Radioresistance Of Colorectal Cancer Research Articles

MicroRNA-298 determines the radio-resistance of colorectal cancer cells by directly targeting human dual-specificity tyrosine(Y)-regulated kinase 1A.

Radiotherapy stands as a promising therapeutic modality for colorectal cancer (CRC); yet, the formidable challenge posed by radio-resistance significantly undermines its efficacy in achieving CRC remission. To elucidate the role played by microRNA-298 (miR-298) in CRC radio-resistance. To establish a radio-resistant CRC cell line, HT-29 cells underwent exposure to 5 gray ionizing radiation that was followed by a 7-d recovery period. The quantification of miR-298 levels within CRC cells was conducted through quantitative RT-PCR, and protein expression determination was realized through Western blotting. Cell viability was assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and proliferation by clonogenic assay. Radio-induced apoptosis was discerned through flow cytometry analysis. We observed a marked upregulation of miR-298 in radio-resistant CRC cells. MiR-298 emerged as a key determinant of cell survival following radiation exposure, as its overexpression led to a notable reduction in radiation-induced apoptosis. Intriguingly, miR-298 expression exhibited a strong correlation with CRC cell viability. Further investigation unveiled human dual-specificity tyrosine(Y)-regulated kinase 1A (DYRK1A) as miR-298's direct target. Taken together, our findings underline the role played by miR-298 in bolstering radio-resistance in CRC cells by means of DYRK1A downregulation, thereby positioning miR-298 as a promising candidate for mitigating radio-resistance in CRC.

The mir-199b-5p encapsulated in adipocyte-derived exosomes mediates radioresistance of colorectal cancer cells by targeting JAG1

Radiotherapy is a key treatment option for colorectal cancer, but its efficacy varies among patients. Our previous studies suggested that adipose tissue may confer the radioresistance of several abdominal tumors, such as pancreatic cancer, biliary cancer, and others. In the present work, the effects of adipocytes in regulating the radiosensitivity of colorectal cancer are explored for the first time. It was found that colony formation was increased and radiation-induced apoptosis decreased in colorectal cancer cells HCT8 and HCT116 co-cultured with adipocytes, which verified the mediation of adipocyte-driven radioresistance in colorectal cancer in vitro. Next, the colorectal cancer cells were incubated with adipocyte-derived exosomes, and a perceptible reduction in radiosensitivity was detected. Furthermore, to investigate the possible mechanisms involved, the exosomes were isolated, the encapsulated microRNAs were extracted and analyzed by small RNA sequencing. Based on bioinformatics analysis and qRT-PCR verification, miR-199b-5p was chosen for functional annotation. It was shown that miR-199b-5p expression was significantly upregulated after 6 Gy irradiation, and overexpressed miR-199b-5p significantly suppressed the radiosensitivity of HCT8 and HCT116 cells. In addition, jagged canonical Notch ligand 1(JAG1) was identified as the target gene of miR-199b-5p by using bioinformatics prediction and dual luciferase reporter gene assay. It was demonstrated that JAG1 conferred the radioresistance of colorectal cancer cells both in vivo and in vitro. Taken together, the present study demonstrates that adipocytes trigger the radioresistance of colorectal cancer cells, probably by targeting JAG1 through an adipocyte-derived exosomal miR-199b-5p.

SUMO-Activating Enzyme Subunit 1 Is Associated with Poor Prognosis, Tumor Progression, and Radio-Resistance in Colorectal Cancer.

