Interlaboratory reproducibility of liquid-based equivocal cervical cytology within a randomized controlled trial framework.

Within a multicentre controlled trial framework, an external quality control (EQC) was scheduled to evaluate the interlaboratory reproducibility of liquid-based cytology. In particular, this EQC intended to evaluate the reproducibility of the ASCUS diagnosis.A selected set of 30 slides (4 within normal limit cases, 16 atypical squamous cells of undetermined significance; 4 low-grade squamous intraepithelial lesions and 6 high-grade squamous intraepithelial lesions) circulated among the 13 laboratories involved in the trial.Kappa values were obtained from the comparison between individual laboratory diagnoses and majority diagnoses with target diagnoses. Specific kappa values resulted moderate to high for HSIL and low to moderate for LSIL and WNL. Meanwhile, the specific kappa for ASCUS was below 0.4 in 12 of 13 participating laboratories. The lack of reproducibility for ASCUS was not a result of the introduction of this new technology but rather to the low reproducibility of the ASCUS category itself stemming from intrinsic uncertainties in the reporting criteria.

Interlaboratory reproducibility of atypical glandular cells of undetermined significance: a national survey.

OBJECTIVE: The aim of this study was to evaluate the inter-laboratory reproducibility for atypical glandular cells (AGC) (The Bethesda System (TBS) 2001) of the laboratories involved in the screening programmes in Italy. METHODS: A set of 35 selected slides were circulated among 167 laboratories involved in local population-based cervical screening programmes. Each laboratory provided one single diagnosis per smear. The smears were read blind to the original diagnosis and to the diagnoses provided by other laboratories. A 'majority' diagnosis was defined for each case and assumed as the reference standard. The diagnosis provided from each laboratory was compared with the majority diagnosis. RESULTS: According to the majority report the 35 slides in the set were classified as negative in nine cases, AGC in eight, adenocarcinoma in eight, and squamous lesion or squamous + glandular lesion in 10. The crude agreement between all pairs of laboratories was 49.43%. K-values were 0.46, 0.21, 0.34, 0.36 and 0.32 for negative, AGC/AIS (adenocarcinoma in situ of endocervix), AdenoCa, Sq/Sq + Gl and all reporting categories respectively. Concordance according to overall K was moderate to substantial in 77% of the participating laboratories. CONCLUSIONS: The present study shows that the AGC category is not easily reproducible. The data confirmed the importance, in a screening scenario, of AGC/AIS diagnoses, but also presented difficulties in differentiating between the two diagnoses. In addition to the results obtained from the circulation of the slides, laboratories which had annually a low number of cervical smears were able to gain experience focused on particular morphological pictures.

Impact of the introduction of organised screening for cervical cancer in Turin, Italy: cancer incidence by screening history 1992-98.

After an organised cervical screening programme was introduced in Turin in 1992, the age-adjusted cervical cancer incidence ratio in 1992-98 was 0.81 (95% confidence interval (CI) 0.59-1.09) for invited vs not invited women and 0.25 (95% CI 0.13-0.50) for attenders vs non attenders. An organised screening programme can further reduce cervical cancer incidence in an area where substantial spontaneous activity was previously present.

Interlaboratory reproducibility of atypical squamous cells of undetermined significance report: a national survey.

The study was aimed at assessing interlaboratory reproducibility in the reporting of cervical smears in the atypical squamous cells of undetermined significance (ASCUS) category. A set of 50 selected slides circulated among 89 laboratories, currently involved in population-based screening programmes for cervical cancer, which provided a diagnostic report according to four main reporting categories based on the 1991 Bethesda system. Interlaboratory agreement was determined according to kappa (K) statistics: overall and weighted K values were determined for each laboratory and for single reporting categories. The results showed a very low reproducibility for the ASCUS category. This finding supports the Bethesda system 1991 recommendation to limit the use of this reporting category and suggests that the clinical response to ASCUS reports should be decided locally, based on the observed positive predictive value for cervical intraepithelial neoplasia 2 or more severe lesions.

Quantitative expression of the human kallikrein gene 9 (KLK9) in ovarian cancer: a new independent and favorable prognostic marker.

