Increased DNA Repair Capacity Research Articles

Analysis of heterochromatin remodeling as a mechanism of radiotherapy (RT) resistance through molecular profiling of radioresistant (RR) head and neck (HNC) and prostate cancers (PCa).

137 Background: Resistance to RT remains a clinical challenge, given our limited understanding of the molecular pathways underpinning radioresistance. Here, we investigated for mechanisms linked to RR using paired experimental and clinico-molecular HNC and PCa datasets for discovery and validation of novel pathways that may be enriched in RR cancers. Methods: For experimental data, we generated RR HNC (FaDu, HK1) and PCa (22Rv1, DU145) cell lines following high-dose X-irradiation (90 Gy in 45 fr). Genotypic characterization of RR and wildtype (WT) cells was performed by genomic and transcriptomic sequencing using WES (100X) and RNAseq (Illumina, CA), respectively, followed by functional characterization by western blot (WB) and immunofluorescence (IF). For clinico-molecular data, we utilized two prospectively recruited RT cohorts, consisting of 311 patients with HNC (n=158) and PCa (n=153). Tumor transcriptomes were profiled using RNAseq and Affymetrix ST array (ThermoFisher, CA), respectively. Comparative analyses of molecular profiles were first performed between RR and WT in vitro models to identify dysregulated pathways that were linked to radioresistance, and subsequently tested for association with disease-free survival (DFS) in the clinical cohorts. Results: Comparative genomic analyses between RR and WT HNC and PCa models revealed an abundance of acquired mutations in the RR compared with WT models, with a higher mutation count observed in PCa than HNC cells (SNV counts: 2,158 [DU145-RR] and 1,387 [22Rv1-RR] vs 396 [FaDu-RR] and 25 [HK1-RR], P<0.0001). Mutational signature analyses indicated a common enrichment of DNA mismatch repair-related mutational signatures (SBS3, SBS14, and SBS21) across the RR HNC and PCa models. Transcriptomic profiling revealed an upregulation of the BAHD1 gene, which is involved in heterochromatin formation, across the 4 RR cell lines, corroborated by an enrichment of heterochromatin-related genesets. These results were consistent with functional characterization of RR versus WT cells by WB and IF indicating increased DNA repair capacity and heterochromatin response post-4 Gy irradiation. We further confirmed the dependency of the RR phenotype on the heterochromatin response with BAHD1 knockdown, resulting in reversal of these cellular responses. Finally, our results were supported by survival analyses indicating an inferior DFS post-RT in patients with HNC and PCa harboring a higher expression of heterochromatin-related geneset (HNC: HRhigh vs low 1.51 ; PCa: HRhigh vs low 1.37 ). Conclusions: Herein, by leveraging on paired experimental and clinico-molecular datasets, we have uncovered a novel BAHD1-dependent heterochromatin response that underpins resistance to RT in HNC and PCa, which may be amendable to systemic agents targeting chromatin remodeling.

Inhibition of AR and DNA-PK for radio-sensitization of AR-driven cancers.

