DNA Damage Repair Ability Research Articles

Quantitative evaluation of DNA damage repair dynamics to elucidate predictors of autism vs. cancer in individuals with germline PTEN variants.

Persons with germline variants in the tumor suppressor gene phosphatase and tensin homolog, PTEN, are molecularly diagnosed with PTEN hamartoma tumor syndrome (PHTS). PHTS confers high risks of specific malignancies, and up to 23% of the patients are diagnosed with autism spectrum disorder (ASD) and/or developmental delay (DD). The accurate prediction of these two seemingly disparate phenotypes (cancer vs. ASD/DD) for PHTS at the individual level remains elusive despite the available statistical prevalence of specific phenotypes of the syndrome at the population level. The pleiotropy of the syndrome may, in part, be due to the alterations of the key multi-functions of PTEN. Maintenance of genome integrity is one of the key biological functions of PTEN, but no integrative studies have been conducted to quantify the DNA damage response (DDR) in individuals with PHTS and to relate to phenotypes and genotypes. In this study, we used 43 PHTS patient-derived lymphoblastoid cell lines (LCLs) to investigate the associations between DDR and PTEN genotypes and/or clinical phenotypes ASD/DD vs. cancer. The dynamics of DDR of γ-irradiated LCLs were analyzed using the exponential decay mathematical model to fit temporal changes in γH2AX levels which report the degree of DNA damage. We found that PTEN nonsense variants are associated with less efficient DNA damage repair ability resulting in higher DNA damage levels at 24 hours after irradiation compared to PTEN missense variants. Regarding PHTS phenotypes, LCLs from PHTS individuals with ASD/DD showed faster DNA damage repairing rate than those from patients without ASD/DD or cancer. We also applied the reaction-diffusion partial differential equation (PDE) mathematical model, a cell growth model with a DNA damage term, to accurately describe the DDR process in the LCLs. For each LCL, we can derive parameters of the PDE. Then we averaged the numerical results by PHTS phenotypes. By performing simple subtraction of two subgroup average results, we found that PHTS-ASD/DD is associated with higher live cell density at lower DNA damage level but lower cell density level at higher DNA damage level compared to LCLs from individuals with PHTS-cancer and PHTS-neither.

Comparison of primordial germ cell differences at different developmental time points in chickens.

Recently, the application in the field of germplasm resource conservation has become an important application of primordial germ cells (PGCs). However, due to the lack of deep understanding of the biological characteristics of PGCs at different time points, there is no systematic scheme for the selection of PGCs at which time points in practical application, which affects the practical application effect of PGCs. This study aims to clarify the differences in PGCs during development. Here, migration experiment, EdU proliferation assay and cell apoptosis assay were conducted to compare the differences in the migration ability, the proliferation ability and the recovery efficiency among female and male PGCs at E3.5, E4.5 and E5.5, which were explained by the following transcriptome sequencing analysis. We found that there were larger differences between female and male PGCs at different embryonic ages, while smaller differences between female and male PGCs at the same embryonic age. Further comparison showed that the cell migration ability of female and male PGCs decreased gradually during development, so female and male PGCs at E3.5 are more suitable for in vitro allotransplantation. At the same time, the proliferation ability of PGCs gradually decreased during development, and cell adhesion and extracellular matrix communication were weakened, indicating that female and male PGCs of E3.5 are more suitable for in vitro long-term culture cell line establishment. Interestingly, female and male PGCs at E5.5 showed strong DNA damage repair ability, thus more suitable for in vitro long-term cryopreservation. This study provides a theoretical basis for systematically selecting PGCs at suitable developmental time points as cell materials for efficient utilization by analyzing the characteristics of female and male PGCs at different developmental time points based on transcriptome.

Oleandrin enhances radiotherapy sensitivity in lung cancer by inhibiting the ATM/ATR-mediated DNA damage response.

