3203 Association Study of the G-Protein B3 Subunit C825T Polymorphism with Disease Progression an Overall Survival in Patients with Head and Neck Squamous Cell Carcinoma Goetz F. Lehnerdt,1 Peter Franz,1 Agnes Bankfalvi,2 Sara Grehl,3 Klaus Jahnke,1,6 Stephan Lang,1,6 Kurt W. Schmid,2,6 Winfried Siffert,4,6 and Ulrich H. Frey4,5 1Department of Otorhinolaryngology, 2Institute of Pathology and Neuropathology, 3Department of Radiotherapy at the West German Cancer Center Essen, 4Institute of Pharmacogenetics, 5Department of Anaesthesiology and Intensive Care Medicine, and 6West-German Cancer Center Essen, Essen, Germany Abstract The T-allele of a common C825T single nucleotide significant genotype-dependent relapse-free interval polymorphism (SNP) in the gene GNB3, encoding the (P = 0.036). In multivariate analysis with stage, G3 subunit of heterotrimeric G-proteins, is associated localization, grade, gender, and smoking habits with a truncated form of the G3 protein that imparts a as covariates, GNB3 825T homozygous patients greater signaling capacity than the alternative C-allele displayed a higher risk for relapse than C825 homozy- encoding a nontruncated protein. We analyzed gous patients (TT versus CC, hazard ratio; 95% the C825T-allele status with regard to disease progres- confidence interval, 1.4-4.8; P =0.002).Thesame sion in patients with head and neck squamous cell genotype effect was found for overall survival, TT carcinoma (HNSCC). The prognostic value of the SNP genotypes were at higher risk for death compared was evaluated in an unselected series of 341 patients with CC genotypes (hazard ratio, 2.6; 95% confidence treated with curative intent for HNSCC including all interval, 1.6-4.3; P < 0.001), and 5-year survival tumor stages with different therapeutic regimens. proportions were 60% for CC, 52% for TC, and 33% Genotype analysis was done by Pyrosequencing using for TT. The GNB3 C825T SNP thus represents a host DNA from paraffin-embedded tissue samples. derived prognostic marker in HNSCC, which allows Genotypes were correlated with relapse-free and identifying high-risk patients, which could benefit overall survival. Proportions of 5-year relapse-free from novel and/or more aggressive therapeutic intervals were 62% for CC, 60% for TC, and 42% regimes. (Cancer Epidemiol Biomarkers Prev 2008; for TT genotypes. Kaplan-Meier curves revealed a 17(11):3203–7) Introduction Head and neck squamous cell carcinoma (HNSCC) is the be sufficiently predicted by these factors (2). Thus, sixth most prevalent neoplasm in the world. Worldwide additional variables would be most desirable to optimize annually, f500,000 cases are reported to be diagnosed the prediction of tumor behavior enabling a more with HNSCC (1). HNSCC develops in the squamous individualized therapeutic approach. epithelial cells of the upper aerodigestive tract and is G-protein hg subunits are of major importance for found in the oral cavity, oropharynx, hypopharynx, and cell migration on stimulation of G-protein –coupled larynx. Due to advances in our understanding and novel receptors. treatment modalities, the 5-year survival rates of We have shown that the 825T-allele of the GNB3 HNSCC have improved to some extent, varying from single nucleotide polymorphism (SNP) is associated site to site, but still the therapeutic outcome in HNSCC with the occurrence of an alternative splice variant represents an oncological challenge. called Gh3s (3). G-protein h subunits belong to the Treatment options of HNSCC include surgery, radio- family of ‘‘propeller proteins’’ and consist of seven therapy, conventional chemoradiotherapy, radiotherapy regular protein domains, each coding for one propeller with new target drugs or a combination of these blade. The splice variant Gh3s lacks the equivalent modalities. Both the therapeutic approach and of one propeller domain. Despite this structural deletion, the patients’ prognosis are currently determined by Gh3s is functionally active and has been associated tumor volume and tumor stage. However, neither the with an increased signal transduction (3, 4). The biological behavior nor the response to therapy can enhanced signal transduction is correlated with increased chemotaxis as shown for migrating cells from 825T-allele carriers (5, 6). In carriers of the GNB3 825T- Received 7/10/08; revised 8/15/08; accepted 8/25/08. allele, stimulation with chemokines or autocoids leads to Grant support: IFORES program, University of Duisburg/Essen, Germany. enhanced chemotaxis in Gh3s-expressing neutrophils Requests for reprints: Goetz Lehnerdt, University Hospital, University of Duisburg- and lymphocytes (6, 7). Our group recently showed that Essen, Hufelandstrahe 55, 45122 Essen. Phone: 49-201-723-2481; Fax: 49-201-723-5903. h E-mail: [email protected] in bladder cancer tumor cells, G 3s is expressed Copyright D 2008 American Association for Cancer Research. exclusively in patients carrying an 825T-allele, whereas doi:10.1158/1055-9965.EPI-08-0616 wild-type Gh3 was associated with the homozygous CC Cancer Epidemiol Biomarkers Prev 2008;17(11). November 2008 Downloaded from cebp.aacrjournals.org on September 28, 2021. © 2008 American Association for Cancer Research. 