Biomarkers Of Radiosensitivity Research Articles

Prognostic and Potential Therapeutic Roles of PRKDC Expression in Lung Cancer.

PRKDC is a key factor involved in the ligation step of the non-homologous end joining pathway. Its dysfunction has proven to be a biomarker for radiosensitivity of cancer cells. However, the prognostic value of PRKDC and its underlying mechanisms have not been clarified yet. In this study, we found that PRKDC overexpressed in lung adenocarcinoma (LUAD) and is significantly related to unfavorable survival, while downregulation of PRKDC is link to inflamed tumor immune signature. Our further in vitro results also showed a potent antitumor efficacy of PRKDC inhibitors alone or combined with cisplatin in human lung cancer cells. This study demonstrated that PRKDC is a potential prognostic biomarker, immunotherapy target, and promising combination candidate for chemotherapy for lung cancer, and highlighted the potential of PRKDC-targeted inhibitors for the treatment of lung cancer.

Current Status of Genetic/Molecular Abnormality Analysis and Prognosis Prediction of Meningioma

Biological molecular studies of meningiomas have also developed with the development of molecular biological methods. In 2013, Clark et al. reported that driver genetic mutations other than NF2, including TRAF7, KLF4, AKT1, and SMO, were associated with meningioma development. In 2017, Sahm et al. proposed a classification of meningiomas based on global methylation status, which was more accurate in predicting prognosis than conventional WHO grading. In 2022, based on this classification, various groups reported an integrated classification that comprehensively included some biological molecular abnormalities, such as DNA mutations, copy number alterations, and RNA sequences. This field is expected to elucidate the mechanism of meningioma development and further research is expected to lead to the development of effective molecularly targeted therapeutics and biomarkers of radiosensitivity in the future. In this article, we summarize the current status and prospects of these biological molecular studies.

Targeted irradiation in an autochthonous mouse model of pancreatic cancer.

The value of radiotherapy in the treatment of pancreatic cancer has been the subject of much debate but limited preclinical research. We hypothesise that the poor translation of radiation research into clinical trials of radiotherapy in pancreatic cancer is due, in part, to inadequate preclinical study models. Here, we developed and refined methods for targeted irradiation in autochthonous mouse models of pancreatic cancer, using a small animal radiotherapy research platform. We tested and optimised strategies for administration of contrast agents, iohexol and the liver imaging agent Fenestra LC, to enable the use of computed tomography imaging in tumour localisation. We demonstrate accurate tumour targeting, negligible off-target effects and therapeutic efficacy, depending on dose, number of fractions and tumour size, and provide a proof of concept that precise radiation can be delivered effectively to mouse pancreatic tumours with a clinically relevant microenvironment. This advance will allow investigation of the radiation response in murine pancreatic cancer, discovery of mechanisms and biomarkers of radiosensitivity or resistance, and development of radiosensitising strategies to inform clinical trials for precision radiotherapy in this disease.

A potential biomarker of radiosensitivity in metastatic hormone sensitive prostate cancer patients treated with combination external beam radiotherapy and radium-223

PurposeThe ADRRAD trial reported the safety and feasibility of the combination of external beam radiotherapy and radium-223 in the treatment of de novo bone metastatic prostate. This study aimed to determine if any biomarkers predictive of response to these treatments could be identified. Experimental design30 patients with newly diagnosed bone metastatic hormone sensitive prostate cancer were recruited to the ADRRAD trial. Blood samples were taken pre-treatment, before cycles 2 to 6 of radium-223, and 8 weeks and 6 months after treatment. Mononuclear cells were isolated and DNA damage was assessed at all timepoints. ResultsDNA damage was increased in all patients during treatment, with bigger increases in foci observed in patients who relapsed late compared to those who relapsed early. Increases in DNA damage during the radium-223 only cycles of treatment were specifically related to response in these patients. Analysis of hematology counts also showed bigger decreases in red blood cell and hemoglobin counts in patients who experienced later biochemical relapse. ConclusionsWhile some patients responded to this combination treatment, others relapsed within one year of treatment initiation. This study identifies a biomarker based approach that may be useful in predicting which patients will respond to treatment, by monitoring both increases in DNA damage above baseline levels in circulating lymphocytes and decreases in red blood cell and hemoglobin counts during treatment.