Despite active clinical trials on the use of Oleandrin alone or in combination with other drugs for the treatment of solid tumors, the potential synergistic effect of Oleandrin with radiotherapy remains unknown. This study reveals a new mechanism by which Oleandrin targets ATM and ATR kinase-mediated radiosensitization in lung cancer. Various assays, including clonogenic, Comet, immunofluorescence staining, apoptosis and Cell cycle assays, were conducted to evaluate the impact of oleandrin on radiation-induced double-strand break repair and cell cycle distribution. Western blot analysis was utilized to investigate alterations in signal transduction pathways related to double-strand break repair. The efficacy and toxicity of the combined therapy were assessed in a preclinical xenotransplantation model. Functionally, Oleandrin weakens the DNA damage repair ability and enhances the radiation sensitivity of lung cells. Mechanistically, Oleandrin inhibits ATM and ATR kinase activities, blocking the transmission of ATM-CHK2 and ATR-CHK1 cell cycle checkpoint signaling axes. This accelerates the passage of tumor cells through the G2 phase after radiotherapy, substantially facilitating the rapid entry of large numbers of inadequately repaired cells into mitosis and ultimately triggering mitotic catastrophe. The combined treatment of Oleandrin and radiotherapy demonstrated superior inhibition of tumor proliferation compared to either treatment alone. Our findings highlight Oleandrin as a novel and effective inhibitor of ATM and ATR kinase, offering new possibilities for the development of clinical radiosensitizing adjuvants.

CD276 enhances sunitinib resistance in clear cell renal cell carcinoma by promoting DNA damage repair and activation of FAK-MAPK signaling pathway.

This study aimed to explore the effect of CD276 expression on the sunitinib sensitivity of clear cell renal cell carcinoma (ccRCC) cell and animal models and the potential mechanisms involved. CD276 expression levels of ccRCC and normal samples were analyzed via online databases and real-time quantitative PCR (RT-qPCR). CD276 was knocked down in ccRCC cell models (sunitinib-resistant 786-O/R cells and sunitinib-sensitive 786-O cells) using shRNA transfection, and the cells were exposed to a sunitinib (2 µM) environment. Cells proliferation was then analyzed using MTT assay and colony formation experiment. Alkaline comet assay, immunofluorescent staining, and western blot experiments were conducted to assess the DNA damage repair ability of the cells. Western blot was also used to observe the activation of FAK-MAPK pathway within the cells. Finally, a nude mouse xenograft model was established and the nude mice were orally administered sunitinib (40mg/kg/d) to evaluate the in vivo effects of CD276 knockdown on the therapeutic efficacy of sunitinib against ccRCC. CD276 was significantly upregulated in both ccRCC clinical tissue samples and cell models. In vitro experiments showed that knocking down CD276 reduced the survival rate, IC50 value, and colony-forming ability of ccRCC cells. Knocking down CD276 increased the comet tail moment (TM) values and γH2AX foci number, and reduced BRCA1 and RAD51 protein levels. Knocking down CD276 also decreased the levels of p-FAK, p-MEK, and p-ERK proteins. Knocking down CD276 effectively improved the sensitivity of ccRCC cell and animal models to sunitinib treatment.

ADAR1 promotes cisplatin resistance in intrahepatic cholangiocarcinoma by regulating BRCA2 expression through A-to-I editing manner.

Aberrant A-to-I RNA editing, mediated by ADAR1 has been found to be associated with increased tumourigenesis and the development of chemotherapy resistance in various types of cancer. Intrahepatic cholangiocarcinoma (iCCA) is a highly aggressive malignancy with a poor prognosis, and overcoming chemotherapy resistance poses a significant clinical challenge. This study aimed to clarify the roles of ADAR1 in tumour resistance to cisplatin in iCCA. We discovered that ADAR1 expression is elevated in iCCA patients, particularly in those resistant to cisplatin, and associated with poor clinical outcomes. Downregulation of ADAR1 can increase the sensitivity of iCCA cells to cisplatin treatment, whereas its overexpression has the inverse effect. By integrating RNA sequencing and Sanger sequencing, we identified BRCA2, a critical DNA damage repair gene, as a downstream target of ADAR1 in iCCA. ADAR1 mediates the A-to-I editing in BRCA2 3'UTR, inhibiting miR-3157-5p binding, consequently increasing BRCA2 mRNA and protein levels. Furthermore, ADAR1 enhances cellular DNA damage repair ability and facilitates cisplatin resistance in iCCA cells. Combining ADAR1 targeting with cisplatin treatment markedly enhances the anticancer efficacy of cisplatin. In conclusion, ADAR1 promotes tumour progression and cisplatin resistance of iCCA. ADAR1 targeting could inform the development of innovative combination therapies for iCCA.

NCAPH serves as a prognostic factor and promotes the tumor progression in glioma through PI3K/AKT signaling pathway.