Many members of the human kallikrein gene family were found to be differentially expressed in various malignancies and some are useful cancer diagnostic/prognostic markers. KLK9 is a newly discovered human kallikrein gene that is expressed in several tissues including thymus, testis, spinal cord, salivary gland, ovary, and skin. Like other kallikreins, the KLK9 gene was found to be regulated by steroid hormones in cancer cell lines. Our purpose is to examine whether quantitative analysis of KLK9 expression has prognostic value in ovarian cancer. We studied the expression of KLK9 by quantitative reverse transcription-PCR in 168 consecutive ovarian tumors of different stages, grades, and histological types, and correlated the expression with clinicopathological parameters, response to chemotherapy, and patients' survival. We found that KLK9 expression was significantly higher in patients with early disease stages (I or II; P = 0.044) and in patients with optimal debulking (P = 0.019). Kaplan-Meier survival curves demonstrated that patients with KLK9-positive tumors have substantially longer progression-free and overall survival (P < 0.001 and P = 0.016, respectively). When the Cox proportional hazard regression analysis was applied to subgroups of patients, KLK9 expression was found to be a significant predictor of progression-free survival in the subgroup of patients with low-grade tumors [hazard ratio (HR), 0.13; P = 0.0015], early stage (HR, 0.099; P = 0.031); and those with optimal debulking (HR, 0.26; P = 0.012). After adjusting for other known prognostic variables, KLK9 retained its independent prognostic value in all of these subgroups of patients. A negative correlation was found between the expression levels of CA125 and KLK9 (rs, 0.350; P = 0.002). Our results indicate that KLK9 is under steroid hormone regulation in ovarian and breast cancer cell lines. Immmunohistochemically, human kallikrein protein (hK9) was localized in the cytoplasm, but not in the nuclei, of the epithelial cells of ovarian cancer tissues. We conclude that KLK9 is a potential new independent favorable prognostic marker for early stage, low-grade, optimally debulked ovarian cancer patients.

[Vulvar Paget's disease. Clinico-pathologic review of the literature]. / La malattia di Paget vulvare. Revisione clinico-patologica della letteratura.

In 1986 the International Society For the Study of Vulvar Disease classified vulvar Paget's disease (VPD) as a non-squamous intraepithelial lesion of the vulva. The clinical multiform aspect of VPD, similar to other dermatological lesions, often delays the execution of a biopsy. Paget's cells could be instead easily identified at histological examination and with histochemical reactions. Underlying adenocarcinomas or stromal invasion are present in about 10% of intraepithelial VPD. Patients with VPD are at risk for a second synchronous or metachronous neoplasia: colo-rectal adenocarcinoma (more frequent in perianal localization of VPD), cervical adenocarcinoma, carcinoma of the transitional epithelium from the renal pelvis to urethra and mammary carcinoma. A wide spectrum of frequency of these associations is reported in the literature (0-45%). Therapy for intraepithelial VPD is wide and deep surgical resection comprising all the skin appendages. However VPD has a high frequency of recurrences (15-62%), often irrespective for radicality of surgical excision. When association with underlying invasive adenocarcinoma or stromal invasion is histologically confirmed, vulvar surgical approach must be integrated with inguino-femoral lymphadenectomy. The role of chemotherapy and radiotherapy in the multimodal approach to extensive or recurring VPD is still controversial. Recurrences or progression of intraepithelal VPD are reported more than 10 years from first surgical resection so that long term follow-up is mandatory.

Recurrent squamous cell carcinoma of the vulva: clinicopathologic determinants identifying low risk patients.

