ORIGINAL RESEARCH * RECHERCHES NOUVELLES
Human papillomavirus and cervical intraepithelial neoplasia in women who subsequently had invasive cancer Deoraj Caussy, PhD; Loraine D. Marrett, PhD; Ann J. Worth, MD; Mary McBride, MSc; William E. Rawls, MD In a retrospective case-control study biopsy specimens of cervical intraepithelial neoplasia (CIN) lesions from 47 women in whom invasive cancer subsequently developed (cases) and from 94 control subjects in whom CIN was diagnosed within 6 months of the diagnosis for the matched case subject but invasive disease did not develop were tested for human papillomavirus (HPV) DNA with tissue in-situ hybridization. There were no significant differences in the frequency of detection of HPV DNA between the two groups. In a cross-sectional survey the prevalence of HPV DNA was found to be 11% in specimens without CIN, 27% in those with CIN I, 49% in those with CIN II and 56% in those with CIN III. The positivity rates for HPV 16/33 DNA increased with the severity of CIN, but this was not observed for HPV 6/11 and 18 DNA. A comparison of the results of the case-control and cross-sectional studies suggested that the younger cohort of women had higher prevalence rates of HPV DNA than the older cohort.
Etude retrospective et comparative des lesions de neoplasie intra-epitheliale du col uterin (NICU) chez 47 femmes (constituant le groupe des sujets) ayant montre par la suite un cancer envahissant et chez 94 temoins oui le diagnostic de NICU avait e pose dans les 6 mois precedent ou suivant le moment ou il avait e pose chez le sujet apparie mais qui ne devaient pas montrer de cancer envahissant. Dans les biopsies on recherche les ADN du papillomavirus humain (PVH) par hybridation tissulaire in situ. La frequence de detection de ces ADN ne differe pas significativement d'un groupe a l'autre. Une etude transversale montre la presence d'ADN-PVH dans 11% des echantillons sans NICU, 27% en cas de NICU I, 49% en cas de NICU II et 56% en cas de NICU III. II existe un rapport direct entre la frequence de positivite de la recherche de l'ADN-PVH 16/33 et la severite de la NICU, ce qui ne s'observe pas pour les ADN-PVH 6/11 et 18. Si on met en regard les resultats de l'etude comparative et ceux de l'etude transversale on peut etre amend a croire que les femmes jeunes ont un plus haut taux de presence d'ADN-PVH que leurs ainees. C
ervical squamous cell carcinoma is generally believed to evolve from various grades of cervical intraepithelial neoplasia (CIN).' The incidence of CIN far exceeds that of invasive cancer; this suggests that only a fraction of the preinvasive lesions progress to cervical cancer.2 Little is known
about the factors contributing to the ultimate invasive potential of a particular CIN lesion, but the human papillomaviruses (HPV) have been implicated in cervical cancer;3 the evidence is based mainly on biologic and clinical observations. HPV 16 DNA can induce cellular transformation in vitro and thus
From the Department of Pathology, McMaster University, Hamilton, Ont., the Ontario Cancer Treatment and Research Foundation, Toronto, the Cancer Control Agency of British Columbia, Vancouver, and the Viral Epidemiology Section, National Cancer Institute, National Institutes ofHealth, Bethesda, Md.