Concurrent chemoradiotherapy is an effective treatment option for patients with low-grade colorectal cancer (CRC) in the local disease stage. At present, the principle of the Taiwan Medical Center is to treat CRC patients with combination radiotherapy and chemotherapy (high-dose 5-FU) for a period of about five weeks prior to surgery. Radical resection of the tumor is performed at least six to eight weeks after concurrent chemoradiotherapy (CCRT). However, this approach fails to produce the desired therapeutic effect in approximately 20% to 30% of patients, and such patients are unnecessarily exposed to the risks of radiation and drug toxicity posed by this therapy. Therefore, it is crucial to explore new biomarkers to predict the prognosis of CRC. SUMO-activating enzyme subunit 1 (SAE1) plays an important role in SUMOylation, a post-translational modification involved in cellular functions, such as cell proliferation, cell cycle, and apoptosis. In our study, to explore the clinical-pathological role of SAE1 protein in CRC, we evaluated the clinical data and paraffin sections from CRC patients. The expression of SAE1 was evaluated using immunohistochemical analysis, and clinical parameters were analyzed using chi-square and Kaplan-Meier survival tests. The results of in vitro proliferation and radiosensitive assays were compared between control groups and SAE1 siRNA groups. Western blotting was also used to detect the expressions of the SAE1, PARP, cyclin D1, p-NF-κB, and NF-κB proteins. Flow cytometry and colony formation assays were used to detect the effect of SAE-1 on radiosensitivity. In vivo, we detected the growth curve in a mouse xenograft model. The results showed that SAE-1 was revealed to be an independent prognostic biomarker of CRC. SAE1 knockdown inhibited CRC proliferation in vitro and in vivo, and led to the cleavage of PARP, downregulation of cyclin D1 protein expression, and downregulation of p-NF-κB/NF-κB. Additionally, SAE1 knockdown promoted radiosensitivity in CRC cells. Therefore, it was inferred that SAE1 may be used as a potential therapeutic target in CRC treatment.

Neuromedin U contributes to radiation resistance in colorectal cancer via YAP/TAZ signaling activation.

Resistance to radiation therapy remains a treatment obstacle for patients with high‑risk colorectal cancer. Neuromedin U (NMU) has been identified as a potential predictor of the response to radiation therapy by RNA sequencing analysis of colorectal cancer tissues obtained from patients. However, the role of NMU in colorectal cancer remains unknown. In order to investigate role of NMU in colorectal cancer, NMU expression was regulated using small interfering RNA or an NMU‑expression pCMV3 vector, and cell counting, wound‑healing and clonogenic assays were subsequently performed. NMU knockdown decreased colorectal cancer cell proliferation and migration, and sensitized the cells to radiation. Conversely, NMU overexpression increased radiation resistance, proliferation and migration of colorectal cancer cells. Furthermore, by western blotting and nuclear fractionation experiments, NMU knockdown inhibited the nuclear translocation of yes‑associated protein (YAP) and transcriptional co‑activator with PDZ‑binding motif (TAZ), resulting from the phosphorylation of these proteins. By contrast, the nuclear translocation of YAP and TAZ was increased following NMU overexpression in colorectal cancer cells. Recombinant NMU regulated YAP and TAZ activity, and the expression of the YAP and TAZ transcriptional target genes AXL, connective tissue growth factor and cysteine‑rich angiogenic inducer 61 in an NMU receptor 1 activity‑dependent manner. These results suggested that NMU may contribute to the acquisition of radioresistance in colorectal cancer by enhancing the Hippo signaling pathway via YAP and TAZ activation.

GSK484, an inhibitor of peptidyl arginine deiminase 4, increases the radiosensitivity of colorectal cancer and inhibits neutrophil extracellular traps.

Colorectal cancer (CRC) is the third most common malignancy and a major cause of cancer-related deaths. Peptidyl arginine deiminase 4 (PAD4 or PADI4) is expressed in neutrophils that, when activated, can drive the formation of neutrophil extracellular traps (NETs). PAD4 has been found to be upregulated in CRC patients and to predict a poor prognosis. This study is aimed at exploring the role of PAD4 inhibitor (GSK484) in NET formation and radioresistance in CRC. Reverse transcriptase quantitative polymerase chain reaction and western blotting were used to measure PAD4 expression in CRC tissues and cells. GSK484, an inhibitor of PAD4, was investigated in the following functional assays in vitro: western blotting, clonogenic survival, colony formation, TUNEL, flow cytometry and transwell assays. Nude mouse xenograft models were applied to evaluate the effect of GSK484 on tumor growth in CRC in vivo. The formation of NETs influenced by GSK484 was also investigated. We showed upregulation of PAD4 mRNA and protein in CRC tissues and cells. High expression of PAD4 was related to a poor prognosis in CRC patients. GSK484 treatment promoted the radiosensitivity of CRC cells and induced cell death by promoting DNA double-strand breaks. Rescue experiments further verified that GSK484 inhibited the effects of PAD4 overexpression in irradiated CRC cells. Moreover, GSK484 injection strengthened the radiosensitivity of CRC and inhibited NET formation in vivo. PAD4 inhibitor GSK484 promotes the radiosensitivity of CRC and inhibits NET formation in vivo and in vitro.

LncRNA OTUD6B-AS1 overexpression promoted GPX4-mediated ferroptosis to suppress radioresistance in colorectal cancer.

