ANTICANCER RESEARCH 27: 563-570 (2007)

The Spectrum of Cervical Induced by Low-risk and Undefined-risk HPVs: Implications for Patient Management

ALINDA D. VÁRNAI1, MAGDOLNA BOLLMANN1, ÁGNES BÁNKFALVI2, HARALD GRIEFINGHOLT1, NATALIE PFENING1, CHRISTOPH SCHMITT3, LÁSZLÓ PAJOR4 and REINHARD BOLLMANN1

1Institute of , Bonn-Duisdorf; 2International Medical College (IMC) of the University of Münster, Münster; 3Institute of Molecular Pathology and Gene Diagnostics (ImoGen GmbH), Bonn-Duisdorf, Germany; 4Institute of Pathology of the Medical School of the University of Pécs, Pécs, Hungary

Abstract. Background: The natural history and carcinogenicity with one of about 15-18 high-risk oncogenic HPV types of rare and novel HPV types is unclear. Materials and Methods: (HR-HPV) (3), but low-risk types (LR-HPV) are also From a total of 5,964 women tested for HPV by PCR and occasionally found in cervical carcinomas. sequence analysis, Pap smears from 293 patients harbouring With the development of PCR-based HPV tests, new mono- with low-risk, undetermined-risk or novel HPV genotypes are detected in an ever increasing number. genotypes were investigated. Results: Sixty-three percent of Furthermore, there is now evidence that even the different HR- patients had ASC-US, 23% LSIL, 9% were negative and 5% HPVs can differ by an order of magnitude in risk for cervical had HSIL in cytology. Of 30 HPV types detected, 19 were of cancer and during different morphological phases of the multi- unknown risk (UR)-types including 3 novel genotypes. Four of step carcinogenesis different HPV types have different the UR-HPVs (HPV 69, 30, 67 and 34) could be assigned as oncogenic potential (3, 4). Therefore, the identification of probable high-risk types and eight as low-risk types based on prevalent HPV type(s) may aid in the stratification of women phylogenetical relationship. Morphology was not discriminative who are at greatest risk for cervical cancer. with regard to HPV type, but non-classical HPV-signs were Despite the realisation that cervical cancer is caused by generally present even in "normal" cytologies. Conclusion: HPV- HPV, the identification of pre-cancerous cervical lesions in typing is important for risk-adapted individual patient cervical screening programmes is still based on cytology management. Women harbouring novel high-risk or probably (Pap smear) the main focus of which is the detection and high-risk HPVs require more intensive care than those bearing treatment of precancerous squamous intraepithelial lesions non high-risk . (SILs) and not HPV-induced epithelial changes. However, according to our present understanding of the natural The human papillomavirus (HPV) has now been conclusively history of HPV infection and its role in cervical identified as the major risk factor for cervical cancer, with carcinogenesis, practically the whole spectrum of squamous not developing in the absence of HPV infection (1, cell abnormalities that encompasses all lesions formerly 2). HPV is associated with a variety of clinical conditions classified as condyloma (koilocytosis), , cervical that range from innocuous lesions to cancer, with most HPV intraepithelial neoplasia and carcinoma in situ can be infections being benign. The mucosotropic anogenital HPV considered as HPV-induced changes that comprise a types are classified as high-risk or low-risk depending on the morphological continuum. Current cytological classification frequency with which they cause cancer. In fact, cervical systems attempt to incorporate these changes into the cancer is only a rare complication of a persistent infection framework of epithelial cell abnormalites, specifically SILs. Although low-risk viruses are more common in low grade squamous intraepithelial lesions/cervical intraepithelial neoplasias grade 1 (LSIL/CIN1) than in high grade SILs and Correspondence to: Dr. Agnes Bánkfalvi, International Medical carcinoma in situ (HSIL/CIN2, CIN3 and CIS), high-risk College (IMC), University of Münster, Gartenstrasse 21, D-48147 viruses predominate in both (2) implicating different risk for Münster, Germany. Tel: +49(0)251 2108639, Fax: +49(0)251 2108640, e-mail: [email protected] persistence or neoplastic progression. Since LR-HPV infections are also capable of producing cytological Key Words: Cervical screening, non-classical HPV sings, HPV- abnormalities up to HSIL (CIN2), yet with negligible risk of genotyping. cancer, the determination of the type of HPV present in

