Cytology Specimens Research Articles

The Impact of Robotic Assistance-Based Navigation Systems on the Adequacy of EBUS-Guided Cytology Specimens - Experience From a Large University Hospital

The Impact of Robotic Assistance-Based Navigation Systems on the Adequacy of EBUS-Guided Cytology Specimens - Experience From a Large University Hospital

Exploring the Efficacy of an Artificial Intelligence Model for Fungal Screening in Bronchoalveolar Lavage Cytology Specimens

Exploring the Efficacy of an Artificial Intelligence Model for Fungal Screening in Bronchoalveolar Lavage Cytology Specimens

Utilization of an artificial intelligence-enhanced, web-based application to review bile duct brushing cytologic specimens: A pilot study.

The authors previously developed an artificial intelligence (AI) to assist cytologists in the evaluation of digital whole-slide images (WSIs) generated from bile duct brushing specimens. The aim of this trial was to assess the efficiency and accuracy of cytologists using a novel application with this AI tool. Consecutive bile duct brushing WSIs from indeterminate strictures were obtained. A multidisciplinary panel reviewed all relevant information and provided a central interpretation for each WSI as being "positive," "negative," or "indeterminate." The WSIs were then uploaded to the AI application. The AI scored each WSI as positive or negative for malignancy (i.e., computer-aided diagnosis [CADx]). For each WSI, the AI prioritized cytologic tiles by the likelihood that malignant material was present in the tile. Via the AI, blinded cytologists reviewed all WSIs and provided interpretations (i.e., computer-aided detection [CADe]). The diagnostic accuracies of the WSI evaluation via CADx, CADe, and the original clinical cytologic interpretation (official cytologic interpretation [OCI]) were compared. Of the 84 WSIs, 15 were positive, 42 were negative, and 27 were indeterminate after central review. The WSIs generated on average 141,950 tiles each. Cytologists using the AI evaluated 10.5 tiles per WSI before making an interpretation. Additionally, cytologists required an average of 84.1 s of total WSI evaluation. WSI interpretation accuracies for CADx (0.754; 95% CI, 0.622-0.859), CADe (0.807; 95% CI, 0.750-0.856), and OCI (0.807; 95% CI, 0.671-0.900) were similar. This trial demonstrates that an AI application allows cytologists to perform a triaged review of WSIs while maintaining accuracy.

Deciphering Breast Origin in Malignant Effusions: The Diagnostic Utility of an MGP, GATA-3, and TRPS-1 Immunocytochemical Panel

Introduction: Defining the origin of metastatic cancer is crucial for establishing an optimal treatment strategy, especially when obtaining sufficient tissue from secondary malignancies is limited. While cytological examination is often used in this diagnostic setting, morphologic analysis alone often fails to differentiate metastases derived from the breast from other primaries. The hormone receptor, human epidermal growth factor receptor-2, gross cystic disease fluid protein 15, and mammaglobin immunohistochemistry are often used to diagnose metastatic breast cancer. However, their effectiveness decreases in estrogen receptor (ER)-negative breast cancers, including the triple-negative breast cancer (TNBC) subtype. Methods: We conducted a comprehensive evaluation of GATA-binding protein 3 (GATA-3), trichorhinophalangeal syndrome type 1 (TRPS-1), and Matrix Gla Protein (MGP) immunochemistry across 140 effusion cytology specimens with metastatic adenocarcinoma derived from various primaries, including the breast, colon, pancreaticobiliary, lung, tubo-ovarian, and stomach. Results: The expression rates of these immunomarkers were significantly higher in metastatic cancers originating from the breast than other primaries. In TNBC, TRPS-1 (80.00%) and MGP (65.00%) exhibited higher positivity rates compared to GATA-3 (40.00%). Additionally, our data suggest that an immunohistochemical panel comprising MGP, GATA-3, and TRPS-1 significantly enhances the detection of metastatic breast cancer in effusion cytology specimens, including TNBC in particular. When considering dual-marker positivity, the diagnostic accuracy was found to be 89.29% across all breast cancer subtypes and 92.93% for TNBC. Conclusions: MGP appears to be a robust marker for identifying metastatic breast cancer in malignant effusions, especially TNBC. MGP notably enhances diagnostic accuracy when incorporated together with GATA-3 and TRPS-1 in an immunohistochemical panel.

Molecular profiling of low-risk papillary thyroid carcinoma (mPTC) on active surveillance.