Radiotherapy is widely employed in colorectal cancer (CRC) treatment but is often compromised by developed radioresistance. This study explored the mechanism of long non-coding RNA ovarian tumor domain containing 6B-antisense RNA1 (lncRNA OTUD6B-AS1) in CRC radioresistance through tripartite motif 16 (TRIM16). CRC and non-cancerous tissues were collected and radioresistant CRC cells were established, with real-time quantitative polymerase chain reaction to determine gene expression in tissues and cells. Radioresistance was evaluated by cell counting kit-8 assay and immunofluorescence (γ-H2AX) and ferroptosis was tested by Western blot assay (ACSL4/GPX4) and assay kits (Fe2+/ROS/MDA/GSH). The association between ferroptosis and lncRNA OTUD6B-AS1-inhibited radioresistance was testified using ferroptosis inhibitor. The subcellular localization of lncRNA OTUD6B-AS1 was tested by the nuclear/cytoplasmic fractionation assay, with RNA immunoprecipitation assay to validate gene interactions. Rescue experiments were conducted to analyze the role of TRIM16 in CRC radioresistance. LncRNA OTUD6B-AS1 and TRIM16 were poorly expressed (P < 0.01) in CRC tissues and cells and further decreased (P < 0.01) in radioresistant CRC cells. OTUD6B-AS1 overexpression decreased cell survival (P < 0.01), increased γ-H2AX levels (P < 0.01), and elevated ferroptosis and oxidative stress (P < 0.01) after X-ray radiation. Ferroptosis inhibitor attenuated radioresistance (P < 0.01) caused by lncRNA OTUD6B-AS1 overexpression. LncRNA OTUD6B-AS1 stabilized TRIM16 mRNA via binding to HuR. TRIM16 knockdown reduced ferroptosis and increased radioresistance (P < 0.05). OTUD6B-AS1 overexpression stabilized TRIM16 via binding to HuR and increased GPX4-mediated ferroptosis, thus attenuating CRC radioresistance. Our study provided a new rationale for the treatment of CRC.

MicroRNAs as Predictive Biomarkers in Patients with Colorectal Cancer Receiving Chemotherapy or Chemoradiotherapy: A Narrative Literature Review.

Colorectal cancer (CRC) is one of the most common malignancies and is associated with high mortality rates worldwide. The underlying mechanism of tumorigenesis in CRC is complex, involving genetic, lifestyle-related, and environmental factors. Although radical resection with adjuvant FOLFOX (5-fluorouracil, leucovorin, and oxaliplatin) chemotherapy and neoadjuvant chemoradiotherapy have remained mainstays of treatment for patients with stage III CRC and locally advanced rectal cancer, respectively, the oncological outcomes of these treatments are often unsatisfactory. To improve patients' chances of survival, researchers are actively searching for new biomarkers to facilitate the development of more effective treatment strategies for CRC and metastatic CRC (mCRC). MicroRNAs (miRs), small, single-stranded, noncoding RNAs, can post-transcriptionally regulate mRNA translation and trigger mRNA degradation. Recent studies have documented aberrant miR levels in patients with CRC or mCRC, and some miRs are reportedly associated with chemoresistance or radioresistance in CRC. Herein, we present a narrative review of the literature on the roles of oncogenic miRs (oncomiRs) and tumor suppressor miRs (anti-oncomiRs), some of which can be used to predict the responses of patients with CRC to chemotherapy or chemoradiotherapy. Moreover, miRs may serve as potential therapeutic targets because their functions can be manipulated using synthetic antagonists and miR mimics.

MiR-7-5p/KLF4 signaling inhibits stemness and radioresistance in colorectal cancer

