Cancer Cell Radioresistance Research Articles

Non-coding RNAs modulation in breast cancer radioresponse: mechanisms and therapeutic implications.

Breast cancer is the most frequent type of cancer in women, with significant incidence and fatality rates. Radiation therapy is an important therapeutic option for breast cancer patients. However, tumor cells' resistance to radiation can limit therapy efficacy, resulting in recurrence and death. Non-coding RNAs (ncRNAs) are aclass of small RNA molecules that do not translate into proteins but can affect the translation of target mRNA. Several investigations on breast cancer have demonstrated abnormal expression of ncRNAs in response to radiation. Non-coding RNAs are essential in controlling numerous processes such as DNA damage response, cancer stem cell pathways, cell cycle regulation, cell death, and inflammation. Dysregulation of ncRNAs after irradiation influences radiosensitivity or radioresistance of breast cancer cells. Understanding the molecular mechanisms underlying Radiation response can lead to innovative treatment ways to reduce breast cancer radioresistance and increase radiotherapy's efficacy. This review summarizes current research on ncRNA dysregulation following irradiation and analyzes ncRNAs' function and mechanism in modifying breast cancer cell radiosensitivity and radioresistance.

Additional considerations in cancer cell radioresistance, integrin αvβ3 and thyroid hormones

ABSTRACT Background The existence of a functional relationship between a certain thyroid hormone analogue and cancer cell radioresistance has been shown by Leith and coworkers. The hormone analogue with relevance to malignant cells’ radioresistance is tetraiodothyroacetic acid (tetrac). Tetrac is the deaminated derivative of L-thyroxine (T4), the principal product of the thyroid gland. Preclinical studies demonstrated that tetrac and chemically modified tetrac (CMT), e.g. a fluorobenzyl-conjugated tetrac analogue, restores radiosensitivity in certain radioresistant tumor cells. Due to their molecular, physico-chemical, and biological properties, actions of CMT analogues are believed to be initiated at the thyroid hormone analogue receptor site on plasma membrane integrin αvβ3. Objective To explore possible molecular mechanisms of the potentially therapeutically beneficial effect of CMT on cancer cells’ sensitivity to radiation, we analyzed actions of CMT analogues on expression of selected sets of genes that have been previously implicated in radioresistance of malignant cells. Discussion and conclusions In the current study, we report that genome-wide gene expression profiling analysis of human glioblastoma (GBM) and acute myelocytic leukemia (AML) cell lines exposed in vitro to noncytotoxic doses of CMT has identified decreased expression of discrete trios of genes each of which was previously linked to cancer cells’ radioresistance. Following the CMT treatment in AML cells, expression of PARP9, PARP15 and STAT3 genes was significantly reduced, while in GBM cells, expression of PRKDC, EGFR and CCNDI was significantly decreased by the drug. Notably, a broader spectrum of genes implicated in cancer cells’ radioresistance was observed in primary patient-derived GBM cells after the CMT treatment. Extensive additional experimental and clinical studies are indicated, including analyses of individual patient tumor genomics and of an array of different tumor types to define the sub-sets of tumors manifesting radioresistance in which tetrac-based agents may be expected to enhance therapeutic effects of radiation.

Cancer radioresistance is characterized by a differential lipid droplet content along the cell cycle

BackgroundCancer radiation treatments have seen substantial advancements, yet the biomolecular mechanisms underlying cancer cell radioresistance continue to elude full understanding. The effectiveness of radiation on cancer is hindered by various factors, such as oxygen concentrations within tumors, cells’ ability to repair DNA damage and metabolic changes. Moreover, the initial and radiation-induced cell cycle profiles can significantly influence radiotherapy responses as radiation sensitivity fluctuates across different cell cycle stages. Given this evidence and our prior studies establishing a correlation between cancer radiation resistance and an increased number of cytoplasmic Lipid Droplets (LDs), we investigated if LD accumulation was modulated along the cell cycle and if this correlated with differential radioresistance in lung and bladder cell lines.ResultsOur findings identified the S phase as the most radioresistant cell cycle phase being characterized by an increase in LDs. Analysis of the expression of perilipin genes (a family of proteins involved in the LD structure and functions) throughout the cell cycle also uncovered a unique gene cell cycle pattern.ConclusionsIn summary, although these results require further molecular studies about the mechanisms of radioresistance, the findings presented here are the first evidence that LD accumulation could participate in cancer cells’ ability to better survive X-Ray radiation when cells are in the S phase. LDs can represent new players in the radioresistance processes associated with cancer metabolism. This could open new therapeutic avenues in which the use of LD-interfering drugs might enhance cancer sensitivity to radiation.