e12593 Background: Breast cancer (BC) is the most diagnosed cancer globally with over 2.2 million new cases and 680,000 deaths annually. Radiotherapy (RT) is essential for curative BC treatment; however, 20% of BC cases are radio-resistant, with the underlying mechanisms remaining poorly understood. Previous studies in prostate cancer (PCa) have shown that androgen receptor (AR) promotes radio-resistance (RT-resistance) through transcriptional regulation and increased DNA repair capacity, including through upregulation of DNA repair proteins DNA-PK and PARP1. Although AR is expressed in 60% of BCs, the implications of AR in BC RT-resistance are not yet fully characterized. We hypothesized that AR may drive RT-resistance in BC through a similar AR-DNA repair loop. Methods: To characterize the repertoire of cofactors that cooperate with AR on the TF complex, we exposed a BC cell line to RT and performed rapid immunoprecipitation mass spectrometry of endogenous proteins (RIME) proteomics. We identified several candidate proteins that interact with AR to further investigate as targets for RT-sensitization. Survival analyses in BC and PCa cell lines were performed to identify RT-sensitizing agents. Molecular mechanisms of RT-sensitization were assessed using ATAC-seq and RNA-seq analyses. Results: Using RIME, we identified several DNA repair proteins including DNA-PK, PARP-1, and XRCC5 as differentially recruited to chromatin-bound AR. This suggests a cooperative role and provides potential druggable targets to improve RT-sensitivity. Based on these results, we also performed a survival analysis in AR-driven BC and PCa cell lines and showed that inhibition of either AR or DNA-PK led to RT-sensitization, and dual inhibition provided an additive effect. We sought to characterize the molecular mechanisms driving the RT-sensitizing effects of AR and DNA-PK inhibition and performed ATAC-seq and RNA-seq on treated and untreated cells. Combining enzalutamide with a DNA-PK inhibitor altered expression of several pathways implicated in RT-sensitization, although these pathways were largely distinct in BC and PCa. Further, we identified distinct genes that may be responsible for this phenotype in AR-driven BC. Additionally, we found a larger effect in chromatin rearrangement after treatment in PCa than BC. Conclusions: Identifying mechanisms of RT-resistance in hormone-driven cancers is essential to develop therapeutic strategies and improve patient outcomes. This study provides key evidence for AR-driven RT-resistance in BC and PCa, and identifies DNA-PK inhibition as a potential drug target. Distinct mechanisms appear to be driving this phenotype across BC and PCa, in our models. Understanding the cooperation of DNA repair and hormone signaling pathways will help support the use of DNA-damaging agents for hormone-driven cancers, and potentially inform rational design of novel drug-RT combinations.

Abstract 2875: Resistance to conventional chemo and radiotherapy associated with increased DNA repair and stem cell-like phenotype in head and neck squamous cell carcinoma expressing HPV E5

Abstract Human papillomavirus-associated head and neck squamous cell carcinoma (HPV+ HNSCC) is one of the fastest-rising cancers in the US. While HPV+ HNSCC patients exhibit better responses to chemoradiation compared to HPV- HNSCC patients, a subset of HPV+ patients do not respond as well and develop recurrent/refractory disease. In this study, we investigated the potential role of HPV E5 in promoting resistance to conventional chemo- and radiotherapy, and the underlying molecular mechanisms. HPV E5 is a small hydrophobic viral oncoprotein with multiple functions including regulation of tumor cell differentiation and apoptosis, modulation of H+ ATPase responsible for acidification of late endosomes, and immune modulation. To test the impact of HPV E5 on therapeutic resistance, we generated HNSCC cell lines and normal keratinocytes expressing HPV16 E5. Upon treatment with radiation (2Gy, 4Gy, 8Gy) and/or cisplatin, HNSCC cells and keratinocytes expressing HPV 16 E5 had increased cell viability, decreased apoptosis, and increased colony-forming capacity compared to empty vector (EV) control lines. Specifically, treatment with 4Gy radiation, 4µM cisplatin, or the combination led to a ~2-fold decrease in % of apoptotic cells in HPV16 E5-expressing HNSCC cells compared to EV and HPV16 E6/E7 HNSCC cells (p <0.05). Examination of DNA damage response and repair pathways indicated that while HPV16 E5 HNSCC cells initially had increased phosphorylation of H2AX 6 hours post treatment, at 24 hours post treatment H2AX phosphorylation in HPV16 E5 cells was significantly decreased compared to EV or HPV16 E6/E7 HNSCC cells. These findings indicate increased DNA repair capacity of HPV16 E5 HNSCC cells. To identify mechanisms underlying this phenotype, we performed RNA-seq and observed enhanced signaling pathways associated with epithelial development and increased expression of stem cell markers in HPV16 E5 HNSCC cells compared to EV control lines. Functionally, we observed a ~2.5-fold increase in sphere-forming capacity and increased expression of markers associated with stem cell phenotype in HPV16 E5 HNSCC cells compared to EV and HPV E6/E7 HNSCC cells. To determine whether these findings translated clinically, 18 HPV+ HNSCC patients from the GSE74927 dataset were grouped according to median expression level of HPV E5 (High vs Low). In support of our hypothesis, HPV E5-High patients exhibited enrichment of pathways associated with extrinsic apoptotic signaling, pluripotency of stem cells, and positive regulation of DNA repair. Taken together these results identify HPV E5 as a potential mediator of resistance to conventional radiation and chemotherapy in HNSCC and suggest that patient tumors expressing HPV E5 may have increased DNA repair capacity and cell survival after cytotoxic therapies. Citation Format: Souvick Roy, Sayuri Miyauchi, Riley Jones, Sangwoo Kim, Andrew B. Sharabi. Resistance to conventional chemo and radiotherapy associated with increased DNA repair and stem cell-like phenotype in head and neck squamous cell carcinoma expressing HPV E5 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2875.

