Comprehensive Mutation Analysis of PIK3CA, p14ARF, p16INK4a and p21Waf1/Cip1 Genes is Suggestive of a Non- Neoplastic Nature of Phenytoin Induced Gingival Overgrowth

  • Swamikannu, Bhuminathan (Department of Prosthodontia, Sree Balaji Dental College and Hospital, Bharath University) ;
  • Kumar, Kishore S. (Department of Orthodontia, Sree Balaji Dental College and Hospital, Bharath University) ;
  • Jayesh, Raghavendra S. (Department of Prosthodontia, Sree Balaji Dental College and Hospital, Bharath University) ;
  • Rajendran, Senthilnathan (Department of Oral and Maxillofacial Surgery, Meenakshiammal Dental College and Hospital) ;
  • Muthupalani, Rajendran Shanmugam (Department of General Medicine, Sree Balaji Dental College and Hospital, Bharath University) ;
  • Ramanathan, Arvind (Human Genetics Laboratory, Sree Balaji Dental College and Hospital, Bharath University)
  • Published : 2013.05.30


Background: Dilantin sodium (phenytoin) is an antiepileptic drug, which is routinely used to control generalized tonic clonic seizure and partial seizure episodes. A few case reports of oral squamous cell carcinomas arising from regions of phenytoin induced gingival overgrowth (GO), and overexpression of mitogenic factors and p53 have presented this condition as a pathology with potential to transform into malignancy. We recently investigated the genetic status of p53 and H-ras, which are known to be frequently mutated in Indian oral carcinomas in GO tissues and found them to only contain wild type sequences, which suggested a non-neoplastic nature of phenytoin induced GO. However, besides p53 and H-ras, other oncogenes and tumor suppressors such as PIK3CA, p14ARF, p16INK4a and $p21^{Waf1/Cip1}$, are frequently altered in oral squamous cell carcinoma, and hence are required to be analyzed in phenytoin induced GO tissues to be affirmative of its non-neoplastic nature. Methods: 100ng of chromosomal DNA isolated from twenty gingival overgrowth tissues were amplified with primers for exons 9 and 20 of PIK3CA, exons $1{\alpha}$, $1{\beta}$ and 2 of p16INK4a and p14ARF, and exon 2 of $p21^{Waf1/Cip1}$, in independent reactions. PCR amplicons were subsequently gel purified and eluted products were sequenced. Results: Sequencing analysis of the twenty samples of phenytoin induced gingival growth showed no mutations in the analyzed exons of PIK3CA, p14ARF, p16INK4a and $p21^{Waf1/Cip1}$. Conclusion: The present data indicate that the mutational alterations of genes, PIK3CA, p14ARF, p16INK4a and $p21^{Waf1/Cip1}$ that are frequently mutated in oral squamous cell carcinomas are rare in phenytoin induced gingival growth. Thus the findings provide further evidence that phenytoin induced gingival overgrowth as a non-neoplastic lesion, which may be considered as clinically significant given the fact that the epileptic patients are routinely administered with phenytoin for the rest of their lives to control seizure episodes.