Non-SMC (Structural Maintenance of Chromosomes) condensin I complex subunit H (NCAPH) has been shown to facilitate progression and predict adverse prognostic outcome in many cancer types. However, the function of NCAPH in gliomas is still unclear. Series of experiments were taken to uncover the function of NCAPH in glioma. The expression of NCAPH and potential mechanism regulating progression of glioma was verified by bioinformatics analysis. Lentiviral transfection was used for establishment of loss-of-function and gain-of-function cell lines. CCK-8 assay and Colony-formation assay were used to evaluate proliferation. Transwell assay and Cell wound healing assay were used to assess migration and invasion. Cell cycle and apoptosis were measured by flow cytometry. Protein and RNA were quantified by WB and RT-PCR, respectively. The nude mice model of glioma was used to evaluate the effect of NCAPH in vivo. The expression of NCAPH increased significantly in glioma tissues and correlated with WHO grade, IDH wild-type and non-1p/19q codeletion. Glioma patients with high expression of NCAPH had an undesirable prognosis. Functionally, upregulated NCAPH promotes the malignant hallmarks of glioma cells in vivo and in vitro. NCAPH correlated with DNA damage repair ability of glioma cells and facilitated the proliferation, invasion, and migration of glioma cells by promoting the PI3K/AKT signaling pathway. This study identifies the important pro-tumor role of NCAPH in glioma and suggests that NCAPH is a potential therapeutic target.

Cryptomphalus aspersa Egg Extract Protects against Human Stem Cell Stress-Induced Premature Senescence.

Cellular senescence is a tightly regulated pathophysiologic process and is caused by replicative exhaustion or external stressors. Since naturally derived bioactive compounds with anti-ageing properties have recently captured scientific interest, we analysed the anti-ageing and antioxidant efficacy of Cryptomphalus aspersa egg extract (CAEE). Its effects on stemness, wound-healing properties, antioxidant defense mechanisms, and DNA damage repair ability of Human Wharton's jelly mesenchymal stem cells (WJ-MSCs) were analysed. Our results revealed that CAEE fortifies WJ-MSCs stemness, which possibly ameliorates their wound-healing ability. Additionally, we show that CAEE possesses a strong antioxidant capacity as demonstrated by the elevation of the levels of the basic antioxidant molecule, GSH, and the induction of the NRF2, a major antioxidant regulator. In addition, CAEE alleviated cells' oxidative stress and therefore prevented stress-induced premature senescence (SIPS). Furthermore, we demonstrated that the prevention of SIPS could be mediated via the extract's ability to induce autophagy, as indicated by the elevation of the protein levels of all basic autophagic molecules and the increase in formation of autophagolysosomes in CAEE-treated WJ-MSCs. Moreover, CAEE-treated cells exhibited decreased Caveolin-1 levels. We propose that Cryptomphalus aspersa egg extract comprises bioactive compounds that can demonstrate strong antioxidant/anti-ageing effects by regulating the Caveolin-1-autophagy-senescence molecular axis.

Enhanced Expression of DNp73 Promotes Drug Resistance and Immune Evasion in Multiple Myeloma By Targeting the MYC and MYCN Pathways