Genomic Correlates of Radiosensitivity in Diffuse Large B Cell Lymphoma

Introduction Radiotherapy (RT) is beneficial for select patients with localized diffuse large B cell lymphoma (DLBCL), in both the early stage and relapsed/refractory settings. Although molecular subtyping plays a key role in classification of DLBCL, genomic predictors of local response to RT have yet to be adequately explored. To our knowledge, there are no existing studies investigating the genomic landscape of DLBCL patients treated with RT. Therefore, we sought to identify genomic signatures of radiosensitivity in localized DLBCL patients. Methods We analyzed an institutional database of DLBCL patients treated with RT for localized measurable disease treated between 2007-2022. All patients underwent next-generation sequencing using an institutional 505-gene panel (MSK-Impact). Clinical and radiographic characteristics were obtained from chart and imaging review. All patients had measurable disease on CT scan at time of RT and included both Ann Arbor stage I/II patients referred for consolidative radiotherapy and those with isolated sites of relapsed/refractory disease. Patients did not receive any systemic therapy between RT start and time of response assessment. The public NCI LymphGen algorithm was used to further classify genomic subgroups. Using Fisher's exact test and logistic regression, we tested for univariable correlation between specific gene alterations and LymphGen subgroups with initial radiation response, defined by Lugano criteria and decrease in maximum SUV. Results We identified 46 patients including 15 (33%) tumors treated in the early-stage setting (13% definitively without prior chemotherapy, 87% as consolidation), while 31 (60%) were treated in the localized relapsed/refractory setting. The majority were GCB subtype (57%) while 28% were non-GCB and 15% had missing immunohistochemistry data. The median number of prior lines of systemic therapy was 1 (0-2) in early stage and 1 (1-7) in localized relapsed/refractory diseases. The median RT dose was 36 Gy (4-54), with 38 (83%) tumors receiving at least 30Gy. The median lesion SUV at the time of RT was 4.11 (1.1-46.0). Complete response (CR) was achieved in 41% of tumors at the time of first imaging assessment (median 3.1 months) with a median of 90.3% (42-100%) decrease in SUV. The most commonly altered individual genes were KMT2D (35%), MYD88 (28%), and TP53 (35%). LymphGen classified 22% of cases as EZB subtype, 13% as MCD, 2% as BN2, 2% as ST2, and the remaining 61% were unclassified. Mutations in ETV6 (13% of tumors) were associated with a higher CR rate in response to RT (83% mutated vs 35% wildtype, p=0.068). Lesions from ETV6 mutated cases had similar SUVs (p=0.74) and sizes (p=0.92) as wildtype cases. Alterations in genes from the LymphGen MCD subgroup were associated with greater reductions in SUV (ETV6 p<0.0001, PIM1 p<0.0001, MYD88 p<0.001, and TNFAIP3 p=0.02) The MCD subgroup trended towards increased odds of CR post-RT (log odds 1.9, p=0.08) and greater reduction of SUV (p=0.07) compared to the EZB subgroup. Conclusions To our knowledge this is the first study evaluating the role of LymphGen subgrouping on radiotherapy response, and the first study to examine genomic alterations in DLBCL patients treated with radiation. Genomic biomarkers of radiosensitivity are needed to identify radioresponsive tumors which may benefit from better integration of RT into existing salvage paradigms such as CAR T cells. Our data suggest that alterations in MCD LymphGen subgroup genes (ETV6, PIM1, MYD88, TNFAIP3) may be associated with better response to radiotherapy. ETV6 is a strong hematopoietic transcription factor known to be altered in many human leukemias and myelodysplastic syndromes. Its role in DLBCL is not well defined, with preclinical studies suggesting a distinct role compared to leukemias. Due to the limited sample size, these findings are hypothesis generating and warrant further study with larger, prospective cohorts to validate the ability of genomic signatures to predict RT response.