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Figure 1. The 293 study patients represent a very strictly selected group out of a total of 5964 women tested for HPV by genotyping in a 5 years period (2000-2005) in our non-academic private institute of pathology. These patients were those who harboured single low-risk (LR), undefined- risk(UR) or novel HPV infections only without having a previous history of higher grade cervical disease (CIN2+) – probably caused by high-risk(HR)-HPV – or previously detected infection with any high-risk HPVs.

abnormal cervical smears is of clinical importance for risk- Materials and Methods adapted individual patient management. Since there is ample literature and sound knowledge on Screening population and procedures. The population served by our HR-HPV infections and their cytohistological correlations, private, non-academic Institute of Pathology comprises about the main objective of our study was to assess the prevalence 30,000 female inhabitants in the Bonn-region in Western Germany and spectrum of Pap smear abnormalities in women who receive annual cervical screening in gynaecological practices. Cytology and/or biopsy material is sent to our institute for routine harbouring infections with low-risk, undetermined-risk and reporting and ancillary testing. The present study population was novel HPV types detected by PCR and sequence analysis selected from a total of 5964 women who were investigated via from a routine office-based screening population. It is a HPV genotyping as an ancillary test to liquid-based cytology pragmatic management study in a pure clinical setting (ThinPrep®, Cytic, Boston, MA, USA) during the 5-year interval focusing on: i) a better understanding of the interrelationship between January 1st 2000 and December 31st 2005. Women were between morphological findings and the presence of less included in the study if they showed infection with only a single characterised or novel HPV types within cervical epithelial type of low-risk, undetermined-risk or novel HPV genotype and had no previous history of cervical dysplasia/carcinoma (≥CIN2) cells; and ii) classifying the unknown-risk and novel HPV or infection with HR-HPV. After excluding all ineligible women types into probable high-risk and probable low-risk categories from the study, 293 patients remained who completely complied based on their phylogenetical relationship with other known with the strict selection criteria. Figure 1 shows a breakdown of HPV species. patient selection. Patients who had more than one specimen tested

564 Várnai et al: Rare and Novel HPV Types-associated Cervical Lesions

Table I. The prevalence of low-risk HPV types according to their Table II. The prevalence of unknown-risk HPV types according to their corresponding cytological diagnoses. corresponding cytological diagnoses.

LR-HPV No. of cases no-ASC ASC-US LSIL HSIL UR-HPV No. of cases no-ASC ASC-US LSIL HSIL types (n) (n) (n) (n) (n) types (n) (n) (n) (n) (n)

6 38 3 21 12 2 30 3 0 1 0 2 11 5 2 0 3 0 32 5 0 4 1 0 40 1 0 1 0 0 34 1 0 0 0 1 42 45 6 28 9 2 62 11 3 6 2 0 43 3 1 1 1 0 67 16 0 8 8 0 54 11 2 7 2 0 69 1 0 1 0 0 61 38 4 26 5 3 71 1 0 1 0 0 70 22 1 13 7 1 74 3 0 3 0 0 72 1 0 1 0 0 83 14 1 9 4 0 81 12 1 8 3 0 84 18 1 12 5 0 CP 6108 1 0 1 0 0 86 3 0 3 0 0 87 2 0 2 0 0 Total 176 20 107 43 8 90 19 1 16 1 1 91 11 0 6 3 2 no-ASC: normal, or inflammatory cytology (Pap I-II); ASC-US: CP8304 4 1 3 0 0 atypical squamous cells of undetermined significance (PapIIw, non- I A09 1 0 1 0 0 classical HPV signs); LR-HPV: low-risk HPVs. JC 9710 1 0 0 1 0 MM 4 1 0 0 1 0 W13B 1 0 0 0 1