Reprint requests to: Dr. William E. Rawls, Department ofPathology, McMaster University, 1200 Main St. W, Hamilton, Ont. L8N 3Z5 -
For prescribing information
see
page 403
CAN MED ASSOC J 1990; 142 (4)
311
has potential oncogenic properties.4 The preferential occurrence of HPV 16 DNA, and to a lesser extent HPV 18 and HPV 33 DNA, in invasive cervical cancers provides the main clinical evidence, but the limitations of the research designs allow two equally plausible but contrasting interpretations.5 First, these viruses may occur secondary to the cancer state as opportunistic pathogens. Second, these viruses could be involved with a part of the neoplastic process that culminates in invasive cancer, in which case the presence of HPV DNA would likely be predictive of CIN lesions that will progress to invasive cancer. We hypothesized that certain types of HPV would predict the progression from various stages of CIN to invasive cervical cancer. If this were true, the occurrence of specific types of HPV should be higher in CIN biopsy specimens from women in whom cervical cancer subsequently developed (cases) than in CIN specimens from women in whom invasive cancer did not develop (control subjects). A casecontrol study was conducted to measure the relative frequency of specific HPV DNA in CIN biopsy specimens from cancer patients and matched control subjects. Since geographic variation has been noted in the association of HPV DNA with cervical neoplasia5 and no information was available on the prevalence of different types of HPV in the women involved in the study, we also conducted a cross-sectional survey of women currently being evaluated for CIN.
interval of at least 2 years between the diagnosis of CIN and invasive cancer was used as an eligibility criterion, since we reasoned that early invasive disease incorrectly diagnosed as CIN would become clinically apparent before this time. Age between 15 and 75 years, inclusion in the CCABC registry and the availability of a preinvasive CIN biopsy specimen for HPV DNA analysis were other eligibility criteria. To select the control group we determined the occurrence of HPV DNA in CIN biopsy specimens from women in whom invasive cervical cancer did not subsequently develop; thus, eligibility included no recorded history of cervical cancer. For a deceased woman to be a control subject she must have been free of cervical cancer and survived for the period between the diagnosis of CIN and of cervical cancer in her matched case. Control subjects had to be between 15 and 75 years of age, have the same CIN stage diagnosed within 6 months of the diagnosis for the matched case, be followed up for at least as long as the matched case and be included in the CCABC registry. CIN biopsy material for HPV DNA analysis must also have been available. Women were excluded if they had undergone partial or complete hysterectomy. A search of the CCABC records of women treated for invasive cervical cancer between 1960 and 1986 revealed that 108 had a recorded diagnosis of CIN 2 or more years before the diagnosis of invasive cancer. Of the 108 records 54 stated that no tissue blocks were available; in 4 the tissue blocks Methods did not contain dysplastic lesions, and in 1 the CIN had been diagnosed in another province. The diagSubjects nosis of invasive cancer could not be confirmed We enrolled subjects who had been evaluated histologically in the review of biopsy specimens from for cervical disease at the Cancer Control Agency of 7 patients. This left 47 patients who fulfilled the British Columbia (CCABC), which offers a free eligibility criteria. Two control subjects were then province-wide cytology service to hospitals, clinics selected for each case. When possible, data were collected on parity, and private practitioners. Follow-up of patients with marital status and number of marriages. The date of cervical neoplasia is virtually complete: written requests are sent to physicians for repeat smears diagnosis, the histologic type and the severity of the annually,6 reports are obtained on all women attend- initial CIN lesion, the date and type of treatment ing colposcopy treatment clinics, which are integrat- administered and information on the dates and ed into the cytology program, and copies of all outcome of follow-up gynecologic examinations were pathology reports mentioning a cancer diagnosis in also recorded. In addition, the date of diagnosis and British Columbia are routinely forwarded to the the histologic type of the invasive cancer, as well as CCABC. The composition of the population served the treatment method and outcome, were noted for by this agency, determined from information provid- the cases. Most information was obtained from the ed by the Provincial Division of Vital Statistics, has computerized records of patients registered with the been detailed previously.7 CCABC; other information was acquired by contacting the family physician by telephone or mail.
