False-negative rate of endoscopic ultrasound-guided fine-needle aspiration for pancreatic solid and cystic lesions with matched surgical resections as the gold standard: one institution's experience.

BACKGROUND: The diagnosis of pancreatic tumors is often complicated because of sampling and interpretive challenges. The current study was performed to determine the rates, types, and causes of diagnostic discrepancies. METHODS: The authors retrospectively reviewed cytology cases from 2004 to 2010 using matched surgical resection cases as the gold standard. RESULTS: A total of 733 cases were divided into 3 categories: 1) positive or suspicious (290 cases); 2) negative or atypical (403 cases); and 3) unsatisfactory (40 cases). Of these cases, 101 fine-needle aspiration (FNA) cases had matched surgical resections including 58 positive diagnoses, 39 negative diagnoses, and 4 unsatisfactory diagnoses. All 19 discrepant cases represented false-negative diagnoses without any false-positive cases noted, which included 2 cases with interpretive errors (10%) and 17 cases with sampling errors (90%). All matched cytology cases were divided into 5 subgroups based on the type of lesion or type of error and were analyzed for sensitivity and specificity. The sampling error rate in cystic lesions (8 of 24; 33%) was significantly higher than that in solid lesions (9 of 73; 12%). The false-negative rate in the interpretive error group (3%) was significantly lower than that in the sampling error group (23%). CONCLUSIONS: The results of the current study confirm that pancreatic endoscopic ultrasound-guided FNA diagnosis has a very low false-positive rate but a relatively high false-negative rate using matched surgical resections as the gold standard. The major cause of a false-negative cytology diagnosis is sampling error and the rate of sampling error in cystic lesions is significantly higher than that in solid lesions.

Effect of Thin Prep(®) imaging system on laboratory rate and relative sensitivity of atypical squamous cells, high-grade squamous intraepithelial lesion not excluded and high-grade squamous intraepithelial lesion interpretations.

INTRODUCTION: Automated screening of Thin Prep(®) Papanicolaou Tests has become increasingly common in clinical practice. Increased productivity has initiated laboratory use of the Thin Prep(®) Imaging System (TIS). Increased sensitivity is a potential additional benefit of TIS. Published studies have shown an increase in discovery of dysplastic cells. This study evaluates the effect of TIS on the incidence of atypical squamous cells high-grade squamous intraepithelial lesion not excluded (ASC-H) and high-grade squamous intraepithelial lesion (HGSIL) results on Thin Prep(®) Pap Tests by comparing TIS-assisted and manual screening findings and the diagnoses on subsequent follow-up in a screening population over a 1-year time period. MATERIALS AND METHODS: A compilation of all ASC-H and HGSIL cases was prepared by conducting a computerized search over a 1-year period (7/06-6/07). The accumulated cases include Thin Prep Pap tests that were both TIS and manually screened. Follow-up results of cytologic and histologic cervical specimens were obtained for a time period extending to 2010. Interpretation utilizing TIS was in place 10 months prior to the study's initiation. RESULTS: During the study period 70,522 Pap tests were performed in our laboratory. One third (33%) of Pap tests were screened with assistance of TIS. Manual screening was performed on 47,380 Pap tests of which 153 (0.32%) were interpreted as ASC-H and 164 (0.35%) were interpreted as HGSIL. During the same time period automated screening (TIS) was performed on 23,111 Pap tests. Interpretation of 62 (0.27%) cases provided an ASC-H result, while 71 (0.31%) were HGSIL. Follow-up cervical dysplasia by colposcopic biopsy and cone biopsy was distributed proportionally between TIS and manual screening for both ASC-H and HGSIL categories. Cervical intraepithelial neoplasia (CIN II/III) was identified on follow-up biopsy of 41% TIS cases and 45% manually screened cases for ASC-H. In the HGSIL subset 71% of TIS cases and 69% manually screened cases showed CIN II/III on follow-up. TIS was 26% less sensitive relative to manual screening for ASC-H cases and 3% less sensitive for HGSIL. CONCLUSION: The similar rate of detection using TIS with an equal percentage of histologic correlation for ASC-H and HGSIL lesions on follow-up histology suggests patients screened by the TIS method are being sent for appropriate follow-up surveillance and treatment. A high-grade or possible high-grade lesion is as likely to be detected by TIS as by a manual screen. The similarities in relative sensitivity and specificity in a direct comparison between manual and TIS screening methodologies indicate that TIS compared to manual screening does not affect detection in patients with high-grade cervical lesions.