Background Multiple myeloma (MM) is an incurable haematological malignancy with high chromosome instability and is highly dependent on the immunosuppressive bone marrow microenvironment. TP53 mutations and deletions are uncommon in MM patients, with other types of mechanisms involved in the abrogation of TP53 pathway activity. DNp73, an inhibitor of the p53 tumour suppressor family, drives drug resistance and cancer progression in several solid malignancies. However, the biological functions of DNp73, the molecular mechanisms underlying these actions and their correlations with aberrant TP53 expression in myelomagenesis remain unclear. In this study, we investigated the role of DNp73 in the drug resistance of MM, and disclosed the corresponding mechanism of how DNp73 promotes the immune escape of MM cells. Aims To disclose the role of DNp73 in promoting drug resistance and immune evasion in MM, and explore the therapeutic strategy based on the mechanism research. Methods We constructed DNp73 overexpression and sh-RNA interference plasmids to obtain stable cell lines. The effects of DNp73 on proliferation and drug sensitivity were determined by CCK8, flow cytometry and xenotransplantation model. To identify mechanisms of drug resistance, we here perform RNA-seq and CHIP-seq analyses in MM cell lines, while Western blot and RT-PCR were used to confirm the results. The DNA damage repair and invasion ability of MM cells were detected by immunofluorescence and transwell assay. To validate the role of DNp73 in immune escape, we also performed phagocytosis assays in vitro. Results Our previous study revealed that the miRNA-15a is a tumour suppressor involved in aggressive MM cell proliferation and drug resistance. Here, we found that miRNA-15a downregulation in MM cells activated NF-κB-p65, which upregulated the expression of DNp73 at the transcriptional level. In addition, the level of miR-15a was negatively correlated with the expression of DNp73 in MM cells of patients (r=-0.672, p<0.05). Enforced expression of DNp73 promoted aggressive proliferation and multidrug resistance in MM cells (p<0.001). Bulk RNA sequencing data showed that the levels of MYCN, MYC and CDK7 were increased in cells with enforced DNp73 expression. Furthermore, a ChIP-qPCR assay revealed that DNp73 bound to the MYCN gene promoter and upregulated MYCN expression. The MM cell immune evasion rate was notable when DNp73 expression was high because of CD47 and PD-L1 upregulation. Blockade of the CD47/SIRPα and PD-1/PD-L1 signalling pathways by the SIRPα-Fc fusion protein IMM01 and monoclonal antibody atezolizumab significantly re-established the anti-MM function of macrophages and T cells in the MM microenvironment, respectively (p<0.01). Notably, DNp73 dysregulation and its pathogenetic function in MM cells were found to be independent of TP53 expression. Conclusion Altogether, our study demonstrates that DNp73 is a potential oncogene involved in MM pathogenesis and promotes MM cell proliferation and immune evasion by targeting the MYC and MYCN pathways.

The impact of female BMI on sperm DNA damage repair ability of oocytes and early embryonic development potential in intracytoplasmic sperm injection cycles.

Obesity adversely influences the quality of oocytes and embryos and can affect DNA repair in embryos, leading to reproductive issues. However, the effects of body mass index (BMI) on DNA repair ability in oocytes during intracytoplasmic sperm injection (ICSI) cycles have not yet been investigated. Therefore, this retrospective study aimed to analyze the influence of sperm DNA damage on embryo development and reproductive outcomes in overweight/obese and normal-weight women in ICSI cycles. A total of 1,141 patients who received the first fresh ICSI cycle treatments were recruited from July 2017 to July 2021. Based on the BMI of the women, all patients were divided into normal weight (18.5≤BMI<25 kg/m2; n=824; 72.22%) and overweight/obese (BMI≥25 kg/m2; n=317; 27.78%) groups. Furthermore, according to the sperm DNA fragmentation index (DFI), these two groups were subdivided into two subgroups: DFI<30% and DFI≥30%. In the normal-weight women group, the embryonic development and reproductive outcomes of ICSI cycles were not statistically different between the two subgroups (DFI<30% and DFI≥30%). However, in the overweight/obese women group, couples with a sperm DFI≥30% had a significantly lower fertilization rate (76% vs. 72.7%; p=0.027), cleavage rate (98.7% vs. 97.2%; p=0.006), and high-quality embryo rate (67.8% vs. 62.6%; p=0.006) than couples with a sperm DFI<30%. When injected sperm with high DFI into the oocytes of overweight/obese women, resulting in lower fertilization, cleavage, and high-quality embryo rates in ICSI cycles, and the decreased early developmental potential of embryos from overweight/obese patients may be caused by the diminished capacity of oocytes to repair sperm DNA damage.

17th International Conference on Malignant Lymphoma, Palazzo dei Congressi, Lugano, Switzerland, 13 - 17 June, 2023.