Case -control study
Cross-sectional survey Women with invasive cancer whose CIN had been diagnosed before the invasive disease was identified were assigned to the case group. An 312
CAN MED ASSOC J 1990; 142 (4)
Biopsy specimens were required for identification in each category of severity of lesions (mild,
moderate and severe dysplasia, as well as carcinoma in situ) in patients attending the colposcopy clinic. At least 40 specimens per histologic grade were estimated to be needed to detect a 10% difference in the respective prevalence rates of HPV. Preliminary analysis revealed similar occurrences of HPV in patients with severe dysplasia and carcinoma in situ; thus, these two groups were pooled and the results presented according to the standard CIN classification. Biopsy specimens were obtained from consecutive patients with recently diagnosed CIN attending the clinic in 1987 over a 3-week period, during which the required numbers of specimens for each severity level were acquired. No attempt was made to select specimens on the basis of koilocytosis or other selection criteria. For control purposes 42 specimens were obtained from women undergoing evaluation for reasons other than cervical neoplasia. The specimens of histologically normal cervices were derived from negative cone biopsy samples, routine histologic biopsy specimens or tissue samples obtained at hysterectomy. Histologic examination of these specimens revealed that 20 patients had normal squamous-columnar cell junctions or unremarkable cervices, 14 had cervicitis, 5 had metaplasia, and 3 had atypia.
Detection of HPV DNA Sections cut from tissue blocks were initially examined to ensure the presence of CIN. Adjacent sections were mounted on slides, deparaffinized, hydrated and then denatured in 0.2 N hydrochloric acid. The presence of HPV DNA was determined with the use of tissue in-situ hybridization.8 Hybridization was carried out in 20% formamide (42°C below the melting temperature of the DNA hybrid) with subsequent washing under stringent conditions (20°C below the melting temperature of the DNA hybrid). The slides were autoradiographed for 3 weeks. Each batch included known positive and
negative sections. Three readers, blinded to the casecontrol status of the women, scored the slides using predetermined criteria; only those samples showing typical grains in the epithelial cells were considered positive. The prevalence of HPV types associated with the lesions in the cross-sectional survey was determined using individual HPV 16, 18 and 33 DNA probes as well as a mixture of HPV 6 and 11 DNA probes. Specimens from the case and control subjects were hybridized with mixtures of HPV 16/33 and 6/11 DNA as well as with HPV 18 DNA.
Statistical analysis All statistical analyses and data handling were done using SAS software (SAS Institute Inc., Carey, NC). The association of HPV with cervical neoplasia or other factors was characterized by contingency table analysis and the Pearson or Mantel-Haenszel chi-squared tests for independence.9 In addition, the log-linear model was used to test whether measured variables other than HPV were also associated with cervical neoplasia.
Results Case -control study Table 1 summarizes selected attributes of the 47 case subjects in whom invasive disease developed 2 or more years after the diagnosis of CIN and the 94 control subjects with CIN in whom invasive cancer
did not develop. The distribution according to the severity of CIN was similar in the two groups; this was expected since the grade of CIN was used as a matching criterion. Age at the time CIN was diagnosed and the prevalence of cervical conization, cautery or cryosurgery were also similar. There were no significant differences between the two groups in the frequencies of detection of specific types of HPV DNA in the CIN biopsy specimens (Table 2).