ALDH1A1 drives prostate cancer metastases and radioresistance by interplay with AR- and RAR-dependent transcription.

Rationale: Current therapies for metastatic osseous disease frequently fail to provide a durable treatment response. To date, there are only limited therapeutic options for metastatic prostate cancer, the mechanisms that drive the survival of metastasis-initiating cells are poorly characterized, and reliable prognostic markers are missing. A high aldehyde dehydrogenase (ALDH) activity has been long considered a marker of cancer stem cells (CSC). Our study characterized a differential role of ALDH1A1 and ALDH1A3 genes as regulators of prostate cancer progression and metastatic growth. Methods: By genetic silencing of ALDH1A1 and ALDH1A3 in vitro, in xenografted zebrafish and murine models, and by comparative immunohistochemical analyses of benign, primary tumor, and metastatic specimens from patients with prostate cancer, we demonstrated that ALDH1A1 and ALDH1A3 maintain the CSC phenotype and radioresistance and regulate bone metastasis-initiating cells. We have validated ALDH1A1 and ALDH1A3 as potential biomarkers of clinical outcomes in the independent cohorts of patients with PCa. Furthermore, by RNAseq, chromatin immunoprecipitation (ChIP), and biostatistics analyses, we suggested the molecular mechanisms explaining the role of ALDH1A1 in PCa progression. Results: We found that aldehyde dehydrogenase protein ALDH1A1 positively regulates tumor cell survival in circulation, extravasation, and metastatic dissemination, whereas ALDH1A3 plays the opposite role. ALDH1A1 and ALDH1A3 are differentially expressed in metastatic tumors of patients with prostate cancer, and their expression levels oppositely correlate with clinical outcomes. Prostate cancer progression is associated with the increasing interplay of ALDH1A1 with androgen receptor (AR) and retinoid receptor (RAR) transcriptional programs. Polo-like kinase 3 (PLK3) was identified as a transcriptional target oppositely regulated by ALDH1A1 and ALDH1A3 genes in RAR and AR-dependent manner. PLK3 contributes to the control of prostate cancer cell proliferation, migration, DNA repair, and radioresistance. ALDH1A1 gain in prostate cancer bone metastases is associated with high PLK3 expression. Conclusion: This report provides the first evidence that ALDH1A1 and PLK3 could serve as biomarkers to predict metastatic dissemination and radiotherapy resistance in patients with prostate cancer and could be potential therapeutic targets to eliminate metastasis-initiating and radioresistant tumor cell populations.

Aldolase A promotes cervical cancer cell radioresistance by regulating the glycolysis and DNA damage after irradiation

ABSTRACT Radioresistance is the major obstacle that affects the efficacy of radiotherapy which is an important treatment for cervical cancer. By analyzing the databases, we found that aldolase A (ALDOA), which is a key enzyme in metabolic reprogramming, has a higher expression in cervical cancer patients and is associated with poor prognosis. We detected the expression of ALDOA in the constructed cervical cancer radioresistance (RR) cells by repetitive irradiation and found that it was upregulated compared to the control cells. Functional assays were conducted and the results showed that the knockdown of ALDOA in cervical cancer RR cells inhibited the proliferation, migration, and clonogenic abilities by regulating the cell glycolysis. In addition, downregulation of ALDOA enhanced radiation-induced apoptosis and DNA damage by causing G2/M phase arrest and further promoted radiosensitivity of cervical cancer cells. The functions of ALDOA in regulating tumor radiosensitivity were also verified by the mouse tumor transplantation model in vivo. Therefore, our study provides new insights into the functions of ALDOA in regulating the efficacy of radiotherapy and indicates that ALDOA might be a promising target for enhancing radiosensitivity in treating cervical cancer patients.