PCL22-186: A Potential Biomarker for Radiosensitivity of Squamous Cell Carcinomas in the Head and Neck

PCL22-186: A Potential Biomarker for Radiosensitivity of Squamous Cell Carcinomas in the Head and Neck

Detection of Radiation-Induced DNA Damage in Breast Cancer Patients by Using Gamma H2AX Biomarker: A Possible Correlation with Their Body Mass Index.

Radiotherapy is one of the most important options for treating breast cancer in humans. The development of biomarkers to monitor radiosensitivity is scarce. The aim of this study is to investigate the γH2AX levels in the human blood samples 0.5 h after radiotherapy compared to the levels before radiotherapy in breast cancer patients in relation to their respective body mass index (BMI). Blood plasma samples were collected from a total of 20 breast cancer patients before and after radiotherapy to measure γH2AX levels with an antibody against γH2AX based on enzyme-linked immunosorbent assay technique. The median BMI of the patients was 30 kg/m2. γH2AX was differentially expressed in breast cancer patients before radiotherapy. γH2AX levels significantly increased in 14 patients after radiotherapy (P = 0.006), whereas γH2AX levels decreased in three patients after radiotherapy, and three patients were excluded. There was no correlation between γH2AX values after radiotherapy and BMI (P = 0.5, r = 0.1). Our results suggest that γH2AX can be used by ELISA technique to measure γH2AX in the blood plasma of breast cancer patients undergoing radiotherapy and can be considered a biomarker of radiosensitivity.

Emerging Role of Aurora A in Radioresistance: A Comprehensive Review

Radiotherapy is one of the most conventional modes of treatment in several cancers. Failure of radiotherapy followed by acquisition of radioresistance is one of the emerging challenges faced by clinical experts. Unusual expression and functional implications of several molecules are observed to facilitate radioresistance. Aurora A, a member of the Aurora kinase (serine/threonine kinase) family, is one such molecule that shows significantly altered expression as well as non-canonical functional crosstalk with other associated factors (cell cycle regulators, signaling molecules, stemness markers, etc.) to favour the adaptations for the acquirement of radioresistance. These mechanisms include progression of cell cycle, stimulatory activation of factors by phosphorylation for enhancing the chance of cellular survivability, and prevention of apoptosis. This review article summarises how Aurora A is responsible for radioresistance in cancer and why this kinase should be considered a negative biomarker of radiosensitivity. This review discloses a wider opportunity in the field of research to find the mechanistic key regulatory pathway of Aurora A, which can be a potential target for enhancing the efficiency of treatment. Further investigations are required to explore the potential of Aurora A inhibitors as reliable radiosensitisers.

Insulin-like growth factor binding protein 3 promotes radiosensitivity of oral squamous cell carcinoma cells via positive feedback on NF-\u03baB/IL-6/ROS signaling

BackgroundEctopic insulin-like growth factor binding protein 3 (IGFBP3) expression has been shown to enhance cell migration and lymph node metastasis of oral squamous cell carcinoma (OSCC) cells. However, OSCC patients with high IGFBP3 expression had improved survival compared with those with low expression. Therefore, we speculated that IGFBP3 expression may play a role in response to conventional OSCC therapies, such as radiotherapy.MethodsWe used in vitro and in vivo analyses to explore IGFBP3-mediated radiosensitivity. Reactive oxygen species (ROS) detection by flow cytometry was used to confirm IGFBP3-mediated ionizing radiation (IR)-induced apoptosis. Geneset enrichment analysis (GSEA) and ingenuity pathway analysis (IPA) were used to analyze the relationship between IGFBP3 and nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling. Assays involving an NF-κB inhibitor, ROS scavenger or interleukin 6 (IL-6) were used to evaluate the NF-κB/IL-6/ROS signaling in IGFBP3-mediated radiosensitivity.ResultsEctopic IGFBP3 expression enhanced IR-induced cell-killing in vitro. In vivo, IGFBP3 reduced tumor growth and increased apoptotic signals of tumor tissues in immunocompromised mice treated with IR. Combined with IR, ectopic IGFBP3 expression induced mitochondria-dependent apoptosis, which was apparent through mitochondrial destruction and increased ROS production. Ectopic IGFBP3 expression enhanced NK-κB activation and downstream cytokine expression. After IR exposure, IGFBP3-induced NF-κB activation was inhibited by the ROS scavenger N-acetyl-L-cysteine (NAC). IGFBP3-mediated ROS production was reduced by the NF-κB inhibitor BMS-345541, while exogenous IL-6 rescued the NF-κB-inhibited, IGFBP3-mediated ROS production.ConclusionsOur data demonstrate that IGFBP3, a potential biomarker for radiosensitivity, promotes IR-mediated OSCC cell death by increasing ROS production through NF-κB activation and cytokine production.

