GLI1 confers resistance to PARP inhibitors by activating the DNA damage repair pathway.

Identifying the mechanisms of action of anticancer drugs is an important step in the development of new drugs. In this study, we established a comprehensive screening platform consisting of 68 oncogenes (MANO panel), encompassing 243 genetic variants, to identify predictive markers for drug efficacy. Validation was performed using drugs that targeted EGFR, BRAF, and MAP2K1, which confirmed the utility of this functional screening panel. Screening of a BRCA2-knockout DLD1 cell line (DLD1-KO) revealed that cells expressing SMO and GLI1 were resistant to olaparib. Gene set enrichment analysis identified genes associated with DNA damage repair that were enriched in cells overexpressing SMO and GLI1. The expression of genes associated with homologous recombination repair (HR), such as the FANC family and BRCA1/2, was significantly upregulated by GLI1 expression, which is indicative of PARP inhibitor resistance. Although not all representative genes of the nucleotide excision repair (NER) pathway were upregulated, NER activity was enhanced by GLI1. The GLI1 inhibitor was effective against DLD1-KO cells overexpressing GLI1 both in vitro and in vivo. Furthermore, the combination therapy of olaparib and GLI1 inhibitor exhibited a synergistic effect on DLD1-KO, suggesting the possible clinical application of GLI1 inhibitor targeting cancer with defective DNA damage repair. This platform enables the identification of biomarkers associated with drug sensitivity, and is a useful tool for drug development.

A molecular glue RBM39-degrader induces synthetic lethality in cancer cells with homologous recombination repair deficiency.

E7820 and Indisulam (E7070) are sulfonamide molecular glues that modulate RNA splicing by degrading the splicing factor RBM39 via ternary complex formation with the E3 ligase adaptor DCAF15. To identify biomarkers of the antitumor efficacy of E7820, we treated patient-derived xenograft (PDX) mouse models established from 42 patients with solid tumors. The overall response rate was 38.1% (16 PDXs), and tumor regression was observed across various tumor types. Exome sequencing of the PDX genome revealed that loss-of-function mutations in genes of the homologous recombination repair (HRR) system, such as ATM, were significantly enriched in tumors that responded to E7820 (p = 4.5 × 103). Interestingly, E7820-mediated double-strand breaks in DNA were increased in tumors with BRCA2 dysfunction, and knockdown of BRCA1/2 transcripts or knockout of ATM, ATR, or BAP1 sensitized cancer cells to E7820. Transcriptomic analyses revealed that E7820 treatment resulted in the intron retention of mRNAs and decreased transcription, especially for HRR genes. This induced HRR malfunction probably leads to the synthetic lethality of tumor cells with homologous recombination deficiency (HRD). Furthermore, E7820, in combination with olaparib, exerted a synergistic effect, and E7820 was even effective in an olaparib-resistant cell line. In conclusion, HRD is a promising predictive biomarker of E7820 efficacy and has a high potential to improve the prognosis of patients with HRD-positive cancers.

Single base substitution signatures 17a, 17b, and 40 are induced by γ-ray irradiation in association with increased reactive oxidative species.

γ-Ray irradiation induces DNA double strand breaks (DSBs) and increases the risk of cancerization. Irradiated cells usually repair DSBs directly, but accumulate replication stress-associated DSBs, increasing the risk of structural variants (SVs). Although single nucleotide variants (SNVs) are also induced, it is still unclear which SNVs are induced by γ-ray irradiation. Here, we show that single base substitution (SBS) 17a, 17b, and 40 signatures were induced by γ-ray irradiation, which is mainly SNV induction in A-T bps. While SNVs induced by genomic instability were usually associated with SVs, SNVs induced by γ-ray irradiation and the associated signatures were not. As reactive oxygen species (ROS) are a possible cause of SBS17a and 17b, ROS were induced upon γ-ray irradiation (1-8 Gy), indicating the association of ROS for the SNV induction. Thus, our results reveal that ROS-associated SNVs are increased by irradiation, and that ROS-associated SNVs are induced independently of SVs.

Genome destabilization-associated phenotypes arising as a consequence of therapeutic treatment are suppressed by Olaparib.

