Elsevier

Oral Oncology

Volume 37, Issue 3, April 2001, Pages 243-250
Oral Oncology

Correlation between p53 gene mutations and circulating antibodies in betel- and tobacco-consuming North Indian population

https://doi.org/10.1016/S1368-8375(00)00092-0Get rights and content

Abstract

Alterations in p53 tumour suppressor gene and its expression may be implicated in the pathogenesis of betel- and tobacco-related oral cancer. There is wide regional variation in betel- and tobacco-consuming habits in different parts of the Indian subcontinent. The purpose of this study was to determine the correlations between p53 gene mutations, protein accumulation and serum antibodies in oral precancer and cancer. We analysed 30 potentially malignant oral lesions (leukoplakia) and 30 oral squamous cell carcinomas (SCCs) from northern India because the betel quid-consuming habits are different from those prevalent in other regions of India. p53 mutations were analysed by polymerase chain reaction amplification of genomic DNA and direct sequencing, p53 protein accumulation by immunohistochemical analysis and circulating p53 antibodies by ELISA. p53 gene mutations, analysed within exons 5–9, were observed in five out of 30 (17%) potentially malignant oral lesions and seven out of 30 (23%) oral SCCs. All the mutations were base substitution mutations. Three missense and two nonsense mutations were observed in potentially malignant oral lesions, while six missense and one nonsense mutations were identified in oral SCCs. The probable hot spots for the mutations were identified at codons 126, 136 and 174, which have not been observed thus far. A good correlation was observed between p53 missense mutation, p53 antibodies and p53 protein accumulation in matched potentially malignant and malignant oral lesions. All the potentially malignant and cancerous lesions harbouring missense mutations showed accumulation of p53 protein and the majority of these patients showed circulating p53 antibodies suggesting that serological detection of p53 antibodies may serve as a surrogate marker for p53 alterations in oral lesions.

Introduction

Carcinoma of the oral cavity is one of the most frequent malignant tumours worldwide [1] with major predominance in south east Asia and India [2]. The development of oral cancer is a multistep process, arising from pre-existing red or white patches termed erythroplakia or leukoplakia (potentially malignant lesions); 5–10% of these potentially malignant lesions undergo transition to frank malignancy. The incidence and mortality from oral cancer are reportedly increasing [2]. Epidemiological studies indicate that intervention studies at an early stage may reduce oral cancer-related deaths. Concerted efforts are therefore required to define multiple markers for early detection of the disease as well as for management and monitoring the efficacy of the treatment. These efforts may also be of value for proactive intervention of high risk groups (potentially malignant lesions).

Several genetic events initiate the progression of normal oral mucosa to squamous cell carcinomas (SCCs); mutations of the p53 tumour suppressor gene play an important role in this process [3]. Numerous studies report specific mutations of the p53 gene in oral SCCs among diverse population groups from several parts of the world (reviewed in [3] and references therein). The importance of alterations of p53 tumour suppressor as a molecular marker in the clinical diagnosis of oral cancer is being extensively investigated [4], [5], [6], [7], [8], [9], [10], [11], [12], [13], [14], [15]. The profile of p53 gene mutations has been associated with several factors, such as prolonged exposure of the oral cavity to various external carcinogens and co-carcinogens that are components of chewable tobacco, and/or betel quid (processed/unprocessed areca nut) [16], [17], [18], [19], [20], [21], [22].

The alternative approach of assessing p53 alteration by detection of p53 protein in tumour cells by immunohistochemistry (IHC) indicates discordancy rates of 30–35% when compared with techniques that determine the gene status [11]. Circulating p53 antibodies in the sera of cancer patients have been reported to be yet another alternative strategy for assessing p53 aberrations in several cancers [23], [24], [25], [26], [27], [28]. Our group and others reported the presence of circulating p53 antibodies in patients with oral leukoplakia and oral SCCs [29], [30]. However, a composite study of the relationship between p53 gene mutations, p53 protein accumulation in the same tumours and circulating anti-p53 antibodies in the tobacco-induced oral cancer patients in the Indian population have not been carried out so far.

We further wanted to assess the ability of p53 antibodies as a surrogate marker for identifying p53 alterations which may be helpful in assessing the ability of p53 to identify the high risk group patients that potentially can benefit from adjutant therapy. Therefore, the aim of the present study was to analyse p53 gene mutations and correlate the specific mutations with intratumoural protein accumulation as well as serum p53-Abs in matched patient samples.

Section snippets

Tissue specimens

Surgical tissue specimens from 30 untreated primary SCCs of the oral cavity and 30 potentially malignant lesions (leukoplakia) were obtained from the Department of Surgical Disciplines, All India Institute of Medical Sciences, New Delhi, India. A pre-tested semi-structured questionaire was used to collect information on the consumption of alcohol, tobacco (chewing or smoking) and use of areca nut in betel quid; betel and areca nut use: moderate=5–10 betel per day for 2–10 years; heavy=over 10

Genetic analysis of p53 in human oral potentially malignant and malignant lesions

The relationships between p53 mutational profile and p53 protein status, circulating p53-Abs as well as clinicopathological parameters of the patients with oral potentially malignant and malignant lesions are shown in Table 1, Table 2, respectively. Genetic analysis of p53 was carried out in 30 potentially malignant lesions and 30 oral SCCs by PCR followed by direct sequencing. Representative DNA sequence scans showing p53 mutations in oral potentially malignant lesions and SCC are shown in

Discussion

p53 gene mutation is one of the most common events in human carcinogenesis and approximately 90% of its mutations are of the missense type [9]. Knowledge of p53 mutations in oral potentially malignant lesions is meagre and their relevance in identifying lesions at high risk of transition to malignancy is not available [15]. The most striking feature of the present study was the correlation observed between p53 missense mutations, p53 antibodies and p53 protein accumulation in matched

Acknowledgements

The authors wish to thank: (1) IGBMC core and oligonucleotide synthesis and DNA sequencing facilities for tremendous help and support; (2) the members of Boh WasylykÆs lab for help and constructive discussion; and (3) members of CEFIPRA for their scientific support. The financial assistance from Indo-French Centre For Promotion of Advanced Research is gratefully acknowledged. S.A. was supported by Senior Research Fellowship from University Grant Commission, India.

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    1

    R.R. and S.A. contributed equally to this study.

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