EXPRESSION OF NUCLEOTIDE EXCISION REPAIR PROTEIN ERCC1 IN TUMOR TISSUE AS A PROGNOSTIC FACTOR IN COLORECTAL CANCER

At present, biomarkers of tumor resistance to platinum drugs in colorectal cancer are of much concern. Relapses and metastases occur in 35 % of patients with colorectal cancer (stages I–III) after adjuvant polychemotherapy with platinum-based anticancer drugs within the first 5 years. Resistance to platinum-based drugs is determined by a number of factors, including increased DNA repair capacity.
 The aim of the study is to evaluate the prognostic role of ERCC1 expression in tumor tissue in colorectal cancer.
 Materials and Methods. The authors studied the expression of the nucleotide excision repair protein ERCC1 in tumor tissue as a prognostic marker for adjuvant chemotherapy FOLFOX/XELOX in patients with colorectal cancer (stages II–IV). The correlation between ERCC1 expression in tumor tissue and chemotherapy effectiveness, clinical stage, age, survival, differentiation, and EGFR pathway mutations (NRAS, KRAS, BRAF) was assessed.
 Results. ERCC1 expression was detected in 46.9 % of tumor tissue samples and in 88.9 % of resection line tissue samples. In patients with ERCC1 overexpression, the median survival was significantly lower than in patients with low ERCC1 expression. In tumors with NRAS and BRAF mutations, no ERCC1 expression was observed. No differences were determined in patient according to the disease stage, tumor differentiation, and mutations.
 Cancer tumors is an independent predictor of FOLFOX/XELOX chemotherapy resistance.

Excessive activation of HOXB13/PIMREG axis promotes hepatocellular carcinoma progression and drug resistance

The transcription factor HOXB13 is bound up with the occurrence, progression and drug fast of many kinds of cancer. Nevertheless, the specific molecular mechanism of HOXB13 in hepatocellular carcinoma (HCC) is still unknown. This provides an obstacle to the exploration of HCC treatments targeting HOXB13. This study found that HOXB13 was up-regulated in HCC tissues. HOXB13 enhanced the multiplication and metastasis of HCC cells. It enhanced HCC cell drug and anoikis resistance. The analysis of HCC RNA seq data indicated that the expression of HOXB13 and PIMREG were positively correlated. Luciferase report assay showed that HOXB13 could activate PIMREG promoter transcription. The results of RT-qPCR and western blot showed that HOXB13 regulated the transcription of PIMREG. Western blot proved that high expression of PIMREG participated in DNA damage repair and cell cycle regulation by up-regulating RAD51, BRCA1, CDC25A, CDC25B and CDC25C and down-regulating HIPK2. This led to a significant increase in DNA repair capacity, accelerated cell cycle progression, and insensitive to DNA damage. Down-regulation of PIMREG in Hep3B cells overexpressing HOXB13 attenuated the phenotype induced by HOXB13. Therefore, HOXB13 functioned through PIMREG instead of directly regulating the transcription of RAD51, BRCA1, CDC25A, CDC25B and CDC25C. The same results were obtained in vivo. It was concluded that HOXB13 affected the expression of cell cycle and DNA repair related factors by up-regulating the transcription of PIMREG, thereby promoting the progression of HCC and enhancing the resistance of HCC to chemotherapeutics.