Malignancy is often associated with therapeutic resistance and metastasis, usually arising after therapeutic treatment. These include radio- and chemo-therapies, which cause cancer cell death by inducing DNA double strand breaks (DSBs). However, it is still unclear how resistance to these DSBs is induced and whether it can be suppressed. Here, we show that DSBs induced by camptothecin (CPT) and radiation jeopardize genome stability in surviving cancer cells, ultimately leading to the development of resistance. Further, we show that cytosolic DNA, accumulating as a consequence of genomic destabilization, leads to increased cGAS/STING-pathway activation and, ultimately, increased cell migration, a precursor of metastasis. Interestingly, these genomic destabilization-associated phenotypes were suppressed by the PARP inhibitor Olaparib. Recognition of DSBs by Rad51 and genomic destabilization were largely reduced by Olaparib, while the DNA damage response and cancer cell death were effectively increased. Thus, Olaparib decreases the risk of therapeutic resistance and cell migration of cells that survive radio- and CPT-treatments.

Echoed induction of nucleotide variants and chromosomal structural variants in cancer cells.

Generally, the number of single-nucleotide variants (SNVs) in somatic cells increases with age, which is expected for replication errors. The number of SNVs in cancer cells, however, is often much higher than that in somatic cells, raising the question of whether cancer cells possess SNV induction pathways. The present study shows that the number of SNVs in cancer cells correlates with the number of chromosomal structural variants (SVs). While Kataegis, localized hypermutations typically arising near SV sites, revealed multiple SNVs within 1 kb, SV-associated SNVs were generally observed within 0.1-1 Mb of SV sites, irrespective of Kataegis status. SNVs enriched within 1 Mb of SV regions were associated with deficiency of DNA damage repair, including HR deficiency-associated single base substitution 3 (SBS3) and exogenous damage-associated SBS7 and SBS36 signatures. We also observed a similar correlation between SVs and SNVs in cells that had undergone clonal evolution in association with genomic instability, implying an association between genomic instability and SV-associated induction of SNVs.

Genomic Instability and Cancer Risk Associated with Erroneous DNA Repair.

Many cancers develop as a consequence of genomic instability, which induces genomic rearrangements and nucleotide mutations. Failure to correct DNA damage in DNA repair defective cells, such as in BRCA1 and BRCA2 mutated backgrounds, is directly associated with increased cancer risk. Genomic rearrangement is generally a consequence of erroneous repair of DNA double-strand breaks (DSBs), though paradoxically, many cancers develop in the absence of DNA repair defects. DNA repair systems are essential for cell survival, and in cancers deficient in one repair pathway, other pathways can become upregulated. In this review, we examine the current literature on genomic alterations in cancer cells and the association between these alterations and DNA repair pathway inactivation and upregulation.

Replication-stress-associated DSBs induced by ionizing radiation risk genomic destabilization and associated clonal evolution.

Exposure to ionizing radiation is associated with cancer risk. Although multiple types of DNA damage are caused by radiation, it remains unknown how this damage is associated with cancer risk. Here, we show that after repair of double-strand breaks (DSBs) directly caused by radiation (dir-DSBs), irradiated cells enter a state at higher risk of genomic destabilization due to accumulation of replication-stress-associated DSBs (rs-DSBs), ultimately resulting in clonal evolution of cells with abrogated defense systems. These effects were observed over broad ranges of radiation doses (0.25-2 Gy) and dose rates (1.39-909 mGy/min), but not upon high-dose irradiation, which caused permanent cell-cycle arrest. The resultant genomic destabilization also increased the risk of induction of single-nucleotide variants (SNVs), including radiation-associated SNVs, as well as structural alterations in chromosomes. Thus, the radiation-associated risk can be attributed to rs-DSB accumulation and resultant genomic destabilization.

Paraovarian Cyst Torsion in a Patient with Rubinstein-Taybi Syndrome: A Case Report.