Introduction: Multiple myeloma (MM) remains an incurable malignancy of plasma cells. The complex bone marrow microenvironment promotes resistance to both current anti-myeloma agents and emerging immunotherapies. Crosstalk between the NF-κB and p53 can play a pivotal role in various hematologic malignancies. DNp73, an inhibitor of the p53 tumor suppressor family, drives drug resistance and cancer progression in several solid malignancies. However, the biological functions and molecular mechanisms of DNp73 in MM remain unclear. In this study, we investigated the role of DNp73 in the drug resistance of MM, and disclosed the corresponding mechanism of how DNp73 promotes the immune escape of MM cells. Methods: We constructed DNp73 overexpression and sh-RNA interference plasmids to obtain stable cell lines. The effects of DNp73 on proliferation and drug sensitivity were determined by flow cytometry and xenotransplantation model. RNA-seq and CHIP-seq were performed to detect the mechanisms of drug resistance in MM cells. The DNA damage repair and invasion ability of MM cells were detected by immunofluorescence and transwell assay. To validate the role of DNp73 in immune escape, we performed phagocytosis assays. Results: Our previous study reported that miR-15a was downregulated in MM cells and correlated with the inferior outcome of MM patients. Further analysis demonstrated that DNp73 was a direct target of p65, loss of miR-15a led to the activation of NF-κB-p65 pathway in MM cells. In addition, the level of miR-15a was negatively correlated with the expression of DNp73 in MM cells of patients (r = −0.672, p < 0.05). After DNp73 was down-regulated, cell proliferation and drug resistance were effectively inhibited. Further in vivo study indicated the tumor volume in DNp73 knockdown group was reduced, and increased sensitivity to the carfilzomib, epirubicin and pomalidomide was observed (p < 0.01). Of note, RNA-seq analysis indicated that DNp73 expression was positively correlated with MYCN, MYC and CDK7 transcriptional programs in MM. GSEA analysis showed that MYC targets and DNA damage repair pathways were significantly enriched in DNp73-OE cells. Compared to control group, DNp73-OE cells were resistant to irradiation-induced cell death (p < 0.01). Meanwhile, DNp73 knockdown significantly reduced the migration and invasion of cells (p < 0.01). CHIP-seq data showed that DNp73 could bind to the promoter region of MYCN. DNp73 overexpression protects MM cells from phagocytosis, treated with anti-human CD47 antibody increased phagocytosis of macrophages (p < 0.01). Conclusions: We demonstrated that miR-15a downregulation activated NF-κB, which promoted DNp73 expression at transcription level. DNp73 promotes drug resistance and immune escape in multiple myeloma by modulating Myc and N-Myc signal pathway. The research was funded by: the CAMS Innovation Fund for Medical Sciences (2022-I2M-1-022), the Natural Science Foundation of China (82170194), the Non-profit Central Research Institute Fund of the Chinese Academy of Medical Sciences (2018RC320012), the Natural Science Foundation of China (81920108006), the Non-profit Central Research Institute Fund of the Chinese Academy of Medical Sciences (2018PT31006) Keywords: multiple myeloma, tumor biology and heterogeneity No conflicts of interests pertinent to the abstract.

Characterization and mechanisms of radioresistant lung squamous cell carcinoma cell lines.

Radiotherapy is an important clinical treatment for patients with lung squamous cell carcinoma (LUSC), and resistance to radiotherapy is an important cause of recurrence and metastasis in LUSC. The aim of this study was to establish and explore the biological characteristics of radioresistant LUSC cells. The LUSC cell lines NCI-H2170 and NCI-H520 were irradiated (4 Gy × 15Fraction). Radiosensitivity, cell apoptosis, cell cycle, and DNA damage repair were measured by clonogenic survival assay, flow cytometry, immunofluorescence for γ-H2AX foci, and Comet assay, respectively. Activation of p-ATM(Ser1981), p-CHK2(Th68), p-DNA-PKcs (Ser2056), and Ku70/Ku80 was measured by western blot. Proteomics was used to explore the differential genes and enriched signaling pathways between radioresistant cell lines and parental lines. In vivo nude mouse xenograft experiments further verified the feasibility of the radioresistant LUSC cell lines. After fractionated irradiation (total dose of 60 Gy), radioresistant cells had decreased radiosensitivity, increased G0/G1 phase arrest, enhanced DNA damage repair ability, and through the ATM/CHK2 and DNA-PKcs/Ku70 pathways regulated double strands break. The upregulated differential genes in radioresistant cell lines were mainly enriched in biological pathways such as cell migration and extracellular matrix (ECM)-receptor interaction. In vivo verification of decreased radiosensitivity of radioresistant cells CONCLUSIONS: Radioresistant LUSC cell lines were established by fractional radiotherapy, which regulates IR-induced DNA damage repair through ATM/CHK2 and DNA-PKcs/Ku70. Tandem Mass Tags (TMT) quantitative proteomics found that the biological process pathway of cell migration and ECM-receptor interaction are upregulated in LUSC radioresistant cells.