Exosomal miR-196a-5p enhances radioresistance in lung cancer cells by downregulating NFKBIA.

Radiation therapy is recognized as an effective modality in the treatment of lung cancer, but radioresistance resulting from prolonged treatment reduces the chances of recovery. MicroRNAs (miRNAs) play a pivotal role in radiotherapy immunity. In this study, we aimed to investigate the mechanism by which miR-196a-5p affects radioresistance in lung cancer. The radioresistant lung cancer cell line A549R26-1 was established by radiation treatment. Cancer-associated fibroblasts (CAFs) and normal fibroblasts (NFs) were observed by microscopy, and the expression levels of CAF-specific marker proteins were detected by immunofluorescence. The shape of the exosomes was observed by electron microscopy. A CCK-8 assay was used to detect cell viability, while clone formation assays were used to detect cell proliferative capacity. Flow cytometry was performed to investigate apoptosis. The binding of miR-196a-5p and NFKBIA was predicted and further verified by the dual luciferase reporter experiment. qRT-PCR and western blotting were used to detect gene mRNA and protein levels. We found that exosomes secreted by CAFs could enhance lung cancer cell radioresistance. Moreover, miR-196a-5p potentially bound to NFKBIA, promoting malignant phenotypes in radioresistant cells. Furthermore, exosomal miR-196a-5p derived from CAFs increased radiotherapy immunity in lung cancer. Exosomal miR-196a-5p derived from CAFs enhanced radioresistance in lung cancer cells by downregulating NFKBIA, providing a new potential target for the treatment of lung cancer.

DNA‑PKcs phosphorylation specific inhibitor, NU7441, enhances the radiosensitivity of clinically relevant radioresistant oral squamous cell carcinoma cells.

Radioresistant cancer cells lead to poor prognosis after radiotherapy. However, the mechanisms underlying cancer cell radioresistance have not been fully elucidated. Thus, the DNA damage response of clinically relevant radioresistant oral squamous cell carcinoma HSC2-R cells, established by long-term exposure of parental HSC2 cells to fractionated radiation, was investigated. The DNA double-strand break (DSB) repair protein-specific inhibitor, NU7441, which targets DNA-dependent protein kinase catalytic subunit (DNA-PKcs) phosphorylation, and IBR2, which targets Rad51, were administered to HSC2 and HSC2-R cells. NU7441 administration eliminated colony formation in both cell lines under 6 Gy X-ray irradiation, whereas IBR2 did not affect colony formation. NU7441 and IBR2 significantly enhanced 6 Gy X-ray irradiation-induced apoptosis in HSC2-R cells. In HSC2-R cells, cell cycle arrest released earlier than in HSC2 cells, and phosphorylated-H2A histone family member X (γH2AX) expression rapidly decreased. Following NU7441 administration, γH2AX expression and the cell percentages of the G2/M phase were not decreased at 48 h after treatment in HSC2-R cells. DNA-PKcs has been demonstrated to regulate non-homologous end-joining (NHEJ) and homologous recombination (HR) repair, and the later phase of DSB repair is dominated by HR. Therefore, the results of the present study indicated that the DSB repair mechanism in HSC2-R cells strongly depends on NHEJ and loss of HR repair function. The present study revealed a potential mechanism underlying the acquired radioresistance and therapeutic targets in radioresistant cancer cells.

Cancer-associated fibroblasts induce growth and radioresistance of breast cancer cells through paracrine IL-6

In breast cancer, the most numerous stromal cells are cancer-associated fibroblasts (CAFs), which are associated with disease progression and chemoresistance. However, few studies have explored the function of CAFs in breast cancer cell radiosensitivity. Here, CAF-derived conditioned media was observed to induce breast cancer cell growth and radioresistance. CAFs secrete interleukin 6 (IL-6) which activates signal transducer and activator of transcription 3 (STAT3) signaling pathway, thus promoting the growth and radioresistance of breast cancer cells. Treatment with an inhibitor of STAT3 or an IL-6 neutralizing antibody blocked the growth and radioresistance induced by CAFs. In in vivo mouse models, tocilizumab (an IL-6 receptor monoclonal antibody) abrogated CAF-induced growth and radioresistance. Moreover, in breast cancer, a poor response to radiotherapy was associated with IL-6 and p-STAT3 expression. These results indicated that IL-6 mediates cross-talk between breast cancer cells and CAFs in the tumor microenvironment. Our results identified the IL-6/STAT3 signaling pathway as an important therapeutic target in breast cancer radiotherapy.