YTHDF1 promotes breast cancer cell growth, DNA damage repair and chemoresistance

Chemoresistance represents a major obstacle to the treatment of human cancers. Increased DNA repair capacity is one of the important mechanisms underlying chemoresistance. In silico analysis indicated that YTHDF1, an m6A binding protein, is a putative tumor promoter in breast cancer. Loss of function studies further showed that YTHDF1 promotes breast cancer cell growth in vitro and in vivo. YTHDF1 facilitates S-phase entry, DNA replication and DNA damage repair, and accordingly YTHDF1 knockdown sensitizes breast cancer cells to Adriamycin and Cisplatin as well as Olaparib, a PARP inhibitor. E2F8 is a target molecule by YTHDF1 which modulates E2F8 mRNA stability and DNA damage repair in a METTL14-dependent manner. These data demonstrate that YTHDF1 has a tumor-promoting role in breast cancer, and is a novel target to overcome chemoresistance.

Efficient DNA Repair Mitigates Replication Stress Resulting in Less Immunogenic Cytosolic DNA in Radioresistant Breast Cancer Stem Cells

Cancer stem cells (CSCs) are a major cause of tumor therapy failure. This is mainly attributed to increased DNA repair capacity and immune escape. Recent studies have shown that functional DNA repair via homologous recombination (HR) prevents radiation-induced accumulation of DNA in the cytoplasm, thereby inhibiting the intracellular immune response. However, it is unclear whether CSCs can suppress radiation-induced cytoplasmic dsDNA formation. Here, we show that the increased radioresistance of ALDH1-positive breast cancer stem cells (BCSCs) in S phase is mediated by both enhanced DNA double-strand break repair and improved replication fork protection due to HR. Both HR-mediated processes lead to suppression of radiation-induced replication stress and consequently reduction of cytoplasmic dsDNA. The amount of cytoplasmic dsDNA correlated significantly with BCSC content (p=0.0002). This clearly indicates that HR-dependent avoidance of radiation-induced replication stress mediates radioresistance and contributes to its immune evasion. Consistent with this, enhancement of replication stress by inhibition of ataxia telangiectasia and RAD3 related (ATR) resulted in significant radiosensitization (SER37 increase 1.7-2.8 Gy, p<0.0001). Therefore, disruption of HR-mediated processes, particularly in replication, opens a CSC-specific radiosensitization option by enhancing their intracellular immune response.

A Global Analysis of Photoreceptor-Mediated Transcriptional Changes Reveals the Intricate Relationship Between Central Metabolism and DNA Repair in the Filamentous Fungus Trichoderma atroviride.

Light provides critical information for the behavior and development of basically all organisms. Filamentous fungi sense blue light, mainly, through a unique transcription factor complex that activates its targets in a light-dependent manner. In Trichoderma atroviride, the BLR-1 and BLR-2 proteins constitute this complex, which triggers the light-dependent formation of asexual reproduction structures (conidia). We generated an ENVOY photoreceptor mutant and performed RNA-seq analyses in the mutants of this gene and in those of the BLR-1, CRY-1 and CRY-DASH photoreceptors in response to a pulse of low intensity blue light. Like in other filamentous fungi BLR-1 appears to play a central role in the regulation of blue-light responses. Phenotypic characterization of the Δenv-1 mutant showed that ENVOY functions as a growth and conidiation checkpoint, preventing exacerbated light responses. Similarly, we observed that CRY-1 and CRY-DASH contribute to the typical light-induced conidiation response. In the Δenv-1 mutant, we observed, at the transcriptomic level, a general induction of DNA metabolic processes and strong repression of central metabolism. An analysis of the expression level of DNA repair genes showed that they increase their expression in the absence of env-1. Consistently, photoreactivation experiments showed that Δenv-1 had increased DNA repair capacity. Our results indicate that light perception in T. atroviride is far more complex than originally thought.