Total 117 7 76 26 7 were considered only once in the evaluation of the association of HPV types with epithelial lesions, and the highest grade of no-ASC: normal, or inflammatory cytology (Pap I-II); ASC-US: cytological abnormality was attached to the associated HPV type. atypical squamous cells of undetermined significance (PapIIw, non- We had a mean follow-up of 23 months (range: 6-84 months) in classic HPV signs); UR-HPVs: unknown-risk HPVs [all types not 243 patients. The mean age of women was 32.8±7.9 (SD) years assigned yet as low-risk, high-risk or probable high-risk type in the (range: 17-81 years). epidemiological classification of HPVs by Munoz et al. (3)].

Cytology. During the 5 years of the study, a total of 300 521 Pap tests (conventional and ThinPrep®) were evaluated at our institute. Cytological diagnoses and specimen adequacy were classified variation and bi/multinucleation); ii) disorders of keratinisation (mild according to the modified Munich (II) Cytological Classification dyskeratosis and parakeratosis); iii) measles cells and abortive (5). Terminology was converted into categories of the Bethesda ; and iv) degenerative changes (macrocytes, cytoplasmic 2001 system for the present publication; Pap I (unsuspicious) and folding and keratohyalin-like granules). Characteristic appearance of Pap II (inflammatory) lesions were considered negative for these minor cytological changes is demonstrated in Figure 2. This intraepithelial lesion or malignancy (no-ASC); Pap IIw expanded cytomorphological HPV diagnosis is routinely performed [(w=wiederholen/repeat): not an official class in the Munich II in our institute for pre-selecting cases prior to performing expensive classification but very often used in daily practice] includes ASC- molecular HPV tests, as described elsewhere (6). US and/or non-classic HPV-signs; Pap IIID lesions with mild dysplasia/CINI were considered low grade squamous intraepithelial Histology. All cases with cytological diagnoses of HSIL were proven lesions (LSIL), whereas Pap IIID with moderate dysplasia and Pap histologically according to standard CIN classification. Histologies IVA (severe dysplasia) were classified as high-grade squamous in lower cytological grades were not consistently available. intraepithelial lesions (HSIL). Smears were analysed by well-trained cytotechnologists who PCR-based HPV DNA detection and genotyping by sequence analysis. have a restricted workload of screening 10 smears per hour in During the 5 years of this study, a total of 8503 HPV DNA tests were accordance with quality assurance guidelines for cytological performed during after informed consent of the women. Detailed examinations issued by the German Medical Association. All cases protocols for the assays have been published elsewhere (7). In short: with borderline or uncertain diagnosis (PapIIw, III/ roughly HPV DNA detection was directly performed on residual material in equating ASC-US, ASC-H) and those with squamous the ThinPrep® collection vial with PCR-based assays using the intraepithelial lesions (≥PapIIID/LSIL-HSIL) were re-reviewed by improved MY09/MY11 consensus primers and the GP5+/6+ general a cytopathologist (MB) for affirmation. primers for amplification of HPV DNA. Reaction parameters In addition to standard cytological screening for the presence of included initial denaturing at 95ÆC for 5 minutes, 40 cycles each at atypical squamous epithelial cells of any grade, classic and particularly 95ÆC for 30 seconds, 37ÆC or 58ÆC for annealing the primer pairs non-classic HPV-induced cytological alterations were also carefully GP5+/6+ or MY09/MY11, respectively. A final extension step was assessed in all cases. The non-classic features of HPV effect included; done at 72ÆC for 5 minutes. The presence of human genomic DNA i) minor nuclear abnormalities (mild hyperchromasia, mild nuclear was verified by PCR amplification of the human globine gene. This

565 ANTICANCER RESEARCH 27: 563-570 (2007)

Figure 2. a) Overview of a Pap smear with minimal cytological abnormalities and non-classic signs of HPV effect. b) Abortive with pycnotic nucleus. c) Macrocyte (left cell) with mild nuclear hyperchromasia, mild nuclear enlargement and fine, uniform granularity of the chromatin. d) Spindle- shaped nucleus with longitudinal nuclear grooves. e) Macrocytic "measle" cell with condensation of the entire cytoplasmic matter into fine granules and empty looking remaining cytoplasm and pycnotic nucl. f) Keratohyalin-like granules (spherical globuli) in the cytoplasma.