Resistance to radiotherapy remains a major unmet clinical obstacle in the treatment of locally advanced rectal cancer. Cancer stem cells (CSCs) are considered to mediate tumor development and radioresistance. However, the role of CSCs in regulating resistance to radiotherapy in colorectal cancer (CRC) remains largely unknown. We established two radioresistant CRC cell lines, HCT116-R and RKO-R, using fractionated irradiation. Analysis using miRNA sequencing and quantitative real-time PCR confirmed lower levels of miR-7-5p in both of the radioresistant cells compared to their parental cells. Subsequently, we validated that miR-7-5p expression was decreased in cancerous tissues from radiotherapy-resistant rectal cancer patients. The Cancer Genome Atlas (TCGA) database analyses revealed that low miR-7-5p expression was significantly correlated with poor prognosis in CRC patients. Overexpression of miR-7-5p led to a rescue of radioresistance and an increase in radiation-induced apoptosis, and attenuated the stem cell-like properties in HCT116-R and RKO-R cells. Conversely, knocking down miR-7-5p in parental HCT116 and RKO cells suppressed the sensitivity to radiation treatment and enhance cancer cell stemness. Stemness-associated transcription factor KLF4 was demonstrated as a target of miR-7-5p. Rescue experiments revealed that miR-7-5p/KLF4 axis could induce radiosensitivity by regulating CSCs in colorectal cancer cells. Furthermore, we used CRC tumor tissues which exhibited resistance to neoadjuvant radiotherapy to establish a patient-derived xenograft (PDX) mouse model. Tail vein injection of magnetic nanoparticles carrying miR-7-5p mimics into the PDX mice significantly inhibited tumor growth with or without irradiation treatment in vivo. Our current studies not only demonstrate an anti-cancer function of miR-7-5p in regulating CSC properties and radiosensitivity in colorectal cancer, but also provide a novel potential strategy for delaying or reverse radiation resistance in preoperative radiotherapy of CRC patients.

Prognostic risk analysis related to radioresistance genes in colorectal cancer.

Radiotherapy (RT) is one of the most important treatments for patients with colorectal cancer (CRC). Radioresistance is the crucial cause of poor therapeutic outcomes in colorectal cancer. However, the underlying mechanism of radioresistance in colorectal cancer is still poorly defined. Herein we established a radioresistant colorectal cancer cell line and performed transcriptomics analyses to search for the underlying genes that contribute to radioresistance and investigate its association with the prognosis of CRC patients. The radioresistant cell line was developed from the parental HCT116 cell by a stepwise increased dose of irradiation. Differential gene analysis was performed using cellular transcriptome data to identify genes associated with radioresistance, from which extracellular matrix (ECM) and cell adhesion-related genes were screened. Survival data from a CRC cohort in the TCGA database were used for further model gene screening and validation. The correlation between the risk score model and tumor microenvironment, clinical phenotype, drug treatment sensitivity, and tumor mutation status were also investigated. A total of 493 different expression genes were identified from the radioresistant and wild-type cell line, of which 94 genes were associated with ECM and cell adhesion-related genes. The five model genes TNFRSF13C, CD36, ANGPTL4, LAMB3, and SERPINA1 were identified for CRC radioresistance via screening using the best model. A ROC curve indicated that the AUC of the resulting prognostic model (based on the 5-gene risk score and other clinical parameters, including age, sex, and tumor stages) was 0.79, 0.77, and 0.78 at 1, 2, and 3 years, respectively. The calibration curve showed high agreement between the risk score prediction and actual survival probability. The immune microenvironment, drug treatment sensitivity, and tumor mutation status significantly differed between the high- and low-risk groups. The risk score model built with five radioresistance genes in this study, including TNFRSF13C, CD36, ANGPTL4, LAMB3, and SERPINA1, showed favorable performance in prognosis prediction after radiotherapy for CRC.

IGFL2-AS1, a Long Non-Coding RNA, Is Associated with Radioresistance in Colorectal Cancer.

Precise prediction of radioresistance is an important factor in the treatment of colorectal cancer (CRC). To discover genes that regulate the radioresistance of CRCs, we analyzed an RNA sequencing dataset of patient-originated samples. Among various candidates, IGFL2-AS1, a long non-coding RNA (lncRNA), exhibited an expression pattern that was well correlated with radioresistance. IGFL2-AS1 is known to be highly expressed in various cancers and functions as a competing endogenous RNA. To further investigate the role of IGFL2-AS1 in radioresistance, which has not yet been studied, we assessed the amount of IGFL2-AS1 transcripts in CRC cell lines with varying degrees of radioresistance. This analysis showed that the more radioresistant the cell line, the higher the level of IGFL2-AS1 transcripts-a similar trend was observed in CRC samples. To directly assess the relationship between IGFL2-AS1 and radioresistance, we generated a CRC cell line stably expressing a small hairpin RNA (shRNA) targeting IGFL2-AS1. shRNA-mediated knockdown of IGFL2-AS1 decreased radioresistance and cell migration in vitro, establishing a functional role for IGFL2-AS1 in radioresistance. We also showed that downstream effectors of the AKT pathway played crucial roles. These data suggest that IGFL2-AS1 contributes to the acquisition of radioresistance by regulating the AKT pathway.

