FANCD2 and FANCI ubiquitination upon DFX treatment is independent of HIF-1. S6. FANCD2 protein down-regulation by DFX is not due to altered protein stability and transcriptional down-regulation may be related to p130 dephosphorylation and HIF-2. S7. FANCD2 protein level decreases in hypoxia and is restored by HPV16-E7 overexpression. S8. FANCI transcriptional down-regulation upon prolonged exposure to DFX or hypoxia is mitigated by HPV16-E7 overexpression. S9. Increased chromosomal aberrations induced by hypoxia in the absence of FANCD2 are not detected by cytogenetic analysis.

Mutation pattern remains unchanged upon miR-155 over-expression in vivo.

Confirmation of polymerase delta mRNA down-regulation.

Development of resistance remains the key obstacle to the clinical efficacy of EGFR tyrosine kinase inhibitors (TKI). Hypoxia is a key microenvironmental stress in solid tumors associated with acquired resistance to conventional therapy. Consistent with our previous studies, we show here that long-term, moderate hypoxia promotes resistance to the EGFR TKI osimertinib (AZD9291) in the non–small cell lung cancer (NSCLC) cell line H1975, which harbors two EGFR mutations including T790M. Hypoxia-induced resistance was associated with development of epithelial–mesenchymal transition (EMT) coordinated by increased expression of ZEB-1, an EMT activator. Hypoxia induced increased fibroblast growth factor receptor 1 (FGFR1) expression in NSCLC cell lines H1975, HCC827, and YLR086, and knockdown of FGFR1 attenuated hypoxia-induced EGFR TKI resistance in each line. Upregulated expression of FGFR1 by hypoxia was mediated through the MAPK pathway and attenuated induction of the proapoptotic factor BIM. Consistent with this, inhibition of FGFR1 function by the selective small-molecule inhibitor BGJ398 enhanced EGFR TKI sensitivity and promoted upregulation of BIM levels. Furthermore, inhibition of MEK activity by trametinib showed similar effects. In tumor xenografts in mice, treatment with either BGJ398 or trametinib enhanced response to AZD9291 and improved survival. These results suggest that hypoxia is a driving force for acquired resistance to EGFR TKIs through increased expression of FGFR1. The combination of EGFR TKI and FGFR1 or MEK inhibitors may offer an attractive therapeutic strategy for NSCLC.Significance:Hypoxia-induced resistance to EGFR TKI is driven by overexpression of FGFR1 to sustain ERK signaling, where a subsequent combination of EGFR TKI with FGFR1 inhibitors or MEK inhibitors reverses this resistance.

H2O2 induces reversible oxidation of classical PTPs.

Supplementary figures 1 to 10 and figure legends.

Supplementary materials, methods, and figure legends.

Hereditary leiomyomatosis and renal cell carcinoma (HLRCC) is an inherited cancer syndrome associated with a highly aggressive form of type 2 papillary renal cell carcinoma (PRCC). Germline inactivating alterations in fumarate hydratase (FH) cause HLRCC and result in elevated levels of reactive oxygen species (ROS). Recent work indicates that FH−/− PRCC cells have increased activation of ABL1, which promotes tumor growth, but how ABL1 is activated remains unclear. Given that oxidation can regulate protein-tyrosine phosphatase (PTP) catalytic activity, inactivation of an ABL-directed PTP by ROS might account for ABL1 activation in this malignancy. Our group previously developed “q-oxPTPome,” a method that globally monitors the oxidation of classical PTPs. In this study, we present a refined q-oxPTPome, increasing its sensitivity by >10×. Applying q-oxPTPome to FH-deficient cell models showed that multiple PTPs were either highly oxidized (including PTPN12) or overexpressed. Highly oxidized PTPs were those with relatively high sensitivity to exogenous H2O2. Most PTP oxidation in FH-deficient cells was reversible, although nearly 40% of PTPN13 was irreversibly oxidized to the sulfonic acid state. Using substrate-trapping mutants, we mapped PTPs to their putative substrates and found that only PTPN12 could target ABL1. Furthermore, knockdown experiments identified PTPN12 as the major ABL1 phosphatase, and overexpression of PTPN12 inhibited ABL1 phosphorylation and HLRCC cell growth. These results show that ROS-induced oxidation of PTPN12 accounts for ABL1 phosphorylation in HLRCC-associated PRCC, revealing a novel mechanism for inactivating a tumor suppressor gene product and establishing a direct link between pathologic PTP oxidation and neoplastic disease.Significance:This work identifies a novel mechanism of activation of the oncogenic kinase ABL1 via ROS-induced, oxidation-mediated inactivation of cognate protein tyrosine phosphatases.

