Cervical cancer is the fifth most common type of cancer worldwide (around 500,000 new cases diagnosed each year) and the second major cause of cancer related to women's deaths. 1,2 It has been estimated that over 95% of cervical cancer is associated with HPV infection, 3 since many cervical carcinomas and pre-invasive lesions contain human papillomavirus DNA sequences. 4 Papillomaviruses are classified as being low-risk or high-risk, depending on their preferential association with benign or malignant lesions. HPV-16 DNA is found in about 50 -60% of human anogenital carcinomas, HPV 18 in about 20% and other HPV genotypes in a further 15%. There are as yet undefined papillomavirus sequences in about 15% of anogenital carcinomas. 5,6 Papillomaviruses (PVs) are nonenveloped particles, characterized by having an icosahedral capsid of about 55 nm diameter. The capsids are composed of 2 virally encoded proteins (the L1 major and the L2 minor capsid proteins) that migrate on SDS-PAGE at about 55 and 75 kDa, respectively. The papillomavirus L1 major capsid protein (L1) is the most abundant virus-encoded protein, being highly conserved amongst different HPV types, having amino-acid homology as high as 80%. L1 presents the ability to self-assemble in virus-like particles (VLPs). 7,8 Different PVs seem to use the same receptor to enter their host cells. 7,9,10 Trypsin treatment of cells markedly reduced their ability to bind to VLPs, suggesting that a surface cell protein was involved in VLP binding. 11,12 Red blood cell invasion by Plasmodium falciparum merozoites, 13-15 reticulocytes by Plasmodium vivax merozoites 14 and hepatic cells by Hepatitis C virus 16 has shown that specific receptorligand interactions mediate microbe binding to host cells. Using a similar methodology, synthetic 125 I-radio-labeled peptides were thus used in receptor-ligand type binding assays to better pin/point VERO and HeLa cell binding activity from high-risk 16 and 18 type L1 amino acid sequences. VERO and HeLa cell high activity binding peptides (HABPs) showed saturable binding, cell receptors being sensitive to enzyme treatment and the majority of bound peptide being surface-exposed. HABPs specifically bound to 69 and 54 kDa HeLa cell surface membrane protein and inhibited HPV-16 VLP binding to both cells. Furthermore, several HABP residues formed part of the VLP surface, becoming the target for neutralizing antibodies.
METHODS
Peptide synthesisTwenty-seven sequential 20-mer peptides, corresponding to HPV type 16 L1, 17 and 29 sequential 20-mer peptides, corresponding to HPV 18 L1, 18 were synthesized by solid-phase multiple peptide system. 19,20 MBHA resin (0.7 meq/g), t-Boc amino-acid and low-high cleavage techniques were used. 21 Peptides were analyzed by MALDI-TOF mass spectrometry and reverse phasehigh performance liquid chromatography (RP-HPLC). These synthesized peptides are shown in 1-letter code in Figures 1a,b. An extra Tyr residue was added to a peptide's C-terminus for radiolabeling purposes when such peptide did not contain Tyr...