Inter-assay precision of clonogenic assays for radiosensitivity in cancer cell line A549

学位記番号：医博甲171

The Role of Vitamin D in Preventing Colorectal Carcinogenesis: A Review of Molecular Mechanisms

Introduction: Colorectal carcinoma is one of the cancers with a high disease burden globally. Previous observational studies have found a connection between colorectal cancer incidence with sunlight exposure and vitamin D levels. Subsequent studies investigated this relationship further and found various anti-tumoral pathways regulated by vitamin D in colorectal tissue. This paper aims to elucidate the actions of those pathways in preventing the malignant transformation of the colorectal cell by reviewing relevant literature. Methods: A search was conducted on several medical literature electronic databases for original research studying the effects of vitamin D treatment on colorectal adenoma and colorectal cancer and its underlying anti-tumoral mechanism. A total of 122 studies were included for evaluation. Results: Twenty-seven studies passed for analysis. These in vitro and in vivo study reveals that vitamin D treatment can suppress cell proliferation, induce apoptosis, maintain cellular differentiation, reduce the pro-inflammatory response, inhibit angiogenesis, and hinder metastatic progression in colorectal cancer and colorectal adenoma cells by regulating associated gene transcription or directly prevents activation of selected signalling pathways. Five studies have also shown that adding calcium to vitamin D treatment increases the anti-tumoral activity of vitamin D through cross-talk between both of their pathways. Conclusion: Vitamin D could potentially impede colorectal cancer transformation and growth through interaction with various signalling pathways and regulating gene transcription. Further clinical studies are needed to confirm whether vitamin D can be used as the basis of targeted colorectal cancer therapy using its inherent anti-tumoral properties

Vitamin D as Radiosensitizer: A Review in Cell Line

Introduction: Vitamin D has been shown to have anti-cancer properties such as antioxidants, anti-proliferative, and cell differentiation. The property of vitamin D as an anticancer agent triggers researchers to find out whether vitamin D is useful as a radiosensitizer. Multiple studies have been carried out on cell lines in various types of cancer, but the benefits of vitamin D as a radiosensitizer still controversial. This paperwork aims to investigate the utilization of Vitamin D3 (Calcitriol) as radiosensitizer in various cell line through literature review.Methods: A systematic search of available medical literature databases was performed on in-vitro studies with Vitamin D as a radiosensitizer in all types of cell lines. A total of 11 in-vitro studies were evaluated.Results: Nine studies in this review showed a significant effect of Vitamin D as a radiosensitizer agent by promoting cytotoxic autophagy, increasing apoptosis, inhibiting of cell survival and proliferation, promoting gene in ReIB inhibition, inducing senescene and necrosis. The two remaining studies showed no significant effect in the radiosensitizing mechanism of Vitamin D due to lack of evidence in-vitro settings.Conclusion: Vitamin D have anticancer property and can be used as a radiosensitizer by imploring various mechanism pathways in various cell lines. Further research especially in-vivo settings need to be evaluated

Tumor microenvironment predicts local tumor extensiveness in PD-L1 positive nasopharyngeal cancer.

Tumor microenvironment have been implicated in many kind of cancers to hold an important role in determining treatment success especially with immunotherapy. In nasopharyngeal cancer, the prognostic role of this immune cells within tumor microenvironment is still doubtful. We conducted a study that included 25 nasopharyngeal cancer biopsy specimens to seek a more direct relationship between tumor infiltrating immune cells and tumor progression. Apart from that, we also checked the PD-L1 protein through immunohistochemistry. The PD-L1 was positively expressed in all our 25 samples with nasopharyngeal cancer WHO type 3 histology. Majority samples have >50% PD-L1 expression in tumor cells. We also found that denser local tumor infiltrating immune cells population have relatively much smaller local tumor volume. The inverse applied, with the mean local tumor volumes were 181.92 cm3 ± 81.45 cm3, 117.13 cm3 ± 88.72 cm3, and 55.13 cm3 ± 25.06 cm3 for mild, moderate, and heavy immune cells infiltration respectively (p = 0.013). Therefore, we concluded that tumor infiltrating immune cells play an important role in tumor progression, hence evaluating this simple and predictive factor may provide us with some valuable prognostic information