Increased DNA repair capacity augments resistance of glioblastoma cells to photodynamic therapy

Photodynamic therapy (PDT) is a clinically approved cancer therapy of low invasiveness. The therapeutic procedure involves administering a photosensitizing drug (PS), which is then activated with monochromatic light of a specific wavelength. The photochemical reaction produces highly toxic oxygen species. The development of resistance to PDT in some cancer cells is its main limitation. Several mechanisms are known to be involved in the development of cellular defense against cytotoxic effects of PDT, including activation of antioxidant enzymes, drug efflux pumps, degradation of PS, and overexpression of protein chaperons. Another putative factor that plays an important role in the development of resistance of cancer cells to PDT seems to be DNA repair; however, it has not been well studied so far. To explore the role of DNA repair and other potential novel mechanisms associated with the resistance to PDT in the glioblastoma cells, cells stably resistant to PDT were isolated from PDT sensitive cells following repetitive PDT cycles. Duly characterization of isolated PDT-resistant glioblastoma revealed that the resistance to PDT might be a consequence of several mechanisms, including higher repair efficiency of oxidative DNA damage and repair of DNA breaks. Higher activity of APE1 endonuclease and increased expression and activation of DNA damage kinase ATM was demonstrated in the U-87 MGR cell line, suggesting and proving that they are good targets for sensitization of resistant cells to PDT.

Impact of DNA repair and reactive oxygen species levels on radioresistance in pancreatic cancer

PurposeRadioresistance in pancreatic cancer patients remains a critical obstacle to overcome. Understanding the molecular mechanisms underlying radioresistance may achieve better response to radiotherapy and thereby improving the poor treatment outcome. The aim of the present study was to elucidate the mechanisms leading to radioresistance by detailed characterization of isogenic radioresistant and radiosensitive cell lines. MethodsThe human pancreatic cancer cell lines, Panc-1 and MIA PaCa-2 were repeatedly exposed to radiation to generate radioresistant (RR) isogenic cell lines. The surviving cells were expanded, and their radiosensitivity was measured using colony formation assay. Tumor growth delay after irradiation was determined in a mouse pancreatic cancer xenograft model. Gene and protein expression were analyzed using RNA sequencing and Western blot, respectively. Cell cycle distribution and apoptosis (Caspase 3/7) were measured by FACS analysis. Reactive oxygen species generation and DNA damage were analyzed by detection of CM-H2DCFDA and γH2AX staining, respectively. Transwell chamber assays were used to investigate cell migration and invasion. ResultsThe acquired radioresistance of RR cell lines was demonstrated in vitro and validated in vivo. Ingenuity pathway analysis of RNA sequencing data predicted activation of cell viability in both RR cell lines. RR cancer cell lines demonstrated greater DNA repair efficiency and lower basal and radiation-induced reactive oxygen species levels. Migration and invasion were differentially affected in RR cell lines. ConclusionsOur data indicate that repeated exposure to irradiation increases the expression of genes involved in cell viability and thereby leads to radioresistance. Mechanistically, increased DNA repair capacity and reduced oxidative stress might contribute to the radioresistant phenotype.

RAD50 deficiency is a predictor of platinum sensitivity in sporadic epithelial ovarian cancers

Intrinsic or acquired resistance seriously limits the use of platinating agents in advanced epithelial ovarian cancers. Increased DNA repair capacity is a key route to platinum resistance. RAD50 is a critical component of the MRN complex, a ‘first responder’ to DNA damage and essential for the repair of DSBs and stalled replication forks. We hypothesised a role for RAD50 in ovarian cancer pathogenesis and therapeutics. Clinicopathological significance of RAD50 expression was evaluated in clinical cohorts of ovarian cancer at the protein level (n = 331) and at the transcriptomic level (n = 1259). Sub-cellular localization of RAD50 at baseline and following cisplatin therapy was tested in platinum resistant (A2780cis, PEO4) and sensitive (A2780, PEO1) ovarian cancer cells. RAD50 was depleted and cisplatin sensitivity was investigated in A2780cis and PEO4 cells. RAD50 deficiency was associated with better progression free survival (PFS) at the protein (p = 0.006) and transcriptomic level (p < 0.001). Basal level of RAD50 was higher in platinum resistant cells. Following cisplatin treatment, increased nuclear localization of RAD50 was evident in A2780cis and PEO4 compared to A2780 and PEO1 cells. RAD50 depletion using siRNAs in A2780cis and PEO4 cells increased cisplatin cytotoxicity, which was associated with accumulation of DSBs, S-phase cell cycle arrest and increased apoptosis. We provide evidence that RAD50 deficiency is a predictor of platinum sensitivity. RAD50 expression-based stratification and personalization could be viable clinical strategy in ovarian cancers.