Genetic instability is a hallmark of cancer; the hypoxic tumor microenvironment has been implicated as a cause of this phenomenon. MicroRNAs (miR) are small nonprotein coding RNAs that can regulate various cellular pathways. We report here that two miRs, miR-210 and miR-373, are up-regulated in a hypoxia-inducible factor-1α–dependent manner in hypoxic cells. Bioinformatics analyses suggested that these miRs could regulate factors implicated in DNA repair pathways. Forced expression of miR-210 was found to suppress the levels of RAD52, which is a key factor in homology-dependent repair (HDR); the forced expression of miR-373 led to a reduction in the nucleotide excision repair (NER) protein, RAD23B, as well as in RAD52. Consistent with these results, both RAD52 and RAD23B were found to be down-regulated in hypoxia, but in both cases, the hypoxia-induced down-regulation could be partially reversed by antisense inhibition of miR-210 and miR-373. Importantly, luciferase reporter assays indicated that miR-210 is capable of interacting with the 3′ untranslated region (UTR) of RAD52 and that miR-373 can act on the 3′ UTR of RAD23B. These results indicate that hypoxia-inducible miR-210 and miR-373 play roles in modulating the expression levels of key proteins involved in the HDR and NER pathways, providing new mechanistic insight into the effect of hypoxia on DNA repair and genetic instability in cancer. [Cancer Res 2009;69(3):1221–9]

Decreased BRCA1 expression in the absence of genetic mutation is observed frequently in sporadic cancers of the breast and other sites, although little is known regarding the mechanisms by which the expression of this gene can be repressed. Here, we show that activating and repressive E2Fs simultaneously bind the BRCA1 promoter at two adjacent E2F sites in vivo, and that hypoxia induces a dynamic redistribution of promoter occupancy by these factors resulting in the transcriptional repression of BRCA1 expression. Functionally, we show that hypoxia is associated with impaired homologous recombination, whereas the nonhomologous end-joining (NHEJ) repair pathway is unaffected under these conditions. Repression of BRCA1 expression by hypoxia represents an intriguing mechanism of functional BRCA1 inactivation in the absence of genetic mutation. We propose that hypoxia-induced decreases in BRCA1 expression and consequent suppression of homologous recombination may lead to genetic instability by shifting the balance between the high-fidelity homologous recombination pathway and the error-prone NHEJ pathway of DNA repair. Furthermore, these findings provide a novel link between E2Fs and the transcriptional response to hypoxia and provide insight into the mechanisms by which the tumor microenvironment can contribute to genetic instability in cancer. (Cancer Res 2005; 65(24): 11597-604)

Supplementary Figure 1 from Hypoxia-Induced Down-regulation of BRCA1 Expression by E2Fs

Supplementary Figure Legend from MicroRNA Regulation of DNA Repair Gene Expression in Hypoxic Stress

Supplementary Figure 3 from Hypoxia-Induced Down-regulation of BRCA1 Expression by E2Fs

Supplementary Figure 1 from MicroRNA Regulation of DNA Repair Gene Expression in Hypoxic Stress

Supplementary Information from Hypoxia-Induced Down-regulation of BRCA1 Expression by E2Fs

Supplementary Figure 2 from Hypoxia-Induced Down-regulation of BRCA1 Expression by E2Fs

PURPOSE Cediranib, a pan-vascular endothelial growth factor receptor inhibitor, suppresses expression of homologous recombination repair (HRR) genes and increases sensitivity to poly-(ADP-ribose) polymerase inhibition in preclinical models. We investigated whether cediranib combined with olaparib improves the clinical outcomes of patients with prostate cancer. METHODS Patients with progressive metastatic castration-resistant prostate cancer (mCRPC) were randomly assigned 1:1 to arm A: cediranib 30 mg once daily plus olaparib 200 mg twice daily or arm B: olaparib 300 mg twice daily alone. The primary end point was radiographic progression-free survival (rPFS) in the intention-to-treat patients. The secondary end points were rPFS in patients with HRR-deficient and HRR-proficient mCRPC. RESULTS In the intention-to-treat set of 90 patients, median rPFS was 8.5 (95% CI, 5.4 to 12.0) and 4.0 (95% CI, 3.2 to 8.5) months in arms A and B, respectively. Cediranib/olaparib significantly improved rPFS versus olaparib alone (hazard ratio [HR], 0.617; 95% CI, 0.392 to 0.969; P = .0359). Descriptive analyses showed a median rPFS of 10.6 (95% CI, 5.9 to not assessed [NA]) and 3.8 (95% CI, 2.33 to NA) months (HR, 0.64; 95% CI, 0.272 to 1.504) among patients with HRR-deficient mCRPC, and 13.8 (95% CI, 3.3 to NA) and 11.3 (95% CI, 3.8 to NA) months (HR, 0.98; 95% CI, 0.321 to 2.988) among patients with BRCA2-mutated mCRPC in arms A and B, respectively. The incidence of grades 3-4 adverse events was 61% and 18% in arms A and B, respectively. CONCLUSION Cediranib combined with olaparib improved rPFS compared with olaparib alone in men with mCRPC. This combination was associated with an increased incidence of grades 3-4 adverse events. BRCA2-mutated subgroups treated with olaparib with or without cediranib were associated with a numerically longer median rPFS.

