Owing to advances in radiotherapy, the physical properties of radiation can be optimized to enable individualized treatment; however, optimization is rarely based on biological properties and, ...therefore, treatments are generally planned with the assumption that all tumours respond similarly to radiation. Radiation affects multiple cellular pathways, including DNA damage, hypoxia, proliferation, stem cell phenotype and immune response. In this Review, we summarize the effect of these pathways on tumour responses to radiotherapy and the current state of research on genomic classifiers designed to exploit these variations to inform treatment decisions. We also discuss whether advances in genomics have generated evidence that could be practice changing and whether advances in genomics are now ready to be used to guide the delivery of radiotherapy alone or in combination.
Oxygen deprivation (hypoxia) in non-small cell lung cancer (NSCLC) is an important factor in treatment resistance and poor survival. Hypoxia is an attractive therapeutic target, particularly in the ...context of radiotherapy, which is delivered to more than half of NSCLC patients. However, NSCLC hypoxia-targeted therapy trials have not yet translated into patient benefit. Recently, early termination of promising evofosfamide and tarloxotinib bromide studies due to futility highlighted the need for a paradigm shift in our approach to avoid disappointments in future trials. Radiotherapy dose painting strategies based on hypoxia imaging require careful refinement prior to clinical investigation. This review will summarize the role of hypoxia, highlight the potential of hypoxia as a therapeutic target, and outline past and ongoing hypoxia-targeted therapy trials in NSCLC. Evidence supporting radiotherapy dose painting based on hypoxia imaging will be critically appraised. Carefully selected hypoxia biomarkers suitable for integration within future NSCLC hypoxia-targeted therapy trials will be examined. Research gaps will be identified to guide future investigation. Although this review will focus on NSCLC hypoxia, more general discussions (eg, obstacles of hypoxia biomarker research and developing a framework for future hypoxia trials) are applicable to other tumor sites.
There is a need to identify patients with early stage breast cancer for whom radiotherapy and its associated toxicities might be avoided. Tutzauer et al. hypothesised that a hypoxia biomarker might ...be used but found that adjuvant radiotherapy following surgery reduces risk of recurrence irrespective of hypoxia status.
Tumor hypoxia is associated with a poor prognosis, hypoxia modification improves outcome, and hypoxic status predicts benefit from treatment. Yet, there is no universal measure of clinical hypoxia. ...The aim of this study was to investigate whether a 26-gene hypoxia signature predicted benefit from hypoxia-modifying treatment in both cancer types.
Samples were available from 157 T2-T4 laryngeal cancer and 185 T1-T4a bladder cancer patients enrolled on the accelerated radiotherapy with carbogen and nicotinamide (ARCON) and bladder carbogen nicotinamide (BCON) phase III randomized trials of radiotherapy alone or with carbogen and nicotinamide (CON) respectively. Customized TaqMan low density arrays (TLDA) were used to assess expression of the 26-gene signature using quantitative real-time PCR. The median expression of the 26 genes was used to derive a hypoxia score (HS). Patients were categorized as TLDA-HS low (≤median) or TLDA-HS high (>median). The primary outcome measures were regional control (RC; ARCON) and overall survival (BCON).
Laryngeal tumors categorized as TLDA-HS high showed greater benefit from ARCON than TLDA-HS low tumors. Five-year RC was 81% (radiotherapy alone) versus 100% (CON) for TLDA-HS high (P=0.009). For TLDA-HS low, 5-year RC was 91% (radiotherapy alone) versus 90% (CON; P=0.90). TLDA-HS did not predict benefit from CON in bladder cancer.
The 26-gene hypoxia signature predicts benefit from hypoxia-modifying treatment in laryngeal cancer. These findings will be evaluated in a prospective clinical trial.
Fulfilling the promise of cancer immunotherapy requires novel predictive biomarkers to characterise the host immune microenvironment. Deciphering the complexity of immune cell interactions requires ...an automated multiplex approach to histological analysis of tumour sections. We tested a new automatic approach to select tissue and quantify the frequencies of cell-cell spatial interactions occurring in the PD1/PD-L1 pathway, hypothesised to reflect immune escape in oropharyngeal squamous cell carcinoma (OPSCC).
Single sections of diagnostic biopsies from 72 OPSCC patients were stained using multiplex immunofluorescence (CD8, PD1, PD-L1, CD68). Following multispectral scanning and automated regions-of-interest selection, the Hypothesised Interaction Distribution (HID) method quantified spatial proximity between cells. Method applicability was tested by investigating the prognostic significance of co-localised cells (within 30 μm) in patients stratified by HPV status.
High frequencies of proximal CD8
and PD-L1
(HR 2.95, p = 0.025) and PD1
and PD-L1
(HR 2.64, p = 0.042) cells were prognostic for poor overall survival in patients with HPV negative OPSCC (n = 31).
The HID method can quantify spatial interactions considered to reflect immune escape and generate prognostic information in OPSCC. The new automated approach is ready to test in additional cohorts and its applicability should be explored in research and clinical studies.
Hypoxia modification improves overall survival in muscle-invasive bladder cancer patients who undergo radiotherapy. There is evidence that hypoxic tumors benefit most from hypoxia modification. The ...study aimed to identify or derive a hypoxia gene signature that predicts benefit from hypoxia-modifying treatment in bladder cancer.
