Mutant p53 Mediates Sensitivity to Cancer Treatment Agents in Oesophageal Adenocarcinoma Associated with MicroRNA and SLC7A11 Expression

TP53 gene mutations occur in 70% of oesophageal adenocarcinomas (OACs). Given the central role of p53 in controlling cellular response to therapy we investigated the role of mutant (mut-) p53 and SLC7A11 in a CRISPR-mediated JH-EsoAd1 TP53 knockout model. Response to 2 Gy irradiation, cisplatin, 5-FU, 4-hydroxytamoxifen, and endoxifen was assessed, followed by a TaqMan OpenArray qPCR screening for differences in miRNA expression. Knockout of mut-p53 resulted in increased chemo- and radioresistance (2 Gy survival fraction: 38% vs. 56%, p < 0.0001) and in altered miRNA expression levels. Target mRNA pathways analyses indicated several potential mechanisms of treatment resistance. SLC7A11 knockdown restored radiosensitivity (2 Gy SF: 46% vs. 73%; p = 0.0239), possibly via enhanced sensitivity to oxidative stress. Pathway analysis of the mRNA targets of differentially expressed miRNAs indicated potential involvement in several pathways associated with apoptosis, ribosomes, and p53 signaling pathways. The data suggest that mut-p53 in JH-EsoAd1, despite being classified as non-functional, has some function related to radio- and chemoresistance. The results also highlight the important role of SLC7A11 in cancer metabolism and redox balance and the influence of p53 on these processes. Inhibition of the SLC7A11-glutathione axis may represent a promising approach to overcome resistance associated with mut-p53.     

Mutation Analysis of Radioresistant Early-Stage Cervical Cancer

Radiotherapy is a definitive treatment for early-stage cervical cancer; however, a subset of this disease recurs locally, necessitating establishment of predictive biomarkers and treatment strategies. To address this issue, we performed gene panel-based sequencing of 18 stage IB cervical cancers treated with definitive radiotherapy, including two cases of local recurrence, followed by in vitro and in silico analyses. Simultaneous mutations in KRAS and SMAD4 (KRAS^mt/SMAD4^mt) were detected only in a local recurrence case, indicating potential association of this mutation signature with radioresistance. In isogenic cell-based experiments, a combination of activating KRAS mutation and SMAD4 deficiency led to X-ray resistance, whereas either of these factors alone did not. Analysis of genomic data from 55,308 cancers showed a significant trend toward co-occurrence of mutations in KRAS and SMAD4. Gene Set Enrichment Analysis of the Cancer Cell Line Encyclopedia dataset suggested upregulation of the pathways involved in epithelial mesenchymal transition and inflammatory responses in KRAS^mt/SMAD4^mt cancer cells. Notably, irradiation with therapeutic carbon ions led to robust killing of X-ray-resistant KRAS^mt/SMAD4^mt cancer cells. These data indicate that the KRAS^mt/SMAD4^mt signature is a potential predictor of radioresistance, and that carbon ion radiotherapy is a potential option to treat early-stage cervical cancers with the KRAS^mt/SMAD4^mt signature.     

Translational Implications for Radiosensitizing Strategies in Rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma of childhood and adolescence that includes FP-RMS, harboring the fusion oncoprotein PAX3/7-FOXO1 and FN-RMS, often mutant in the RAS pathway. Risk stratifications of RMS patients determine different prognostic groups and related therapeutic treatment. Current multimodal therapeutic strategies involve surgery, chemotherapy (CHT) and radiotherapy (RT), but despite the deeper knowledge of response mechanisms underpinning CHT treatment and the technological improvements that characterize RT, local failures and recurrence frequently occur. This review sums up the RMS classification and the management of RMS patients, with special attention to RT treatment and possible radiosensitizing strategies for RMS tumors. Indeed, RMS radioresistance is a clinical problem and further studies aimed at dissecting radioresistant molecular mechanisms are needed to identify specific targets to hit, thus improving RT-induced cytotoxicity.     

抗辐射菌Deinococcus radiodurans一种新的DNA修复基因pprA的分子克隆测序及特性研究(英文)

抗辐射菌Ｄｅｉｎｏｃｏｃｕｓｒａｄｉｏｄｕｒａｎｓ具有显著的ＤＮＡ修复能力，包括对丝裂霉素（ＭＣ），紫外线（ＵＶ）及γ射线。通过化学诱导野生型菌株ＫＤ８３０１而得到的突变体ＫＨ３１１１，对ＭＣ，ＵＶ及γ射线敏感，对链霉素（Ｓｍ）具有抗性。ＫＨ３１１１的辐射抗性能够通过转染野生型抗辐射菌ＫＤ８３０１基因组中一个已被克隆的ＤＮＡ片断而得到恢复。转染实验表明，突变体具有很好的自然转化能力，即不缺乏重组修复能力。本研究确定了ＫＨ３１１１内发生突变的基因及其核苷酸序列，该基因（ｏｒｆ１４４ｂ）所编码的蛋白质由２８４个氨基酸组成，与其它已知的蛋白质不具有同源性，即它是一种新的蛋白质。通过ＤＮＡ测序分析，ＫＤ８３０１和ＫＨ３１１１的ｏｒｆ１４４ｂ序列只是一个碱基的改变，即由Ｇ突变为Ａ，所编码的第１４９位氨基酸由甘氨酸变为谷氨酰胺。说明甘氨酸是ＤＮＡ修复基因ｏｒｆ１４４ｂ所编码的蛋白质中的一个重要残基。用ＫＤ８３０１的ｏｒｆ１４４ｂ基因转染ＫＨ３１１１，得到转染体ＫＨ３１１２，其对ＭＣ，ＵＶ及γ射线都具有同母本（ＫＤ８３０１）一样的抗性。在抗辐射菌中，这种对多种ＤＮＡ损伤剂产生抗性所需要的基因（ｏｒｆ１４４ｂ），我们命名为     