3204 Predictive Value of C825T SNP in HNSCC genotype (8). Moreover, 825T allele carriers showed a determined by Pyrosequencing as described (9). shorter time to metastasis compared with carriers of the Genotyping was done blinded, with the genotyping homozygous CC genotype (8). laboratory being unaware of any clinical data. The aim of the present study was to investigate GNB3 Healthy Blood Donors. The control group consisted of whether the C825T SNP of the gene is related 255 age- and sex-matched healthy Caucasian individuals to the clinical outcome in patients with HNSCC. who were randomly recruited at the local Institute of Genotyping of this SNP may offer the opportunity to Transfusion Medicine, University Hospital of Essen. define subgroups of HNSCC patients who may benefit The control sample consisted of 161 males and 94 females, from novel therapeutic options. and the mean age was 56.7 F 4.4 y. Details of this control group have been published previously (10). Materials and Methods Statistical Analysis and Presentation of Data. The genotyping results of the present study were correlated The study was strictly done according to the Declaration with overall survival (OS; defined as time from first of Helsinki and approved by the local Ethics Committee diagnosis to death or loss of follow-up) and time to relapse of the University Hospital of Essen. (defined as time from last day of therapy to day of diagnosis of tumor relapse). Kaplan-Meier plots and the Patients. The study group consisted of 341 patients log-rank test for trend were used to retrospectively (340 Caucasians, 1 Asian) with HNSCC diagnosed and evaluate the relationship between tumor localization and treated between 1995 and 2001 at the West German GNB3 genotypes. Log-rank tests for GNB3 genotypes Cancer Center Essen. Relevant clinico-pathologic were adjusted for tumor localization. Multivariate models data [American Joint Committee on Cancer (AJCC) of clinical follow up were established using clinical and and tumor-node-metastasis tumor stage, tumor site, pathologic variables known as predictors of prognosis histologic grading, therapeutic regimens, 5-y follow-up, including the genotypes of the C825T polymorphism. relapse, and cause of death] were extracted from the A backward stepwise Cox proportional hazard model was patients’ files or collected by telephone interviews with used to calculate hazard ratios, 95% confidence interval, the patients or their family doctors. HNSCC cases that and P values (11). Contingency tables and the Pearson’s m2 had been defined as palliative by the West German test were used to compare categorical variables Cancer Center Essen interdisciplinary tumor board using GNB3 genotypes as indicated. Because the GNB3 (the vast majority classified as tumor stage IVC) were polymorphism displays a gene-dose effect (3, 12), a excluded from this study. linear ANOVA was used for comparison of parametric Genotyping. For genotyping of the C825T polymor- continuous variables where appropriate. Nonparametric phism of the GNB3 gene, genomic DNA was isolated linear variables were compared using the Kruskal-Wallis from several 10- to 20-Am-thick sections (overall weight test. Differences were regarded significant at a P value of not exceeding 25 mg) from routinely processed paraffin <0.05. All statistical analyses was done using SPSS 15.0 blocks. Genotypes of the GNB3 polymorphism were (SPSS). Continuous variables are given as means F SD. Table 1. C 825 T genotype distribution Total CC CT TT P n (%) total 341 170 (49.8) 144 (42.2) 27 (7.9) Oral cavity 20 5 (25.0) 12 (60.0) 3 (15.0) Oropharynx 108 64 (59.3) 35 (32.4) 9 (8.3) Hypopharynx 49 23 (46.9) 25 (51.0) 1 (2.0) Larynx 150 73 (48.7) 66 (44.0) 11 (7.3) Multiple sites 14 5 (35.7) 6 (42.9) 3 (21.4) 0.033 Mean age (y F SD) 61.8 F 12.1 61.2 F 10.7 62.1 F 13.8 63.6 F 10.7 0.315 Median follow up, mo (range) 61 (3-143) 65 (3-128) 57 (6-143) 38 (6-97) 0.018* Gender (m/f) 286/55 137/33 129/15 20/7 0.496 Smoking, n (%) 321 (90.0) 152 (49.5) 131 (42.7) 24 (7.8) 0.724 Pack years 46.6 F 22.5 45.4 F 21.2 46.2 F 23.2 56.0 F 25.7 0.101 AJCC I 53 22 (41.5) 27 (50.9) 4 (7.5) II 50 25 (50.0) 21 (42.0) 4 (8.0) III 55 27 (49.1) 22 (40.0) 6 (10.9) IVA 157 82 (52.2) 62 (39.5) 13 (8.3) IVB 23 12 (52.2) 11 (47.8) 0 (0) IVC 3 2 (66.7) 1 (33.3) 0 (0) 0.866 Grade 1 21 8 (38.1) 11 (52.4) 2 (9.5) 2 246 116 (47.2) 109 (44.3) 21 (8.5) 3-4 74 46 (62.2) 24 (32.4) 4 (5.4) 0.170 NOTE: C825T genotype distribution with regard to tumor site, demographic characteristics, primary therapy, tumor stage (AJCC), and histologic grading in 341 patients with HNSCC.