Developing Tumor Radiosensitivity Signatures Using LncRNAs.

Long non-coding RNAs (lncRNAs) are involved in diverse biological processes, including DNA damage repair, and are of interest as potential biomarkers of radiosensitivity. We investigated whether lncRNA radiosensitivity signatures could be derived for use in cancer patients treated with radiotherapy. Signature development involved radiosensitivity measurements for cell lines and primary tumor samples, and patient outcome after radiotherapy. A 10-lncRNA signature trained on radiosensitivity measurements in bladder cell lines showed a trend towards independent validation. In multivariable analyses, patients with tumors classified as radioresistant by the lncRNA signature had poorer local relapse-free survival (P = 0.065) in 151 patients with muscle-invasive bladder cancer who underwent radiotherapy. An mRNA-based radiosensitivity index signature performed similarly to the lncRNA bladder signature for local relapse-free survival (P = 0.055). Pathway analysis showed the lncRNA signature associated with molecular processes involved in radiation responses. Knockdown of one of the lncRNAs in the signature showed a modest increase in radiosensitivity in one cell line. An alternative approach involved training on primary cervical tumor radiosensitivity or local control after radiotherapy. Both approaches failed to generate a cervix lncRNA radiosensitivity signature, which was attributed to the age of samples in our cohorts. Our work highlights challenges in validating lncRNA signatures as biomarkers in archival tissue from radiotherapy cohorts, but supports continued investigation of lncRNAs for a role in radiosensitivity.

Alteration in Expression of Trim29, TRIM37, TRIM44, and β-Catenin Genes After Irradiation in Human Cells with Different Radiosensitivity.

Introduction: Radiotherapy is a crucial component of treatment for ∼70% of all cancer patients. The identification of effective biomarkers of radiosensitivity (RS) is a fundamental goal of radiobiology. The authors hypothesize that the RS of human normal and tumoral cells is correlated by the level of expression of TRIM29, TRIM37, TRIM44, and β-catenin genes. Materials and Methods: Clonogenic assay was performed and RS of four cell lines was determined by survival fraction at 2 Gy. To determine the level of gene expression 6 and 24 h after irradiation, RNA was extracted from each cell line, and expression of the above-mentioned genes in cell lines with different RS was determined by real-time polymerase chain reaction (PCR). Results: The clonogenic assay showed that human dermal fibroblasts (fibroblast) and HT-29 (colorectal) cells are radioresistant, while human foreskin fibroblasts (fibroblast) and QU-DB (lung) cells are radiosensitive. Analysis of the real-time PCR data, 6 h after irradiation, showed that the increase and decrease of the expression of TRIM29 and TRIM37 genes were directly correlated with the RS of normal and tumor cells. At 24 h postirradiation, a considerable difference was only observed in the expression of the β-catenin gene. Conclusion: This study showed that the TRIM29 and TRIM37 genes are involved in the cell response to radiation and proposed that these genes may be biomarkers for predicting RS in normal and tumoral cell lines.

Predictive value of CD8/FOXP3 ratio combined with PD-L1 expression for radiosensitivity in patients with squamous cell carcinoma of the larynx receiving definitive radiation therapy.