Cellient™ automated cell block versus traditional cell block preparation: a comparison of morphologic features and immunohistochemical staining.

Traditional cell block (TCB) sections serve as an important diagnostic adjunct to cytologic smears but are also used today as a reliable preparation for immunohistochemical (IHC) studies. There are many ways to prepare a cell block and the methods continue to be revised. In this study, we compare the TCB with the Cellient™ automated cell block system. Thirty-five cell blocks were obtained from 16 benign and 19 malignant nongynecologic cytology specimens at a large university teaching hospital and prepared according to TCB and Cellient protocols. Cell block sections from both methods were compared for possible differences in various morphologic features and immunohistochemical staining patterns. In the 16 benign cases, no significant morphologic differences were found between the TCB and Cellient cell block sections. For the 19 malignant cases, some noticeable differences in the nuclear chromatin and cellularity were identified, although statistical significance was not attained. Immunohistochemical or special stains were performed on 89% of the malignant cases (17/19). Inadequate cellularity precluded full evaluation in 23% of Cellient cell block IHC preparations (4/17). Of the malignant cases with adequate cellularity (13/17), the immunohistochemical staining patterns from the different methods were identical in 53% of cases. The traditional and Cellient cell block sections showed similar morphologic and immunohistochemical staining patterns. The only significant difference between the two methods concerned the lower overall cell block cellularity identified during immunohistochemical staining in the Cellient cell block sections.

High-risk HPV testing in women 30 years or older with negative Papanicolaou tests: initial clinical experience with 18-month follow-up.

Cervical screening with combined cytology and high-risk human papillomavirus (HR-HPV) detection has been approved for women 30 years or older. We investigated the clinical use of cotesting for women with negative Papanicolaou tests. Follow-up cytology, HR-HPV test, and biopsy findings were identified during an 18-month period. In 1 year, 2,719 cotests from 2,686 women were identified; 146 were positive for HR-HPV. Among women with positive HR-HPV testing, 120 had follow-up, including 70 with repeated cotesting, and 3 had high-grade dysplasia identified (2.5% of women with follow-up). In 1,334 women with initial double-negative cotest results who had repeated cytologic testing within 18 months, 2 high-grade dysplasias were found (0.1%). The vast majority of cotest results are double-negative. Among tests that show HR-HPV positivity, the prevalence of underlying high-grade dysplasia is low. About half of all women who undergo cotesting receive follow-up that is not in accord with published guidelines.

ThinPrep Pap test of endocervical adenocarcinoma with lymph node metastasis: report of a case in a 17-year-old woman.

Endocervical adenocarcinoma is an uncommon malignancy that is composed of multiple subtypes and accounts for approximately 15% of all cervical cancers. In this article, we describe the cytomorphology and differential diagnosis of an AJCC clinical stage IIIb, FIGO IB2 endocervical adenocarcinoma in a 17-year-old woman in a ThinPrep Pap test.The patient was a 17-year-old G0P0 white woman with no significant past medical history and no prior history of cervical dysplasia. She presented to her physician with a putrid vaginal discharge. A sample was sent to cytology that was interpreted as atypical endocervical cells, favor neoplasia. A subsequent cervical biopsy was diagnosed as endocervical adenocarcinoma with villoglandular features and ultimately, a hysterectomy with lymph node dissection was performed. The final diagnosis was endocervical adenocarcinoma with metastasis to three pelvic lymph nodes.The cytomorphology of endocervical adenocarcinoma on ThinPrep Pap test is similar to that described for conventionally-processed Pap smears. This difficult diagnosis should be considered on a ThinPrep Pap test, regardless of age when the characteristic cytomorphology is observed. On a cytology sample, it is advisable to state atypical endocervical cells, adenocarcinoma in situ, or endocervical adenocarcinoma without providing a specific subtype even if there is a predominance of features for a particular subtype.