KCNE4 expression is correlated with the pathological characteristics of colorectal cancer patients and associated with the radioresistance of cancer cells.

Colorectal cancer (CRC) is a common malignancy, and radioresistance limits the effectiveness of radiotherapy for rectal cancer. This study is performed to investigate the role and regulatory mechanism of Potassium Voltage-Gated Channel Subfamily E Regulatory Subunit 4 (KCNE4) in the radioresistance of CRC cells. Immunohistochemical staining results of KCNE4 in normal tissues and CRC tissues were obtained from the Human Protein Atlas (HPA) database. The UALCAN database was used for analyzing KCNE4 mRNA expression in normal tissue samples and CRC tissue samples and its relationship with tumor stage. The relationship of KCNE4 expression with prognosis was analyzed utilizing the data of GEPIA database. LinkedOmics database was searched to analyze the co-expressed gene sets of KCNE4 in CRC, and to analyze the signaling pathways related with KCNE4 in CRC. GO and KEGG enrichment analyses were carried out on the co-expressed genes of KCNE4 with DAVID database. Ionizing radiation (IR)-resistant cell lines (HCT116/IR and HT29/IR) were established; cell viability was assessed via cell counting kit-8 (CCK-8) and EdU assays, and terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay was performed for detecting cell apoptosis. Western blotting was carried out to detect the expressions of p-p85 and p-AKT. KCNE4 was highly expressed in CRC tissues and linked to advanced tumor stage, lymph node metastasis and poor prognosis of CRC patients. KCNE4 overexpression promoted HCT116/IR cell proliferation and inhibited the apoptosis, while KCNE4 knockdown suppressed HT29/IR cell proliferation and facilitated the apoptosis. Furthermore, high KCNE4 expression was associated with the activation of the PI3K/AKT signal pathway. KCNE4 is associated with the clinicopathological characteristics of CRC patients, and its high expression level contributes to the radioresistance of cancer cells via activating the PI3K/AKT signal pathway.

SHF confers radioresistance in colorectal cancer by the regulation of mitochondrial DNA copy number.

Altered mitochondrial function contributes greatly to pathogenesis and progression of colorectal cancer. In this study, we report a functional pool of Src homology 2 domain-containing F (SHF) in mitochondria controlling the response of colorectal cancer cells to radiation therapy. We found that elevated expression of SHF in cancer cells is essential for promoting mitochondrial function by increasing mitochondrial DNA copy number, thus reducing the sensitivity of colorectal cancer cells to radiation. Mechanistically, SHF binds to mitochondrial DNA and promotes POLG/SSBP1-mediated mitochondrial DNA synthesis. Importantly, SHF loss-mediated radiosensitization was phenocopied by depletion of mitochondrial DNA. Thus, our data demonstrate that mitochondrial SHF is an important regulator of radioresistance in colorectal cancer cells, identifying SHF as a promising therapeutic target to enhance radiotherapy efficacy in colorectal cancer.

PRDM15 interacts with DNA-PK-Ku complex to promote radioresistance in rectal cancer by facilitating DNA damage repair

Neoadjuvant radiotherapy is a standard treatment for locally advanced rectal cancer, however, resistance to chemoradiotherapy is one of the main obstacles to improving treatment outcomes. The goal of this study was to explore the role of PRDM15 involved in the radioresistance of colorectal cancer and to clarify the underlying mechanism. In present study, we demonstrated that, after DNA damage, PRDM15 was upregulated and localized to DNA damage sites, co-localizing with γ-H2AX. Knockdown of PRDM15 inhibited DNA damage repair and increased radiosensitivity in colorectal cancer cells. Mechanistically, PRDM15 promoted DNA repair by interacting with DNA-PKcs and Ku70/Ku80 complex. In preclinical models of rectal cancer, knockdown of PRDM15 sensitized cell derived xenograft and patient derived xenograft to radiotherapy. In 80 rectal cancer patients treated with neoadjuvant chemoradiotherapy, higher PRDM15 expression was observed associated with weaker tumor regression and poorer prognosis. Our findings revealed that inhibiting PRDM15 was potent to overcome radioresistance through abrogating DNA repair in colorectal cancer cells. Additionally, the expression level of PRDM15 could be applied to predict radiotherapy responsiveness and the outcome of neoadjuvant radiotherapy in rectal cancer patients.