Dual role of Nrf2 in cancer: molecular mechanisms, cellular functions and therapeutic interventions

Nrf2 regulates oxidative stress, which is essential for cellular function. Fundamental initiation of Nrf2 in many malignancies increases prosurvival genes & endorses tumour cell propagation via metabolic reprogramming, suppression of tumour programmed cell death, & increased cancer stem cell self-renewal potential. More specifically, Nrf2 has been associated with cancer cell chemoresistance, radioresistance & inflammation-induced carcinogenesis. METHODS AND RESULTS: Many Nrf2 inhibitors have been revealed for tumour treatment and targeting Nrf2 could be an effective cancer therapeutic method. Before spreading, cancer cells adapt to their surroundings. Cancer cells usually have mutations in tumor suppressor genes. In a variety of malignancies, somatic mutations & other anomalies in the Nrf2 genes, as well as renowned cancer suppressor genes including TP53, CDKN2A, PTEN & PIK3CA, have been found. In tumour cells, somatic mutations in the Nrf2 genes, as well as additional mechanisms that affect Nrf2 binding, and produce aberrant Nrf2 activation. Uncontrolled Nrf2 causes tumour cells to become resistant to antineoplastic drugs & reactive oxygen species (ROS), as well as guiding them toward metabolic reprogramming. CONCLUSIONS: As a result, Nrf2 has been studied as potential malignancy treatment target. We covered the pathways, mechanisms, and dual characteristics of Nrf2 in malignancy in this article. We also discussed how Nrf2 inhibitors are targeted against cancer in this review.

Dose-enhancement of MCF 7 cell line radiotherapy using silica-iron oxide nanocomposite

Cancer radiotherapy is one of the most effective regimens of cancer treatments, but cancer cell radioresistance remains a concern. Radiosensitizers can selectively improve the efficacy of radiotherapy and reduce inherent damage. The purpose of this work is to evaluate the effect of silica-coated iron oxide magnetic nanoparticles (SIONPs) as a radiosensitizer and compare their therapeutic effect with that of Iron oxide magnetic nanoparticles (IONPs). IONPs and SIONPs were characterized using several physical techniques such as a transmission electron microscope (TEM) and Vibrating sample magnetometer (VSM). MTT and DNA double-strand breaks (Comet) assays have been used to detect the cytotoxicity, cell viability, and DNA damage of MCF-7 cells, which were treated with different concentrations of prepared nanoparticles and exposed to an X-ray beam. In this study, an efficient radiosensitizer, SIONPs, was successfully prepared and characterized. With 0.5 Gy dose, dose enhancement factor (DEF) values of cells treated with 5 and 10 μg/ml of IONPs were 1 and 1.09, respectively, while those treated with SIONPs at these concentrations had DEF of 1.21 and 1.32, respectively. Results demonstrated that SIONPs provide a potential for improving the radiosensitivity of breast cancer.

Increased glucose influx and glycogenesis in lung cancer cells surviving after irradiation