Cystic fibrosis (CF) is caused by mutations in the CF transmembrane conductance regulator (CFTR) gene. We sought to correct the multiple organ dysfunction of the F508del CF-causing mutation using systemic delivery of peptide nucleic acid gene editing technology mediated by biocompatible polymeric nanoparticles. We confirmed phenotypic and genotypic modification in vitro in primary nasal epithelial cells from F508del mice grown at air-liquid interface and in vivo in F508del mice following intravenous delivery. In vivo treatment resulted in a partial gain of CFTR function in epithelia as measured by in situ potential differences and Ussing chamber assays and correction of CFTR in both airway and GI tissues with no off-target effects above background. Our studies demonstrate that systemic gene editing is possible, and more specifically that intravenous delivery of PNA NPs designed to correct CF-causing mutations is a viable option to ameliorate CF in multiple affected organs.

Lessons Learned Background Cediranib, a vascular endothelial growth factor receptor inhibitor, suppresses expression of BRCA1/2 and RAD51 inducing homologous recombination DNA repair deficiency (HRD) in several cancer cell lines and xenograft models [1]. Olaparib provides a clinical benefit in patients with metastatic pancreatic adenocarcinoma (mPDAC) with germline BRCA mutation (gBRCAmt) [2]. We hypothesized that cediranib induces HRD in the absence of gBRCAmt and synergizes with olaparib, resulting in an objective response in patients with mPDAC. Methods Patients with mPDAC with at least one prior systemic chemotherapy were enrolled. Patients with known gBRCAmt were excluded. Patients took cediranib 30 mg daily and olaparib 200 mg twice daily, orally. The primary endpoint was objective response (OR) rate. Results Nineteen patients received the study drugs. Seven patients came off treatment before the first restaging scan: six because of clinical progression and one because of an adverse event. No OR was observed. Six patients had stable disease (SD) as a best overall response. The median duration of SD was 3.1 months. The median overall survival was 3.4 months. Common treatment-related adverse events were fatigue, hypertension, and diarrhea. Conclusion Cediranib and olaparib combination did not result in clinically meaningful activity in patients with mPDAC without gBRCAmt.

The move from reading to writing the human genome offers new opportunities to improve human health. The United States National Institutes of Health (NIH) Somatic Cell Genome Editing (SCGE) Consortium aims to accelerate the development of safer and more-effective methods to edit the genomes of disease-relevant somatic cells in patients, even in tissues that are difficult to reach. Here we discuss the consortium’s plans to develop and benchmark approaches to induce and measure genome modifications, and to define downstream functional consequences of genome editing within human cells. Central to this effort is a rigorous and innovative approach that requires validation of the technology through third-party testing in small and large animals. New genome editors, delivery technologies and methods for tracking edited cells in vivo, as well as newly developed animal models and human biological systems, will be assembled—along with validated datasets—into an SCGE Toolkit, which will be disseminated widely to the biomedical research community. We visualize this toolkit—and the knowledge generated by its applications—as a means to accelerate the clinical development of new therapies for a wide range of conditions., This Perspective discusses how the Somatic Cell Genome Editing Consortium aims to accelerate the implementation of safe and effective genome-editing therapies in the clinic.