Assessment of B7-H3 as a Potent Biomarker for Predicting the Radiosensitivity of IDH-Wildtype Glioblastoma

<h3>Purpose/Objective(s)</h3> Glioblastoma (GBM) is the most aggressive primary malignant brain tumor, and the IDH-wildtype GBM, accounting for about 90% of GBM cases, is characterized by poor prognosis and response to the standard chemoradiotherapy. B7-H3, a B7-family immunoregulatory protein, has been reported to correlate with poor survival in various solid tumors and to enhance radioresistance in colorectal cancer. However, the effect of B7-H3 on chemoradioresistance and prognosis in IDH-wildtype GBM remains unexplored. Therefore, this study analyzes the potential of B7-H3 expression as a predictive biomarker for chemoradiosensitivity and a prognostic factor in IDH-wildtype GBM. <h3>Materials/Methods</h3> 65 patients histopathologically diagnosed with IDH-wildtype GBM undergoing standard chemoradiotherapy (Stupp Protocol) at a single institution from 2015 to 2019 were involved in this retrospective study. Patients were divided into the chemoradioresistant group and chemoradiosensitive group based on the short-term treatment outcome. Patient tumor tissue samples were subjected to immunohistochemistry for B7-H3 expression. Receiver operating characteristic (ROC) analysis was exploited to estimate the diagnostic value. Kaplan-Meier curves, the log-rank test and Cox proportional hazard model were used to test for the prognostic association of B7-H3 expression <h3>Results</h3> B7-H3 expression in patients resistant to chemoradiotherapy (n=18) was significantly higher than that in patients sensitive to chemoradiotherapy (n=47) (p=0.048, t-test). ROC curves showed that the area under the curve (AUC) for B7-H3 expression was 0.778 (95% CI = 0.670-0.926; p=0.001). Patients with high B7-H3 expression demonstrated significantly shorter median progression-free survival (PFS) and median overall survival (OS) than patients with low B7-H3 expression (median PFS = 10 vs. 21 months, p=0.047; median OS = 17 vs. 31 months, p=0.023). univariable Cox proportional hazard model displayed an increased risk of death for patients with high B7-H3 expression (hazard ratio=5.926; 95% CI=1.669-21.044; p=0.006). <h3>Conclusion</h3> Our results suggest that B7-H3 expression might serve as a potential biomarker for chemoradiosensitivity prediction in IDH-wildtype GBM. Moreover, the data demonstrate B7-H3 expression as a prognostic factor candidate for IDH-wildtype GBM. This study implies previously unrecognized B7-H3 contribution to chemoradioresistance, encouraging future studies to explore further the mechanisms behind.

A novel long noncoding RNA SP100-AS1 induces radioresistance of colorectal cancer via sponging miR-622 and stabilizing ATG3

Although radiotherapy is an essential modality in the treatment of colorectal cancer (CRC), the incidence of radioresistance remains high clinically. Long noncoding RNAs (lncRNAs) reportedly play critical roles in CRC radioresistance by regulating genes or proteins at the transcriptional or post-translational levels. This study aimed to identify novel lncRNAs involved in radioresistance. We found that SP100-AS1 (lncRNA targeting antisense sequence of SP100 gene) was upregulated in radioresistant CRC patient tissues using RNA-seq analysis. Importantly, knockdown of SP100-AS1 significantly reduced radioresistance, cell proliferation, and tumor formation in vitro and in vivo. Mechanistically, mass spectrometry and bioinformatics analyses were used to identify the interacting proteins and microRNAs of SP100-AS1, respectively. Moreover, SP100-AS1 was found to interact with and stabilize ATG3 protein through the ubiquitination-dependent proteasome pathway. In addition, it could serve as a sponge for miR-622, which targeted ATG3 mRNA and affected autophagic activity. Thus, lncRNA SP100-AS1 could act as a radioresistance factor in CRC patients via RNA sponging and protein stabilizing mechanisms. In conclusion, the present study indicates that SP100-AS1/miR-622/ATG3 axis contributes to radioresistance and autophagic activity in CRC patients, suggesting it has huge prospects as a therapeutic target for improving CRC response to radiation therapy.