Purpose Lung cancer is considered as one of the most frequent malignancies worldwide. Radiotherapy is the main treatment modality applied for locally advanced disease, but remnant surviving cancer tissue results in disease progression in the majority of irradiated lung carcinomas. Metabolic reprogramming is regarded as a cancer hallmark and is associated with resistance to radiation therapy. Here, we explored metabolic alterations possibly related to cancer cell radioresistance. Materials and methods We compared the expression of metabolism-related enzymes in the parental A549 lung cancer cell line along with two new cell lines derived from A549 cells after recovery from three (A549-IR3) and six (A549-IR6) irradiation doses with 4 Gy. Differential GLUT1 and GYS1 expression on proliferation and radioresistance were also comparatively investigated. Results A549-IR cells displayed increased extracellular glucose absorption, and enhanced mRNA and protein levels of the GLUT1 glucose transporter. GLUT1 inhibition with BAY-876, suppressed cell proliferation and the effect was significantly more profound on A549-IR3 cells. Protein levels of molecules associated with aerobic or anaerobic glycolysis, or the phosphate pentose pathway were similar in all three cell lines. However, glycogen synthase 1 (GYS1) was upregulated, especially in the A549-IR3 cell line, suggestive of glycogen accumulation in cells surviving post irradiation. GYS1-gene silencing repressed the proliferation capacity of A549, but this increased their radioresistance. The radio-protective effect of the suppression of proliferative activity induced by GYS1 silencing did not protect A549-IR3 cells against further irradiation. Conclusions These findings indicate that GYS1 activity is a critical component of the metabolism of lung cancer cells surviving after fractionated radiotherapy. Targeting the glycogen metabolic reprogramming after irradiation may be a valuable approach to pursue eradication of the post-radiotherapy remnant of disease.

Peroxiredoxin IV plays a critical role in cancer cell growth and radioresistance through the activation of the Akt/GSK3 signaling pathways

High levels of redox enzymes have been commonly observed in various types of human cancer, although whether and how the enzymes contribute to cancer malignancy and therapeutic resistance have yet to be understood. Peroxiredoxin IV (Prx4) is an antioxidant with bona fide peroxidase and molecular chaperone functions. Here, we report that Prx4 is highly expressed in prostate cancer patient specimens, as well as established prostate cancer cell lines, and that its levels can be further stimulated through the activation of androgen receptor signaling. We used lentivirus-mediated shRNA knockdown and CRISPR-Cas9 based KO techniques to establish Prx4-depleted prostate cancer cells, which showed delayed cell cycle progression, reduced rate of cell proliferation, migration, and invasion compared to control cells. In addition, we used proteome profiler phosphokinase arrays to identify signaling changes in Prx4-depleted cells; we found that loss of Prx4 results in insufficient phosphorylation of both Akt and its downstream kinase GSK3α/β. Moreover, we demonstrate that Prx4-depleted cells are more sensitive to ionizing radiation as they display compromised ability to scavenge reactive oxygen species and increased accumulation of DNA damage. In mouse xenograft models, we show depletion of Prx4 leads to significant suppression of tumor growth, and tumors formed by Prx4-depleted cells respond more effectively to radiation therapy. Our findings suggest that increased levels of Prx4 contribute to the malignancy and radioresistance of prostate cancer through the activation of Akt/GSK3 signaling pathways. Therefore, strategies targeting Prx4 may be utilized to potentially inhibit tumor growth and overcome radioresistance in prostate cancer.

Numb/Notch/PLK1 signaling pathway mediated hyperglycemic memory in pancreatic cancer cell radioresistance and the therapeutic effects of metformin

BackgroundDiabetes mellitus has presented a positive role in the progression of pancreatic cancer and “Hyperglycemic memory” could be an important cause for diabetic damage. While limited information is available regarding the role of “hyperglycemic memory” in the pancreatic cancer and its radio-resistance. We therefore investigated correlation among hyperglycemic memory, Numb and metformin in pancreatic cancer radio-resistance. MethodsHigh glucose and hyperglycemic memory models were established in vitro and in vivo. Western blot, real-time PCR were accordingly used to detect Numb /Notch/ Polo-like kinase1 (PLK1) signaling at the level of molecular, cellular and experimental animal model, respectively. The apoptosis rate was evaluated by TUNEL assay and Capase-3 activity. The therapeutic effect of metformin was revealed by detecting the level of Numb / Notch /PLK1 through Western blot and real-time PCR. ResultsInactivation of Numb promotes the pancreatic cancer radio-resistance through hyperglycemic memory and metformin could suppress the radio-resistance by activating Numb in vitro and in vivo. In addition, PLK1 and Notch signaling pathway (Notch1, HEY1) elevated in pancreatic cancer radio-resistance condition, which was induced by hyperglycemic memory. Moreover, Numb overexpression or metformin could suppress Notch pathway to alleviate pancreatic cancer radio-resistance. ConclusionsOur data demonstrated that Numb might be a promising target for the improvement of hyperglycemic memory damage and the effect of metformin deserved urgent attention on pancreatic cancer radio-resistance therapy.