reaction served as a positive control. PCR products were purified were analysed on an ABI Prism 310 automated sequencer (Applied using the High Pure PCR product purification kit (Roche Biosystems); results were compared with documented virus sequences Diagnostics, Mannheim, Germany) according to the manufacturers available in the GenBank database using the BLAST program (Blast, instructions. The sequence of one strand of the purified PCR Pittsboro, NC, USA). Classification of HPV types was based on the fragments was determined with the BigDye Terminator sequencing epidemiological risk assessment of human papillomaviruses in the kit (Applied Biosystems, Foster City, CA, USA) using 3-5 pmol of development of cervical cancer (3) and according to their GP5+ or MY09 as the sequencing primers. Sequencing reactions phylogenetical origin (8), respectively.

566 Várnai et al: Rare and Novel HPV Types-associated Cervical Lesions

Table III. Phylogenetical grouping, corresponding epidemiological risk- Table IV. Phylogenetical grouping, epidemiological risk-classification and classification and cytological findings in monoinfections with low-risk cytological findings in monoinfections with probably high-risk, unknown- HPVs from the alpha genus. risk and novel anogenital HPVs from the alpha genus.

Genus/ HPV type No. of no-ASC ASC-US LSIL HSIL Genus/ HPV type No. of no-ASC ASC-US LSIL HSIL type (species) cases (n) (n) (n) (n) (n) type (species) cases (n) (n) (n) (n) (n)

known low-risk HPV types probable high-risk HPV types and probable low-risk types (based on genetical relationship or (based on genetical relationship or rare association with cancer) no proven association with cancer) ·5 69 1 0 1 0 0 ·1 32 5 0 4 1 0 ·6 30 3 0 1 0 2 42 45 6 28 9 2 ·9 67 16 0 8 8 0 ·3 61 38 4 26 5 3 ·11 34 1 0 0 0 1 72 1 0 1 0 0 ? MM 4*** 1 0 0 1 0 81 12 1 8 3 0 unknown-risk HPV types 62 11 3 6 2 0 ·15 71 1 0 1 0 0 83 14 1 9 4 0 90 19 1 16 1 1 84 18 1 12 5 0 novel HPV types 86 3 0 3 0 0 ? I A09 1 0 1 0 0 87 2 0 2 0 0 ? JC 9710 1 0 0 1 0 ·7 70* 21 1 13 7 1 ? W13B 1 0 0 0 1 ·8 40 1 0 1 0 0 43 3 1 1 1 0 Types written in black were classified epidemiologically by Munoz et 91 11 0 6 3 2 al. (3). Types written in red and blue have not been classified according ·10 6 38 3 21 12 2 to risk for cervical cancer, yet. Types written in green are novel HPV 11 5 2 0 3 0 genotypes. ***MM4 has been found in carcinomas in a few women 74 3 0 3 0 0 from different geographical areas and thus it was proposed to have ·13 54 11 2 7 2 0 high oncogenic potential (10). ? CP 6108 1 0 1 0 0 ? CP 8304** 4 1 3 0 0