Chimeric antigen receptor (CAR) T cell and immune-cell therapies are revolutionizing how patients are treated in the clinic by targeting tumor-specific antigens. Specifically, advances in antigen targeting of CD20 and CD19 with CAR T cell therapy has increased overall survival of this patient population and has influenced to expand T cell repertoire to novel antigen targets. However, limitations to these cell-based therapies are the lack of development against solid tumor antigens, partly due to the paucity of antigen candidates that are sufficiently specific to epithelial cancers. Therefore, technologies that can mark tumor cells for immune recognition could prove beneficial in the treatment of these solid cancers. This project aims to artificially deliver an antigen to tumor cells by harnessing a universal property of tumors: acidic microenvironment. The pH low insertion peptide (pHLIP) is a technology that targets solid tumors based on the low pH produced by the Warburg effect. pHLIP-conjugates have undergone extensive development for drug delivery, with one notable candidate CBX12 entering phase I clinical trials in 2021. Our system uses a pHLIP-conjugated SIINFEKL peptide (derived from ovalbumin) to serve as our model antigen system, with SIINFEKL-targeting T cells (OT1) that can be used as a tool to investigate antigen delivery, recognition, and functionality. We have observed through immunofluorescence that B16 F10 mouse melanoma cells treated in vitro with the 1uM pHLIP-SIINFEKL construct under acidic conditions can recruit OT1 cells and induce a T cell synapse. Preliminary in vivo experiments in C57BL/6-Tg OT1 mice bearing B16 F10 flank tumors showed that treatment with pHLIP-SIINFEKL by intravenous administration led to a suppression of tumor growth compared to controls, with approximately 50% smaller fold change in tumor volume. Cumulatively, these initial results suggest that pHLIP-SIINFEKL can be delivered to tumors to recruit OT1 cells to engage in antigen recognition and lead to efficacy in tumor growth delay in vivo. This technology has the capability to provide a novel approach to decorate solid tumors with engineered antigens for enhanced immune recognition and anti-tumor efficacy. Citation Format: Annali M. Yurkevicz, Yanfeng Liu, Peter M. Glazer. Utilizing a pH sensitive peptide (pHLIP) for tumor targeted delivery of an immunogenic peptide motif [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 2715.

Oncogenic RAS mutations are associated with tumor resistance to radiation therapy. Cell-cell interactions in the tumor microenvironment (TME) profoundly influence therapy outcomes. However, the nature of these interactions and their role in Ras tumor radioresistance remain unclear. Here we use Drosophila oncogenic Ras tissues and human Ras cancer cell radiation models to address these questions. We discover that cellular response to genotoxic stress cooperates with oncogenic Ras to activate JAK/STAT non-cell autonomously in the TME. Specifically, p53 is heterogeneously activated in Ras tumor tissues in response to irradiation. This mosaicism allows high p53-expressing Ras clones to stimulate JAK/STAT cytokines, which activate JAK/STAT in the nearby low p53-expressing surviving Ras clones, leading to robust tumor re-establishment. Blocking any part of this cell-cell communication loop re-sensitizes Ras tumor cells to irradiation. These findings suggest that coupling STAT inhibitors to radiotherapy might improve clinical outcomes for Ras cancer patients., Dong, Vadla et al. find that oncogenic Ras and p53 cooperatively stimulate JAK/STAT cytokines to promote non-autonomous tissue overgrowth in Drosophila. Further, blocking STAT signalling prevents the growth-inducing effect of p53 overexpression and ionising irradiation in human cancer lines in culture and in mouse xenografts, suggesting a therapeutic route.

Pediatric GIST (gastrointestinal stromal tumors) are mostly KIT/ PDGFRA-WT and harbor mutations in SDH (succinate dehydrogenase), making them TKI (tyrosine kinase inhibitor) resistant due to the absence of gain-of-function tyrosine kinase mutations. Due to rarity of GIST in children, there have been few clinical trials available for patients with advanced disease, resulting in limited treatment options and a lack of pediatric-specific consensus guidelines. Here, we report the case of a patient with progressive, metastatic SDHB-mutant GIST that displayed a significant clinical response with a combination of Olaparib and TMZ (temozolomide).

Deregulation of metabolism and disruption of genome integrity are hallmarks of cancer1. Increased levels of the metabolites 2-hydroxyglutarate, succinate and fumarate occur in human malignancies owing to somatic mutations in the isocitrate dehydrogenase-1 or -2 (IDH1 or IDH2) genes, or germline mutations in the fumarate hydratase (FH) and succinate dehydrogenase genes (SDHA, SDHB, SDHC and SDHD), respectively2-4. Recent work has made an unexpected connection between these metabolites and DNA repair by showing that they suppress the pathway of homology-dependent repair (HDR)5,6 and confer an exquisite sensitivity to inhibitors of poly (ADP-ribose) polymerase (PARP) that are being tested in clinical trials. However, the mechanism by which these oncometabolites inhibit HDR remains poorly understood. Here we determine the pathway by which these metabolites disrupt DNA repair. We show that oncometabolite-induced inhibition of the lysine demethylase KDM4B results in aberrant hypermethylation of histone 3 lysine 9 (H3K9) at loci surrounding DNA breaks, masking a local H3K9 trimethylation signal that is essential for the proper execution of HDR. Consequently, recruitment of TIP60 and ATM, two key proximal HDR factors, is substantially impaired at DNA breaks, with reduced end resection and diminished recruitment of downstream repair factors. These findings provide a mechanistic basis for oncometabolite-induced HDR suppression and may guide effective strategies to exploit these defects for therapeutic gain.