Carbonic anhydrase IX and human papillomavirus as diagnostic biomarkers of cervical dysplasia/neoplasia in women with a cytologic diagnosis of atypical glandular cells: a Gynecologic Oncology Group study in United States.

High-risk human papillomavirus (H-HPV) infection is strongly linked to cervical neoplasia, but its role in detecting glandular lesions (GLs) is unclear. In the cervix, carbonic anhydrase IX (CA-IX) is expressed in cervical neoplasia, but rarely in the benign cervix. The diagnostic utility of these biomarkers was evaluated in women with a cytologic diagnosis of atypical glandular cells (AGC). H-HPV was detected using hybrid capture 2 (HC2) in liquid-based cytology, and CA-IX immunoreactivity was studied on conventional Pap smears. Of 403 patients, 111 (28%) were positive for significant cervical lesions (SCLs) including CIN2, CIN3, adenocarcinoma in situ or invasive carcinoma. CA-IX testing alone (n = 403) had a sensitivity of 75, 95 or 65% for SCLs, significant GLs or squamous lesions (SLs), respectively, with a specificity of 88% and a false negative rate (FNR defined as 1 minus negative predictive value) of 10%. Testing for H-HPV (n = 122) had a sensitivity of 97, 100 or 96% for SCLs, GLs or SLs, respectively, with a specificity of 87% and a FNR of 1%. The combination of CA-IX and H-HPV testing (n = 122), collectively, had the same sensitivity, specificity and FNR for SCLs, GLs or SLs as H-HPV testing alone. The conclusions of our study are that both H-HPV and CA-IX testing are useful diagnostic markers for GLs. However, H-HPV testing is a better diagnostic marker for SLs. The combination of CA-IX with H-HPV testing does not improve the diagnostic accuracy for cervical neoplasia in women with AGC diagnosis over that of H-HPV testing alone.

Confocal microscopy of unfixed breast needle core biopsies: a comparison to fixed and stained sections.

BACKGROUND: Needle core biopsy, often in conjunction with ultrasonic or stereotactic guided techniques, is frequently used to diagnose breast carcinoma in women. Confocal scanning laser microscopy (CSLM) is a technology that provides real-time digital images of tissues with cellular resolution. This paper reports the progress in developing techniques to rapidly screen needle core breast biopsy and surgical specimens at the point of care. CSLM requires minimal tissue processing and has the potential to reduce the time from excision to diagnosis. Following imaging, specimens can still be submitted for standard histopathological preparation. METHODS: Needle core breast specimens from 49 patients were imaged at the time of biopsy. These lesions had been characterized under the Breast Imaging Reporting And Data System (BI-RADS) as category 3, 4 or 5. The core biopsies were imaged with the CSLM before fixation. Samples were treated with 5% citric acid and glycerin USP to enhance nuclear visibility in the reflectance confocal images. Immediately following imaging, the specimens were fixed in buffered formalin and submitted for histological processing and pathological diagnosis. CSLM images were then compared to the standard histology. RESULTS: The pathologic diagnoses by standard histology were 7 invasive ductal carcinomas, 2 invasive lobular carcinomas, 3 ductal carcinomas in-situ (CIS), 21 fibrocystic changes/proliferative conditions, 9 fibroadenomas, and 5 other/benign; two were excluded due to imaging difficulties. Morphologic and cellular features of benign and cancerous lesions were identified in the confocal images and were comparable to standard histologic sections of the same tissue. CONCLUSION: CSLM is a technique with the potential to screen needle core biopsy specimens in real-time. The confocal images contained sufficient information to identify stromal reactions such as fibrosis and cellular proliferations such as intra-ductal and infiltrating carcinoma, and were comparable to standard histologic sections of the same tissue. Morphologic and cellular features of benign and cancerous lesions were identified in the confocal images. Additional studies are needed to 1.) establish correlation of the confocal and traditional histologic images for the various diseases of the breast; 2.) validate diagnostic use of CSLM and; 3.) further define features of borderline lesions such as well-differentiated ductal CIS vs. atypical hyperplasia.