Table 1: Selected attributes of women with cervical intraepithelial neoplasia (CIN) who either had (cases) or did not have (matched controls) invasive cervical cancer
Attribute Grade of CIN, no. (and %) of subjects 1
II III Mean age (and standard deviation [SD] ) at diagnosis, yr Conization, no. (and %) of subjects Cautery or cryosurgery, no. (and %) of subjects
Case
Control
group
group
8 (17) 9 (19) 30 (64)
19 (20) 10 (10) 65 (69)
0.20 0.39 0.27
38.5 (13) 28 (60)
36.1 (11) 60 (64)
0.29 0.75
2 (4)
11 (12)
0.23
p value
CAN MED ASSOCJ 1990; 142(4)
313
-.M
Cross-sectional survey
the two groups the prevalence of HPV showed a similar trend of increasing rates with increasing severity of CIN; HPV 16 and 33 were the most prevalent types detected. However, the overall positivity rates for HPV were considerably lower among the case-control subjects than among those in the cross-sectional survey. The ages at diagnosis were lower for women in the cross-sectional survey than for those in the case-control study, and the differences were statistically significant in the CIN II (p = 0.018) and the CIN III (p < 0.001) groups. There was about a 10-year difference between the mean year of diagnosis of CIN I and CIN II in the two groups, whereas 5 years separated the mean year of diagnosis of CIN III. These observations suggest that the women represented in the two samples might not be comparable regarding HPV exposure. We also compared the rates of detection of HPV among women with CIN diagnosed in the remote versus the recent past (Table 4). The positivity rate among the biopsy specimens collected between 1961 and 1981 (3%) was lower than the rate among the specimens collected between 1982 and 1984 (38%), even though they were assayed in the same batch. The positivity rate among the specimens collected between 1982 and 1984 in the case-control study was not significantly different from the rate for 1987 in the cross-sectional survey (38% v. 46%), whereas the rate among specimens collected in 1961-81 was
The positivity rates of HPV in the CIN specimens from the case and control subjects were lower than the rates reported by others.'0"-2 However, this was not true for the women examined in the crosssectional survey. The prevalence rates of HPV DNA were 11% in specimens without CIN, 27% in those with CIN I, 49% in those with CIN II and 56% in those with CIN III. The rates of HPV 16 and HPV 33 DNA steadily increased from 10% in specimens from normal cervices to 54% in CIN III specimens (Table 3). However, the prevalence rates of both HPV 6/11 and 18 in all disease categories were similar to those observed in biopsy specimens of normal cervices. Statistical analysis revealed no association between the age of the patients and the positivity rates; thus, the trends for HPV 16 and HPV 33 DNA appeared to reflect differing rates of virus expression in lesions of differing severity.
Comparison of case-control and cross-sectional survey subjects To find possible explanations for the difference in the positivity rates of HPV in the CIN specimens between women in the cross-sectional survey and those in the case-control study we compared the age at diagnosis and the year of diagnosis (Table 3). In -.
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Human papillomavirus and cervical intraepithelial neoplasia in women who subsequently had invasive cancer Deoraj Caussy, PhD; Loraine D. Marrett, PhD; Ann J. Worth, MD; Mary McBride, MSc; William E. Rawls, MD In a retrospective case-control study biopsy specimens of cervical intraepithelial neoplasia (CIN) lesions from 47 women in whom invasive cancer subsequently developed (cases) and from 94 control subjects in whom CIN was diagnosed within 6 months of the diagnosis for the matched case subject but invasive disease did not develop were tested for human papillomavirus (HPV) DNA with tissue in-situ hybridization. There were no significant differences in the frequency of detection of HPV DNA between the two groups. In a cross-sectional survey the prevalence of HPV DNA was found to be 11% in specimens without CIN, 27% in those with CIN I, 49% in those with CIN II and 56% in those with CIN III. The positivity rates for HPV 16/33 DNA increased with the severity of CIN, but this was not observed for HPV 6/11 and 18 DNA. A comparison of the results of the case-control and cross-sectional studies suggested that the younger cohort of women had higher prevalence rates of HPV DNA than the older cohort.