FLI1 regulates radiotherapy resistance in nasopharyngeal carcinoma through TIE1-mediated PI3K/AKT signaling pathway

BackgroundRadiotherapy resistance is the main cause of treatment failure in nasopharyngeal carcinoma (NPC), which leads to poor prognosis. It is urgent to elucidate the molecular mechanisms underlying radiotherapy resistance.MethodsRNA-seq analysis was applied to five paired progressive disease (PD) and complete response (CR) NPC tissues. Loss-and gain-of-function assays were used for oncogenic function of FLI1 both in vitro and in vivo. RNA-seq analysis, ChIP assays and dual luciferase reporter assays were performed to explore the interaction between FLI1 and TIE1. Gene expression with clinical information from tissue microarray of NPC were analyzed for associations between FLI1/TIE1 expression and NPC prognosis.ResultsFLI1 is a potential radiosensitivity regulator which was dramatically overexpressed in the patients with PD to radiotherapy compared to those with CR. FLI1 induced radiotherapy resistance and enhanced the ability of DNA damage repair in vitro, and promoted radiotherapy resistance in vivo. Mechanistic investigations showed that FLI1 upregulated the transcription of TIE1 by binding to its promoter, thus activated the PI3K/AKT signaling pathway. A decrease in TIE1 expression restored radiosensitivity of NPC cells. Furthermore, NPC patients with high levels of FLI1 and TIE1 were correlated with poor prognosis.ConclusionOur study has revealed that FLI1 regulates radiotherapy resistance of NPC through TIE1-mediated PI3K/AKT signaling pathway, suggesting that targeting the FLI1/TIE1 signaling pathway could be a potential therapeutic strategy to enhance the efficacy of radiotherapy in NPC.

Establishment of Human Lung Adenocarcinoma Radioresistant Cell Lines and the Mechanism of Radioresistance

Radiotherapy is one of the most common treatments for lung cancer, and about 40%-50% of patients after radiotherapy will appear uncontrolled or recurrence in the case of local tumors. Radioresistance is the predominant cause of local therapeutic failure. Nevertheless, the lack of in vitro radioresistance models is an influential factor obstructing the study of its mechanism. Therefore, the establishment of radioresistant cell lines, H1975DR and H1299DR, was beneficial to explore the mechanism of radioresistance in lung adenocarcinoma. The radioresistant cell lines of H1975DR and H1299DR were obtained from H1975 and H1299 cells irradiated with equal doses of X-rays; Clonogenic assays were performed to compare the clone-forming ability of H1975 vs H1975DR cells, H1299 vs H1299DR cells, then fitting cell survival curve by linear quadratic model; The comet assay was employed to examine DNA damage repair and calculate the percentage of DNA tails; The optical microscopy, CCK-8, flow cytometry, Transwell invasion assays were used to compare biological characteristics such as cell morphology, cell proliferation, apoptosis level, cycle distribution, cell migration and invasion; Western blot was carried out to measure the protein expression of DNA damage repair factors, such as DNA-PKcs, 53BP1, RAD51, and p-ATM. After five months of continuous irradiation and stable culture, radioresistant cell lines H1975DR and H1299DR were obtained. The cell proliferation activity, clone formation ability and DNA damage repair ability of the two radioresistant cell lines were significantly improved under X-ray irradiation. The proportion of the G2/M phase was markedly decreased and the proportion of the G0/G1 phase was increased. Cell migration and invasion ability were significantly enhanced. Relative expression levels of p-DNA-PKcs (Ser2056), 53BP1 in the nonhomologous end-joining (NHEJ) repair pathway and p-ATM (Ser1981), RAD51 in the homologous recombination (HR) repair pathway were higher than those in H1975 and H1299. H1975 and H1299 cell lines can be able to differentiate into lung adenocarcinoma radioresistant cell lines H1975DR and H1299DR by equal dose fractional irradiation, which provided an in vitro cytological model for the study of radiotherapy resistance mechanism of lung cancer patients.

Epigenetic-Metabolic Interplay in the DNA Damage Response and Therapeutic Resistance of Breast Cancer.