Pulmonary metastases from uterine neoplasms after long tumour-free interval: four cases and review of the literature.

AIMS: Uncommon cases of lung metastasis from different types of uterine neoplasms with a long tumour-free interval after hysterectomy are reported. METHODS AND RESULTS: Four cases were retrieved from our surgical pathology archives. Case 1 was a 68-year-old woman who had three pulmonary nodules 23 years after hysterectomy for low-grade endometrial stromal sarcoma (LGESS). The nodules obtained with video-assisted thoracic surgeries were consistent with metastatic LGESS. Case 2 was a 36-year-old woman who had numerous bilateral pulmonary nodules 6 years after hysterectomy for leiomyoma. A transthoracic biopsy revealed benign metastasising leiomyoma. Case 3 was a 77-year-old woman who had a large lung mass with satellite nodules 17 years after hysterectomy with bilateral salpingo-oophorectomy and subsequent radiotherapy for endometrial endometrioid adenocarcinoma (EEA). The biopsied and resected lung tumour was consistent with metastatic EEA. Case 4 was a 51-year-old woman who underwent total hysterectomy and subsequent radiotherapy for endocervical adenocarcinoma 12 years ago and lung lobectomy for metastatic disease 8 years ago. She then developed two pulmonary lesions 14 months ago, and these resected after radiotherapy were metastatic endocervical adenocarcinoma. CONCLUSIONS: A review of the literature revealed that late pulmonary metastasis from uterine neoplasms is rare but not negligible. Immunohistochemical studies and molecular tests, together with detailed clinical information and imaging findings, are important for rendering a diagnosis.

Identification of novel methylation markers in cervical cancer using restriction landmark genomic scanning.

Aberrant methylation of CpG islands in gene promoters often represents an early clonal event in carcinogenesis. Accordingly, defining methylation profiles may be useful for developing marker panels for early detection or predicting the risk of cancer precursors. To identify specific genes frequently methylated in cervical cancer, we conducted methylation profiling of 20 primary human cervical cancers using NotI-based restriction landmark genomic scanning (RLGS). Of 2,172 RLGS fragments analyzed (average, 1,753 CpG islands per patient), 186 RLGS fragments were lost in at least one tumor and 40 were lost in three or more. Methylation was identified in 19 (95%) of 20 tumor samples compared with normal DNA. Bisulfite sequencing was conducted to confirm RLGS results. Of the confirmed markers frequently methylated, we developed Methylight assays for two corresponding genes, nucleolar protein 4 (NOL4), and lipoma HMGIC fusion partner-like protein 4 (LHFPL4), which were methylated in 85% and 55% of cancers, respectively. Using these assays, we further confirmed frequent CpG island methylation in the original cancers and in another independent series of 15 cervical cancers. We also showed methylation at a reduced frequency in a set of carefully reviewed cytology specimens demonstrating cells exfoliated from cancer precursor lesions. In summary, we identified, for the first time, NOL4 and LHFPL4 as novel methylation targets specific for cervical cancer. Inclusion of NOL4 and LHFPL4 in evaluating methylation panels for early detection, risk prediction, and etiologic research on cervical cancer is warranted.

Use of the ThinPrep Imaging System does not alter the frequency of interpreting Papanicolaou tests as atypical squamous cells of undetermined significance.

