A novel set of polymerase chain reaction (PCR) primers , designated SPF1 and SPF2 and located in the L1 region , was developed for universal detection of human papillomavirus (HPV). A short PCR fragment (SPF) of only 65 pb was synthesized. SPF amplimers were detected in a microtiter-based hybridization system , using a mixture of oligonucleotide probes. The SPF system allowed detection of at least 43 different HPV genotypes. The clinical performance of the novel SPF system was assessed in three different patient groups. 1) Analysis of 534 cervical scrapes , obtained from treated patients , showed that the detection rate in 447 (83.7%) scrapes with normal cytology was significantly higher using the SPF system as compared with the universal primer set GP5؉/6؉ (P < 0.001). 2) The SPF assay detected HPV DNA in 299 (98.4%) of 304 scrapes with cytological dyskaryosis. 3) The SPF system detected HPV DNA in 100% of 184 formalin-fixed, paraffin-embedded cervical carcinoma specimens. In conclusion , the novel SPF system permitted universal and highly sensitive detection of HPV DNA in diverse clinical materials and may improve the molecular diagnosis and epidemiology of this important virus. (Am J Pathol 1998, 153:1731-1739)Cervical cancer is the second most important cause of cancer-related mortality in women after breast cancer, 1 annually affecting an estimated 500,000 women worldwide. 2 Smoking, age of first sexual contact, number of sexual partners, and immunological status have been identified as significant risk factors for cervical carcinoma. [3][4][5][6] Infection with human papillomavirus (HPV) also is considered to be an important risk factor. 7,8 HPV is an epitheliotropic virus and contains a circular doublestranded DNA genome of approximately 7900 bp. To date, at least 77 distinct HPV genotypes have been described that can be classified into a cutaneous and a mucosal group, as established by their associated clinical manifestations. 8 Among the mucosal genotypes that infect the anogenital tract, potentially high-risk HPV genotypes 16,18,31,33,35,39, 45, 51, 52, 56, 58, 66, and 69 have been defined based on their prevalence in cervical intraepithelial neoplasia (CIN) lesions and cervical carcinomas. 8 The HPV genotypes 6,11,34,40, 42, 43, 44, 53, 54, 55, 59, 61, 62, 67, 68, 71, and 74 are classified as putative low-risk factors. By definition, infection with high-risk HPV genotypes favors an increased risk for cancer. 9 -11 However, the distinction between high-risk and low-risk genotypes is a moot point as low-risk HPV genotypes have also been found in carcinomas. 9 As HPV cannot be cultured in vitro and serological assays are still ineffective, diagnosis of HPV injury is based on the use of molecular tools. Direct dot-spot detection and in situ hybridization assays have been described, 12,13 but these methods are tedious and appear to lack sensitivity and specificity. DNA amplification methods, such as the polymerase chain reaction (PCR), permit more sensitive detection of the viral DNA. Besides type-specif...

Human papillomavirus (HPV) can be detected by amplification of viral DNA. A novel PCR primer set generating a short PCR fragment (SPF PCR) was used for amplification of a fragment of only 65 bp from the L1 region and permitted ultrasensitive detection of a broad spectrum of HPV genotypes. The intra- and intertypic sequence variations of the 22-bp interprimer region of this amplimer were studied. Among 238 HPV sequences from GenBank and clinical specimens, HPV genotypes were correctly identified based on the 22-bp sequence in 232 cases (97.2%). Genotype-specific probes for HPV genotypes 6, 11, 16, 18, 31, 33 to 35, 39, 40, 42 to 45, 51 to 54, 56, 58, 59, 66, 68, 70, and 74 were selected, and a reverse hybridization line probe assay (LiPA) (the INNO-LiPA HPV prototype research assay) was developed. This LiPA permits the use of amplimers generated by the SPF as well as the MY 09/11 primers. The assay was evaluated with a total of 1,354 clinical specimens, comprising cervical scrapes (classifications ranging from normal cytology to severe dyskaryosis) and formalin-fixed, paraffin-embedded cervical carcinoma samples. LiPA results were highly concordant with sequence analysis of the SPF amplimer, genotype-specific PCR, and sequence analysis of amplimers generated by MY 09/11 primers. The sensitivity of the SPF primers was higher than that of the GP5+/6+ primers over a broad range of HPV types, especially when multiple HPV genotypes were present. In conclusion, the SPF LiPA method allows extremely sensitive detection of HPV DNA as well as reliable identification of HPV genotypes in both cervical smears and paraffin-embedded materials.

Imiquimod is effective in the treatment of vulvar intraepithelial neoplasia. (Current Controlled Trials number, ISRCTN11290871 [controlled-trials.com].).

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BACKGROUND:In the management of early stage cervical cancer, knowledge about the prognosis is critical. Although many factors have an impact on survival, their relative importance remains controversial. This study aims to develop a prognostic model for survival in early stage cervical cancer patients and to reconsider grounds for adjuvant treatment. METHODS: A multivariate Cox regression model was used to identify the prognostic weight of clinical and histological factors for disease-specific survival (DSS) in 710 consecutive patients who had surgery for early stage cervical cancer (FIGO [International Federation of Gynecology and Obstetrics] stage IA2-IIA). Prognostic scores were derived by converting the regression coefficients for each prognostic marker and used in a score chart. The discriminative capacity was expressed as the area under the curve (AUC) of the receiver operating characteristic. RESULTS:The 5-year DSS was 92%. Tumor diameter, histological type, lymph node metastasis, depth of stromal invasion, lymph vascular space invasion, and parametrial extension were independently associated with DSS and were included in a Cox regression model. This prognostic model, corrected for the 9% overfit shown by internal validation, showed a fair discriminative capacity (AUC, 0.73). The derived score chart predicting 5-year DSS showed a good discriminative capacity (AUC, 0.85). CONCLUSIONS: In patients with early stage cervical cancer, DSS can be predicted with a statistical model. Models, such as that presented here, should be used in clinical trials on the effects of adjuvant treatments in high-risk early cervical cancer patients, both to stratify and to include patients. Cancer 2011;117:768-76.

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We have compared the efficacies of three general primer pairs for the detection of human papillomavirus (HPV) DNA in formaldehyde-fixed paraffin-embedded carcinomas. The use of these primer pairs leads to underestimates of the HPV prevalence (GP5/6, 61.1%; CPI/IIG, 57.4%; MY09/11, 46.9%; combined, 72.8%). The efficacy of each primer pair seemed to be inversely correlated to the length of the amplimer produced. By using newly developed type-specific primer pairs (amplimer length, approximately 100 bp), an increase in HPV DNA detection (87.6%) was found.