Types written in black were classified epidemiologically by Munoz et al. (3). Types written in red have not been classified epidemiologically et al. (3). There was no difference in the mean age of the according to risk for cervical cancer. *Risk assignment of HPV 70 is patients in the different groups with LR- or UR-HPV controversial; it belongs to the high-risk ·7 species, but was infections (32.97 and 32.89 years, respectively). characterised as low-risk epidemiologically by Munoz et al. (3). None of the patients had cervical carcinoma; 15 women **CP8304 has been proposed to be non-high-risk by Meyer et al. (10). (5%) had HSIL in cytology – of these, 2 were histologically diagnosed as carcinoma in situ, 8 as CIN3 and 7 as CIN2 lesions. Seven HSILs were associated with UR-HPVs (2 x HPV 30, 1 x HPV34, 1x HPV 90, 2 x HPV 91 and 1 x Statistical analyses. Statistical analyses were performed using the SPSS program (Version 12.0). Descriptive statistics for continuous W13B) and 8 with LR-HPVs (2 x HPV6, 2 x HPV 42, 3x measures are given as the mean with standard deviation (SD) and HPV 61 and 1 x HPV 70). range, frequencies and prevalences are given in row numbers and The most frequent cytological diagnosis was ASC-US percentage. (n=183; 63%); 107 of the cases (59%) were associated with LR-HPVs and 76 cases (41%) with UR-HPVs. The second Results largest cytological group was LSIL (n=69; 23%); about two- thirds of the cases (43/69) were associated with LR-HPVs The prevalence of different HPV types and their and one-third (26/69) with UR-HPVs. Twenty-seven cases corresponding cytological diagnoses is shown in Tables I (9%) were negative for intraepithelial lesion or malignancy and II. Overall, 30 different HPV types were detected in the (no-ASC) indicative of reactive, inflammatory or atrophic 293 study patients: 11 low-risk types (LR-HPVs) and 19 changes. About three-quarters (20/27) of the cytologically undetermined-risk types (UR-HPVs); the latter group "negative" cases were associated with LR-HPVs and one- included 3 novel HPV genotypes. LR-HPVs contained HPV quarter (7/27) with UR-HPVs. 6, 11, 40, 42, 43, 54, 61, 70, 72, 81 and CP6108; UR-HPVs The most prevalent HPV types in the study population contained all other types detected that are not assigned as were: HPV 42 (n= 45; 15%), HPV 6 and HPV 61 (both low-risk, high-risk or probable high-risk types yet in the n=38; 13%), HPV 70 (n=21; 7%), HPV 90 (n= 19; 6.5%) epidemiological classification of HPVs published by Munoz and HPV 84 (n=18; 6.1%).