BACKGROUND: The identification of prognostic factors in the recurrence of vulvar squamous cell carcinoma is crucial for less invasive treatments. METHODS: The authors studied 101 patients treated for primary invasive squamous cell carcinoma of the vulva. Selected pathologic variables were observed in a standardized manner during treatment, and their association with disease free survival was investigated using the Cox model. Independent prognostic factors were selected by a stepwise procedure. The absolute survival of patient groups determined on the basis of such factors was computed by the product limit method. RESULTS: The median follow-up was 3.1 years (range, 56 days to 15.5 years). Recurrences developed in 33 patients. The independent recurrence predictors were as follows: International Federation of Gynecology and Obstetrics (FIGO) Stage IVA (vs. IB, II, or III) (risk ratio [RR]adjusted for other independent factors, 7.39), tumor multifocality (RR, 4.10), lymphovascular space involvement (LVSI) (RR, 2.96), the presence of associated vulvar intraepithelial neoplasia (VIN) Grade 2 or 3 (RR, 3.34), and the involvement of resection margins (RR, 4.88). By ignoring the FIGO stage and lymph node status, the independent predictors were then as follows: greatest tumor dimension < 2.5 cm, 2.5-4 cm (RR, 2.86), or > 4 cm (RR, 5.98); tumor multifocality (RR, 3.36); LVSI (RR, 4.19); the presence of VIN 2 or 3 (RR, 3.06); and the involvement of surgical margins (RR, 2.78). No recurrences were observed in 119 at-risk years among patients with unifocal tumors < 2.5 cm in greatest dimension, free surgical margins, no LVSI, and no associated VIN 2 or 3. CONCLUSIONS: The presence of associated VIN 2 or 3 was revealed to be a previously unidentified independent prognostic factor for recurrence. Subjects at low risk of recurrence could be identified even without consideration of lymph node status.

[The role of human papillomavirus in cyto-histological practice: distribution and prevalenceof hig-risk strains (16, 18, 31, 33, and 35) in intraepithelial lesions and neoplasia of the uterine cervix]. / Il ruolo del papillomavirus umano (HPV) nella pratica cito-istologica: distribuzione e prevalenza dei ceppi ad alto rischio (16, 18, 31, 33 e 35) nelle lesioni intraepiteliali e nelle neoplasie della cervice uterina.

INTRODUCTION: Many studies have already shown the association of persistent infection of human high risk papillomavirus (HPV) with the development of pre-invasive and invasive cervical disease. MATERIALS AND METHODS: We evaluated the use of high risk HPV testing in a study of about 1908 women, aged 29-78, who attending, from 1996 to 1998, the Sant'Anna Hospital in Turin for routine, second level smears and histopathological diagnosis. We considered all cervical lesions: ASCUS, LSIL, HSIL, squamous and adeno invasive cancers. HPV testing was performed by polymerase chain reaction (PCR) using L1 consensus primers which can detect almost all infections (high and low risk types). The most important high risk HPV types (16, 18, 31, 33 and 35) were tested using specific primers. RESULTS: The prevalence of high risk HPV was: ASCUS 42.2%, LSIL 39%, HSIL 73.5%, squamous invasive cancers 98.3% and adeno 100%. In addition HPV 16 is the most represented type in all lesions: ASCUS 40%, LSIL 62%, HSIL 71.2% squamous invasive cancers 73.3% and adeno 50.6%. In addition we study the mean age of cervical cancer onset compared with the different high risk HPV types. We found that HPV 18 related cancer occurs in younger women (mean age 41 years; range 39-42). CONCLUSIONS: The addition of high risk HPV testing to cytology may improve early identification of women at risk for cervical cancer.

Membranous hypertrophy of the posterior fourchette as a cause of dyspareunia and vulvodynia.

Twenty-one women were treated surgically for entry dyspareunia and vulvodynia. The ages of the patients ranged from 18 to 39 years (mean, 24.5). Physical examination showed the presence of membranous hypertrophy of the posterior fourchette with consequent stricture of the vaginal introitus in all the patients. Eighty percent of the patients had erythema and tenderness of the vestibule, particularly in the posterior part. The histologic findings were somewhat enigmatic and quite unimpressive, frequently suggestive of chronic nonspecific inflammation; in only two cases were histologic changes suggestive of human papillomavirus infection observed. All the patients underwent excision of the posterior part of the vestibule with vaginal advancement under general anesthesia. Follow-up showed elimination of the symptoms in 19 patients and an improvement in the symptoms in the remaining 2.

Peritumoral blood and lymphatic vessel invasion as a prognostic indicator in stage-I breast-carcinoma.

