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BACKGROUND: The objective of this study was to analyze the diagnostic performance of a newly established immunocytochemical dual-stain protocol, which simultaneously detects p16INK4a and Ki-67 expression in cervical cytology samples, for identifying high-grade cervical intraepithelial neoplasia (CIN2+) in women with Papanicolaou (Pap) cytology results categorized as atypical squamous cells of undetermined significance (ASCUS) or low-grade squamous intraepithelial lesions (LSIL). METHODS: Residual liquid-based cytology material from 776 retrospectively collected ASCUS/LSIL cases that were available from a recent study evaluating p16 cytology and HPV testing were subjected to p16/Ki-67 dual staining. The presence of 1 or more double-immunoreactive cell(s) was regarded as a positive test outcome, irrespective of morphology. Test results were correlated to histology follow-up. RESULTS: Sensitivity of p16/Ki-67 dual-stain cytology for biopsy-confirmed CIN2+ was 92.2% (ASCUS) and 94.2% (LSIL), while specificity rates were 80.6% (ASCUS) and 68.0% (LSIL), respectively. Similar sensitivity/specificity profiles were found for both age groups of women aged

We analyzed the performance of p16INK4a immunocytochemistry on a series of 810 retrospectively collected atypical squamous cells of undetermined significance (ASC-US) and low-grade squamous intraepithelial lesion (LSIL) cases with available biopsy follow-up data, including 94 cases of cervical intraepithelial neoplasia (CIN) 2 and 128 cases of CIN 3. Human papillomavirus (HPV) testing was performed from the same residual liquid-based cytologic specimen, and results for both tests were correlated with histologic follow-up data. Sensitivity values for high-grade CIN (HGCIN) confirmed on biopsy within 6 months were 92.6% (ASC-US) and 92.2% (LSIL) for cytotechnologists’ reviews of p16 cytology and 90.1% (ASC-US) and 95.7% (LSIL) for HPV testing. Sensitivity rates of initial pathologists’ reviews were slightly lower, 76.4% to 80.1%, with levels comparable to cytotechnologists’ results after adjudication. The specificity of p16 cytology for HGCIN detection was significantly higher than for HPV testing for cytotechnologists and pathologists: 63.2% to 71.1% (p16 cytology) vs 37.8% for HPV in ASC-US (P < .001) and 37.3% to 53.3% (p16 cytology) vs 18.5% for HPV in LSIL (P < .001). This evaluation of the diagnostic performance of p16 cytology confirms the potential of this stain for the efficient triage of ASC-US and LSIL cytologic results.

Peripheral T-cell lymphomas (PTCLs) are a heterogeneous group of aggressive non-Hodgkin lymphomas. Angioimmunoblastic T-cell lymphoma (AITL) is a common subtype of PTCL, and patients with AITL typically have a poor prognosis with limited treatment options. Clinical studies have demonstrated the activity of romidepsin, a structurally unique, potent, bicyclic class 1 selective histone deacetylase inhibitor, in patients with relapsed or refractory AITL. In the case presented herein, we describe a patient treated with single-agent romidepsin at first diagnosis of AITL, resulting in complete remission for over 2 years and leading to the use of maintenance dosing. The patient eventually underwent a successful autologous stem cell transplant. This case illustrates the successful use of romidepsin for the long-term treatment of a patient with AITL in a clinical setting. Maintenance dosing may be an option for patients who have an extended response to romidepsin in order to optimize outcomes and to prolong time to the next subsequent line of therapy. In our case, the patient was able to remain in complete remission for more than 1 year while receiving maintenance dosing of romidepsin.