  1. Abbas T, Dutta A (2009). p21 in cancer: intricate networks and multiple activities. Nat Rev Cancer, 9, 400-14.
  2. Bialer M, Johannessen SI, Levy RH, et al (2013). Progress report on new antiepileptic drugs: a summary of the Eleventh Eilat Conference (EILAT XI). Epilepsy Res, 103, 2-30.
  3. Brodie MJ, Kwan P (2012). Current position of phenobarbital in epilepsy and its future. Epilepsia, 53, 40-6.
  4. Brunet L, Miranda J, Farre M, Berini L, Mendieta C (1996). Gingival enlargement induced by drugs. Drug Saf, 15, 219-31.
  5. Carl GF, Smith DB (1983). The effect of chronic phenytoin treatment on tissue folate concentrations and on the activities of the methyl synthetic enzymes in the rat. J Nutr, 113, 2368-74.
  6. Cornacchio AL, Burneo JG, Aragon CE (2011). The effects of antiepileptic drugs on oral health. J Can Dent Assoc, 77, b140.
  7. Correa JD, Queiroz-Junior CM, Costa JE, Teixeira AL, Silva TA (2011). Phenytoin-induced gingival overgrowth: a review of the molecular, immune, and inflammatory features. ISRN Dent, 2011, 497850.
  8. Das N, Dhanawat M, Shrivastava SK (2012). An overview on antiepileptic drugs. Drug Discov Ther, 6, 178-93.
  9. Dayal, Reddy R, Anuradha Bhat K (2000). Malignant potential of oral submucous fibrosis due to intraoral trauma. Indian J Med Sci, 54, 182-7.
  10. Fuller KL, Wang YY, Cook MJ, Murphy MA, D'Souza WJ (2013). Tolerability, safety, and side effects of levetiracetam versus phenytoin in intravenous and total prophylactic regimen among craniotomy patients: A prospective randomized study. Epilepsia, 54, 45-57.
  11. Gil J, Peters G (2006). Regulation of the INK4b-ARF-INK4a tumour suppressor locus: all for one or one for all. Nat Rev Mol Cell Biol, 7, 667-77.
  12. Kanno CM, Oliveira JA, Garcia JF, Castro AL, Crivelini MM (2008). Effects of cyclosporin, phenytoin, and nifedipine on the synthesis and degradation of gingival collagen in tufted capuchin monkeys (Cebus apella): histochemical and MMP-1 and -2 and collagen I gene expression analyses. J Periodontol, 79, 114-22.
  13. Kato T, Okahashi N, Kawai S, et al (2005). Impaired degradation of matrix collagen in human gingival fibroblasts by the antiepileptic drug phenytoin. J Periodontol, 76, 941-50.
  14. Kim WY, Sharpless NE (2006). The regulation of INK4/ARF in cancer and aging. Cell, 127, 265-75.
  15. Kresty LA, Mallery SR, Knobloch TJ, et al (2002). Alterations of p16(INK4a) and p14(ARF) in patients with severe oral epithelial dysplasia. Cancer Res, 62, 5295-300.
  16. Lissowska J, Pilarska A, Pilarski P, et al (2003). Smoking, alcohol, diet, dentition and sexual practices in the epidemiology of oral cancer in Poland. Eur J Cancer Prev, 12, 25-33.
  17. Lucchesi JA, Cortelli SC, Rodrigues JA, Duarte PM (2008). Severe phenytoin-induced gingival enlargement associated with periodontitis. Gen Dent, 56, 199-203
  18. McLoughlin P, Newman L, Brown A (1995). Oral squamous cell carcinoma arising in phenytoin-induced hyperplasia. Br Dent J, 178, 183-4.
  19. Murugan AK, Hong NT, Fukui Y, Munirajan AK, Tsuchida N (2008). Oncogenic mutations of the PIK3CA gene in head and neck squamous cell carcinomas. Int J Oncol, 32, 101-11.
  20. Murugan AK, Munirajan AK, Tsuchida N (2012). Ras oncogenes in oral cancer: the past 20 years. Oral Oncol, 48, 383-92.
  21. Pahor M, Guralnik JM, Ferrucci L, et al (1996). Calcium-channel blockade and incidence of cancer in aged populations. Lancet, 348, 493-97.
  22. Pandey AK, Gupta S (2012). Psychiatric symptomatology, scholastics, and phenytoin. Indian J Psychiatry, 54, 286-7.
  23. Qiu W, Schonleben F, Li X, et al (2006). PIK3CA mutations in head and neck squamous cell carcinoma. Clin Cancer Res, 12, 1441-6.
  24. Sailasree R, Abhilash A, Sathyan KM, et al (2008). Differential roles of p16INK4A and p14ARF genes in prognosis of oral carcinoma. Cancer Epidemiol Biomarkers Prev, 17, 414-20.
  25. Saito K, Mori S, Tanda N, Sakamoto S (1999). Expression of p53 protein and Ki-67 antigen in gingival hyperplasia induced by nifedipine and phenytoin. J Perio, 70, 581-86.
  26. Samuels Y, Wang Z, Bardelli A, et al (2004) High frequency of mutations of the PIK3CA gene in human cancers. Science, 304, 554.
  27. Subramani T, Senthilkumar K, Periasamy S, Rao S (2012). Expression of angiotensin II and its receptors in cyclosporineinduced gingival overgrowth. J Periodont Res, doi: 10.1111/jre.12020
  28. Tandon S, Tudur-Smith C, Riley RD, Boyd MT, Jones TM (2010). A systematic review of p53 as a prognostic factor of survival in squamous cell carcinoma of the four main anatomical subsites of the head and neck. Cancer Epidemiol Biomarkers Prev, 19, 574-87.
  29. Tulloch JK, Carr RR, Ensom MH (2012). A systematic review of the pharmacokinetics of antiepileptic drugs in neonates with refractory seizures. J Pediatr Pharmacol Ther, 17, 31-44.
  30. Vaccarezza GF, Antunes JL, Michaluart-Junior P (2010). Recurrent sores by ill-fitting dentures and intra-oral squamous cell carcinoma in smokers. J Public Health Dent, 70, 52-7.
  31. Varga E, Tyldesley WR (1991). Carcinoma arising in cyclosporininduced gingival hyperplasia. Br Dent J, 171, 26-7.

Cited by

  1. mutations in South Indian oral cancers vol.23, pp.5, 2017,