The active surveillance (AS) program for papillary thyroid carcinoma (≤ 1 cm) at low-risk (mPTC) showed a low percentage of progression. The aim of this study was to find a molecular signature of cases that showed disease progression during AS, which would allow their early identification. We performed next generation sequencing of 95 fine needle aspiration cytology specimens from cases prospectively enrolled in the AS program to analyze key somatic driver alterations or gene fusions implicated in PTC tumorigenesis. TERT promoter analysis was performed using Sanger sequencing or droplet digital PCR. BRAF p.V600E was found in 66.3% (63/95) of mPTC and was the most common somatic alteration, followed by RAS oncogene mutations detected in 3.2% of mPTC (3/95: 2 NRAS and 1 KRAS) and gene fusions detected in 3.2% of mPTC (3/95: 1 RET-PTC1, 1 TFG-NTRK1, 1 ALK imbalance). No TERT promoter mutations (C228T and C250T) were found in the analyzed mPTC (84/95). The comparison between the molecular profile and the clinical outcome of the mPTC (stable versus progressive disease) showed no correlation (p-value=0.6) and did not identify a molecular signature able to identify progressive mPTC. The molecular profile of mPTC is like that of bigger PTC with the exception that none of them showed a TERT promoter mutation. The identification of the most common driver mutations, such as BRAF, RAS, or gene fusions, is not helpful for the early identification of mPTC that will show disease progression during follow-up in the AS program.

Deep learning model utilizing fluorescence lifetime imaging microscopy and confidence learning for predicting endometrial cancer risk

This study introduces a novel diagnostic approach for the prediction of endometrial cancer risk, which involves minimally invasive exfoliation cytology specimens from 63 participants, staining with the viscosity-sensitive fluorescent probe DCVJ, and detection by fluorescence lifetime imaging microscopy (FLIM). Initially, DCVJ-stained tissue sections from cancerous and normal regions were utilized to confirm the elevated viscosity in cancerous lesions. Subsequently, DCVJ-stained aspirated cell samples were detected by FLIM to capture viscosity characteristics and cellular morphology. A total of 2348 FLIM images collected from 42 individuals, were utilized for training purposes. A deep learning model, incorporating confidence learning, was developed and demonstrated a sensitivity of 84.6% and specificity of 75.0% in predicting endometrial cancer risk. This study highlights the potential of the deep learning-FLIM model for the prediction of endometrial cancer risk.

Cytologic Findings and Ancillary Tests Results of Sclerosing Pneumocytoma: Our Institutional Experience.

Sclerosing pneumocytoma (SP) is a rare benign tumor and a potential diagnostic pitfall. Our aim was to review the cytologic features of our surgically diagnosed SP cases including the clinical, immunohistochemical and available molecular findings. A computerized search from 2013 to 2020 for surgical cases with corresponding cytology specimens diagnosed as SP was performed. The clinical data, cytology, and surgical specimens were collated for analysis. Six cytology specimens were collected. All were female (mean age = 35). Three have incidental lung nodules and three with cough. Cytologic findings showed variable architectural pattern (papillary, solid, singly scattered, acinar/rosette-like) and cellular heterogeneity (surface, stromal, epithelioid, plasmacytoid cells). Atypia was identified in 4/6 cases. The original cytology diagnoses were negative = 1, SP = 2 and adenocarcinoma = 3. The latter diagnoses were amended to SP after review of the surgical specimens. The three false positive cases on review have cytologic features mimicking adenocarcinoma. Immunohistochemical stains showed tumor cells (surface and stromal) were positive for TTF-1, and EMA with only the surface cells positive for pancytokeratin and Napsin A. Though two cases sent for molecular testing were negative for AKT1 or CTNNB1 exon 3 mutation, our panel did not evaluate AKT1 exon 4. SP is a diagnostic pitfall with 50% initially misdiagnosed as adenocarcinoma. Integrating the clinical/radiologic findings, cytologic features, and performance of immunohistochemistry on cell block are helpful in avoiding misdiagnosis. Molecular testing for recurrent mutations, if present, could be helpful for diagnosis and possible therapy options. However, routinely used molecular testing may not always capture relevant molecular markers for SP.

Comparison of Claudin-4, BerEP4, Carcinoembryonic Antigen and MOC31 in Serous Fluids Metastases Demonstrate High Sensitivity of Claudin-4 at Low Cellularity.

