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Human papillomavirus genotyping by multiplex pyrosequencing in cervical cancer patients from India

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Abstract

Cervical cancer is a leading cause of cancer-related deaths among women in India. Human papillomavirus (HPV) infection is the causative agent of cervical cancer; and infection with the high-risk genotypes, predominantly HPV16 and 18, is the biggest risk factor. Vaccines targeting HPV16 and 18 have been found to confer protection in large-scale clinical trials. HPV genotyping has traditionally been carried out to screen the population “at risk” using indirect methods based on polymerase chain reaction (PCR) using consensus primers combined with various DNA hybridization techniques, and often followed by the sequencing of candidate products. Recently, a high-throughput and direct method based on DNA sequencing has been described for HPV genotyping using multiplex pyrosequencing. We present a pilot study on HPV genotyping of cervical cancer and non-malignant cervical samples using multiplex pyrosequencing. Using genomic DNA from cell lines, cervical biopsies, surgical tissues or formalin-fixed, paraffin-embedded tissue samples, we could successfully resolve 6 different HPV types out of the 7 tested, with their prevalence found to be in agreement with earlier reports. We also resolved coinfections with two different HPV types in several samples. An HPV16 genotype with a specific and recurrent sequence variation was observed in 8 cancer samples and one non-malignant sample. We find this technique eminently suited for high-throughput applications, which can be easily extended to large sample cohorts to determine a robust benchmark for HPV genotypes prevalent in India.

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References

  • Bosch F X, Lorincz A, Munoz N, Meijer C J and Shah K V 2002 The causal relation between human papillomavirus and cervical cancer; J. Clin. Pathol. 55 244–265

    Article  CAS  Google Scholar 

  • Choi Y D, Jung W W, Nam J H, Choi H S and Park C S 2005 Detection of HPV genotypes in cervical lesions by the HPV DNA chip and sequencing; Gynecol. Oncol. 98 369–375

    Article  CAS  Google Scholar 

  • Clifford G M, Smith J S, Plummer M, Munoz N and Franceschi S 2003 Human papillomavirus types in invasive cervical cancer worldwide: a meta-analysis; Br. J. Cancer 88 63–73

    Article  CAS  Google Scholar 

  • Cope J U, Hildesheim A, Schiffman M H, Manos M M, Lorincz A T, Burk R K, Glass A G, Greer C et al 1997 Comparison of the hybrid capture tube test and PCr for detection of human papillomavirus DNA in cervical specimens; J. Clin. Microbiol. 35 2262–2265

    CAS  PubMed  PubMed Central  Google Scholar 

  • Coutlee F, Gravitt P, Kornegay J, Hankins C, Richardson H, Lapointe N, Voyer H and Franco E 2002 Use of PGMY primers in L1 consensus PCR improves detection of human papillomavirus DNA in genital samples; J. Clin. Microbiol. 40 902–907

    Article  CAS  Google Scholar 

  • de Roda Husman A M, Walboomers J M, van den Brule A J, Meijer C J and Snijders P J 1995 The use of general primers GP5 and GP6 elongated at their 3′ ends with adjacent highly conserved sequences improves human papillomavirus detection by PCR; J. Gen. Virol. 76 (Pt 4) 1057–1062

    Article  Google Scholar 

  • Elahi E and Ronaghi M 2004 Pyrosequencing: a tool for DNA sequencing analysis; Methods Mol. Biol. 255 211–219

    CAS  PubMed  Google Scholar 

  • Franceschi S, Rajkumar T, Vaccarella S, Gajalakshmi V, Sharmila A, Snijders P J, Munoz N, Meijer C J and Herrero R 2003 Human papillomavirus and risk factors for cervical cancer in Chennai, India: a case-control study; Int. J. Cancer 107 127–133

    Article  CAS  Google Scholar 

  • Franceschi S, Rajkumar R, Snijders P J, Arslan A, Mahe C, Plummer M, Sankaranarayanan R, Cherian J, Meijer C J and Weiderpass E 2005 Papillomavirus infection in rural women in southern India; Br. J. Cancer 92 601–606

    Article  CAS  Google Scholar 

  • Frazer I H 2004 Prevention of cervical cancer through papillomavirus vaccination; Nat. Rev. Immunol. 4 46–54

    Article  CAS  Google Scholar 

  • Gharizadeh B, Kalantari M, Garcia C A, Johansson B and Nyren P 2001 Typing of human papillomavirus by pyrosequencing; Lab. Invest. 81 673–679