Roles of DNA polymerase ζ in the radiotherapy sensitivity and oxidative stress of lung cancer cells.

Previous reports stated that DNA polymerase ζ is highly expressed in lung cancer tissues. The present study aimed to investigate the roles and underlying mechanism of DNA polymerase ζ in lung cancer cell radioresistance. The A549 and HCC827 cells were used to evaluate the effects of different doses of radiation on the mRNA and protein expressions of DNA polymerase ζ. Multiple assays, including PCR, western blot, flow cytometry, and EdU have been used to investigate the roles of DNA polymerase ζ on the cellular behaviors of A549 and HCC827 cells. The mRNA and protein expression of REV7L in A549 and HCC827 cells were significantly increased after 6Gy irradiation compared to the control group. In A549 and HCC827 cells, over-expression of polymerase ζ decreased the radiosensitivity, inhibited cell apoptosis as well as reduced oxidative stress; while the knockdown of polymerase ζ showed the opposite effects. Knockdown of Polymerase ζ in xenograft animal models significantly decreased tumor size and induced oxidative stress of tumor tissues after irradiation. Our results provided a theoretical basis for targeting DNA polymerase ζ to improve the radiosensitivity of the lung cancer cells to radiotherapy.

Targeting senescence-like fibroblasts radiosensitizes non-small cell lung cancer and reduces radiation-induced pulmonary fibrosis.

Cancer cell radioresistance is the primary cause of the decreased curability of non–small cell lung cancer (NSCLC) observed in patients receiving definitive radiotherapy (RT). Following RT, a set of microenvironmental stress responses is triggered, including cell senescence. However, cell senescence is often ignored in designing effective strategies to resolve cancer cell radioresistance. Herein, we identify the senescence-like characteristics of cancer-associated fibroblasts (CAFs) after RT and clarify the formidable ability of senescence-like CAFs in promoting NSCLC cell proliferation and radioresistance through the JAK/STAT pathway. Specific induction of senescence-like CAF apoptosis using FOXO4-DRI, a FOXO4-p53–interfering peptide, resulted in remarkable effects on radiosensitizing NSCLC cells in vitro and in vivo. In addition, in this study, we also uncovered an obvious therapeutic effect of FOXO4-DRI on alleviating radiation-induced pulmonary fibrosis (RIPF) by targeting senescence-like fibroblasts in vivo. In conclusion, by targeting senescence, we offer a strategy that simultaneously decreases radioresistance of NSCLC and the incidence of RIPF.

Decreased mitochondrial membrane potential is an indicator of radioresistant cancer cells

AimsTo overcome radioresistant cancer cells, clinically relevant radioresistant (CRR) cells were established. To maintain their radioresistance, CRR cells were exposed 2 Gy/day of X-rays daily (maintenance irradiation: MI). To understand whether the radioresistance induced by X-rays was reversible or irreversible, the difference between CRR cells and those without MI for a year (CRR-NoIR cells) was investigated by the mitochondrial function as an index. Main methodsRadiation sensitivity was determined by modified high density survival assay. Mitochondrial membrane potential (Δψm) was determined by 5,5′,6,6′-tetrachloro-1,1′, tetraethylbenzimidazolocarbo-cyanine iodide (JC-1) staining. Rapid Glucose-Galactose assay was performed to determine the shift in their energy metabolism from aerobic glycolysis to oxidative phosphorylation in CRR cells. Involvement of prohibitin-1 (PHB1) in Δψm was evaluated by knockdown of PHB1 gene followed by real-time PCR. Key findingsCRR cells that exhibited resistant to 2 Gy/day X-ray lost their radioresistance after more than one year of culture without MI for a year. In addition, CRR cells lost their radioresistance when the mitochondria were activated by galactose. Furthermore, Δψm were increased and PHB1 expression was down-regulated, in the process of losing their radioresistance. SignificanceOur finding reveled that tune regulation of mitochondrial function is implicated in radioresistance phenotype of cancer cells. Moreover, as our findings indicate, though further studies are required to clarify the precise mechanisms underlying cancer cell radioresistance, radioresistant cells induced by irradiation and cancer stem cells that are originally radioresistant should be considered separately, the radioresistance of CRR cells is reversible.