Claudin-4 has been described as a highly sensitive immunocytochemical marker for detection of metastatic carcinoma cells in effusion cytology specimens. This study aims to challenge the performance of claudin-4 in different types of malignancies and low cellularity specimens, by comparison with other markers in a large cohort of carcinomatous effusion specimens. Cell block preparations from peritoneal and pleural fluid specimens were retrieved, with malignant (carcinoma) diagnoses confirmed by review of hospital diagnosis code and pathology reports. Claudin-4, BerEP4, CEA, and MOC31 immunocytochemistry were performed and scored by expression proportion and intensity. Tumor cellularity was assessed for subgroup analysis of low cellularity specimens. Totally 147 specimens (70 pleural, 77 peritoneal) of 68 lung, 62 breast, 9 gynecological, and 7 gastrointestinal carcinomas were retrieved. The average proportion expression of claudin-4 was highest (89.6%, vs. CEA 40.5%, BerEp4 18.6%, MOC31 16.8%) and the percentage of strong expression was highest for claudin-4 (72.1%). Expression levels of claudin-4 were consistently higher than other markers in subgroups of all primary sites. The difference was more significant for low cellularity specimens. High (≥50%) proportion expression was seen for 96.61% of cases for claudin-4 (vs. BerEp4 8.77%, CEA 46.55%, MOC31 8.77%, p < 0.001). These factors contributed to a low concordance between claudin-4 and BerEp4, CEA and MOC31 (K = 0.010-0.043). Claudin-4 is more sensitive than CEA, BerEp4 and MOC31, suitable for low cellularity specimens of most types of metastatic carcinoma and is a robust immunocytochemical marker for carcinoma that can be used solitarily.

Clinicopathological features of metastatic melanoma in effusion cytology of serosal cavity

Objective: To investigate the clinical, cytomorphology, immunocytochemical and molecular features of metastatic melanoma in serosal cavity effusion. Methods: Cytological specimens of 14 patients with melanoma in the chest and abdomen were collected from 2017 to 2023, at the Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China. SOX10, S-100 protein, PRAME, BRAF V600E, HMB45, and Melan A were detected by immunocytochemical methods. Fourteen cases were tested for routine antibody combinations, including Claudin4, HEG1, Calretinin, CD68, etc. Four of the patients had biopsy or surgical samples of metastatic solid lesions of primary sites, and further next-generation sequencing (NGS) or amplification refractory mutation system (ARMS)-PCR molecular test was performed. In addition, 30 cases of serosal effusion samples were collected as control groups (10 cases of benign mesothelial cell reactive hyperplasia, 10 cases of mesothelioma, and 10 cases of metastatic lung adenocarcinoma). Results: Among the 14 cases of melanoma, there were 7 males and 7 females, with ages ranging from 35 to 86 years, and an average age of 57 years, there 10 cases aged ≥50 years. The tumor cells in the serosal effusion varied in morphology and degree of atypia. SOX10 was positive in all 14 cases (14/14), S-100 protein was positive in 10 cases (10/14), PRAME was positive in 12 cases (12/14), BRAF V600E was positive in 10 cases (10/14), HMB45 was positive in 12 cases (12/14), and Melan A was positive in 13 cases (13/14). In 4 patients with histological correlation, the cytological and histological expression of SOX10, BRAF V600E, and PRAME was positive in all 4 cases (4/4); S-100 protein was positive in 2 cases (2/4); and HMB45 and Melan A were positive in 3 cases (3/4). Using NGS or ARMS-PCR, missense mutations of BRAF V600E were detected in all 4 patients; TERT promoter mutations was detected in 1 case; and CDKN2A terminating mutations and MSI1 deletion mutations were detected in the other case. SOX10, S-100, HMB45, Melan A, PRAME and BRAF V600E were all negative in 30 control samples of serosal cavity effusion. Conclusion: By observing the morphology of tumor cells, immunocytochemical test of several combination markers, especially the expression of SOX10, BRAF V600E and PRAME, can help to improve the positive diagnosis rate of melanoma in serous cavity effusion.

Application of the international system for reporting serous fluid cytopathology on pleural effusion cytology with paired pleural biopsy: A new insight and novel approach on risk of malignancy.