Between 1978 and 1986 we treated 318 consecutive patients with stage I breast cancer. Median follow-up time is 8.5 years. In all cases the invasion of peritumoral lymphatic (LVI) and blood vessels (BVI) was studied by the hematoxylin-eosin staining method. Different ten-year survival (86% vs 67%) and disease-free survival (82% vs 61%) probabilities were found in patients with or without LVI. BVI showed low sensitivity and specificity, therefore vessel invasion was reassessed on 190 specimens with the immunoperoxidase technique using the Ulex Europeus Type 1 lectine. A significant correlation between the vessel staining intensity and the presence of intraluminal metastases was found. With this method a better sensitivity was obtained, but it was associated with a loss of specificity and prognostic significance.

Tissue cea and tpa expression in stage-I breast-cancer.

Tumoral Carcino Embryonic Antigen (CEA) and Tissue Polypeptide Antigen (TPA) expression was investigated by immunoperoxidase technique on paraffin embedded tissues of 191 patients with infiltrating breast cancer and negative axillary nodes. TPA was almost always detected at high levels. CEA staining was evenly distributed, but high CEA positivity was associated with higher TPA expression. Lower levels of both markers were found in medullar carcinomas as well as in cancers larger than 1 cm. Higher estrogen and progesterone receptor content was associated with higher expression of tissue CEA and TPA. After a median follow-up of 9 years, neither tissue CEA nor tissue TPA show any prognostic value.

Biologic behavior of vulvar intraepithelial neoplasia. Histologic and clinical parameters.

The aim of this study was to evaluate the role by which different factors, such as human papillomavirus (HPV) infection, age, dystrophic alterations, focal nature and size of the lesion, influence the biologic behavior of vulvar intraepithelial neoplasia (VIN). Sixty-nine cases of VIN were investigated (28 VIN 1, 9 VIN 2, 32 VIN 3). Follow-up was possible in 58 cases, with a mean of 31 months; no treatment was given to 3 patients, while 55 were treated either medically or surgically. Eighty-four percent of the patients were cured, recurrences were found in 11%, and 5% of the patients showed progression of the disease to carcinoma. The ratio between medical and surgical treatment was the same among the cured, recurred and progressed groups of patients. No differences with regard to focal nature of the lesion, presence of HPV infection or dystrophic alterations were observed between the three groups of patients. Only the mean age was higher in patients who showed progression of the lesion to carcinoma.

Histologic parameters of vulvar invasive carcinoma and lymph node metastases.

We evaluated seven histologic parameters (tumor diameter, histologic grading, depth of stromal invasion, vascular invasion, pattern of invasion, lymphoplasmocytic infiltration and amount of necrosis) of 50 cases of vulvar invasive carcinoma to assess their correlation with groin lymph node metastases. Of 50 patients, 25 had groin lymph node metastases. No lymph node metastasis was found in four cases with depth of invasion < or = 2.0 mm. Among the 31 patients with vascular invasion, 23 (74%) had positive nodes, whereas lymph nodes were metastatic only in two of the 19 patients (10%) without vascular invasion. At univariate analysis, performed with Fisher's exact method, all the parameters considered, except pattern of invasion and amount of necrosis, were significantly associated (P < .05) with lymph node metastases. However, after adjustment by multiple logistic regression for the variables statistically significant at univariate level, only the presence of vascular invasion was significantly associated with nodal involvement and tumor diameter was borderline, whereas the effect of the other variables was almost completely explained by confounding.

In situ hybridization for human papillomavirus as a method of predicting the evolution of cervical intraepithelial neoplasia.

