L-Myc Polymorphism in Head and Neck Nonmelanoma Skin and Lower Lip Cancers

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L-Myc Polymorphism in Head and Neck Nonmelanoma Skin and Lower Lip Cancers ORIGINAL ARTICLE L-myc Polymorphism in Head and Neck Nonmelanoma Skin and Lower Lip Cancers Aydın Go¨zu¨, MD; Arzu Ergen, PhD; Deniz Dayıcıog˘lu, MD; I˙lhan Yaylım, PhD; Zafer O¨ zsoy, MD; Turgay I˙sbir, PhD Objective: To evaluate the presence of L-myc gene varia- rose gel electrophoresis were used to determine the L-myc tions as a genetic predisposition to head and neck non- oncogene genotypes. melanoma skin cancer (HNNMSC) and lower lip cancer Results: The presence of the LS genotype was found to (LLC). be significantly increased in the study group, whereas the LL genotype was not detected. The S allele was also more Design: A case-control study. frequent in the study group. The SS genotype was found to correlate with aggressive tumor behavior in patients with Setting: An academic institute laboratory. HNNMSC and a family history of cancer. Patients with LLC displayed significantly less of the SS genotype. Participants: Twenty-four patients with HNNMSC and 27 with LLC were compared with 51 age- and sex- Conclusions: The L-myc gene polymorphism may help matched control subjects. detect and prevent HNNMSC and LLC in susceptible in- dividuals. It may also contribute to estimation of tumor behavior in patients with HNNMSC. Main Outcome Measures: Polymerase chain reac- tion restriction fragment length polymorphism and aga- Arch Otolaryngol Head Neck Surg. 2008;134(7):725-728 ANCER OF THE SKIN IS THE noma skin cancer (HNNMSC) and lower lip most common form of ma- cancer (LLC). Proto-oncogenes are nor- lignant disease, and the mal cellular genes involved in the regula- skin of the head and neck tion of cellular proliferation that lead to neo- is the site most frequently plastic cell proliferation when they have Cinvolved. The number of basal cell carci- mutations or are overexpressed. They nomas (BCCs) and squamous cell carcino- mostly display a very broad tumor spec- mas (SCCs) is believed to be rising at a rate trum, whereas some tend to be activated pri- of 5% per year.1 Basal cell carcinoma is the marily in certain cancer types. The myc fam- most common type of skin cancer. Lip can- ily of oncogenes, which includes c-myc, cer is a form of oral cancer at the junction N-myc, and L-myc, has been proved to be between the oral cavity and the skin. The amplified late in the progression of many lips are the most common site of cancer in human tumors involving head and neck the oral cavity, and the lower lip is more fre- cancers and nonmelanoma skin cancers and quently affected than the upper lip. Can- is generally associated with an aggres- cers arising from the vermilion of the lip can sively malignant phenotype.3-9 be considered to be a unique group of tu- Since cloning of the L-myc gene (Gen- mors because they derive from a modified bank M19720) in 1985,3 many studies have and external mucosal tissue that is ex- investigated the possible role of the L- posed to different environmental factors myc gene in various cancers. The L-myc than other sites of the oral cavity.2 Squa- protein is involved in the tissue-specific mous cell carcinoma is the most common regulation of cell growth, and alterations malignant neoplasm of the lower lip. in the expression of L-myc may partici- Author Affiliations: The risk of BCC and SCC is associated pate in malignant transformation.3,4 The Department of Plastic and mainly with long-term exposure to UV ra- L-myc EcoRI polymorphism is a noncod- Reconstructive Surgery, Vakif diation but also with geographic location, ing variation in the second intron of the Gureba Research and Education race, immune status, and other as-yet- L-myc gene, resulting in short (S) and long Hospital (Drs Go¨zu¨ , Dayıcıog˘lu, ¨ undetermined genetic factors. Early detec- (L) alleles. It is the first genetic variation and Ozsoy), and Institute of tion of high-risk populations for preven- found to be associated with prognosis in Experimental Medical Research, 5 Department of Molecular tion of disease and estimation of biological cancer. Individuals carrying the S allele Medicine, Istanbul University tumor behavior for planning treatment fa- tend to have a poor prognosis and in- (Drs Ergen, Yaylım, and I˙sbir), vor the need for molecular studies of the ge- creased risk of several tumor types, al- Istanbul, Turkey. netic changes in head and neck nonmela- though controversial results have been re- (REPRINTED) ARCH OTOLARYNGOL HEAD NECK SURG/ VOL 134 (NO. 7), JULY 2008 WWW.ARCHOTO.COM 725 ©2008 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/27/2021 Marker 1 2 3 4 5 6 Table 2. Distribution of Allelic Frequencies Participants, 267 bp No. (%) Relative Risk P (95% Confidence 