Etude retrospective et comparative des lesions de neoplasie intra-epitheliale du col uterin (NICU) chez 47 femmes (constituant le groupe des sujets) ayant montre par la suite un cancer envahissant et chez 94 temoins oui le diagnostic de NICU avait e pose dans les 6 mois precedent ou suivant le moment ou il avait e pose chez le sujet apparie mais qui ne devaient pas montrer de cancer envahissant. Dans les biopsies on recherche les ADN du papillomavirus humain (PVH) par hybridation tissulaire in situ. La frequence de detection de ces ADN ne differe pas significativement d'un groupe a l'autre. Une etude transversale montre la presence d'ADN-PVH dans 11% des echantillons sans NICU, 27% en cas de NICU I, 49% en cas de NICU II et 56% en cas de NICU III. II existe un rapport direct entre la frequence de positivite de la recherche de l'ADN-PVH 16/33 et la severite de la NICU, ce qui ne s'observe pas pour les ADN-PVH 6/11 et 18. Si on met en regard les resultats de l'etude comparative et ceux de l'etude transversale on peut etre amend a croire que les femmes jeunes ont un plus haut taux de presence d'ADN-PVH que leurs ainees. C
ervical squamous cell carcinoma is generally believed to evolve from various grades of cervical intraepithelial neoplasia (CIN).' The incidence of CIN far exceeds that of invasive cancer; this suggests that only a fraction of the preinvasive lesions progress to cervical cancer.2 Little is known
about the factors contributing to the ultimate invasive potential of a particular CIN lesion, but the human papillomaviruses (HPV) have been implicated in cervical cancer;3 the evidence is based mainly on biologic and clinical observations. HPV 16 DNA can induce cellular transformation in vitro and thus
From the Department of Pathology, McMaster University, Hamilton, Ont., the Ontario Cancer Treatment and Research Foundation, Toronto, the Cancer Control Agency of British Columbia, Vancouver, and the Viral Epidemiology Section, National Cancer Institute, National Institutes ofHealth, Bethesda, Md.
Reprint requests to: Dr. William E. Rawls, Department ofPathology, McMaster University, 1200 Main St. W, Hamilton, Ont. L8N 3Z5 -
For prescribing information
see
page 403
CAN MED ASSOC J 1990; 142 (4)
311
has potential oncogenic properties.4 The preferential occurrence of HPV 16 DNA, and to a lesser extent HPV 18 and HPV 33 DNA, in invasive cervical cancers provides the main clinical evidence, but the limitations of the research designs allow two equally plausible but contrasting interpretations.5 First, these viruses may occur secondary to the cancer state as opportunistic pathogens. Second, these viruses could be involved with a part of the neoplastic process that culminates in invasive cancer, in which case the presence of HPV DNA would likely be predictive of CIN lesions that will progress to invasive cancer. We hypothesized that certain types of HPV would predict the progression from various stages of CIN to invasive cervical cancer. If this were true, the occurrence of specific types of HPV should be higher in CIN biopsy specimens from women in whom cervical cancer subsequently developed (cases) than in CIN specimens from women in whom invasive cancer did not develop (control subjects). A casecontrol study was conducted to measure the relative frequency of specific HPV DNA in CIN biopsy specimens from cancer patients and matched control subjects. Since geographic variation has been noted in the association of HPV DNA with cervical neoplasia5 and no information was available on the prevalence of different types of HPV in the women involved in the study, we also conducted a cross-sectional survey of women currently being evaluated for CIN.
interval of at least 2 years between the diagnosis of CIN and invasive cancer was used as an eligibility criterion, since we reasoned that early invasive disease incorrectly diagnosed as CIN would become clinically apparent before this time. Age between 15 and 75 years, inclusion in the CCABC registry and the availability of a preinvasive CIN biopsy specimen for HPV DNA analysis were other eligibility criteria. To select the control group we determined the occurrence of HPV DNA in CIN biopsy specimens from women in whom invasive cervical cancer did not subsequently develop; thus, eligibility included no recorded history of cervical cancer. For a deceased woman to be a control subject she must have been free of cervical cancer and survived for the period between the diagnosis of CIN and of cervical cancer in her matched case. Control subjects had to be between 15 and 75 years of age, have the same CIN stage diagnosed within 6 months of the diagnosis for the matched case, be followed up for at least as long as the matched case and be included in the CCABC registry. CIN biopsy material for HPV DNA analysis must also have been available. Women were excluded