A 26-Gene Hypoxia Signature Predicts Benefit from Hypoxia-Modifying Therapy in Laryngeal Cancer but Not Bladder Cancer

Abstract

Purpose: 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 hypoxiamodifying treatment in both cancer types.Experimental Design: 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).Results: 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 ¼ 0.009). For TLDA-HS low, 5-year RC was 91% (radiotherapy alone) versus 90% (CON; P ¼ 0.90). TLDA-HS did not predict benefit from CON in bladder cancer.Conclusion: The 26-gene hypoxia signature predicts benefit from hypoxia-modifying treatment in laryngeal cancer. These findings will be evaluated in a prospective clinical trial.

Low Cancer Stem Cell Marker Expression and Low Hypoxia Identify Good Prognosis Subgroups in HPV(−) HNSCC after Postoperative Radiochemotherapy: A Multicenter Study of the DKTK-ROG

“…To date, the potential of hypoxia gene signatures to discriminate pretreatment tumor specimen regarding their hypoxiainduced gene expression levels has not been tested in the postoperative setting (15,16,18). However, there is some evidence from an experimental study of our laboratory, which showed that pretreatment hypoxia impacts local tumor control after radiotherapy also when radiotherapy was applied under homogenous anoxic conditions (20).…”

“…Samples were then kept at 4 C for a maximum of 18 hours and subjected to the nCounter system and processed as described elsewhere (27). Raw counts were logarithmized and then normalized to the mean of the internal level of reference genes ACTR3, NDFIP1, RPL37A, B2M, GNB2L1, RPL11, POLR2A or to the reference genes of the corresponding hypoxia gene signatures (Supplementary Table S1B), respectively (15,16). cDNA generation and RT-PCR RNA was converted to cDNA using the High Capacity cDNA Reverse Transcription Kit (Applied Biosystems, Life Technologies).…”

“…Because of low RNA yield, each cDNA was preamplified for genes of interest using the pooled TaqMan gene expression assays according to the manufacturer's protocol (10 cycles). TaqMan gene expression assays were used for both hypoxia classifier and reference genes (15,16) as well as for CD44. RT-PCR was performed on an ABI StepOne Plus Real-Time PCR System (Life Technologies).…”

“…Several gene classifiers have been developed as a measure of tumor hypoxia in biopsies routinely taken for diagnosis (15)(16)(17)(18). Toustrup and colleagues developed a 15-gene classifier for patients with HNSCC who received primary radiotherapy (15).…”

“…Another highly prognostic hypoxia metagene signature was developed by Buffa and colleagues for patients with head and neck, breast, and lung cancers (18). The 26-gene classifier by Eustace and colleagues, that is based on this metagene signature (18), has been shown to predict the benefit from hypoxia-modifying treatment in laryngeal cancer (16). However, as the gene classifiers are based on the measurement of hypoxia-induced gene expression and not on hypoxia itself, the classification by the gene signature does not necessarily correlate with hypoxia assessed by PET (13,19).…”