Mitochondrial dysfunction induces radioresistance in colorectal cancer by activating [Ca2+]m-PDP1-PDH-histone acetylation retrograde signaling

Mitochondrial retrograde signaling (mito-RTG) triggered by mitochondrial dysfunction plays a potential role in regulating tumor metabolic reprogramming and cellular sensitivity to radiation. Our previous studies showed phos-pyruvate dehydrogenase (p-PDH) and PDK1, which involved in aerobic glycolysis, were positively correlated with radioresistance, but how they initiate and work in the mito-RTG pathway is still unknown. Our further genomics analysis revealed that complex I components were widely downregulated in mitochondrial dysfunction model. In the present study, high expression of p-PDH was found in the complex I deficient cells and induced radioresistance. Mechanistically, complex I defects led to a decreased PDH both in cytoplasm and nucleus through [Ca2+]m-PDP1-PDH axis, and decreased PDH in nucleus promote DNA damage repair (DDR) response via reducing histone acetylation. Meanwhile, NDUFS1 (an important component of the complex I) overexpression could enhance the complex I activity, reverse glycolysis and resensitize cancer cells to radiation in vivo and in vitro. Furthermore, low NDUFS1 and PDH expression were validated to be correlated with poor tumor regression grading (TRG) in local advanced colorectal cancer (CRC) patients underwent neoadjuvant radiotherapy. Here, we propose that the [Ca2+]m-PDP1-PDH-histone acetylation retrograde signaling activated by mitochondrial complex I defects contribute to cancer cell radioresistance, which provides new insight in the understanding of the mito-RTG. For the first time, we reveal that NDUFS1 could be served as a promising predictor of radiosensitivity and modification of complex I function may improve clinical benefits of radiotherapy in CRC.

Radiation-Induced Overexpression of TGFβ and PODXL Contributes to Colorectal Cancer Cell Radioresistance through Enhanced Motility.

The primary cause of colorectal cancer (CRC) recurrence is increased distant metastasis after radiotherapy, so there is a need for targeted therapeutic approaches to reduce the metastatic-relapse risk. Dysregulation of the cell-surface glycoprotein podocalyxin-like protein (PODXL) plays an important role in promoting cancer-cell motility and is associated with poor prognoses for many malignancy types. We found that CRC cells exposed to radiation demonstrated increased TGFβ and PODXL expressions, resulting in increased migration and invasiveness due to increased extracellular matrix deposition. In addition, both TGFβ and PODXL were highly expressed in tissue samples from radiotherapy-treated CRC patients compared to those from patients without this treatment. However, it is unclear whether TGFβ and PODXL interactions are involved in cancer-progression resistance after radiation exposure in CRC. Here, using CRC cells, we showed that silencing PODXL blocked radiation-induced cell migration and invasiveness. Cell treatment with galunisertib (a TGFβ-pathway inhibitor) also led to reduced viability and migration, suggesting that its clinical use may enhance the cytotoxic effects of radiation and lead to the effective inhibition of CRC progression. Overall, the results demonstrate that downregulation of TGFβ and its-mediated PODXL may provide potential therapeutic targets for patients with radiotherapy-resistant CRC.