142 bp 125 bp LSValue Interval) Control group (n=102) 60 (59) 42 (41) .008 2.12 (1.21-3.71) Study group (n=102) 41 (40) 61 (60) LL SS LL LL – LS Figure. Direct visualization of polymerase chain reaction products by means TCA-GGA-AGC-TTG-AG-3Ј in a volume of 50 µL containing of ethidium bromide staining. A 267–base pair (bp) L-myc fragment was 3mM magnesium chloride, 50mM potassium chloride, 10mM amplified, cleaved with EcoRI, and electrophoresed on 2% agarose gel. Results of 5 representative control subjects are indicated. Lanes 1, 3, and 4 Tris hydrochloride (pH, 8.4), 0.5mM of each deoxynucleotide show the LL homozygote; lane 2, the SS homozygote; lane 5, no polymerase triphosphate (MBI Fermentas, Burlington, Ontario, Canada), chain reaction product; and lane 6, the LS heterozygote. The 3 genotypes and1UofTaq polymerase (MBI Fermentas). Amplification was were the LL homozygote, appearing as a 267-bp fragment; the LS performed using a DNA thermal cycler (MBI Fermentas) for heterozygote, with 267-, 142-, and 125-bp fragments; and the SS 30 cycles, with denaturation steps at 94°C for 30 seconds, an- homozygote, with 142- and 125-bp fragments. nealing at 50°C for 1 minute, and extension at 74°C for 1 minute. The polymerase chain reaction product exhibited a 267–base pair (bp) fragment. The amplification fragment was digested Table 1. Distribution of L-myc Genotypes with 5 U of EcoRI (MBI Fermentas) at 37°C for 1 hour. The digested DNA fragments were separated by means of gel elec- Participants, No. (%) trophoresis on 2% agarose gel in 1X Tris borate EDTA buffer P and DNA visualized by means of ethidium bromide staining. LL LS SS Value The responsible L-myc restriction fragment length polymor- Control group (n=51) 20 (39) 20 (39) 11 (22) .001 phism alleles were identified in each sample. Study group (n=51) 0 41 (80) 10 (20) Statistical analysis was performed using SPSS for Windows 10.0 software (SPSS Inc, Chicago, Illinois). Descriptive statis- tical methods (mean [SD]) and quantitative variables were com- 10 pared using the 2-tailed t test. The ␹2 and Fischer exact tests ported. The literature includes no studies, to our Ͻ knowledge, regarding the relation of the L-myc polymor- were used to compare qualitative variables, and P .05 was con- sidered statistically significant. phism with either HNNMSC or LLC. We aimed to evalu- ate them together because of their anatomical and etio- logic proximity. RESULTS METHODS The polymorphic L-myc locus was analyzed by means of polymerase chain reaction–restriction fragment length polymorphism for 51 patients and 51 control subjects. Thirty-six men and 15 women aged 26 to 85 years (mean, 60.92 years) were included in the study. They were selected from The 3 genotypes revealed were the LL homozygote, ap- among previously untreated patients with biopsy-confirmed head pearing as a 267-bp fragment; the LS heterozygote, with and neck skin cancer (n=24) and patients with LLC (n=27) 267-, 142-, and 125-bp fragments; and the SS homozy- who underwent surgery in the Plastic and Reconstructive Sur- gote, with 142- and 125-bp fragments (Figure). gery Department at Vakif Gureba Research and Education Hos- The presence of the LS genotype in the study group (41 pital between January 2, 2005, and April 30, 2006. The skin patients [80%]) was found to be higher than that in the cancer subgroup included 10 patients with BCC, and all the control group (20 patients [39%]). The LL genotype was other patients were diagnosed as having SCC. Fifty-one age- present in 20 patients in the control group [39%], whereas and sex-matched control subjects without any disease history it was not detected in the study group (Table 1). The dif- were included in the study. Two standardized questionnaires ference in genotypic distribution between the groups was were administered for each subgroup to record the clinical char- acteristics of patients. The institutional review board of Vakif significant (P= .001). Allelic frequencies also displayed sig- Gureba Research and Education Hospital approved the study, nificant differences between them: the S allele was more and informed consent was obtained from all the participants. frequent in the study group (Table 2). When tumor (LLC Skin tumors were classified as clinically aggressive if they and HNNMSC) and histopathologic (SCC and BCC) sub- exceeded 2 cm in 1 surface dimension; invaded muscle, bone, groups were compared with the control group sepa- or cartilage; or were metastatic to lymph nodes. Lower lip tu- rately, the LS genotype dominance was significant in each mors were classified in accordance with the TNM staging sys- subgroup (PϽ.01 for the LLC, HNNMSC, and SCC sub- 11 tem. All the frozen or formalin-fixed tumor tissue samples were groups and P=.05 for the BCC subgroup).
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