Characterization of the mechanism of Scutellaria baicalensis on reversing radio-resistance in colorectal cancer

Scutellaria baicalensis (SB) has been shown to improve the therapeutic effects of colorectal cancer (CRC) and perform well for reversing radio-resistance in different cancers. However, its potential function and mechanism related to radio-resistance in CRC has not been explored. A radio-resistant human CRC cell line (HCT116R) was applied. A network pharmacological analysis was performed to reveal the potential mechanism of SB for reversing radio-resistance in CRC, and computational pathological analysis was applied to indicate the clinicopathological significance of the key targets. Then, our hypothesis was further verified by molecular docking. The network pharmacology analysis showed that wogonin is the key compound of SB for reversing the radio-resistance of CRC. A Kyoto Encyclopedia of Genes and Genomes analysis showed that the genes for SB that reverse radio-resistance in CRC are mainly involved in steroid hormone biosynthesis. An enrichment analysis pointed out that Sulfotransferase family 2B member 1 (SULT2B1) is a potentially vital gene. SULT2B1 was demonstrated as being highly expressed in CRC and upregulated in radio-resistant rectal tissues or cell lines. A CCK-8 and clone formation test showed that the viability and clone formation ability of HCT116R were significantly decreased by wogonin combined with radiotherapy, compared to radiotherapy alone. By contrast, flow cytometry revealed that the apoptosis of HCT116R was significantly increased when wogonin treatment combined with radiotherapy, compared with radiotherapy alone. Molecular docking verification indicated that SULT2B1 and wogonin have a good binding ability. Taken together, SULT2B1 may be the potential drug target in treating radio-resistant CRC. Wogonin may be the core compound of SB for reversing radio-resistance in CRC by targeting SULT2B1.

Abstract 5240: S-phase cyclinA associated protein in the ER(SCAPER)rs3812908 T&amp;gt;C promotes the nuclear translocation of CyclinA2 and enhances DNA damage repair in radioresistance of colorectal cancer

Abstract Radiotherapy resistance is still an obstacle of improving the survival of patients with locally advanced colorectal cancer. In this study, whole-exome sequencing and targeted sequencing were preformed on colorectal cancer tumor tissue samples which were colloected from multiple centers, and the pathologic complete response predictive model PGS-LARC(Prediction Genotype Signature for Locally Advanced Rectal Cancer) was established. A candidate SNV-SCAPER rs3812908 T&amp;gt;C was screened using this model. To investigate the relationship between SCAPER rs3812908 T&amp;gt;C and radiosensitivity of colorectal cancer, targeted sequencing was performed and the results revealed that proportion of CC genotype was significantly higher in resistant sequencing cohort than in sensitive ones (30% vs. 22%). And Immunohistochemistry (IHC) results revealed that SCAPER protein expressed less in CC genotype colorectal cancer tissues than in TT genotype. Here, the cell proliferation results showed that SCAPER-KD cells and SCAPER rs3812908 CC cells were more radioresistant than SCAPER-NC and TT cells. Furthermore, the immunofluorescence assay showed SCAPER rs3812908 T&amp;gt;C promoted translocation of S phase associated protein-Cyclin A2 into nucleus and western blotting results showed that the phosphorylation of ERK and JNK(p-ERK and p-JNK) increased significantly in SCAPER CC cells, and the protein p-ATR, p-ATM, p-CDC25C and CDC25A related to homologous recombination repair were also increased in SCAPER-KD and SCAPER CC cells.Taken together, the SCAPER gene rs3812908 T&amp;gt;C plays an important role in radioresistance of colorectal cancer. The SNV of SCAPER rs3812908 T&amp;gt;C significantly promotes Cyclin A2 nuclear translocation, activates the MAPKs pathway, and increases homologous recombination repair, and finally leads to radioresistance. Citation Format: Hai-na Yu, Ting Jiang, Shuang Liu, Yan Yuan, Shao-yan Xi, Zhi-wei Guo, Yuan-hong Yuan, Yan Li, Wei-wei Xiao. S-phase cyclinA associated protein in the ER(SCAPER)rs3812908 T&amp;gt;C promotes the nuclear translocation of CyclinA2 and enhances DNA damage repair in radioresistance of colorectal cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5240.

RIOK1 mediates p53 degradation and radioresistance in colorectal cancer through phosphorylation of G3BP2.