Clinical Perspectives of ERCC1 in Bladder Cancer.

ERCC1 is a key regulator of nucleotide excision repair (NER) pathway that repairs bulky DNA adducts, including intrastrand DNA adducts and interstrand crosslinks (ICLs). Overexpression of ERCC1 has been linked to increased DNA repair capacity and platinum resistance in solid tumors. Multiple single nucleotide polymorphisms (SNPs) have been detected in ERCC1 gene that may affect ERCC1 protein expression. Platinum-based treatment remains the cornerstone of urothelial cancer treatment. Given the expanding application of neoadjuvant and adjuvant chemotherapy in locally advanced bladder cancer, there is an emerging need for biomarkers that could distinguish potential responders to cisplatin treatment. Extensive research has been done regarding the prognostic and predictive role of ERCC1 gene expression and polymorphisms in bladder cancer. Moreover, novel compounds have been recently developed to target ERCC1 protein function in order to maximize sensitivity to cisplatin. We aim to review all the existing literature regarding the role of the ERCC1 gene in bladder cancer and address future perspectives for its clinical application.

The E3 Ubiquitin Ligase NEDD4L Targets OGG1 for Ubiquitylation and Modulates the Cellular DNA Damage Response.

8-Oxoguanine DNA glycosylase (OGG1) is the major cellular enzyme required for the excision of 8-oxoguanine DNA base lesions in DNA through the base excision repair (BER) pathway, and therefore plays a major role in suppressing mutagenesis and in controlling genome stability. However, the mechanism of regulation of cellular OGG1 protein, particularly in response to oxidative stress, is unclear. We have purified the major E3 ubiquitin ligase responsible for OGG1 ubiquitylation from human cell extracts, and identify this as E3 ubiquitin-protein ligase NEDD4-like (NEDD4L). We demonstrate that recombinant NEDD4L stimulates ubiquitylation of OGG1 in vitro, particularly on lysine 341, and that NEDD4L and OGG1 interact in U2OS cells. Depletion of NEDD4L in U2OS cells has no impact on the stability and steady-state protein levels of OGG1, however, OGG1 stability is enhanced in response to oxidative stress induced by ionizing radiation. Furthermore, ubiquitylation of OGG1 by NEDD4L in vitro inhibits its DNA glycosylase/lyase activity. As a consequence of prolonged OGG1 stability and increased excision activity in the absence of NEDD4L, cells display increased DNA repair capacity but conversely that this decreases cell survival post-irradiation. This effect can be reproduced following OGG1 overexpression, suggesting that dysregulation of OGG1 increases the formation of lethal intermediate DNA lesions. Our study therefore highlights the importance of balancing OGG1 protein levels and BER capacity in maintaining genome stability.

Predictive and Prognostic Value of DNA Damage Response Associated Kinases in Solid Tumors.

Dysfunctional DNA repair with subsequent genome instability and high mutational burden represents a major hallmark of cancer. In established malignant tumors, increased DNA repair capacity mediates resistance to DNA-damaging therapeutics, including cytotoxic drugs, radiotherapy, and selected small molecules including inhibitors of poly (ADP-ribose) polymerase (PARP), Ataxia Telangiectasia Mutated (ATM), ataxia telangiectasia and Rad3-related protein (ATR), and Wee1 kinase (Wee1). In addition, DNA repair deficiency is not only associated with sensitivity to selected anticancer drugs, but also with increased mutagenicity and increased neoantigen load on tumor cells, resulting in increased immunogenicity and improved response to CTLA4- or PD-(L)1 targeting monoclonal antibodies. DNA damage response (DDR) is composed of complex signalling pathways, including the sensing of the DNA damage, signal transduction, cellular response pathways to DNA damage, and activation of DNA repair. DNA double strand breaks (DSBs) are the most dangerous form of DNA damage. Tumor cells are characterised by frequent accumulation of DSBs caused by either endogenous replication stress or the impact of cancer treatment, most prominently chemotherapy and radiotherapy. Therefore, response of cancer cells to DSBs represents a crucial mechanism for how tumors respond to systemic treatment or radiotherapy, and how resistance develops. Ample clinical evidence supports the importance of DDR associated kinases as predictive and prognostic biomarkers in cancer patients. The ATM-CHK2 and ATR-CHK1-WEE1 pathways initiate DNA DSB repair. In the current review, we focus on major DDR associated kinases including ATM, ATR, CHK1, CHK2, and WEE1, and discuss their potential prognostic and predictive value in solid malignancies.