Rubinstein-Taybi syndrome is an extremely rare autosomal dominant genetic disorder that occurs in 1/125,000 and is characterized by distinctive facial appearance, short stature, mild to severe mental retardation, and higher risk for cancer. In addition, variable organ anomalies had been reported. Paraovarian cyst causing torsion of the ipsilateral fallopian tube is less common, with an estimated incidence of 1/1,500,000, but it can adversely affect tubal function. It occurs mainly in women in the reproductive age and is very rare in prepubescent girls. Here, we described the successful treatment of an extremely rare case of paraovarian cyst causing torsion of the ipsilateral fallopian tube in a patient with Rubinstein-Taybi syndrome. A 14-year-old girl with Rubinstein-Taybi syndrome was referred to our hospital for abdominal pain. Her medical history was unremarkable, except for moderate hirsutism and keloid scar. Physical examination revealed tenderness in the lower abdominal midline. The preoperative diagnosis was torsion of a left ovarian cyst. An exploratory laparoscopy was performed because of acute abdominal pain and revealed a left fallopian tube that was twisted twice due to an ipsilateral paraovarian cyst. The huge paraovarian cyst required laparotomy cystectomy, and the left ovary was preserved. Her postoperative course was uncomplicated. Preoperative diagnosis of paraovarian cysts can be difficult. The moderate hirsutism seen in our patient suggested the presence of a large paraovarian cyst due to androgen receptor-mediated effects. Therefore, Rubinstein-Taybi syndrome patients with hirsutism should be screened and assessed by pediatric surgeons for the presence of paraovarian cysts.

Hypertensive Cardiomyopathy with Congestive Heart Failure in an Infant with Unilateral Wilms Tumor: A Case Report.

Wilms tumor (WT) is the most common malignant kidney tumor in children. High blood pressure is seen in up to 55% of children with WT. However, hypertensive cardiomyopathy with congestive heart failure due to WT is remarkably rare, with only several cases reported worldwide. In this report, a pediatric case of WT with hypertension causing hypertensive cardiomyopathy and congestive heart failure is presented. An 8-month-old male child with abdominal distension was seen by his primary physician. He was referred to our hospital for further examination and treatment. Abdominal contrast-enhanced computed tomography demonstrated a weakly enhancing, large abdominal mass, which was larger than 12 cm. Two-dimensional transthoracic echocardiography showed a diffuse hypokinetic left ventricle. The patient was diagnosed with cardiomyopathy caused by hypertension. Open surgical resection of the mass was successfully performed. His postoperative course was uncomplicated, and the patient was successfully discharged. The plasma renin activity was maintained at a high level even after left nephrectomy, suggesting that the right kidney was likely the source of renin secretion. Mechanical compression of the right renal blood vessels by a greatly enlarged left kidney can cause right renal ischemia, which activates renin excretion. Nephrectomy can be an effective treatment for a WT patient with hypertension causing hypertensive cardiomyopathy, and then cardiac function will be improved within several weeks. We recommend routine echocardiography surveillance in patients with WT. This report can help pediatric surgeons become more familiar with cardiomyopathy caused by WT.

Genomic destabilization and its associated mutagenesis increase with senescence-associated phenotype expression.

Cancer develops through multiple rounds of clonal evolution of cells with abrogated defense systems. Such clonal evolution is triggered by genomic destabilization with associated mutagenesis. However, what increases the risk of genomic destabilization remains unclear. Genomic instability is usually the result of erroneous repair of DNA double-strand breaks (DSB); paradoxically, however, most cancers develop with genomic instability but lack mutations in DNA repair systems. In this manuscript, we review current knowledge regarding a cellular state that increases the risk of genomic destabilization, in which cells exhibit phenotypes often observed during senescence. In addition, we explore the pathways that lead to genomic destabilization and its associated mutagenesis, which ultimately result in cancer.

Resveratrol and its Related Polyphenols Contribute to the Maintenance of Genome Stability.

Genomic destabilisation is associated with the induction of mutations, including those in cancer-driver genes, and subsequent clonal evolution of cells with abrogated defence systems. Such mutations are not induced when genome stability is maintained; however, the mechanisms involved in genome stability maintenance remain elusive. Here, resveratrol (and related polyphenols) is shown to enhance genome stability in mouse embryonic fibroblasts, ultimately protecting the cells against the induction of mutations in the ARF/p53 pathway. Replication stress-associated DNA double-strand breaks (DSBs) that accumulated with genomic destabilisation were effectively reduced by resveratrol treatment. In addition, resveratrol transiently stabilised the expression of histone H2AX, which is involved in DSB repair. Similar effects on the maintenance of genome stability were observed for related polyphenols. Accordingly, we propose that polyphenol consumption can contribute to the suppression of cancers that develop with genomic instability, as well as lifespan extension.

Genomic-Destabilization-Associated Mutagenesis and Clonal Evolution of Cells with Mutations in Tumor-Suppressor Genes.