The risk of malignancy (ROM) remains an area of interest for further evaluation in reporting systems including in International System for reporting serous fluid cytopathology (TIS), which is a standardized system for reporting effusion cytology. Herein, we report our findings in further investigation of ROM in TIS by studying on paired pleural effusion specimens and corresponding pleural biopsies with emphasis on negative for malignancy, and atypia of undetermined significance categories. The Johns Hopkins Hospital pathology database was retrospectively searched for patients with a pleural biopsy (PBX) and a paired pleural effusion (PF) cytology specimens over a 4-year period. We employed the TIS categories. The following statistical parameters were evaluated: sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), and ROM. A total of 223 patient cases were included. Effusions TIS reclassification and ROM were as follows: 1.8% non-diagnostic (ROM 75%), 75.8% negative for malignancy (ROM 23%), 4.9% atypical cells of undetermined significance (ROM 45%), 2.2% suspicious for malignancy (ROM 80%), and 15.2% malignant (ROM 100%). Overall accuracy, sensitivity, specificity, PPV and NPV were calculated and were 79.4%, 45%, 97.7%, 91.2% and 77%, respectively. Among, discordant cases diagnosed negative for malignancy on PF and positive for malignancy on PBX, there were significant number of lymphomas, mesotheliomas, and sarcomas. Lung cancer was the most common carcinoma; however, rare types of carcinomas were noted. Cells blocks and immunohistochemistry (IHC) studies were utilized to confirm either malignant conditions or rule out malignancy in both cell blocks and histology biopsies. This study demonstrates the high specificity and ROM for 'malignant' and 'suspicious for malignancy' categories in the TIS reporting system and highlights the modest negative predictive value for the 'negative for malignancy' category. Although Tissue biopsies are usually considered as 'gold standard', any definitive diagnosis of malignancy of body fluid should be considered positive for malignancy in further clinical decision-making.

Expression and hypermethylation of JAM and EPB41L3 in cervical squamous cell carcinoma: Clinical significance and applications.

This study aimed to explore the expression and hypermethylation of EPB41L3 and JAM3 in cervical squamous cell carcinoma (CSCC) and to investigate their clinical significance. JAM3 and EPB41L3 mRNA expression was analyzed using a public database, and protein expression was detected using immunohistochemistry. The methylation status of JAM3 and EPB41L3 was detected in CSCC tissues and cervical cytological specimens using a quantitative methylation-specific PCR (qMSP). JAM3 and EPB41L3 mRNA were downregulated in CSCC. The JAM3 protein was positively detected in 39.4% of CSCC tissues and frequently expressed in those with lower FIGO stage and no lymph node metastasis. EPB41L3 was expressed in 18.9% of CSCC tissues. The hypermethylation of JAM3 was detected in 52.3% of CSCC tissues and related to higher FIGO stage and lymph node metastasis. EPB41L3 hypermethylation was detected in 72.7% of CSCC tissues and related to older ages and lymph node metastasis. In cervical cytological specimens, no methylation of JAM3 and EPB41L3 was found in normal or inflamed cervical epithelial cells. The methylation of JAM3 was detected in 0%, 8.3%, and 6.3% of ASCUS, LSIL, and HSIL samples, while EPB41L3 was detected in 12.5%, 42.9%, and 71.4%, respectively. The sensitivity of the combination of JAM3 and EPB41L3 methylation detection in ASCUS, LSIL, and HSIL was 8.3%, 15.6%, and 85.7%, respectively. The specificity of the combination of JAM3 and EPB41L3 methylation detection was 100%. Downregulation of JAM3 and EPB41L3 by hypermethylation was detected in CSCC. JAM3 and EPB41L3 hypermethylation are potential biomarkers for cervical cancer screening.

Quality Assurance in Immunocytochemistry: A Review and Practical Considerations.

Cytological samples play a critical role in diagnosing advanced-stage tumors and those arising in difficult-to-reach anatomical sites such as the pancreatobiliary tract, lung, thyroid, suprarenal, pelvis, and others such as salivary glands. These samples are often the only available material for accurate diagnosis and for performing ancillary studies, such as immunocytochemistry (ICC) or the detection of molecular biomarkers. While the use of immunohistochemistry is well established and standardized on formalin-fixed-paraffin-embedded histological tissue, in cytological samples, it presents unique challenges. Methods used for obtaining and processing these specimens are complex and are not standardized among laboratories. Moreover, there is also diversity in the types of cytological samples potentially suitable for ICC. This review explores the current landscape of ICC practices in European and North American laboratories, highlighting variability in methods and the need for standardization to ensure reliable results and reproducibility of ICC on cytological specimens.