Little is known about immune-related radiosensitivity in patients with squamous cell carcinoma of the larynx (SCC-L) treated with radiation therapy (RT). We retrospectively reviewed 91 patients with SCC-L treated with RT or chemoradiation therapy and performed immunohistochemical examination to analyze PD-L1 level on tumor cells, CD4+ tumor-infiltrating lymphocytes (TILs), CD8+ TIL, and FOXP3+ TIL using pretreated biopsy specimens. The association between these immune-related parameters and radiosensitivity was evaluated. Multivariate analyses showed that high CD8/FOXP3 ratio combined with negative PD-L1 expression was an independent and significant favorable predictive factor for local control, compared with the other groups. We showed that high CD8/FOXP3 ratio combined with negative PD-L1 expression might be a useful biomarker of radiosensitivity in patients with SCC-L receiving definitive RT. We propose that coassessment of CD8/FOXP3 ratio and PD-L1 expression level in tumor cells can help predict potential radiosensitivity in patients with SCC-L.

Inhibition of Delta-like Ligand 4 enhances the radiosensitivity and inhibits migration in cervical cancer via the reversion of epithelial\u2013mesenchymal transition

BackgroundConcurrent chemoradiotherapy is the common first-line treatment for patients with advanced cervical cancer. However, radioresistance remains a major clinical challenge, which results in recurrence and poor survival. Many studies have shown the potential of Delta-like Ligand 4 (DLL4) as a novel prognostic biomarker and therapeutic target in many solid tumors. Previously, we have found that high DLL4 expression in tumor cells may predict the pelvic lymph node metastasis and poor prognosis in patients with cervical cancer. In our present study, we further studied the effects of DLL4 on the biological behavior and radiosensitivity of cervical cancer cells.MethodsThe expression of DLL4 and epithelial–mesenchymal transition (EMT) phenotype markers in cervical cancer cell lines or tissues were detected using Western blotting, and the expression of DLL4 mRNA in cervical cancer cell lines or tissues was detected using Quantitative real-time PCR. The effect of DLL4 on cell proliferation, migration, and radiosensitivity was evaluated using the CCK8 assay, flow cytometry, Transwell assays for cell invasion and migration, and Immunofluorescence staining in vitro.ResultsThe expression of DLL4 in radiotherapy-resistant SiHa cells was significantly higher than that in radiotherapy-sensitive Me-180 cells. Furthermore, downregulation of DLL4 enhanced the radiosensitivity of SiHa and Caski cells via the inhibition of cell proliferation, promotion of radiation-induced apoptosis, and inhibition of the DNA damage repair. Moreover, downregulation of DLL4 inhibited the EMT and reduced the proliferation, invasion, and migration ability in SiHa and Caski cells. Consistent with the DLL4 expression in the cell lines, the expression of DLL4 in the tissues of the radioresistant group was also higher than that of the radiosensitive group.ConclusionsDownregulation of DLL4 inhibited the progression and increased the radiosensitivity in cervical cancer cells by reversing EMT. These results indicated the promising prospect of DLL4 against the radioresistance and metastasis of cervical cancer and its potential as a predictive biomarker for radiosensitivity and prognosis in patients with cervical cancer patients receiving concurrent chemoradiotherapy (cCRT).

Evaluation of the influence of chronic low-dose radiation on DNA repair gene polymorphisms [XRCC1, XRCC3, PRKDC (XRCC7), LIG1, NEIL1] in individuals from normal and high level natural radiation areas of Kerala Coast