RIO Kinase 1 (RIOK1) is involved in various pathologies, including cancer. However, the role of RIOK1 in radioresistance of colorectal cancer (CRC) remains largely unknown. In this study, we reported that RIOK1 was overexpressed in rectal cancer tissue with weaker tumor regression after neoadjuvant chemoradiotherapy (neoCRT). Moreover, higher RIOK1 expression predicted a poor prognosis in patients with rectal cancer. Blockade of RIOK1 using Toyocamycin, a pharmacological inhibitor of RIOK1, or by knocking down its expression, decreased the resistance of CRC cells to radiotherapy in vitro and in vivo. A mechanistic study revealed that RIOK1 regulates radioresistance by suppressing the p53 signaling pathway. Furthermore, we found that RIOK1 and Ras-GAP SH3 domain binding protein 2 (G3BP2) interact with each other. RIOK1 phosphorylates G3BP2 at Thr226, which increases the activity of G3BP2. RIOK1-mediated phosphorylation of G3BP2 facilitated ubiquitination of p53 by murine double minute 2 protein (MDM2). Altogether, our study revealed the clinical significance of RIOK1 in CRC, and therapies targeting RIOK1 might alleviate the CRC tumor burden in patients.

Skp2 stabilizes Mcl-1 and confers radioresistance in colorectal cancer

Overexpression of Skp2 plays a critical role in tumorigenesis and correlates with poor prognosis in human malignancies. Thus, Skp2 has been proposed as an attractive target for anti-tumor interventions. The expression of Skp2 in human colorectal cancer (CRC) and the role of Skp2 in tumorigenic properties and irradiation sensitivities of CRC cells were examined by anchorage-dependent and -independent growth assays, immunoblot, flow cytometry, immunohistochemical staining, ubiquitination analysis, co-immunoprecipitation assay, CRISPR-Cas9-based gene knockout, and xenograft experiments. Skp2 is highly expressed in CRC patient tissues. Blocking Skp2 expression reduces the tumorigenic properties of CRC cells in vitro and in vivo. Depletion of Skp2 confers sensitivity to irradiation of CRC cells. Skp2 deficiency enhances irradiation-induced intrinsic apoptosis by facilitating E3 ligase FBW7-mediated Mcl-1 ubiquitination and degradation. Knockout of Skp2 sensitizes CRC cells to irradiation treatments in vivo. Our findings indicate that Skp2 stabilizes Mcl-1, and targeting Skp2 in combination with traditional radiotherapy might be efficacious in treating CRC.

FOXQ1-mediated SIRT1 upregulation enhances stemness and radio-resistance of colorectal cancer cells and restores intestinal microbiota function by promoting \u03b2-catenin nuclear translocation

BackgroundResistance of colorectal cancer (CRC) cells to radiotherapy considerably contributes to poor clinical outcomes of CRC patients. Microarray profiling in this study revealed the differentially expressed forkhead box Q1 (FOXQ1) in CRC, and thus we aimed to illustrate the role of FOXQ1 in CRC by modulating stemness and radio-resistance of CRC cells.MethodsCRC and adjacent normal tissues were collected from CRC patients, and the correlation between FOXQ1 expression and CRC prognosis was analyzed. Subsequently, we determined the expression of FOXQ1, sirtuin 1 (SIRT1) and β-catenin in CRC tissues and cell lines. The binding affinity between FOXQ1 and SIRT1 and that between SIRT1 and β-catenin were validated with luciferase reporter gene, Co-IP and ChIP assays. Following a metagenomics analysis of CRC intestinal microbiota, the effects of the FOXQ1/SIRT1/β-catenin axis on CRC stem cell phenotypes and radio-resistance was evaluated in vitro and in vivo through manipulation of gene expression. Besides, mouse feces were collected to examine changes in intestinal microbiota.ResultsFOXQ1 was highly expressed in CRC tissues and cells and positively correlated with poor prognosis of CRC patients. FOXQ1 overexpression contributed to resistance of CRC cells to radiation. Knockdown of FOXQ1 inhibited the stemness of CRC cells and reversed their radio-resistance. FOXQ1 enhanced the transcriptional expression of SIRT1, and SIRT1 enhanced the expression and nuclear translocation of β-catenin. Knockdown of FOXQ1 repressed SIRT1 expression, thus reducing the stemness and radio-resistance of CRC cells. Moreover, FOXQ1 knockdown suppressed CRC xenograft formation in xenograft-bearing nude mice through inhibiting SIRT1 and β-catenin to reduce the content of pathological bacteria that were up-regulated in CRC.ConclusionFOXQ1-mediated SIRT1 upregulation augments expression and nuclear translocation of β-catenin and benefits CRC-related intestinal pathological bacterial, thereby enhancing the stemness and radio-resistance of CRC cells.