RNF8 promotes high linear energy transfer carbon-ion-induced DNA double-stranded break repair in serum-starved human cells

The cell-killing effect of radiotherapy largely depends on unrepaired DNA double-stranded breaks (DSBs) or lethal chromosome aberrations induced by DSBs. Thus, the capability of DSB repair status is critically important in cancer cell-killing effect after ionizing radiation. Here, we investigate the involvement of the DNA damage signaling factors ataxia telangiectasia mutated (ATM) and ring finger protein (RNF) 8 and RNF168 in quiescent G0/G1 cells, which occupy the majority population in tumors, after high-linear-energy-transfer (LET) carbon-ion irradiation. Interestingly, ATM inhibition caused a substantial DSB repair defect after high-LET carbon-ion irradiation. Similarly, RNF8 or RNF168 depletion causes a substantial DSB repair defect. ATM inhibition in RNF8-depleted cells did not have an additive effect, suggesting that ATM and RNF8 function in the same pathway. Importantly, we found that RNF8 RING mutant show similar DSB repair defect, suggesting the requirement of ubiquitin ligase activity in this repair pathway. RNF8 FHA domain, which is required for the interaction with MDC1, is also required for DSB repair in this axis. Furthermore, depletion of p53 binding protein 1 (53BP1), which is an important downstream factor in RNF8-dependent DSB repair, is also required for this repair. Importantly, either ATM inhibition or RNF8 depletion increased the frequency of chromosomal breaks, but reduced dicentric chromosome formation, demonstrating that ATM/RNF8 is required for the rejoining of DSB ends for dicentric chromosome formation. Finally, we show that RNF8 depletion augmented radiosensitivity after high-LET carbon-ion irradiation. Taken together, these results suggest that the inhibition of RNF8 activity or its downstream pathway may augment the efficacy of high-LET carbon-ion therapy

Expression of non‑homologous end joining factor, Ku80, is negatively correlated with PD‑L1 expression in cancer cells after X‑ray irradiation

The growing importance of antitumour immunity by cancer immunotherapy has prompted studies on radiotherapy‑induced immune response. Previous studies have indicated that programmed cell death‑1 ligand (PD‑L1) expression is regulated by DNA damage signalling. However, PD‑L1 up‑regulation after radiotherapy has not been fully investigated at the clinical level, particularly in the context of expression of DNA repair factors. The present study examined the correlation of mRNA expression between PD‑L1 and non‑homologous end joining (NHEJ) factors using The Cancer Genome Atlas database analysis. Among NHEJ factors, Ku80 mRNA expression was negatively correlated with PD‑L1 mRNA expression levels in several types of cancer (colon adenocarcinoma, breast invasive carcinoma, skin cutaneous melanoma, lung adenocarcinoma, head and neck squamous cell carcinoma, uterine corpus endometrial carcinoma, cervical squamous cell carcinoma and endocervical adenocarcinoma). To verify the negative correlation in clinical samples, the present study analysed whether Ku80 expression levels affected PD‑L1 up‑regulation after radiotherapy using cervical squamous cell carcinoma samples. Quantitative evaluation using software analysis of immunohistochemically stained slides revealed that patients with low Ku80 positivity in biopsy specimens demonstrated increased PD‑L1 expression levels after 10 Gy irradiation (Spearman\u27s rank correlation coefficient=‑0.274; P=0.017). Furthermore, PD‑L1 induction levels in tumour cells after 10 Gy of irradiation were significantly inversely correlated with Ku80 expression levels (Spearman\u27s rank correlation coefficient=‑0.379; P<0.001). The present study also confirmed that short interfering RNA‑mediated Ku80 depletion was associated with greater X‑ray‑induced PD‑L1 up‑regulation in HeLa cells. These results indicated that radiotherapy could enhance PD‑L1 induction in tumour cells with low Ku80 expression in a clinical setting. Furthermore, these data highlighted Ku80 as a potential predictive biomarker for immune checkpoint therapy combined with radiotherapy

DNA double-strand break repair pathway regulates PD-L1 expression in cancer cells