Background: Single Nucleotide Polymorphisms (SNPs) at DNA repair genes are considered as potential biomarkers of radio-sensitivity. The coastal belt of Kerala in south west India has a patchy distribution of monazite in its beach sand that contains Th-232 and its decay products. Thus, radiation levels in this area vary from <1.0mGy to 45.0mGy/year. The areas with external gamma radiation dose >1.5mGy/year are considered as High-Level Natural Radiation Areas (HLNRA) and ≤ 1.5mGy/year are Normal Level Natural Radiation Area (NLNRA).Objective: In the present study, an attempt was made to evaluate the influence of chronic low dose radiation exposure on DNA repair gene polymorphisms in NLNRA and HLNRA population of Kerala coast.Materials and methods: Genomic DNA was isolated from venous blood samples of 246 random, healthy individuals (NLNRA, N = 104; HLNRA, N = 142) and genotyping of five SNPs such as X-ray repair cross complementing 1(XRCC1 Arg399Gln), X-ray repair cross complementing 3 (XRCC3 Thr241Met], Protein kinase, DNA-activated, catalytic subunit (PRKDC) (X-ray repair cross-complementing group 7, XRCC7 G/T), nei like DNA glycosylase 1 (NEIL1 G/T) and DNA ligase 1 (LIG1 A/C) was carried out using PCR based restriction fragment length polymorphism (PCR-RFLP) followed by silver staining.Results: Our results showed no significant difference in genotype frequencies in HLNRA vs NLNRA at three of the five SNPs studied i.e. XRCC1 Arg399Gln (χ2(2) = 5.85, p = .054), XRCC3 Thr241Met (χ2(1) = 0.71, p = .339), PRKDC (XRCC7 G/T) (χ2(2) = 3.72, p = .156), whereas significant difference was observed at NEIL1 G/T (χ2(2) =8.71, p = .013) and LIG1 A/C (χ2(2) = 7.66, p = .022). The odds of heterozygote to homozygote genotypes in HLNRA relative to NLNRA at XRCC1 Arg399Gln (OR = 1.96, 95% CI: 1.13–3.40), XRCC3 Thr241Met (OR = 0.73, 95% CI: 0.41–1.31), PRKDC (XRCC7 G/T), (OR = 0.81; 95% CI: 0.48–1.38), NEIL1 G/T (OR = 0.54; 95% CI: 0.31–0.96) and LIG1 A/C (OR = 1.62; 95% CI: 0.97–2.69) was also not significantly different in HLNRA vs NLNRA, except at XRCC1 and NEIL1.Conclusion: The genotype frequencies at three of these SNPs i.e. XRCC1 Arg399Gln, XRCC3 Thr241Met and PRKDC (XRCC7 G/T) were similar, whereas NEIL1 G/T and LIG1 A/C showed significant difference between HLNRA and NLNRA population. However, further research using more number of SNPs in a larger cohort is required in this study area.

Human papillomavirus 16 promotes microhomology-mediated end-joining

Squamous cell carcinomas (SCCs) arising from aerodigestive or anogenital epithelium that are associated with the human papillomavirus (HPV) are far more readily cured with radiation therapy than HPV-negative SCCs. The mechanism behind this increased radiosensitivity has been proposed to be secondary to defects in DNA repair, although the specific repair pathways that are disrupted have not been elucidated. To gain insight into this important biomarker of radiosensitivity, we first examined genomic patterns reflective of defects in DNA double-strand break repair, comparing HPV-associated and HPV-negative head and neck cancers (HNSCC). Compared to HPV-negative HNSCC genomes, HPV+ cases demonstrated a marked increase in the proportion of deletions with flanking microhomology, a signature associated with a backup, error-prone double-strand break repair pathway known as microhomology-mediated end-joining (MMEJ). Then, using 3 different methodologies to comprehensively profile double-strand break repair pathways in isogenic paired cell lines, we demonstrate that the HPV16 E7 oncoprotein suppresses canonical nonhomologous end-joining (NHEJ) and promotes error-prone MMEJ, providing a mechanistic rationale for the clinical radiosensitivity of these cancers.

Long non-coding RNA transcribed from pseudogene PPIAP43 is associated with radiation sensitivity of small cell lung cancer cells.