Dual Knockdown of Musashi RNA-Binding Proteins MSI-1 and MSI-2 Attenuates Putative Cancer Stem Cell Characteristics and Therapy Resistance in Ovarian Cancer Cells

In ovarian cancer, therapy resistance mechanisms complicate cancer cell eradication. Targeting Musashi RNA-binding proteins (MSI) may increase therapeutic efficacy. Database analyses were performed to identify gene expression associations between MSI proteins and key therapy resistance and cancer stem cell (CSC) genes. Then, ovarian cancer cells were subjected to siRNA-based dual knockdown of MSI-1 and MSI-2. CSC and cell cycle gene expression was investigated using quantitative polymerase chain reaction (qPCR), western blots, and flow cytometry. Metabolic activity and chemoresistance were assessed by MTT assay. Clonogenic assays were used to quantify cell survival post-irradiation. Database analyses demonstrated positive associations between MSI proteins and putative CSC markers NOTCH, MYC, and ALDH4A1 and negative associations with NOTCH inhibitor NUMB. MSI-2 expression was negatively associated with the apoptosis regulator p21. MSI-1 and MSI-2 were positively correlated, informing subsequent dual knockdown experiments. After MSI silencing, CSC genes were downregulated, while cell cycle progression was reduced. Metabolic activity was decreased in some cancer cells. Both chemo- and radioresistance were reduced after dual knockdown, suggesting therapeutic potential. Dual knockdown of MSI proteins is a promising venue to impede tumor growth and sensitize ovarian cancer cells to irradiation and chemotherapy.     

重离子辐照微生物效应及诱变育种进展

重离子束作为一种高效的、典型的诱变辐射源广泛用于生物育种。不同质量数、电荷数和不同能量的重离子束作用于生物细胞有特殊的过程和作用,其先后经历物理、化学和生物学三个阶段,最终引发终点生物学效应,如辐射低剂量超敏感现象,活性氧代谢变化等。本文重点探讨重离子束辐照微生物体后出现的主要终点生物效应,以及近年来重离子辐照微生物诱变育种在工农业等方面取得的成果。     

A Century of Fractionated Radiotherapy: How Mathematical Oncology Can Break the Rules

Radiotherapy is involved in 50% of all cancer treatments and 40% of cancer cures. Most of these treatments are delivered in fractions of equal doses of radiation (Fractional Equivalent Dosing (FED)) in days to weeks. This treatment paradigm has remained unchanged in the past century and does not account for the development of radioresistance during treatment. Even if under-optimized, deviating from a century of successful therapy delivered in FED can be difficult. One way of exploring the infinite space of fraction size and scheduling to identify optimal fractionation schedules is through mathematical oncology simulations that allow for in silico evaluation. This review article explores the evidence that current fractionation promotes the development of radioresistance, summarizes mathematical solutions to account for radioresistance, both in the curative and non-curative setting, and reviews current clinical data investigating non-FED fractionated radiotherapy.     

Hypersensitivity and Induced Radioresistance in Chinese Hamster Cells Exposed to Radiations with Different LET Values

We study the impact of radiation LET on manifestation of HRS/IRR response in Chinese hamster cells ovary cells exposed to radiations used in radiotherapy. Earlier we have investigated this response to carbon ions (455 MeV/amu) in the pristine Bragg curve plateau and behind the Bragg peak, ⁶⁰Co γ-rays, and 14.5 MeV neutrons. Now we present results of cytogenetic metaphase analysis in plateau-phase CHO-K1 cells irradiated with scanning beam protons (83 MeV) at doses < 1 Gy and additional data for 14.5 MeV neutrons. Dose curves for frequency of total chromosome aberrations (CA, protons), paired fragments (protons, neutrons), aberrant cells (neutrons) had typical HRS/IRR structure: HRS region (up to 0.1 and 0.15 Gy), IRR region (0.1–0.6 Gy and 0.15–0.35 Gy) for protons and neutrons, respectively, and regular dose dependence. Taken together with previous results, the data show that LET increase shifts the HRS upper border (from 0.08–0.1 Gy for γ-rays, protons and plateau carbons to 0.12–0.15 Gy for “tail” carbons and neutrons). The IRR regions shortens (0.52–0.4 γ-rays and protons, 0.25 plateau carbons, 0.2 Gy “tail” carbons and neutrons). CA level of IRR increases by 1.5–2.5 times for carbons as compared to γ-rays and protons. Outside HRS/IRR the yield of CA also enhanced with LET increase. The results obtained for different LET radiations suggest that CHO-K1 cells with G1-like CA manifested the general feature of the HRS/IRR phenomena.