Accumulating evidence suggests that exogenous cellular stress induces PD-L1 upregulation in cancer. A DNA double-strand break (DSB) is the most critical type of genotoxic stress, but the involvement of DSB repair in PD-L1 expression has not been investigated. Here we show that PD-L1 expression in cancer cells is upregulated in response to DSBs. This upregulation requires ATM/ATR/Chk1 kinases. Using an siRNA library targeting DSB repair genes, we discover that BRCA2 depletion enhances Chk1-dependent PD-L1 upregulation after X-rays or PARP inhibition. In addition, we show that Ku70/80 depletion substantially enhances PD-L1 upregulation after X-rays. The upregulation by Ku80 depletion requires Chk1 activation following DNA end-resection by Exonuclease 1. DSBs activate STAT1 and STAT3 signalling, and IRF1 is required for DSB-dependent PD-L1 upregulation. Thus, our findings reveal the involvement of DSB repair in PD-L1 expression and provide mechanistic insight into how PD-L1 expression is regulated after DSBs

Analysis of radiotherapy‑induced alteration of CD8+ T cells and PD‑L1 expression in patients with uterine cervical squamous cell carcinoma

Abstract. Radiotherapy induces an immune response in the cancer microenvironment that may influence clinical outcome. The present study aimed to analyse the alteration of CD8+ T‑cell infiltration and programmed death‑ligand 1 (PD‑L1) expression following radiotherapy in clinical samples from patients with uterine cervical squamous cell carcinoma. Additionally, the current study sought to analyse the association between these immune responses and clinical outcomes. A total of 75 patients who received either definitive chemoradiotherapy or radiotherapy were retrospectively analyzed. CD8+ T‑cell infiltration and PD‑L1 expression were determined by immunohistochemistry using biopsy specimens before radiotherapy (pre‑RT) and after 10 Gy radiotherapy (post‑10 Gy). The PD‑L1+ rate was significantly increased from 5% (4/75) pre‑RT to 52% (39/75) post‑10 Gy (P<0.01). Despite this increase in the PD‑L1+ rate post‑10 Gy, there was no significant association between both pre‑RT and post‑10 Gy and overall survival (OS), locoregional control (LC) and progression‑free survival (PFS). On the other hand, the CD8+ T‑cell infiltration density was significantly decreased for all patients (median, 23.1% pre‑RT vs. 16.9% post‑10 Gy; P=0.038); however, this tended to increase in patients treated with radiotherapy alone (median, 17.7% pre‑RT vs. 24.0% post‑10 Gy; P=0.400). Notably, patients with high CD8+ T‑cell infiltration either pre‑RT or post‑10 Gy exhibited positive associations with OS, LC and PFS. Thus, the present analysis suggested that CD8+ T‑cell infiltration may be a prognostic biomarker for patients with cervical cancer receiving radiotherapy. Furthermore, immune checkpoint inhibitors may be effective in patients who have received radiotherapy, since radiotherapy upregulated PD‑L1 expression in cervical cancer specimens

Mutational analysis of uterine cervical cancer that survived multiple rounds of radiotherapy.

Radiotherapy is an essential component of cancer therapy. Despite advances in cancer genomics, the mutation signatures of radioresistant tumors have not yet been fully elucidated. To address this issue, we analyzed a unique set of clinical specimens from a uterine cervical cancer that repeatedly locally recurred after multiple rounds of radiotherapy. Exon sequencing of 409 cancer-related genes in the treatment-naïve tumor and the tumors that recurred after initial and secondary radiotherapy identified (i) activating mutations in and , and putative inactivating mutations in , as trunk mutation signatures that persisted over the clinical course; and (ii) mutations in , , and as acquired mutation signatures observed only in recurrent tumors after radiotherapy. Comprehensive mining of published genomics data pertaining to radiosensitivity revealed that simultaneous mutations in and , which have not been described previously in uterine cervical cancer, are associated with cancer cell radioresistance. The association between this mutation signature and radioresistance was validated by isogenic cell-based experiments. These results provide proof-of-principle for the analytical pipeline employed in this study, which explores clinically relevant mutation signatures for radioresistance, and demonstrate that this approach is worth pursuing with larger cohorts in the future