Small cell lung cancer (SCLC) is a highly lethal disease. Although radiation therapy is effective for the majority of patients with SCLC, patient sensitivity to radiation varies. The lack of biomarkers impedes advances in targeting radiation-sensitive patients. In this study, the changes in transcription patterns of SCLC cell lines were evaluated with or without 2 Gy gamma radiation. The results demonstrated that peptidyl-prolyl cis-trans isomerase A pseudogene 43 (PPIAP43) transcription was increased 2-fold in cells irradiated with 2 Gy gamma radiation compared with unirradiated cells in pre-reported radio-sensitive sensitive cell lines H69, H128, H146, H209 and H187. These cells shared 259 upregulated and 96 downregulated RNA transcripts following radiation. Pre-reported less sensitive cell lines H526, D53, D114 and D153 in which PPIAP43 was not upregulated 2-fold following irradiation with 2 Gy gamma radiation compared with unirradiated cells, shared 3 upregulated and 9 downregulated RNA transcripts. The RNA transcript of PPIAP43 was aligned with the mRNA of peptidyl-prolyl cis-trans isomerase A (PPIA) at 2 sections (3,732 to 3,917 and 5,327 to 5,657 of the PPIA gene) and the sequences were shown to be 96 and 94% similar, respectively. Therefore, PPIAP43 may act as a sponge for microRNAs which bind with the RNA of PPIA. Therefore, PPIAP43 RNA transcription may serve as a potential biomarker of radio-sensitivity of SCLC.

Robustness of Clonogenic Assays as a Biomarker for Cancer Cell Radiosensitivity.

Photon radiation therapy is a major curative treatment for cancer. However, the lack of robust predictive biomarkers for radiosensitivity precludes personalized radiation therapy. Clonogenic assays are the gold standard method for measuring the radiosensitivity of cancer cells. Although a large number of publications describe the use of clonogenic assays to measure cancer cell radiosensitivity, the robustness of results from different studies is unclear. To address this, we conducted a comprehensive detailed literature search of 256 common cancer cell lines and identified the eight cell lines most-frequently examined for photon sensitivity using clonogenic assays. Survival endpoints and experimental parameters from all 620 relevant experiments were compiled and analyzed. We found that the coefficients of variation for SF2 (surviving fraction after 2 Gy irradiation) and for D10 (dose that yields a surviving fraction of 10%) were below 30% for all cell lines, indicating that SF2 and D10 have acceptable inter-assay precision. These data support further analysis of published data on clonogenic assays using SF2 and D10 as survival endpoints, which facilitates robust identification of biological profiles representative of cancer cell sensitivity to photons.

Residual gammaH2AX foci in head and neck squamous cell carcinomas as predictors for tumour radiosensitivity: Evaluation in pre-clinical xenograft models and clinical specimens

Background and purposePredictive biomarkers can be instrumental to treatment individualisation of cancer patients and improve therapy outcome. Residual γH2AX foci represent a promising biomarker to predict tumour radiosensitivity. In this pre-clinical study, the slope of the dose–response curve was evaluated for its predictive relevance in head and neck squamous cell carcinoma xenografts (HNSCC). Additionally, the feasibility of the translated assay was tested in a clinical setting in patient derived HNSCC samples, and associations between residual γH2AX foci and clinical parameters were analysed. Materials and methodsSeven HNSCC xenografts models (FaDu, SAS, SKX, UT-SCC-5, UT-SCC-14, UT-SCC-45, XF354) were used. Tumour bearing NMRI nude mice were randomly distributed to five treatment arms (0–8 Gy). Residual γH2AX foci (24 h post irradiation) were counted by visual scoring in a micromilieu dependent manner (assessed with BrdU and pimonidazole). The local tumour control values measured as TCD50 (tumour control dose 50%) have previously been published. Patient derived HNSCC biopsies were cultivated ex vivo for 24 h including 4 h of pimonidazole and BrdU treatment, subsequently irradiated with 0–8 Gy and fixed after 24 h. ResultsIn the pre-clinical study, the dose–response curve slopes negatively correlated with the tumour control dose after fractionated irradiation (TCD50,fx, R2 = 0.63, p = 0.032) and after single dose irradiation under homogeneous hypoxia (TCD50,SD,clamp, R2 = 0.66, p = 0.027). The γH2AX assay in clinical HNSCC samples showed a dose–response relationship, with the values of the slopes ranging from 0.099 Gy−1 to 0.920 Gy−1 (coefficient of variation = 52.8%). Slopes derived from patients were in the same ranges as the sensitive, moderate and resistant models of the pre-clinical study. Statistical analysis revealed a significant negative correlation between the slope and the patients’ age (R2 = 0.65, p = 0.001). ConclusionThese results further support the promise of the slope of the residual γH2AX foci dose–response as a biomarker for radiosensitivity. In the clinical samples, the variation in the slopes reveals patients’ specific repair capacities, which could hold potential value for treatment individualisation.