if they had undergone partial or complete hysterectomy. A search of the CCABC records of women treated for invasive cervical cancer between 1960 and 1986 revealed that 108 had a recorded diagnosis of CIN 2 or more years before the diagnosis of invasive cancer. Of the 108 records 54 stated that no tissue blocks were available; in 4 the tissue blocks Methods did not contain dysplastic lesions, and in 1 the CIN had been diagnosed in another province. The diagSubjects nosis of invasive cancer could not be confirmed We enrolled subjects who had been evaluated histologically in the review of biopsy specimens from for cervical disease at the Cancer Control Agency of 7 patients. This left 47 patients who fulfilled the British Columbia (CCABC), which offers a free eligibility criteria. Two control subjects were then province-wide cytology service to hospitals, clinics selected for each case. When possible, data were collected on parity, and private practitioners. Follow-up of patients with marital status and number of marriages. The date of cervical neoplasia is virtually complete: written requests are sent to physicians for repeat smears diagnosis, the histologic type and the severity of the annually,6 reports are obtained on all women attend- initial CIN lesion, the date and type of treatment ing colposcopy treatment clinics, which are integrat- administered and information on the dates and ed into the cytology program, and copies of all outcome of follow-up gynecologic examinations were pathology reports mentioning a cancer diagnosis in also recorded. In addition, the date of diagnosis and British Columbia are routinely forwarded to the the histologic type of the invasive cancer, as well as CCABC. The composition of the population served the treatment method and outcome, were noted for by this agency, determined from information provid- the cases. Most information was obtained from the ed by the Provincial Division of Vital Statistics, has computerized records of patients registered with the been detailed previously.7 CCABC; other information was acquired by contacting the family physician by telephone or mail.
Case -control study
Cross-sectional survey Women with invasive cancer whose CIN had been diagnosed before the invasive disease was identified were assigned to the case group. An 312
CAN MED ASSOC J 1990; 142 (4)
Biopsy specimens were required for identification in each category of severity of lesions (mild,
moderate and severe dysplasia, as well as carcinoma in situ) in patients attending the colposcopy clinic. At least 40 specimens per histologic grade were estimated to be needed to detect a 10% difference in the respective prevalence rates of HPV. Preliminary analysis revealed similar occurrences of HPV in patients with severe dysplasia and carcinoma in situ; thus, these two groups were pooled and the results presented according to the standard CIN classification. Biopsy specimens were obtained from consecutive patients with recently diagnosed CIN attending the clinic in 1987 over a 3-week period, during which the required numbers of specimens for each severity level were acquired. No attempt was made to select specimens on the basis of koilocytosis or other selection criteria. For control purposes 42 specimens were obtained from women undergoing evaluation for reasons other than cervical neoplasia. The specimens of histologically normal cervices were derived from negative cone biopsy samples, routine histologic biopsy specimens or tissue samples obtained at hysterectomy. Histologic examination of these specimens revealed that 20 patients had normal squamous-columnar cell junctions or unremarkable cervices, 14 had cervicitis, 5 had metaplasia, and 3 had atypia.
Detection of HPV DNA Sections cut from tissue blocks were initially examined to ensure the presence of CIN. Adjacent sections were mounted on slides, deparaffinized, hydrated and then denatured in 0.2 N hydrochloric acid. The presence of HPV DNA was determined with the use of tissue in-situ hybridization.8 Hybridization was carried out in 20% formamide (42°C below the melting temperature of the DNA hybrid) with subsequent washing under stringent conditions (20°C below the melting temperature of the DNA hybrid). The slides were autoradiographed for 3 weeks. Each batch included known positive and
negative sections. Three readers, blinded to the casecontrol status of the women, scored the slides using predetermined criteria; only those samples showing typical grains in the epithelial cells were considered positive. The prevalence of HPV types associated with the lesions in the cross-sectional survey was determined using individual HPV 16, 18 and 33 DNA probes as well as a mixture of HPV 6 and 11 DNA probes. Specimens from the case and control subjects were hybridized with mixtures of HPV 16/33 and 6/11 DNA as well as with HPV 18 DNA.