To investigate the test performance of a commercially available detection kit for human papillomavirus (HPV), the relationship between the detection of HPV by dot filter hybridization (DFH) and by standard morphologic methods was studied. Four hundred two cervical samples taken from 381 patients referred to a colposcopy clinic were examined. Human papillomavirus DNA sequences were identified and typed using commercially available anti-sense RNA probes. Simultaneous cytologic smears were obtained in 289 patients, directed biopsy samples in 284, and both smears and biopsy samples in 171 samples. Human papillomavirus DNA was detected in 164 specimens (41%), of which 24 (15%) were type 6/11, 74 (45%) were type 16/18, 39 (24%) were type 31/33/35, and 27 (16%) were untyped due to the presence of multiple positive signals. Viral types 16/18 and 31/33/35 were eight and six times more frequent in cervical intraepithelial neoplasia (CIN) II/CIN III lesions than in condyloma/CIN I, respectively. When the cytologic diagnosis was considered the standard of reference, the results of DFH for the detection of HPV were concordant in 167 (56%) paired samples. The sensitivity of DFH was 48% and the specificity was 77%. The distribution of the morphologic diagnoses in the group of false-negative results and true-positive results was similar. When the histologic diagnosis was considered the standard of reference, the efficiency of DFH was 62%, the sensitivity was 59%, and the specificity was 79%. In the subgroup of 118 samples with simultaneous smear and biopsy and at least one positive examination, 42 (36%) were positive by all three methods, 42 (36%) by two, and 34 (29%) by one, including 6 (5%) by DFH alone. Fifteen cases more were detected by the complementary use of DFH and cytology than with cytology alone. The results demonstrated that the sets of patients positive for HPV when detected by DFH or by morphologic methods were not identical but rather overlapped. The detection of HPV may be slightly improved by using DFH in addition to conventional examinations. A significant number of HPV-positive patients without a morphologic lesion and patients with low-grade lesions had HPV 16/18 or 31/33/35, suggesting a possible role for typing in establishing a risk profile. However, given uncertainties in understanding the biology of HPV-associated lesions, the role, if any, of clinical testing for HPV by DFH remains to be defined.

The present investigation was undertaken to study the cellular localization and kinetics of synthesis of CA 125 in the endometrium. CA 125 was localized by immunohistochemistry to the infranuclear region of epithelial cells during the proliferative phase and to the apical luminal border during the secretory phase. In gestational endometrium, both the cytoplasm and the apical luminal border of epithelial cells were intensely positive. No staining was seen in endometrial stromal cells during the normal cycle or in decidualized endometria. Results obtained from in vitro cultures of separated glandular and stromal cells were similar to those obtained by immunohistochemistry. That is, epithelial cells released between 5 and 25 times more CA 125 into the culture medium than did stromal cells. The release of CA 125 was highest in epithelial and stromal cells obtained during the early secretory phase. CA 125 concentrations were markedly elevated in endometrial aspirations obtained during the secretory phase or in endometria with crumbling stroma compared to plasma levels. Plasma levels of CA 125 were slightly elevated during menses. These results suggest that CA 125 is an exocrine product of endometrial epithelial cells. Plasma levels of CA 125 may be of endometrial origin only when the membrane barriers, which normally prevent its entry into the circulation, are damaged.

We performed a prospective study to analyze the functional results following elective laparoscopic sigmoidectomy for computed tomography (CT)-proven diagnosis of acute diverticulitis and review the literature. Forty-three of 45 available patients (96%) who had laparoscopic sigmoidectomy for CT-proven acute diverticulitis answered, after a mean time of 40 months, a questionnaire exploring new abdominal symptoms, bowel function, and the patient's own judgement of the surgical outcome. Surgical technique aimed at removing all the sigmoid by taking down the splenic flexure and do a colorectal anastomosis. Four patients (9%) complained of new abdominal pain. Bowel function was reported as better for 24 patients (56%), unchanged for 16 patients (37%), and worse for 3 (7%). Twenty patients (47%) considered their final result as excellent to good, 17 patients (40%) as satisfying, and 6 patients (13%) as mediocre. Male gender, absence of preoperative history compatible with an irritable bowel syndrome, length of resected sigmoid and residual acute inflammation on histology are statistically predictive of a better postoperative degree of satisfaction. After elective laparoscopic sigmoidectomy for CT-proven diverticulitis, a great majority of patients are very satisfied with their postoperative general comfort.

EDITOR—Coste et al evaluated different techniques in gynaecological cytology,1 but I could not find any comments in the paper about whether replacement of the conventional Pap smear by liquid based methods should be reconsidered, although this was highlighted in the accompanying paragraph for This week in the BMJ , “New cervical smear tests …