ALDH-1-positive cells exhibited a radioresistant phenotype that was enhanced with hypoxia in cervical cancer

BackgroundWe have previously found there was a small subpopulation of cells with cancer stem cell-like phenotype ALDH-1 in cervical cancer. Radiotherapy has been applied in most of the cervical cancer. However,the mechanisms underlying radioresistance still remained elusive. Our study is to explore whether ALDH+ cell promotes radioresistance by hypoxia.MethodsCells were respectively cultured in hypoxia and normoxia environment and analyzed for marker stability, and cell cycle distribution. Results: Cell growth, apoptosis, cell cycle, sphere formation were affected by hypoxia. ALDH-1 and CHK2 were upregulated after hypoxia.ConclusionsHere we show that ALDH-1 positive cells contribute to cervical carcinoma radioresistance through preferential activation of the DNA damage checkpoint response and an increase in DNA repair capacity. The fraction of these cells is enriched after radiation in cervical carcinoma.

Glioblastomas located in proximity to the subventricular zone (SVZ) exhibited enrichment of gene expression profiles associated with the cancer stem cell state.

Conflicting results have been reported in the association between glioblastoma proximity to the subventricular zone (SVZ) and enrichment of cancer stem cell properties. Here, we examined this hypothesis using magnetic resonance (MR) images derived from 217 The Cancer Imaging Archive (TCIA) glioblastoma subjects. Pre-operative MR images were segmented automatically into contrast enhancing (CE) tumor volumes using Iterative Probabilistic Voxel Labeling (IPVL). Distances were calculated from the centroid of CE tumor volumes to the SVZ and correlated with gene expression profiles of the corresponding glioblastomas. Correlative analyses were performed between SVZ distance, gene expression patterns, and clinical survival. Glioblastoma located in proximity to the SVZ showed increased mRNA expression patterns associated with the cancer stem-cell state, including CD133 (P = 0.006). Consistent with the previous observations suggesting that glioblastoma stem cells exhibit increased DNA repair capacity, glioblastomas in proximity to the SVZ also showed increased expression of DNA repair genes, including MGMT (P = 0.018). Reflecting this enhanced DNA repair capacity, the genomes of glioblastomas in SVZ proximity harbored fewer single nucleotide polymorphisms relative to those located distant to the SVZ (P = 0.003). Concordant with the notion that glioblastoma stem cells are more aggressive and refractory to therapy, patients with glioblastoma in proximity to SVZ exhibited poorer progression free and overall survival (P < 0.01). An unbiased analysis of TCIA suggests that glioblastomas located in proximity to the SVZ exhibited mRNA expression profiles associated with stem cell properties, increased DNA repair capacity, and is associated with poor clinical survival.

Mechanisms of Multidrug Resistance in Cancer Chemotherapy.

Cancer is one of the main causes of death worldwide. Despite the significant development of methods of cancer healing during the past decades, chemotherapy still remains the main method for cancer treatment. Depending on the mechanism of action, commonly used chemotherapeutic agents can be divided into several classes (antimetabolites, alkylating agents, mitotic spindle inhibitors, topoisomerase inhibitors, and others). Multidrug resistance (MDR) is responsible for over 90% of deaths in cancer patients receiving traditional chemotherapeutics or novel targeted drugs. The mechanisms of MDR include elevated metabolism of xenobiotics, enhanced efflux of drugs, growth factors, increased DNA repair capacity, and genetic factors (gene mutations, amplifications, and epigenetic alterations). Rapidly increasing numbers of biomedical studies are focused on designing chemotherapeutics that are able to evade or reverse MDR. The aim of this review is not only to demonstrate the latest data on the mechanisms of cellular resistance to anticancer agents currently used in clinical treatment but also to present the mechanisms of action of novel potential antitumor drugs which have been designed to overcome these resistance mechanisms. Better understanding of the mechanisms of MDR and targets of novel chemotherapy agents should provide guidance for future research concerning new effective strategies in cancer treatment.