    Article  CAS  Google Scholar 

  • Gharizadeh B, Ghaderi M, Donnelly D, Amini B, Wallin KL and Nyren P 2003 Multiple-primer DNA sequencing method; Electrophoresis 24 1145–1151

    Article  CAS  Google Scholar 

  • Gharizadeh B, Oggionni M, Zheng B, Akom E, Pourmand N, Ahmadian A, Wallin K L and Nyren P 2005 Type-specific multiple sequencing primers: a novel strategy for reliable and rapid genotyping of human papillomaviruses by pyrosequencing technology; J. Mol. Diagn. 7 198–205

    Article  CAS  Google Scholar 

  • Gharizadeh B, Zheng B, Akhras M, Ghaderi M, Jejelowo O, Strander B, Nyren P, Wallin K L and Pourmand N 2006 Sentinel-base DNA genotyping using multiple sequencing primers for high-risk human papillomaviruses; Mol. Cell Probes 20 230–238

    Article  CAS  Google Scholar 

  • Harper D M, Franco E L, Wheeler C M, Moscicki A B, Romanowski B, Roteli-Martins C M, Jenkins D, Schuind A, Costa Clemens S A and Dubin G 2006 Sustained efficacy up to 4.5 years of a bivalent L1 virus-like particle vaccine against human papillomavirus types 16 and 18: follow-up from a randomised control trial; Lancet 367 1247–1255

    Article  CAS  Google Scholar 

  • Jacobs M V, de Roda Husman A M, van den Brule A J, Snijders P J, Meijer C J and Walboomers J M 1995 Group-specific differentiation between high-and low-risk human papillomavirus genotypes by general primer-mediated PCR and two cocktails of oligonucleotide probes; J. Clin. Microbiol. 33 901–905

    CAS  PubMed  PubMed Central  Google Scholar 

  • Klaassen C H, Prinsen C F, de Valk H A, Horrevorts A M, Jeunink M A and Thunnissen F B 2004 DNA microarray format for detection and subtyping of human papillomavirus; J. Clin. Microbiol. 42 2152–2160

    Article  CAS  Google Scholar 

  • Legood R, Gray A M, Mahe C, Wolstenholme J, Jayanth K, Nene B M, Shashtri S S, Malvi S G, Muwonge R, Budukh A and Sankaranarayanan R 2005 Screening for cervical cancer in India: how much will it cost? A trial based analysis of the cost per case detected; Int. J. Cancer 117 981–987

    Article  CAS  Google Scholar 

  • Lie A K, Risberg B, Borge B, Sandstad B, Delabie J, Rimala R, Onsrud M and Thoresen S 2005 DNA-versus RNA-based methods for human papillomavirus detection in cervical neoplasia; Gynecol. Oncol. 97 908–915

    Article  CAS  Google Scholar 

  • Menon M M, Simha M R and Doctor V M 1995 Detection of human papillomavirus (HPV) types in precancerous and cancerous lesions of cervix in Indian women: a preliminary report; Indian J. Cancer 32 154–159

    CAS  PubMed  Google Scholar 

  • Moscicki A B, Schiffman M, Kjaer S and Villa L L 2006 Chapter 5: updating the natural history of HPV and anogenital cancer; Vaccine 24(Suppl 3) S42–S51

    Article  Google Scholar 

  • Munirajan A K, Kannan K, Bhuvarahamurthy V, Ishida I, Fujinaga K, Tsuchida N and Shanmugam G 1998 The status of human papillomavirus and tumor suppressor genes p53 and p16 in carcinomas of uterine cervix from India; Gynecol. Oncol. 69 205–209

    Article  CAS  Google Scholar 

  • Munoz N, Bosch F X, de Sanjose S, Herrero R, Castellsague X, Shah K V, Snijders P J and Meijer CJ 2003 Epidemiologic classification of human papillomavirus types associated with cervical cancer; N. Engl. J. Med. 348 518–527

    Article  Google Scholar 

  • Nelson J H, Hawkins G A, Edlund K, Dvander M, Kjellberg L, Wadell G, Dillner J, Gerasimova T et al 2000 A novel and rapid PCR-based method for genotyping human papillomaviruses in clinical samples; J. Clin. Microbiol. 38 688–695

    CAS  PubMed  PubMed Central  Google Scholar 

  • Pretet J L, Jacquard A C, Carcopino X, Charlot J F, Bouhour D, Kantelip B, Soubeyrand B, Leocmach Y, Mougin C and Riethmuller D 2008a Human papillomavirus (HPV) genotype distribution in invasive cervical cancers in France: EDITH study; Int. J. Cancer 122 424–427