The performance of the ThinPrep Papanicolaou test (TP)(CYTYC Corp., Boxborough, MA) was evaluated in a low- risk outpatient population in order to assess its efficacy in routine clinical use. A total of 18,246 cervical cytology samples were prepared using TP during a 6-month evaluation period from January through June 1997. Results of TP interpretation, assessed for specimen quality and diagnostic information as reported according to the Bethesda system terminology, were reviewed and compared with corresponding results from 2 historical control series of 35,347 and 35,409 conventional cervical smears (Papanicolaou smears). Archived conventional smears obtained during the first 6 months of 1995 and 1996 from the same general patient population by the same group of gynecologists and general practitioners were selected for analysis. Positive cytologic diagnoses in the TP series were reviewed and cytologic-histologic correlations were analyzed when biopsy material was available. The results demonstrated that the proportion of cytologic examinations classified as “satisfactory” was significantly greater in the TP series as compared to conventional controls (88.8% vs 71.1%, P < .0001). There was a significant increase in the number of TP examinations with an adequate endocervical component as compared to conventional controls (90.2% vs 86.9%, P < .005) and a significant decrease in the number of TP examinations that were classified as “satisfactory but limited by” either poor fixation or obscuring factors, such as blood, neutrophils, and contaminants (1.1% TP vs 14.1% nonTP, P < .0001). The overall sensitivity of detection for abnormal cells as indicated by the number of positive diagnoses was increased in the TP series (5.7% vs 2.4%, P < .01). The cytologic diagnoses of ASCUS, LSIL, and HSIL were respectively 1.6, 3.6, and 3 times more frequent in the TP series than in controls. Cytologic-histologic correlations were available for 110 (27%) of the 403 cytologic diagnoses of LSIL and for 90 (83%) of the 121 cytologic diagnoses of HSIL. The presence of a histologic lesion (LSIL or HSIL) was confirmed in 71 (65%) biopsies for LSIL and in 81 (90%) biopsies for HSIL. In conclusion, both sample quality and diagnostic sensitivity were significantly improved using the ThinPrep Papanicolaou test under routine conditions in a low-risk population.

The clinical and pathological findings are reported concerning a 73-year-old woman with progressive metastatic carcinoma of the left breast, who developed interfollicular Hodgkin's disease within the right cervical lymph nodes. Metastatic carcinoma and Hodgkin's disease were present concurrently as a "collision tumor' in the lymph nodes examined. To the best of the authors' knowledge, this is the first description of a collision tumor with these particular histologic components in this tissue. The factors influencing the coexistence and the order of appearance of the two pathologies in the present case remain unknown.

The kinetics of various specific and nonspecific immunologic responses were examined in BALB/c mice infected with 17X nonlethal Plasmodium berghei yoelii (a self-limiting infection). The sequence of events after infection was characterized by rapid sensitization of splenic T cells to malaria antigen and polyclonal B cell activation, followed by a period of depressed splenic proliferative responses in vitro to mitogens (PHA and LPS) and malaria (specific) antigen. At the same time, suppressed primary in vitro splenic PFC responses to trinitrophenyl-aminoethylcarbamylmethyl-Ficoll (TNP-F) were seen. This suppression was an active process requiring adherent cells. During this period, levels of antimalarial antibody also increased exponentially. As the infection was cleared, splenic malaria antigen-specific proliferative responses were again observed and splenic PFC and in vitro mitogen responses returned to preinfection levels after variable periods of time. Both splenic proliferative responses to malaria antigen and antimalarial antibody responses remained persistently elevated. In addition, some responses were examined in mice infected with 17X lethal P. b. Yoelii (a fatal infection); in comparison to the early responses of mice infected with the nonlethal substrain, there was a decrease and delay in the development of a splenic T cell response to malaria antigen and a blunted antimalarial antibody response.

The estrogen receptor (ER) content of 31 surgically removed breast tumors (26 duct carcinomas, one lobular carcinoma, one papillary carcinoma, one colloid carcinoma, one duct carcinoma in situ, and one atypical fibroadenoma) was determined by a commercially available immunocytochemical method (Abbott Laboratories, ER-ICA) on cytologic material obtained by fine needle aspiration biopsy (FNAB) of surgical specimens. Immunocytochemical staining of cells by a peroxidasaantiperoxidase technique was evaluated on the basis of the percentage of positive cells and the intensity of staining. An immunostaining score for cytologic (ISCYTO) and histologic (IS-HISTO) material was defined and a threshold of positivity determined to facilitate the serniquantitation of results and the comparison of cases. The results of immunostaining of cytologic material were compared with the evaluation of ER in corresponding tissue samples as determined by the radioligand binding assay using the dextran-coated charcoal procedure (ER-DCC) and by ER-ICA using cryostat sections of frozen tissue. The sensitivity, specificity, predictive value of a positive test, and test efficiency of ER-ICA in cytologic material as compared to ER-DCC was 96%, 831, 96% and 93%, respectively. The ISCYTO was significantly correlated with the IS-HISTO in corresponding histologic material (r = 0.72, P < 0.001). In conclusion, the combination of ER-ICA with FNAB represents a useful new technique for the evaluation of ER which may be applied to small primary tumors, tumor recurrences, and metastases. Cancer 60:1163-1172, 1987. HE DETERMINATION OF the presence of estrogen reT ceptors (ER) by biochemical means is now standard practice in the evaluation of the hormono-dependence of carcinoma of the breast.' This information is correlated with the response to endocrine therapy of individual tumors and may be of prognostic significance.2-6 Recently, monoclonal antibodies to the ER protein estrophilin have been developed, thereby providing a complementary method to determine ER status in histologic mate~ial.~-~ Preliminary studies suggest that the presence of estrophilin as demonstrated by immunocytochemical means is closely correlated with the identification of ER by biochemical techniques and with the response of breast cancer patients to hormonal therapy .9-1