Enhanced DNA-repair capacity and resistance to chemically induced carcinogenesis upon deletion of the phosphatase regulator NIPP1

Nuclear Inhibitor of PP1 (NIPP1) is a conserved regulatory subunit of protein phosphatase PP1. The selective deletion of NIPP1 in mouse liver parenchymal cells or skin epidermal cells culminates in a late-onset hyperproliferation of a subset of resident progenitor cells. Although a hyperplastic phenotype is usually tumor promoting, we show here that the absence of NIPP1 conferred a strong resistance to chemically induced hepatocellular or skin carcinoma. The ablation of NIPP1 did not affect the metabolism of the administered mutagens (diethylnitrosamine or 7,12-dimethylbenz[a]anthracene), but reduced the conversion of mutagen-induced covalent DNA modifications into cancer-initiating mutations. This reduced sensitivity to mutagens correlated with an enhanced DNA-damage response and an augmented expression of rate-limiting DNA-repair proteins (MGMT in liver, XPD and XPG in skin), hinting at an increased DNA-repair capacity. Our data identify NIPP1 as a repressor of DNA repair and as a promising target for novel cancer prevention and treatment therapies.

MicroRNA-140 impedes DNA repair by targeting FEN1 and enhances chemotherapeutic response in breast cancer.

An increased DNA repair capacity is associated with drug resistance and limits the efficacy of chemotherapy in breast cancers. Flap endonuclease 1 (FEN1) participates in various DNA repair pathways and contributes to cancer progression and drug resistance in chemotherapy. Inhibition of FEN1 serves as a potent strategy for cancer therapy. Here, we demonstrate that microRNA-140 (miR-140) inhibits FEN1 expression via directly binding to its 3' untranslated region, leading to impaired DNA repair and repressed breast cancer progression. Overexpression of miR-140 sensitizes breast cancer cells to chemotherapeutic agents and overcomes drug resistance in breast cancer. Notably, ectopic expression of FEN1 abates the effects of miR-140 on DNA damage and the chemotherapy response in breast cancer cells. Furthermore, the transcription factor/repressor Ying Yang 1 (YY1) directly binds to the miR-140 promoter and activates miR-140 expression, which is attenuated in doxorubicin resistance. Our results demonstrate that miR-140 acts as a tumor suppressor in breast cancer by inhibiting FEN1 to repress DNA damage repair and reveal miR-140 to be a new anti-tumorigenesis factor for adjunctive breast cancer therapy. This novel mechanism will enhance the treatment effect of chemotherapy in breast cancer.

Combined treatment with cisplatin and the tankyrase inhibitor XAV-939 increases cytotoxicity, abrogates cancer-stem-like cell phenotype and increases chemosensitivity of head-and-neck squamous-cell carcinoma cells

Cancer stem-like cells (CSCs) were reported to be linked with tumorigenesis, metastasis and resistant to chemo and radiotherapy in head and neck squamous cell carcinoma (HNSCC). In this study we investigated the role of CSCs in chemoresistance and abrogation of CSC mediated chemoresistance by combinatorial treatment with cisplatin and small molecule tankyrase inhibitor XAV-939. Two cisplatin-resistant HNSCC cells were generated by stepwise dose incremental strategy. We evaluated the chemoresistance, sphere forming capacity, extent of DNA damage and repair capacity in parental and cisplatin-resistant HNSCC cells. Furthermore, the abrogation of CSC mediated chemoresistance was evaluated in HNSCC cells with XAV-939 alone and in combination with cisplatin. It was observed that cisplatin-resistant HNSCC cell lines exhibited increase in chemoresistance, CSC phenotype and increased DNA repair capacity. We observed that combination of cisplatin and XAV-939 acts synergistically to abrogate chemoresistance by increasing DNA damage. Molecular docking study also revealed similar binding region that could contribute towards synergy predictions between cisplatin and XAV939. In conclusion, this study elucidated that combination of cisplatin and XAV-939 exerted cytotoxic and genotoxic effect to abrogate CSC mediated chemoresistance in HNSCC in synergistic manner.