BACKGROUND: Automated screening of Papanicolaou tests (Pap tests) improves the productivity of cytopathology laboratories. The ThinPrep Imaging System (TIS) has been widely adopted primarily for this reason for use on ThinPrep Pap tests (TPPT). However, TIS may also influence the interpretation of Pap tests, leading to changes in the frequency of various interpretive categories. The effect of the TIS on rates of TPPT interpretation as atypical squamous cells of undetermined significance (ASC-US) is of concern because any shift in the frequency of ASC-US will alter the sensitivity and specificity of the Pap test. We have sought to determine whether automated screening of TPPT has altered ASC-US rates in our institution when compared with manual screening (MS) of TPPT. METHODS: A computerized search for all ASC-US with reflex Human Papillomavirus (HPV) testing over a one-year-period (7/1/06 to 6/30/07) was conducted. Cases included both TPPT screened utilizing TIS and screened manually. HPV test results for both groups were recorded. Pertinent follow-up cervical cytology and histology results were retrieved for the period extending to 11/30/07. Automated screening was in clinical use for 10 months prior to the start of the study. RESULTS: Automated screening was performed on 23,103 TPPT, of which 977 (4.23%) were interpreted as ASC-US. Over the same period, MS was performed on 45,789 TPPT, of which 1924 (4.20%) were interpreted as ASC-US. Reflex HPV testing was positive for high risk (HR) types in 47.4% of the TIS cases and 50.2% of MS cases. Follow-up cervical dysplasia found by colposcopy was also distributed proportionally between the two groups. Cervical intraepithelial neoplasia (CIN) was found on follow-up biopsy of 20.1% of the TIS cases (5.2% CIN 2/3) and 21.2% of MS cases (5.1% CIN 2/3). None of these differences were statistically significant. CONCLUSION: Use of the ThinPrep Imaging System did not appreciably change ASC-US rates or follow-up reflex HPV test results in our laboratory. This demonstrates that the benefits of automated screening may be obtained without increasing the rate of referral to colposcopy for ASC-US follow-up.

Utility of anti-L523S antibody in the diagnosis of benign and malignant serous effusions.

BACKGROUND: Immunohistochemistry is helpful in distinguishing metastatic carcinoma from atypical mesothelial cells; however, it is not useful in differentiating atypical mesothelial cells from malignant mesothelial cells. K homolog domain containing protein overexpressed in cancer (KOC), a member of the insulin-like growth factor mRNA-binding protein (IMP) family, also known as L523S and IMP3, is expressed during embryogenesis and in various malignancies. Using a mouse monoclonal antibody (L523S) against KOC, KOC expression was investigated in malignant tumors and reactive mesothelial cells in serous effusions. METHODS: Seventy-six cases with paraffin-embedded pleural, pericardial, and peritoneal serous effusion cell blocks including 60 malignant serous effusions (11 malignant pleural mesotheliomas and 49 metastatic carcinomas) and benign pleural effusions (14 cases with reactive mesothelial cells and 2 cases with atypical cells with uncertain significance) were selected for immunohistochemical analysis with L523S, calretinin, and CK5/6. RESULTS: Immunohistochemical studies showed that positive staining for KOC of variable degrees of intensity was observed in 47 of 60 cases in malignant serous effusions including 10 of 11 mesotheliomas and 36 of 49 metastatic carcinomas. The associated reactive mesothelial cells were negative for KOC but positive for calretinin and CK5/6. All 11 malignant mesotheliomas exhibited positivity for calretinin, and 9 of 11 cases had CK5/6 staining. In addition, 16 cases that were originally diagnosed either as pleural effusions with reactive mesothelial cells (14) or atypical cells with uncertain significance (2) were also tested for KOC expression. Interestingly, 3 of 16 cases exhibited various degrees of positivity for KOC, 2 of which were diagnosed as lung adenocarcinoma with a recurrence after tumor resection and 1 as malignant pleural mesothelioma. CONCLUSIONS: Anti-L523S antibody is a useful marker for the detection of malignant cells in serous effusions and it can have significant utility in differentiating reactive mesothelial cells from malignant mesothelioma and metastatic carcinoma in combination with calretinin and CK5/6 staining.

Evaluation of candidate methylation markers to detect cervical neoplasia.