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After re-classifying our data according to the infections. However, many HPV infections remain phylogenetical grouping of known HPVs [9], 12 of 19 UR- suppressed for a long time without showing these prominent HPVs could be assigned either as LR types (HPV 32, 62, 83, cytological alterations, or striking colposcopical evidence of 84, 86, 87, 91, 74) or probable high-risk types (HPV 69, 30, disease, but are detectable with HPV DNA by sensitive 67, 34) as demonstrated in Tables III and IV. PCR-based methods. Reported rates of HPV-positive Concerning cytomorphology, all negative smears (no- "normal" specimens encompass 25% (13), 29% (14), 36% ASC) and ASC-US cases showed at least one but generally (15) and 44% (16), respectively. In our present study, 27 more minor non-classical cytological alterations indicative cases (9%) were classified as "normal" (no-ASC) in the strict of HPV effect. These included in decreasing order of sense of showing no signs of cytological atypia. However, frequency: mild nuclear changes, disorders of keratinisation, using our recently published expanded cytological abortive koilocytes and "measles" cells as well as classification of HPV-related alterations (6), minor non- degenerative changes, as published recently by our group classic HPV-signs, especially mild nuclear changes, could be (6) and demonstrated in Figure 2. The majority of LSIL and seen in all these specimens. Similar observations were made HSIL cases showed a combination of both classic by others as well (12, 17, 18). These results suggest that the (koilocytosis and dyskeratosis) and non-classic HPV-signs. rate of "normal" cytology with a positive HPV test would be No cytological progression of lesions was observed in radically reduced by careful assessment of non-classic HPV- many patients in a mean follow-up period of 23 months effects. (range: 6-84 months). Intriguingly, it was not possible in any of these studies to discriminate between HR-HPV and non-HR-HPV positive Discussion cases cytomorphologically. This is of particular note, especially in women with minimally abnormal Pap tests, According to current knowledge, cervical cancer develops because patients with HR-HPV positive "normal" smears from pre-cancerous cervical lesions (CINs) over the course are at increased risk for developing CIN3 (19, 20), whereas of many years, following a well defined path with the probability of progression to HSIL in women who identifiable intermediate stages (11). In many respects this screen positive for LR-HPVs is extremely low (21). Thus, process mirrors the natural course of other HPV-associated HPV typing may have clinical importance for risk adapted benign diseases. Changes caused by HR-HPV infections and individual patient management. those associated with common LR-HPV types are well The most common diagnosis in our present study was characterised because of their outstanding clinical ASCUS (63%) referring to cells with nuclear atypia of significance and high prevalence. There are, however, a which 59% were associated with LR-HPV types and 41% growing number of new or rare HPV types with yet with UR-HPV types. Because of the general presence of unknown oncogenic potential and poorly characterized one or more non-classic HPV-related signs in the ASCUS phenotypic appearance of infected cells. smears and the epidemiological evidence that infection with In our present strictly selected collective, none of the 293 HR-HPV types – undistinguishable morphologically from cases with single non-high-risk HPV infection only was non-HR-HPVs – is associated with cervical neoplasia, mild diagnosed as a carcinoma and only 15 (5%) HSILs were cytological HPV-related changes should be viewed with classified by cytological assessment. This, and the finding of greater concern and possibly added to the ASCUS no disease progression during follow-up, supports the well- definition. This is further supported by the prospective known low oncogenic potential of LR-HPVs and affirms follow-up in the ASCUS/LSIL Triage Study that suggested a that the 19 unknown-risk types detected in the present study similar risk for subsequent CIN grade 2 or 3, approximately (see Table I and Table II) are probably alike. However, in 12% over 2 years, among women with HPV-positive our screening population, 2 verrucous carcinomas and one ASCUS and LSIL indicating that these two cytological transitional cell cervical carcinoma were detected after findings are clinically equivalent (22). persistent infections with HPV 6, 11 and 42, respectively Of the 69 LSIL cases (23%) in the present study, 39% [authors unpublished data]. These women were ineligible were associated with UR-HPV types and 61% with LR- for the present study. HPV types. It was not possible to predict, based on In current cytological classification systems, HPV cytological appearance, which HPV type was responsible infections are diagnosed on the basis of classic cytopathic for the infection, nor was it possible to distinguish effects of HPV, such as koilocytosis, dyskeratosis, and between a "dysplasia" (neoplastic transformation) and a "rasinoid-shape" nuclear atypia. While these cytological or "simple HPV infection" of a productive type. The few histological changes are, when strictly used, specific, they HSIL cases, (15; 5%) were associated in almost equal are only moderately sensitive for diagnosis of HPV (12). frequency with LR-HPVs (n=8) and UR-HPVs (n=7) in These are the morphological correlates of productive viral the present setting.