The development of cancer is driven by genomic instability and mutations. In general, cancer develops via multiple steps. Each step involves the clonal evolution of cells with abrogated defense systems, such as cells with mutations in cancer-suppressor genes. However, it remains unclear how cellular defense systems are abrogated and the associated clonal evolution is triggered and propagated. In this manuscript, we review current knowledge regarding mutagenesis associated with genomic destabilization and its relationship with the clonal evolution of cells over the course of cancer development, focusing especially on mechanistic aspects.

Replication stress triggers microsatellite destabilization and hypermutation leading to clonal expansion in vitro.

Mismatch repair (MMR)-deficient cancers are characterized by microsatellite instability (MSI) and hypermutation. However, it remains unclear how MSI and hypermutation arise and contribute to cancer development. Here, we show that MSI and hypermutation are triggered by replication stress in an MMR-deficient background, enabling clonal expansion of cells harboring ARF/p53-module mutations and cells that are resistant to the anti-cancer drug camptothecin. While replication stress-associated DNA double-strand breaks (DSBs) caused chromosomal instability (CIN) in an MMR-proficient background, they induced MSI with concomitant suppression of CIN via a PARP-mediated repair pathway in an MMR-deficient background. This was associated with the induction of mutations, including cancer-driver mutations in the ARF/p53 module, via chromosomal deletions and base substitutions. Immortalization of MMR-deficient mouse embryonic fibroblasts (MEFs) in association with ARF/p53-module mutations was ~60-fold more efficient than that of wild-type MEFs. Thus, replication stress-triggered MSI and hypermutation efficiently lead to clonal expansion of cells with abrogated defense systems.

Usefulness of the HyperEye Medical System in Obstructive Colitis Proximal to Colon Carcinoma.

Obstructive colitis (OC) is a nonspecific inflammatory condition that occurs at the proximal side of a completely or partially stenotic lesion typically caused by colorectal cancer. Impaired blood flow caused by these stenotic changes in the colon or rectum results in this condition. During surgery for sigmoid colon carcinoma with OC, complete surgical removal of the OC lesions is required. However, it is difficult to anticipate the range of OC before surgery. Diagnosing the potential ischemia during surgery would decrease the need for re-operation. This is the first report of HyperEye Medical System (HEMS) angiography for surgery of colon cancer with OC. We report a case of sigmoid colon carcinoma in which HEMS angiography was used and found to be useful for real-time detection of the OC lesion.

Mismatch repair dependence of replication stress-associated DSB recognition and repair.

Most cancers develop with one of two types of genomic instability, namely, chromosomal instability (CIN) or microsatellite instability (MSI). Both are induced by replication stress-associated DNA double-strand breaks (DSBs). The type of genomic instability that arises is dependent on the choice of DNA repair pathway. Specifically, MSI is induced via a PolQ-dependent repair pathway called microhomology-mediated end joining (MMEJ) in a mismatch repair (MMR)-deficient background. However, it is unclear how the MMR status determines the choice of DSB repair pathway. Here, we show that replication stress-associated DSBs initially targeted by the homologous recombination (HR) system were subsequently hijacked by PolQ-dependent MMEJ in MMR-deficient cells, but persisted as HR intermediates in MMR-proficient cells. PolQ interacting with MMR factors was effectively loaded onto damaged chromatin in an MMR-deficient background, in which merged MRE11/γH2AX foci also effectively formed. Thus, the choice of DNA repair pathway according to the MMR status determines whether CIN or MSI is induced.

Onset of deaminase APOBEC3B induction in response to DNA double-strand breaks.

Deamination of 5-methyl cytosine is a major cause of cancer-driver mutations in inflammation-associated cancers. The deaminase APOBEC3B is expressed in these cancers and causes mutations under replication stress; however, the mechanisms by which APOBEC3B mediates deamination and its association with genomic disorders are still unclear. Here, we show that APOBEC3B is stabilized to induce deamination reaction in response to DNA double-strand breaks (DSBs), resulting in the formation of long-lasting DSBs. Uracil, the major deamination product, is subsequently targeted by base excision repair (BER) through uracil-DNA glycosylase 2 (UNG2); hence late-onset DSBs arise as by-products of BER. The frequency of these delayed DSBs was increased by treatment of cells with a PARP inhibitor, and was suppressed following knock-down of UNG2. The late-onset DSBs were induced in an ATR-dependent manner. Those secondary DSBs were persistent, unlike DSBs directly caused by γ-ray irradiation. Overall, these results suggest that the deaminase APOBEC3B is induced in response to DSBs, leading to long-lasting DSB formation in addition to mutagenic 5me-C>T transition induction.