Alkylating agents and 32P have been widely employed in the treatment of patients with essential thrombocythemia (ET). During a four-month period, we observed 3 cases of ET that had transformed into leukemia. Two patients had been treated with uracil mustard: One developed acute myelogenous leukemia 79 months after institution of therapy, and the other patient developed chronic myelomonocytic leukemia 24 months after the start of therapy. The third patient had been treated with busulfan, and ET evolved into myelofibrosis and eventually into acute undifferentiated leukemia with myelofibrosis. The patient who developed acute myelogenous leukemia was asymptomatic at the time of diagnosis of ET but was treated because his platelet count was greater than 1,000,000/mm3. He died 1 month after leukemic transformation, during induction chemotherapy. The other 2 patients presented with symptoms referable to their thrombocythemia. Review of the English literature revealed 12 other definite or probable cases of ET with leukemic transformation, all but 1 having been treated with alkylating agents and/or 32P. We propose that the natural history of ET may be similar to that of polycythemia vera, with evolution into leukemia being an unusual occurrence except in the setting of previous chemotherapy. Therefore, the current practice of treating asymptomatic patients with ET may not be justified, since administration of alkylating agents or 32P may increase the risk of subsequent development of leukemia.

Infection of BALB/c mice with Leishmania tropica NIH S strain resulted in splenic enlargement, hypergammaglobulinemia, and polyclonal activation of B lymphocytes as measured by the splenic plaque-forming cell response (PFC) to trinitrophenyl (TNP) and sheep erythrocytes (SRBC). The peak anti-SRBC PFC response occurred 5 weeks after infection; both direct and indirect (facilitated) plaques were significantly increased. The in vitro primary immune response to trinitrophenyl haptenated lipopolysaccharide (TNP-LPS), as enumerated by the anti-TNP PFC response, was also increased on a per-spleen basis beginning 3 weeks after infection. The properties of a lysate of L. tropica promastigotes (LTL) was studied to determine whether polyclonal B-cell activation was related to a parasite-derived mitogen. A B-cell mitogen was identified in LTL which stimulated the proliferation of spleen cells in vitro from uninfected control and congenitally athymic (T-cell-deficient) but not from μ-suppressed (B-cell-deficient) animals. Preliminary characterization of the mitogen material indicated that it was a nonpyrogenic, heat-labile peptide or protein and was probably not bacterial lipopolysaccharide (LPS).

To characterize further lymphomas which arise in chronically immunosuppressed allotransplant patients, we studied the clinical and histological features of seven cases of nonHodgkin's lymphoma and determined the immunological phenotype in four cases, which developed in a series of 182 patients who underwent cardiac transplantation at Stanford University Hospital. Clinical features which were correlated with the development of lymphoma included patient age, pretransplant diagnosis, and the number of transplants. All cases of nonHodgkin's lymphoma presented as solitary or multiple localized extranodal lesions while none presented in lymph nodes. Extranodal sites of involvement included brain, lung, and soft tissues of the thigh at the site of antilymphocyte serum injections. Histologically, five of seven lymphomas were classified as high grade, large cell, immunoblastic and two of seven as intermediate grade, large noncleaved cell. The immunological phenotype of the neoplasm from four patients was determined by reaction with immunoglobulin light and heavy chain reagents and monoclonal antibodies to T cell and B cell antigens. All lymphomas from four patients which were tested stained for Ia antigen (HLA-DR) but did not stain for immunoglobulin or the T cell antigens detected in this study.

We report a modified method to prepare cytologic specimens for the evaluation of estrogen receptors by immunocytochemistry. Cytologic samples were obtained by needle aspiration of tumor masses (nine patients) and body cavity fluids (nine patients). The cytologic material was divided, and slides for immunostaining were prepared according to two different techniques: (1) fixation of cells in suspension (modified method); and (2) fixation of cells adherent to slides (standard method). The results after immunostaining demonstrated that the number of cells adhering to slides prepared according to the modified method was greatly increased as compared with the number of cells remaining attached to slides prepared according to the standard method. There was no loss of cell groups or aggregates using the modified method. The antigenicity of estrophilin and the cell morphology were well-preserved for up to 30 days. With this modified method, cytologic samples may be stored and conveniently processed in batches, thereby improving the utilization of laboratory resources.