Abstract PO-030: The harmala alkaloid harmine as a novel cancer cell radiosensitizer

Abstract Radiation therapy resistance remains a persistent clinical challenge in the treatment of multiple different cancer types. Better appreciation of novel molecular targets to improve radiation therapy efficacy is needed. TWIST1 is an epithelial-mesenchymal transcription factor whose expression is linked to therapy resistance, including radioresistance, in cancer. We have shown previously that the harmala alkaloid harmine degrades TWIST1 and possess anti-tumor activity. We hypothesized that harmine may radiosensitize cancer cells via downregulation of TWIST1. We examined the effects of harmine combined with radiation (RT) on two TWIST1 overexpressing tumor cell lines, A549 human non-small cell lung cancer (NSCLC) cells and the LMT mouse-derived hepatocellular cancer cell line (HCC), both with doxycycline-inducible Twist1 expression. We also used the LM mouse-derived HCC cell line without Twist1 expression as a control. In vitro assays for clonogenic survival, apoptosis, cell cycle distribution, γH2AX foci kinetics, and protein expression in pathways important for cancer cell survival and radioresistance were performed. Harmine alone exhibited anti-cancer activity irrespective of Twist1 expression as shown by decreased clonogenic survival. Harmine caused a supra-additive radiosensitization in all cancer cell lines with radiation enhancement ratios between 1.05 – 3.04. In vivo we are evaluating tumor growth delay and effects of combined harmine and RT treatment in a NSCLC hind-flank tumor growth model. Mechanistic elucidation of these results, studies on normal cells, and in vivo validation are ongoing. Citation Format: Caleb R.P. Smack, Audrey Lafargue, Christine Lam, Francesca A. Carrieri, Ismaeel Siddiqui, Hailun Wang, Phuoc T. Tran. The harmala alkaloid harmine as a novel cancer cell radiosensitizer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-030.

Abstract PO-079: Peroxiredoxin 4 contributes to radiation resistance of prostate cancer cells through the activation of the PI3K/AKT signaling pathway

Abstract The peroxiredoxin (Prx) family of proteins functions as major cellular antioxidants that mediate oxidative signaling under physiological conditions as well as scavenge extra hydrogen peroxide in the context of oxidative stress. Prx4 is one of Prx family members which is mainly localized in endoplasmic reticulum (ER), but can be also found in cytosol and extracellular space. In addition to its function as a peroxidase, Prx4 has a specific role in oxidative protein folding in ER. However, the specific function of Prx4 has not been well defined. Previous studies indicate that Prxs are frequently upregulated in various types of human cancer. However, the role of Prx4 in prostate cancer has not been well defined. The purpose of this study is to examine the expression of Prx4 in prostate cancer and to explore its functional significance in cancer radiation resistance and recurrence. Bioinformatic tools were used to evaluate genetic alterations of PRDX4 gene as well as the levels of its transcripts in prostate normal and cancer populations. Kaplan-Meier survival analysis was used to explore the association of Prx4 levels with the prognosis of prostate cancer patients. Western blot and immunohistochemistry were used to evaluate the expression of Prx4 protein in cell lines and patient specimens. Loss of Prx4 in cells was achieved by knock down using lentiviral shRNA or knockout using CRISPR-Cas9 techniques. Cell proliferation, survival, and protein profiler kinase arrays were used to examine the differences between control and Prx4-depleted cells with or without ionizing radiation. We demonstrated that PRDX4 gene is frequently amplified in prostate cancer and the level of its transcript is highly elevated. Patients with Prx4 at higher quartile have significantly reduced probability of survival compared with those in lower quartile. Prostate cancer cells express much higher levels of Prx4 than normal epithelial cells. Moreover, Prx4 is upregulated by the activation of AR-dependent signaling, and depletion of Prx4 sensitizes prostate cancer cells to radiation-induced cell death. Mechanistically, Prx4 contributes to prostate cancer radiation resistance through the activation of PI3K/AKT signaling pathway. A combination of bioinformatic, histochemical, cellular, and molecular methods reveals that Prx4 plays a critical role in prostate cancer cell proliferation, radio-resistance and reoccurrence. Citation Format: Na Ding, Hong Jiang, Pratik Thapa, Yanning Hao, Aziza Alshahrani, Shengming Yang, Vivek M. Rangnekar, Xiaoqi Liu, Qiou Wei. Peroxiredoxin 4 contributes to radiation resistance of prostate cancer cells through the activation of the PI3K/AKT signaling pathway [abstract]. In: Proceedings of the AACR Virtual Special Conference on Radiation Science and Medicine; 2021 Mar 2-3. Philadelphia (PA): AACR; Clin Cancer Res 2021;27(8_Suppl):Abstract nr PO-079.