Statistical analysis All statistical analyses and data handling were done using SAS software (SAS Institute Inc., Carey, NC). The association of HPV with cervical neoplasia or other factors was characterized by contingency table analysis and the Pearson or Mantel-Haenszel chi-squared tests for independence.9 In addition, the log-linear model was used to test whether measured variables other than HPV were also associated with cervical neoplasia.
Results Case -control study Table 1 summarizes selected attributes of the 47 case subjects in whom invasive disease developed 2 or more years after the diagnosis of CIN and the 94 control subjects with CIN in whom invasive cancer
did not develop. The distribution according to the severity of CIN was similar in the two groups; this was expected since the grade of CIN was used as a matching criterion. Age at the time CIN was diagnosed and the prevalence of cervical conization, cautery or cryosurgery were also similar. There were no significant differences between the two groups in the frequencies of detection of specific types of HPV DNA in the CIN biopsy specimens (Table 2).
Table 1: Selected attributes of women with cervical intraepithelial neoplasia (CIN) who either had (cases) or did not have (matched controls) invasive cervical cancer
Attribute Grade of CIN, no. (and %) of subjects 1
II III Mean age (and standard deviation [SD] ) at diagnosis, yr Conization, no. (and %) of subjects Cautery or cryosurgery, no. (and %) of subjects
Case
Control
group
group
8 (17) 9 (19) 30 (64)
19 (20) 10 (10) 65 (69)
0.20 0.39 0.27
38.5 (13) 28 (60)
36.1 (11) 60 (64)
0.29 0.75
2 (4)
11 (12)
0.23
p value
CAN MED ASSOCJ 1990; 142(4)
313
-.M
Cross-sectional survey
the two groups the prevalence of HPV showed a similar trend of increasing rates with increasing severity of CIN; HPV 16 and 33 were the most prevalent types detected. However, the overall positivity rates for HPV were considerably lower among the case-control subjects than among those in the cross-sectional survey. The ages at diagnosis were lower for women in the cross-sectional survey than for those in the case-control study, and the differences were statistically significant in the CIN II (p = 0.018) and the CIN III (p < 0.001) groups. There was about a 10-year difference between the mean year of diagnosis of CIN I and CIN II in the two groups, whereas 5 years separated the mean year of diagnosis of CIN III. These observations suggest that the women represented in the two samples might not be comparable regarding HPV exposure. We also compared the rates of detection of HPV among women with CIN diagnosed in the remote versus the recent past (Table 4). The positivity rate among the biopsy specimens collected between 1961 and 1981 (3%) was lower than the rate among the specimens collected between 1982 and 1984 (38%), even though they were assayed in the same batch. The positivity rate among the specimens collected between 1982 and 1984 in the case-control study was not significantly different from the rate for 1987 in the cross-sectional survey (38% v. 46%), whereas the rate among specimens collected in 1961-81 was
The positivity rates of HPV in the CIN specimens from the case and control subjects were lower than the rates reported by others.'0"-2 However, this was not true for the women examined in the crosssectional survey. The prevalence rates of HPV DNA were 11% in specimens without CIN, 27% in those with CIN I, 49% in those with CIN II and 56% in those with CIN III. The rates of HPV 16 and HPV 33 DNA steadily increased from 10% in specimens from normal cervices to 54% in CIN III specimens (Table 3). However, the prevalence rates of both HPV 6/11 and 18 in all disease categories were similar to those observed in biopsy specimens of normal cervices. Statistical analysis revealed no association between the age of the patients and the positivity rates; thus, the trends for HPV 16 and HPV 33 DNA appeared to reflect differing rates of virus expression in lesions of differing severity.
Comparison of case-control and cross-sectional survey subjects To find possible explanations for the difference in the positivity rates of HPV in the CIN specimens between women in the cross-sectional survey and those in the case-control study we compared the age at diagnosis and the year of diagnosis (Table 3). In -.
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