    Article  CAS  Google Scholar 

  • Pretet J L, Jacquard A C, Carcopino X, Monnier-Benoit S, Averous G, Soubeyrand B, Leocmach Y, Mougin C and Riethmuller D 2008b Human papillomavirus genotype distribution in high grade cervical lesions (CIN 2/3) in France: EDITH study; Int. J. Cancer 122 428–432

    Article  CAS  Google Scholar 

  • Roden R and Wu T C 2006 How will HPV vaccines affect cervical cancer?; Nat. Rev. Cancer 6 753–763

    Article  CAS  Google Scholar 

  • Ronaghi M 2001 Pyrosequencing sheds light on DNA sequencing; Genome Res. 11 3–11

    Article  CAS  Google Scholar 

  • Sankaranarayanan R, Chatterji R, Shastri S S, Wesly R S, Basu P, Mahe C, Mowonge R, Sgigneurin D, et al 2004 Accuracy of human papillomavirus testing in primary screening of cervical neoplasia: results from a multicenter study in India; Int. J. Cancer 112 341–347

    Article  CAS  Google Scholar 

  • Sankaranarayanan R, Nene B M, Dinshaw K A, Mahe C, Jayanth K, Shastri S S, Malvi S G, Chinoy R, et al 2005 A cluster randomized controlled trial of visual, cytology and human papillomavirus screening for cancer of the cervix in rural India; Int. J. Cancer 116 617–623

    Article  CAS  Google Scholar 

  • Schmiedeskamp M R and Kockler D R 2006 Human papillomavirus vaccines; Ann. Pharmacother. 40 1344–1352

    Article  CAS  Google Scholar 

  • Shastri S S, Dinshaw K, Amin G, Goswami S, Patil S, Chinoy R, Kane S, Kelkar R et al 2005 Concurrent evaluation of visual, cytological and HPV testing as screening methods for the early detection of cervical neoplasia in Mumbai, India; Bull. World Health Organ. 83 186–194

    PubMed  PubMed Central  Google Scholar 

  • Smits H L, Bollen L J, Tjong A H S P, Vonk J, Van Der Velden J, Ten Kate F J, Kaan J A, Mol B W and Ter Schegget J 1995 Intermethod variation in detection of human papillomavirus DNA in cervical smears; J. Clin. Microbiol. 33 2631–2636

    CAS  PubMed  PubMed Central  Google Scholar 

  • Sowjanya A P, Jain M, Poli U R, Padma S, Das M, Shah K V, Rao B N, Devi R R, Gravitt P E and Ramakrishna G 2005 Prevalence and distribution of high-risk human papilloma virus (HPV) types in invasive squamous cell carcinoma of the cervix and in normal women in Andhra Pradesh, India; BMC Infect. Dis. 5 116

    Article  Google Scholar 

  • Trottier H and Franco EL 2006 The epidemiology of genital human papillomavirus infection; Vaccine (Suppl 1) 24 S1–S15

    Google Scholar 

  • Unger E R 2000 In situ diagnosis of human papillomaviruses; Clin. Lab. Med. 20 289–301, vi

    Article  CAS  Google Scholar 

  • van den Brule A J, Pol R, Fransen-Daalmeijer N, Schouls L M, Meijer C J and Snijders P J 2002 GP5+/6+ PCR followed by reverse line blot analysis enables rapid and high-throughput identification of human papillomavirus genotypes; J. Clin. Microbiol. 40 779–787

    Article  Google Scholar 

  • Ylitalo N, Bergstrom T and Gyllensten U 1995 Detection of genital human papillomavirus by single-tube nested PCR and type-specific oligonucleotide hybridization; J. Clin. Microbiol. 33 1822–1828

    CAS  PubMed  PubMed Central  Google Scholar 

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Author notes

  1. Chandan Kumar-Sinha

    Present address: Michigan Center for Translational Pathology, Department of Pathology, University of Michigan, Ann Arbor, Michigan, 48109, USA

Authors and Affiliations

  1. Advanced Center for Treatment, Research and Education in Cancer (ACTREC), Tata Memorial Centre, Navi Mumbai, India

    Cheryl M. Travasso, Mona Anand, Mansi Samarth, Aditi Deshpande & Chandan Kumar-Sinha

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  1. Cheryl M. Travasso
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  2. Mona Anand
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  3. Mansi Samarth
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Correspondence to Chandan Kumar-Sinha.

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Travasso, C.M., Anand, M., Samarth, M. et al. Human papillomavirus genotyping by multiplex pyrosequencing in cervical cancer patients from India. J Biosci 33, 73–80 (2008). https://doi.org/10.1007/s12038-008-0023-x

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