568 Várnai et al: Rare and Novel HPV Types-associated Cervical Lesions

Though indistinguishable morphologically, the risks of viral least 20 other genotypes and the theoretical concern that persistence and neoplastic progression to CIN3/carcinoma additional HPV genotypes could expand to fill the gap left differ markedly with HPV type, with genetically related types after effective vaccination against only HPV 16 and 18. appearing to act most alike (3). Accordingly, phylogenetic Therefore, further research is needed to understand the grouping may predict the natural history and carcinogenicity pathobiology of papillomavirus infection, in which each HPV of individual HPV types (9) even if they occur rarely and thus type should be considered as a separate sexually transmitted cannot be evaluated epidemiologically. infection posing different risk for cervical cancer. In the present study, 8 out of 19 unknown-risk HPVs could be classified as LR-types (HPV 32, 62, 83, 84, 86, 87, Conclusion 91, 74) based on their common genetic origin (see Table III). Additionally, our present findings on the relative high Our present findings highlight the importance of type- prevalence of HPV 90 (n=19; 6.5%), which belongs to specific HPV testing and the use of expanded cytological species ·15 together with HPV 71, and its association with classification of HPV signs from the lowest end of HPV- 16 ASC-US, 1 normal, 1 LSIL and 1 HSIL cases suggest a related cellular changes and upwards. Non-classic signs of low oncogenic potential of this type as well. From the HPV-effect should be viewed with greater concern and remaining types, CP8304 has also been detected in smears possibly added to the ASC-US definition. The efficiency of with normal cytologies but not in cancer (10, 15, 23). Our cytology-based cervical cancer screening programs could be present data also support the classification of CP8304 as a increased by including objective individual risk factors for low-risk type. Risk assignment of HPV 70 is controversial; it women with normal or equivalent cytology, or with cervical belongs to the high-risk ·7 species, but was characterised as diseases at the lower end of squamous intraepithelial low-risk epidemiologically by Munoz et al. (3). In our lesions, such as the detection and risk assignment of HPV- present study, HPV 70 was found in 21 women (1 normal type(s) according to epidemiological classifications or cytology, 13 ASC-US, 7 LSIL and 1 HSIL). The majority of phylogenetical origin. these infections has been persisting for over years indicating that the immune system cannot readily eliminate this HPV Acknowledgements type and the development of associated higher grade We would like to thank Marianna Szendy and Dr. Ante Trosic for pathologic changes is rare and slow. The species IA 09, JC the excellent technical assistance and pre-screening. 9710 and W 13B, found in a small number of patients only, represent novel genotypes with an as yet undetermined References oncogenic risk. Intriguingly, four viruses from the UR-types (HPV 69, 30, 1 Walboomers JMM, Jacobs MV, Manos MM, Bosch FX, 67, 34) could be assigned as probable high-risk species Kummer JA, Shah KV, Snijders PJ, Peto J, Meijer CJ and based on their genetic relationship with other HR-HPVs. Of Munoz N: Human papillomavirus is a necessary cause of them, HPV 67 occurred in 16 women (8 ASC-US and 8 invasive cervical cancer worldwide. J Pathol 189: 12-19, 1999. 2 Bosch FX, Lorincz A, Munoz N Meijer CJ and Shah KV: The LSIL). This type belongs phylogenetically to species ·9, causal relation between Human Papillomavirus and cervical which comprises the most carcinogenic HPV types including cancer. J Clin Pathol 55: 244-265, 2002. HPV16. Also MM44, which was detected only in 1 LSIL in 3 Munoz N, Bosch FX, de Sanjose S, Herrero R, Castellsague X, our present setting, has been found by other investigators in Shah KV, Snijders PJ and Meijer CJ; International Agency for a few cervical carcinomas in women from different Research on Cancer Multicenter Cervical Cancer Study Group: geographical areas indicating its high oncogenic potential Epidemiologic classification of human papillomavirus types (10). HPV 30 was detected in one laryngeal carcinoma by associated with cervical cancer. N Engl J Med 348: 518-527, 2003. Kahn and co-workers (24) and it was found in two women 4 Bulkmans NW, Bleeker MC, Berkhof J, Voohosrt FJ, Snijders with HSIL/carcinoma in situ in our present study. Thus, the PJ and Meijer CP: Prevalence of types 16 and 33 is increased detection of probable high-risk genotypes in LSIL and lower in high-risk human papillomavirus positive women with cervical cytologies can serve as a prognostic indicator as to the intraepithelial neoplasia grade 2 or worse. Int J Cancer 117: chance of progression to HSIL and cancer, and such 177-181, 2005. patients would possibly benefit from more intensive 5 Soost HJ: Befundwiedergabe in der gynäkologischen management than women with other HPV types. Zytodiagnostik – Münchener Normenklatur II. Gynäkol Prax The prophylactic and therapeutic targeting of HR-HPV 14: 433-438, 1990. 6 Bollmann M, Bánkfalvi A, Trosic A, Speich N, Schmitt C and infections, particularly HPV 16 and 18, is the subject of Bollmann R: Can we detect cervical human papillomavirus much current research. However, it is important to note that (HPV) infection by cytomorphology alone? Diagnostic value of together HPV 16 and 18 currently account for about 70% of non-classic cytological signs of HPV effect in minimally cervical cancer cases (2). This leaves ~30% caused by at abnormal Pap tests. 16: 13-21, 2005.

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