Twenty-four women with cervical condylomata which were immunohistochemically positive for human papillomavirus (PV-Ag) (15 with CIN 1 and 9 with CIN 2) were followed for a period of 2-65 months. Fifty-seven biopsies were studied by the in situ hybridization (ISH) procedure for the detection of HPV 6/11 and 16/18 DNA. ISH positivity was found in 13/24 cases (54.2%); HPV 16/18 was evident in 7/9 CIN 2 (77.8%) as against 3/15 CIN 1 (20%) (P = 0.017) and in 8/13 cases with koilocytosis affecting up to 2/3 of the epithelial thickness (61.5%) as against 2/11 cases with koilocytosis affecting more than 2/3 of the epithelial layer (18.2%) (P = 0.03). Progression to CIN 3 occurred in 4 cases (2 CIN 1 and 2 CIN 2), the degree of dysplasia remained static in 5 cases (1 CIN 1 and 4 CIN 2) and regression occurred in 15 cases (9 CIN 1 and 6 CIN 2). The immunoperoxidase (IP) positive staining for PV-Ag persisted in 5/24 cases and disappeared in 19/24; 6/13 ISH positive cases maintained ISH positive and 7/13 became negative. The progression of dysplasia was significantly related to disappearance of the IP positivity (P less than 0.0001), to the ISH positivity (P = 0.05), to the persistence of ISH positivity (P = 0.008) and to HPV 16/18 positivity (P = 0.01). We believe that ISH positivity for HPV 16/18 in CIN 1 or 2 with low degrees of koilocytosis and conversion from PV-Ag positive to negative indicate a high risk of progression to CIN 3.

Topographic distribution of groin lymph nodes. A study of 50 female cadavers.

There is a discrepancy between anatomy textbooks' description of groin node position and Way's technique of lymphadenectomy. On the one hand, anatomic studies have demonstrated that the deep femoral nodes are on the medial side of the femoral vein, lying on the deep portion of the fascia lata, and can be seen easily through the opening of the fossa ovalis. On the other hand, the standard technique of deep femoral lymphadenectomy consists of removing the fat lying lateral to the femoral artery through the incision and detachment of the fascia lata from the sartorius to adductor longus muscle. With the aim of demonstrating that a correct deep femoral lymphadenectomy does not require removal of the fascia lata, we dissected Scarpa's triangles in 50 female cadavers. The examination of 100 specimens demonstrated that the deep femoral nodes are always situated within the opening of the fossa ovalis, and no lymph nodes are distal to the lower margin of the fossa ovalis, under the fascia cribrosa. These findings suggest that deep femoral lymphadenectomy can be performed without removing the fascia lata.

Deep femoral lymphadenectomy with preservation of the fascia lata. Preliminary report on 42 invasive vulvar carcinomas.

Forty-two patients with primary invasive vulvar carcinoma were treated with radical vulvectomy and deep femoral lymphadenectomy with preservation of the fascia lata and cribriform fascia. The rationale for using this technique was based on anatomic knowledge of the topographic distribution of groin lymph nodes, which was confirmed by the study of 50 cadavers. The preliminary data show that the number of superficial and deep femoral lymph nodes removed from the 42 patients (mean number of nodes, 20; range, 8-32) was similar to the number reported in anatomy books. In addition, the five-year actuarial survival rate, 70%, was comparable to that in the literature. These preliminary results suggest that the surgical technique used in this study is as radical an oncologic procedure as Way's classic groin lymphadenectomy, which consists of removing the fascia lata and cribriform fascia.

Vulvar intraepithelial neoplasia. A clinicopathologic study of 60 cases.

Sixty cases of vulvar intraepithelial neoplasia (VIN) were analyzed clinicopathologically (24 VIN I, 9 VIN II, 27 VIN III). The ages of the patients ranged from 21 to 83 years (mean, 53.7). Colposcopic examinations showed the presence of white areas in 29 cases, red areas in 9, acetowhite areas in 6 and other alterations in 13. One-third of the lesions were multifocal. Pruritus and burning were present in 65% of the cases. Fifty-one percent of the cases showed histologic changes suggestive of human papillomavirus (HPV) infection; the mean age of those patients was significantly lower than that of patients without HPV infection. In 15 cases of VIN, HPV DNA testing was performed with Southern blot hybridization; in three (20%) of those specimens HPV 16 episomal DNA was identified. Epithelial alterations surrounding the areas of VIN were found in 24 cases (40%)-23 squamous cellular hyperplasias and 1 lichen sclerosus. Different types of treatment were performed according to the different grades of VIN: medical therapy, diathermocoagulation, local excision, hemivulvectomy and total vulvectomy. Follow-up was possible in 52 cases, with a mean of 33 months (range, 3-98). Two cases of VIN I showed progression of disease over 12-24 months.