Published hypoxia signatures were tested and a new one derived by analyzing bladder cancer transcriptomic data from public databases. Tumor samples were available from the BCON phase III randomized trial of radiotherapy alone or with carbogen and nicotinamide (CON). Gene expression data were generated for 151 tumors using Affymetrix Human 1.0 Exon ST arrays and used for independent validation.
A 24-gene signature was derived, which was prognostic in four of six independent surgical cohorts (
= 679; meta HR, 2.32; 95% CI, 1.73-3.12;
< 0.0001). The signature was also prognostic in BCON patients receiving radiotherapy alone (
= 75; HR for local relapse-free survival, 2.37; 95% CI, 1.26-4.47;
= 0.0076). The signature predicted benefit from CON (
= 76; HR, 0.47; 95% CI, 0.26-0.86;
= 0.015). Prognostic significance (
= 0.017) and predictive significance (
= 0.058) remained after adjusting for clinicopathologic variables. A test for interaction between hypoxia status and treatment arms was significant (
= 0.0094).
A 24-gene hypoxia signature has strong and independent prognostic and predictive value for muscle-invasive bladder cancer patients. The signature can aid identification of patients likely to benefit from the addition of carbogen and nicotinamide to radiotherapy.
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Hypoxia is common in non-small cell lung cancer (NSCLC) and an attractive therapeutic target. As hypoxia-targeting treatments are effective in patients with the most hypoxic tumours, we aimed to ...develop a lung adenocarcinoma (LUAD) hypoxia-related gene expression signature. RNAseq was used to identify genes significantly differentially expressed under hypoxia (1% O
) in four LUAD cell lines. Identified genes were used for unsupervised clustering of a TCGA-LUAD training dataset (n = 252) and in a machine learning approach to build a hypoxia-related signature. Thirty-five genes were upregulated in common in three of the four lines and reduced in the training cohort to a 28-gene signature. The signature was prognostic in the TCGA training (HR 2.12, 95% CI 1.34-3.37, p = 0.0011) and test (n = 250; HR 2.13, 95% CI 1.32-3.45, p = 0.0016) datasets. The signature was prognostic for overall survival in a meta-analysis of nine other datasets (n = 1257; HR 2.08, 95% CI 1.60-2.70, p < 0.0001). The 28-gene LUAD hypoxia related signature can be taken forward for further validation using a suitable gene expression platform.
Hypoxia and a suppressive tumour microenvironment (TME) are both independent negative prognostic factors for muscle-invasive bladder cancer (MIBC) that contribute to treatment resistance. Hypoxia has ...been shown to induce an immune suppressive TME by recruiting myeloid cells that inhibit anti-tumour T cell responses. Recent transcriptomic analyses show hypoxia increases suppressive and anti-tumour immune signalling and infiltrates in bladder cancer. This study sought to investigate the relationship between hypoxia-inducible factor (HIF)-1 and -2, hypoxia, and immune signalling and infiltrates in MIBC. ChIP-seq was performed to identify HIF1α, HIF2α, and HIF1β binding in the genome of the MIBC cell line T24 cultured in 1% and 0.1% oxygen for 24 h. Microarray data from four MIBC cell lines (T24, J82, UMUC3, and HT1376) cultured under 1%, 0.2%, and 0.1% oxygen for 24 h were used. Differences in the immune contexture between high- and low-hypoxia tumours were investigated using in silico analyses of two bladder cancer cohorts (BCON and TCGA) filtered to only include MIBC cases. GO and GSEA were used with the R packages "limma" and "fgsea". Immune deconvolution was performed using ImSig and TIMER algorithms. RStudio was used for all analyses. Under hypoxia, HIF1α and HIF2α bound to ~11.5-13.5% and ~4.5-7.5% of immune-related genes, respectively (1-0.1% O
). HIF1α and HIF2α both bound to genes associated with T cell activation and differentiation signalling pathways. HIF1α and HIF2α had distinct roles in immune-related signalling. HIF1 was associated with interferon production specifically, whilst HIF2 was associated with generic cytokine signalling as well as humoral and toll-like receptor immune responses. Neutrophil and myeloid cell signalling was enriched under hypoxia, alongside hallmark pathways associated with Tregs and macrophages. High-hypoxia MIBC tumours had increased expression of both suppressive and anti-tumour immune gene signatures and were associated with increased immune infiltrates. Overall, hypoxia is associated with increased inflammation for both suppressive and anti-tumour-related immune signalling and immune infiltrates, as seen in vitro and in situ using MIBC patient tumours.
There is increasing evidence supporting the role of genetic variants in the development of radiation-induced toxicity. However, previous candidate gene association studies failed to elucidate the ...common genetic variation underlying this phenotype, which could emerge years after the completion of treatment. We performed a genome-wide association study on a Spanish cohort of 741 individuals with prostate cancer treated with external beam radiotherapy (EBRT). The replication cohorts consisted of 633 cases from the UK and 368 cases from North America. One locus comprising TANC1 (lowest unadjusted P value for overall late toxicity=6.85×10(-9), odds ratio (OR)=6.61, 95% confidence interval (CI)=2.23-19.63) was replicated in the second stage (lowest unadjusted P value for overall late toxicity=2.08×10(-4), OR=6.17, 95% CI=2.25-16.95; Pcombined=4.16×10(-10)). The inclusion of the third cohort gave unadjusted Pcombined=4.64×10(-11). These results, together with the role of TANC1 in regenerating damaged muscle, suggest that the TANC1 locus influences the development of late radiation-induced damage.
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