OBJECTIVE: Studies of cervical cancer and its immediate precursor, cervical intraepithelial neoplasia 3 (CIN3), have identified genes that often show aberrant DNA methylation and therefore represent candidate early detection markers. We used quantitative PCR assays to evaluate methylation in five candidate genes (TNFRSF10C, DAPK1, SOCS3, HS3ST2 and CDH1) previously demonstrated as methylated in cervical cancer. METHODS: In this analysis, we performed methylation assays for the five candidate genes in 45 invasive cervical cancers, 12 histologically normal cervical specimens, and 23 liquid-based cervical cytology specimens confirmed by expert review as unequivocal demonstrating cytologic high-grade squamous intraepithelial lesions, thus representing the counterparts of histologic CIN3. RESULTS: We found hypermethylation of HS3ST2 in 93% of cancer tissues and 70% of cytology specimens interpreted as CIN3; hypermethylation of CDH1 was found in 89% of cancers and 26% of CIN3 cytology specimens. Methylation of either HS3ST2 or CDH1 was observed in 100% of cervical cancer tissues and 83% of CIN3 cytology specimens. None of the five genes showed detectable methylation in normal cervical tissues. CONCLUSION: Our data support further evaluation of HS3ST2 and CDH1 methylation as potential markers of cervical cancer and its precursor lesions.

Myometrial dysplasia (atypical myometrial hyperplasia).

Although precursor lesions are well known for cervical and endometrial neoplasms, precursor lesions are not currently recognized for the most common tumor of the uterus-leiomyomas. Myometrial hyperplasia has been recently described and evaluated by morphometry, but its relationship to uterine leiomyomas has not been systematically explored. Myometrial dysplasia (atypical myometrial hyperplasia) has not been previously recognized. We herein report a case of myometrial dysplasia with immunostains for proliferation marker MIB-1 (Ki-67) and for p53. The paradoxical rarity of myometrial dysplasia is considered in comparison to the striking frequency of uterine leiomyomas.

Scientific issues related to the cytology proficiency testing regulations.

The member organizations of the Cytology Education and Technology Consortium believe there are significant flaws in current cytology proficiency testing regulations. The most immediate needed modifications include lengthening the required testing interval, utilizing stringently validated and continuously monitored slides, changing the grading scheme, and changing the focus of the test from the individual to laboratory level testing. Integration of new computer-assisted and located-guided screening technologies into the testing protocols is necessary for the testing protocol to be compliant with the law.

Should women with atypical squamous cells, cannot exclude high-grade squamous intraepithelial lesion, receive reflex human papillomavirus-DNA testing?

BACKGROUND: The 2001 American Society for Colposcopy and Cervical Pathology Consensus Guidelines recommend that women who have Papanicolaou (Pap) smears diagnosed as atypical squamous cells (ASC), cannot exclude high-grade squamous intraepithelial lesion (HSIL) (ASC-H) should be referred for immediate colposcopic examination. The objective of this pilot study was to evaluate whether reflex human papillomavirus (HPV)-DNA testing performed on smears diagnosed as ASC-H may obviate the need for immediate colposcopic examination. METHODS: All ThinPrep Pap smears that were diagnosed as ASC-H or atypical squamous metaplastic cells (ASMT) between 2001-2003 and that had HPV-DNA testing and subsequent histologic and/or cytologic follow-up were evaluated. Those smears that were diagnosed as ASMT were reviewed and reclassified under the 2001 Bethesda System as either ASC of undetermined significance (ASCUS) or ASC-H. Smears that were diagnosed as ASCUS were excluded from the study. RESULTS: The study included of 48 smears that were diagnosed as ASC-H. All patients with biopsy-proven HSIL had positive high-risk (HR)-HPV results (100% negative predictive value). Approximately 80% of patients with ASC-H who had biopsy-proven, low-grade intraepithelial lesion on follow-up had positive HR-HPV results. Among the patients who had ASC-H with negative follow-up, 50% had positive HR-HPV results, and 50% had negative HR-HPV results. CONCLUSIONS: Among patients with ASC-H, a negative HR-HPV result was found to be an excellent predictor of the absence of HSIL. The results of this pilot study suggested that HPV-DNA testing may serve as a means to better select which patients with ASC-H on Pap smear should undergo colposcopic examination. This approach potentially may reduce medical costs and eliminate the need for routine colposcopic examination among patients with ASC-H Pap smears.