Therapy resistance is imposing a daunting challenge on effective clinical management of breast cancer. Although the development of resistance to drugs is multifaceted, reprogramming of energy metabolism pathways is emerging as a central but heterogenous regulator of this therapeutic challenge. Metabolic heterogeneity in cancer cells is intricately associated with alterations of different signaling networks and activation of DNA damage response pathways. Here we consider how the dynamic metabolic milieu of cancer cells regulates their DNA damage repair ability to ultimately contribute to development of therapy resistance. Diverse epigenetic regulators are crucial in remodeling the metabolic landscape of cancer. This epigenetic-metabolic interplay profoundly affects genomic stability of the cancer cells as well as their resistance to genotoxic therapies. These observations identify defining mechanisms of cancer epigenetics-metabolism-DNA repair axis that can be critical for devising novel, targeted therapeutic approaches that could sensitize cancer cells to conventional treatment strategies.

A lncRNA MEG3 variant enhances telomerase activity by increasing DNA damage repair ability in human liver cancer stem cells

A lncRNA MEG3 variant enhances telomerase activity by increasing DNA damage repair ability in human liver cancer stem cells

Insufficient Dose of ERCC8 Protein Caused by a Frameshift Mutation Is Associated With Keratoconus With Congenital Cataracts.

The purpose of this study was to identify a new candidate gene for keratoconus and congenital cataracts and further investigate its underlying pathogenic mechanisms. This study, using a Chinese family with keratoconus and congenital cataracts, 262 patients with sporadic keratoconus, and 20 patients with sporadic congenital cataract as subjects, used clinical and genetic analysis and in vitro cell experiments to detect genetic mutations and further investigate the underlying pathogenic mechanisms. We found that a novel frameshift mutation of ERCC8 (NM_000082.3: c.394-398del, p. L132Nfs*6) is responsible for familial keratoconus with congenital cataracts. This mutation showed co-segregation with the phenotype in the family. This was revealed in another patient with sporadic keratoconus, absent in the 210 unrelated health controls, and considered to be "disease-causing." ERCC8 was expressed both in the cornea and lens. Through an in vitro cell experiment, we further demonstrated that the mutant proteins of ERCC8 were degraded and could lead to an insufficient dose of the ERCC8 protein. An insufficient dose reduced the DNA damage repair ability of human corneal fibroblast (HTK) and lens epithelial cells (HLEC) treated with hydrogen peroxide, leading to both cells showing higher DNA damage levels. In addition, it decreased cell viability, resulting in decreased collagen expression in HTK and increased apoptosis in HLEC via aberrant activation of the unfolded protein response. All these results suggested that ERCC8 plays an important role in the normal function of corneal stromal and lens epithelial cells. Our study showed that ERCC8 is a new gene associated with keratoconus and congenital cataracts.

Supramolecular Hydrogel-Wrapped Gingival Mesenchymal Stem Cells in Cutaneous Radiation Injury.

Radiation-induced skin wound/dermatitis is one of the common side effects of radiotherapy or interventional radiobiology. Gingiva-derived mesenchymal stem cells (GMSCs) were indicated to have therapeutic potentials in skin diseases. However, stem cells are prone to spread and difficult to stay in the skin for a long time, limiting their curative effects and application. This study investigated the therapeutic efficacy of Nap-GDFDFpDY (pY-Gel) self-assembled peptide hydrogel-encapsulated GMSCs to treat 137Cs γ-radiation-induced skin wounds in mice. The effects were evaluated by skin damage score, hind limb extension measurement and histological and immunohistochemical analysis. In vivo studies showed that pY-Gel self-assembled peptide hydrogel-encapsulated GMSCs could effectively improve wound healing in irradiated skin tissues. In addition, it was found that GMSCs conditioned medium (CM) could promote the proliferation, migration and DNA damage repair ability of skin cells after irradiation in human keratinocyte cell line HaCaT and normal human dermal fibroblasts (HFF). Mechanistically, GMSCs-CM can promote the expression of epidermal growth factor receptor (EGFR), signal transducers and activators of transcription 3 (STAT3) and matrix metalloproteinases (MMPs), suggesting that activation of the EGFR/STAT3 signaling pathway may be involved in the repair of skin cells after exposure to radiations. In conclusion, pY-Gel self-assembled peptide hydrogel-encapsulated GMSCs have a beneficial therapeutic effect on radiation-induced cutaneous injury and may serve as a basis of novel cells therapeutic approach.

Biochemical and Structural Study of RuvC and YqgF from Deinococcus radiodurans.