Outcomes Prediction in Pre-Operative Radiation Therapy Locally Advanced Rectal Cancer: Leucocyte Assessment as Immune Biomarker

Leukocytes are hypothesized to reflect the inflammatory tumor microenvironment. We aimed to validate their prognostic significance in a large cohort of patients treated with pre-operative radiation for locally advanced rectal cancer (RC). Clinical records from consecutive patients treated in a single institution between 2004 and 2015 with curative-intent radiotherapy were retrospectively analyzed. Classical prognosis factors of RC and peripheral immune markers based on lymphocytes and neutrophil counts were studied. From 2004 to 2015, 257 RC patients with available biological data underwent a pre-operative radiotherapy, with a median age of 66 years. The median rectal EQD2 was 49.2 Gy. Most of patients experienced concurrent chemotherapy (n=245, 95.4%), mainly with 5-FU (83.3%). Clear surgical margins (i.e. complete resection) were achieved in 234 patients (91.1%). A complete (Mandard TRG1: n=35, 13.6%) or almost complete pathological response (Mandard TRG2: n=56, 21.8%) were achieved in 91 patients (35.4%). With a median follow-up of 46.1 months, 8 patients (3.1%) experienced local relapse, 38 (14.8%) experienced metastases and 45 (17.5%) died. Elevated pre-radiation neutrophil to lymphocyte ratio (NLR>2.8) was identified as an independent predictive factor of increased local relapse, of decreased progression-free survival and overall survival in multivariate analysis. Elevated NLR was marginally associated with incomplete pathological response in multivariate analysis, suggesting a possible value as a biomarker of radio-sensitivity. Pre-radiation NLR is a simple and robust biomarker for risk stratification in locally advanced RC patients undergoing pre-operative radiotherapy, and might select the subpopulation eligible to treatment intensification or to neoadjuvant chemotherapy.

Developing Predictive or Prognostic Biomarkers for Charged Particle Radiotherapy.

The response to radiotherapy can vary greatly among individuals, even though advances in technology allow for the highly localized placement of therapeutic doses of radiation to a tumor. This variability in patient response to radiation is biologically driven, but the individuality of tumor and healthy tissue biology are not used to create individual treatment plans. Biomarkers of radiosensitivity, whether intrinsic or from hypoxia, would move radiation oncology from precision medicine to precise, personalized medicine. Charged particle radiotherapy allows for even greater dose conformity, but the biological advantages of charged particle radiotherapy have not yet been cultivated. The development of biomarkers that would drive biologically based clinical trials, identify patients for whom charged particles are most appropriate, or aid in particle-selection strategies could be envisioned with appropriate biomarkers. Initially, biomarkers for low-linear energy transfer (LET) radiation responses should be tested against charged particles. Biomarkers of tumor radioresistance to low-LET radiations could be used to identify patients for whom the enhanced relative biological effectiveness (RBE) of charged particles would be more effective compared with low-LET radiations and those for whom specific DNA-repair inhibitors, in combination with charged particles, may also be appropriate. Furthermore, heavy charged particles can overcome the radioresistance of hypoxic tumors when used at the appropriate LET. Biomarkers for hypoxia could identify hypoxic tumors and, in combination with imaging, define hypoxic regions of a tumor for specific ion selection. Moreover, because of the enhanced RBE for charged particles, the risk for adverse healthy tissue effects may be greater, even though charged particles have greater tumor conformality. There are many validated healthy-tissue biomarkers available to test against charged particle exposures. Lastly, newer biological techniques, as well as newer bioinformatic and computational methods, are rapidly changing the landscape for biomarker identification, validation, and clinical trial design.