Sensitization of Cancer Cells to Radiation and Topoisomerase I Inhibitor Camptothecin Using Inhibitors of PARP and Other Signaling Molecules.

Radiation and certain anticancer drugs damage DNA, resulting in apoptosis induction in cancer cells. Currently, the major limitations on the efficacy of such therapies are development of resistance and adverse side effects. Sensitization is an important strategy for increasing therapeutic efficacy while minimizing adverse effects. In this manuscript, we review possible sensitization strategies for radiation and anticancer drugs that cause DNA damage, focusing especially on modulation of damage repair pathways and the associated reactions.

Laparoscopy-assisted distal gastrectomy is feasible also for elderly patients aged 80 years and over: effectiveness and long-term prognosis.

BACKGROUND: Elderly patients usually have concurrent ailments, and the safety and effectiveness of laparoscopy-assisted distal gastrectomy (LADG) for these patients have been controversial. This study aimed to evaluate whether laparoscopy-assisted distal gastrectomy is safe and effective for elderly patients aged 80 years and over, as well as to clarify their long-term prognosis. METHODS: A total of 31 patients aged 80 years and over who underwent LADG in our hospital were retrospectively reviewed. Peri- and postoperative data were compared with those of 38 patients aged 65 years and younger. The median follow-up period of the elderly and younger group was 56.0 and 63.0 months, respectively, and their prognosis was examined. RESULTS: There were significant differences between the two groups in preoperative respiratory and renal functions, hemoglobin, and nutritional index. Significant differences in postoperative complications were seen only in pneumonia and delirium. There were no hospital deaths, but the 3-year and 5-year overall survival rates were significantly lower in the elderly group than in the non-elderly group. However, in the elderly group, only one patient died of gastric cancer recurrence, whereas four died of cardiovascular disease and three died of pneumonia during follow-up. Therefore, the recurrence-free survival rate was not significantly different between the groups. CONCLUSIONS: LADG seems to be safe and effective even for elderly patients, and their prognosis was satisfactory. However, careful monitoring for cardiovascular and pulmonary disease should be observed during the follow-up period.

Acute respiratory distress syndrome after gastrointestinal surgery.

BACKGROUND/AIMS: Acute respiratory distress syndrome (ARDS) occasionally occurs after gastrointestinal surgery involving severe inflammation such as diffuse peritonitis. Management of this condition has been difficult and effective therapies have not yet been established. In the present study the management for ARDS after gastrointestinal surgery was evaluated. METHODOLOGY: A total of 15 patients developed ARDS after gastrointestinal operations performed in our institution. The mean patient age was 75.4±11.1 years. Onset of ARDS occurred ≤24 hours postoperatively in 12 patients and 3- 11 days postoperatively in 3 patients. Treatment for ARDS comprised continuous hemodiafiltration (CHDF), high-dose glucocorticoid therapy or administration of a neutrophil elastase inhibitor (sivelestat). RESULTS: Four patients died 3-45 days after onset of ARDS (mortality rate, 26.6%). CHDF was performed in 12 patients, 8 patients received highdose glucocorticoid therapy and 11 patients received sivelestat. No differences in severity scores and clinical data were noted between survivors and non-survivors. PaO2/FiO2 ratio was significantly lower in non-survivors than in survivors from 5 days after starting treatment, whereas no difference was apparent at the onset of ARDS. CONCLUSIONS: Multimodal therapies for ARDS were effective. Longitudinal fluctuation in PaO2/ FiO2 ratio after starting treatment appears to offer a prognostic factor for ARDS.

A case of mucoid carcinoma of the sigmoid colon with features of a submucosal tumor.

An 82-year-old man visited a medical clinic for dyschezia. His serum carcinoembryonic antigen (CEA) level was 16ng/ml. Abdominal computed tomography (CT) revealed an 8×7cm tumor in the sigmoid colon. Endoscopy showed that the sigmoid colon was insufficiently dilated but no mucosal lesions were present. Histological examination of biopsy specimens did not show malignant cells. Sigmoidectomy was performed due to suspected submucosal tumor. The final pathological diagnosis was mucoid carcinoma with features of a submucosal tumor which had no mucosal lesions. Mucoid carcinoma with features of a submucosal tumor in the colorectum is rare. Most cases are associated with ulcer lesions or scar lesions on the mucosal surface of tumor because it arises from epithelial cells.