ABSTRACTDeinococcus radiodurans possesses robust DNA damage response and repair abilities, and this is mainly due to its efficient homologous recombination repair system, which incorporates an uncharacterized Holliday junction (HJ) resolution process. D. radiodurans encodes two putative HJ resolvase (HJR) homologs: RuvC (DrRuvC) and YqgF (DrYqgF). Here, both DrRuvC and DrYqgF were identified as essential proteins for the survival of D. radiodurans. The crystal structures and the biochemical properties of DrRuvC and DrYqgF were also studied. DrRuvC crystallized as a homodimer, while DrYqgF crystallized as a monomer. DrRuvC could preferentially cleave HJ at the consensus 5′-(G/C)TC↓(G/C)-3′ sequence and could prefer using Mn2+ for catalysis in vitro, which would be different from the preferences of the other previously characterized RuvCs. On the other hand, DrYqgF was identified as a Mn2+-dependent RNA 5′–3′ exo/endonuclease with a sequence preference for poly(A) and without any HJR activity.

Inhibition of Ciliogenesis Enhances the Cellular Sensitivity to Temozolomide and Ionizing Radiation in Human Glioblastoma Cells

ObjectiveTo investigate the function of primary cilia in regulating the cellular response to temozolomide (TMZ) and ionizing radiation (IR) in glioblastoma (GBM). MethodsGBM cells were treated with TMZ or X-ray/carbon ion. The primary cilia were examined by immunostaining with Arl13b and γ-tubulin, and the cellular resistance ability was measured by cell viability assay or survival fraction assay. Combining with cilia ablation by IFT88 depletion or chloral hydrate and induction by lithium chloride, the autophagy was measured by acridine orange staining assay. The DNA damage repair ability was estimated by the kinetic curve of γH2AX foci, and the DNA-dependent protein kinase (DNA-PK) activation was detected by immunostaining assay. ResultsPrimary cilia were frequently preserved in GBM, and the induction of ciliogenesis decreased cell proliferation. TMZ and IR promoted ciliogenesis in dose- and time-dependent manners, and the suppression of ciliogenesis significantly enhanced the cellular sensitivity to TMZ and IR. The inhibition of ciliogenesis elevated the lethal effects of TMZ and IR via the impairment of autophagy and DNA damage repair. The interference of ciliogenesis reduced DNA-PK activation, and the knockdown of DNA-PK led to cilium formation and elongation. ConclusionPrimary cilia play a vital role in regulating the cellular sensitivity to TMZ and IR in GBM cells through mediating autophagy and DNA damage repair.

Homologous Recombination Pathway Alternation Predicts Prognosis of Colorectal Cancer With Chemotherapy.

Background: Chemotherapy is the basic treatment for colorectal cancer (CRC). However, colorectal cancer cells often develop resistance to chemotherapy drugs, leading to recurrence and poor prognosis. More and more studies have shown that the Homologous recombination (HR) pathway plays an important role in chemotherapy treatment for tumors. However, the relationship between HR pathway, chemotherapy sensitivity, and the prognosis of CRC patients is still unclear. Methods: We collected 35 samples of CRC patients after chemotherapy treatment from Guangxi Medical University Cancer Hospital, then collected mutation data and clinical prognosis data from the group. We also downloaded Mondaca-CRC, TCGA-CRC cohorts for chemotherapy treatment. Result: We found that HR mutant-type (HR-MUT) patients are less likely to experience tumor metastasis after receiving chemotherapy. Additionally, our univariate and multivariate cox regression models showed that HR-MUT can be used as an independent predictor of the prognosis of chemotherapy for CRC patients. The KM curve showed that patients with HR-MUT CRC had significantly prolonged overall survival (OS) time (log-rank p = 0.017; hazard ratio (HR) = 0.69). Compared to HR mutant-type (HR-WT), HR-MUT has a significantly lower IC50 value with several chemotherapeutic drugs. Pathway enrichment analysis further revealed that the HR-MUT displayed a significantly lower rate of DNA damage repair ability, tumor growth, metastasis activity, and tumor fatty acid metabolism activity than HR-WT, though its immune response activity was notably higher. Conclusion: These findings indicate that HR-MUT may be a relevant marker for CRC patients receiving chemotherapy, as it is closely related to improving OS time and reducing chemotherapy resistance.