The Advantage of Targeted Next-Generation Sequencing over qPCR in Testing for Druggable EGFR Variants in Non-Small-Cell Lung Cancer.

The emergence of targeted therapies in non-small-cell lung cancer (NSCLC), including inhibitors of epidermal growth factor receptor (EGFR) tyrosine kinase, has increased the need for robust companion diagnostic tests. Nowadays, detection of actionable variants in exons 18-21 of the EGFR gene by qPCR and direct DNA sequencing is often replaced by next-generation sequencing (NGS). In this study, we evaluated the diagnostic usefulness of targeted NGS for druggable EGFR variants testing in clinical NSCLC material previously analyzed by the IVD-certified qPCR test with respect to DNA reference material. We tested 59 NSCLC tissue and cytology specimens for EGFR variants using the NGS 'TruSight Tumor 15' assay (Illumina) and the qPCR 'cobas EGFR mutation test v2' (Roche Diagnostics). The sensitivity and specificity of targeted NGS assay were evaluated using the biosynthetic and biological DNA reference material with known allelic frequencies (VAF) of EGFR variants. NGS demonstrated a sufficient lower detection limit for diagnostic applications (VAF < 5%) in DNA reference material; all EGFR variants were correctly identified. NGS showed high repeatability of VAF assessment between runs (CV% from 0.02 to 3.98). In clinical material, the overall concordance between NGS and qPCR was 76.14% (Cohen's Kappa = 0.5933). The majority of discordant results concerned false-positive detection of EGFR exon 20 insertions by qPCR. A total of 9 out of 59 (15%) clinical samples showed discordant results for one or more EGFR variants in both assays. Additionally, we observed TP53 to be a frequently co-mutated gene in EGFR-positive NSCLC patients. In conclusion, targeted NGS showed a number of superior features over qPCR in EGFR variant detection (exact identification of variants, calculation of allelic frequency, high analytical sensitivity), which might enhance the basic diagnostic report.

Myxoid liposarcoma diagnosed on fine needle aspiration cytology: There is more to it than meets the eye.

Liposarcoma is a rare mesenchymal neoplasm commonly involving deep soft tissues and the retroperitoneum. Among the various types of liposarcoma, myxoid liposarcoma is the most frequently encountered in adolescents and young adults, with a predilection for lower extremities. Fine needle aspiration allows easy assessment and rapid on-site evaluation for distinguishing benign from malignant lipomatous lesions. Here, we present a case of myxoid liposarcoma in the calf region of a 19-year-old boy, diagnosed via fine needle aspiration cytology, and subsequently confirmed by histopathological examination after surgical resection. The intention behind this case report is to highlight the cytological features of myxoid liposarcoma and to improve understanding of this tumor entity, aiming to prevent misdiagnosis by inexperienced pathologists when evaluating cytology specimens.

Cytomorphologic comparison of upper urinary tract urothelial carcinomas and renal cell carcinomas on urine cytology.

Compared to urothelial carcinomas (UCs), the cytomorphology of renal cell carcinomas (RCCs) is underdescribed. This study aims to investigate whether UCs and RCCS of the upper urinary tract can be differentiated cytologically, and to identify distinguishing cytomorphological features. Consecutive urine cytology specimens with atypical/C3, suspicious/C4 or malignant/C5 diagnoses matched with a nephrectomy or ureterectomy specimen with UC or RCC over a 15-year period were reviewed for cellularity, architecture, background composition and cytomorphologic features. Totally 132 specimens were retrieved, comprising 24 RCCs and 108 UCs. Clear cell RCC (CCRCC) (n = 18) was the most common RCC. Urine cytology specimens from UC showed a trend of higher cellularity (p = 0.071) against RCC and was significant in subgroup analysis with CCRCC (p < .001). Epithelial structures in sheets, tubules, and papillae were exclusive in specimens of UC (p < .05). For background features, squamous cells were more common for RCC (p = .006) including CCRCC (p = .003), whereas polymorphs (p = .011) and necrotic material (p = .010) were associated with UC. Average nuclear size was larger and nuclear size variation (p < .001) and nuclear-cytoplasmic ratio (p = .001) were greater in UC (p = .001) than RCC. Comparing RCC to high-grade UCs only, nuclear-cytoplasmic ratio maintained statistical significance (p = .006) while average nuclear size showed a trend (p = .063). A clean background free of tumor necrosis and polymorphs, and the lack of complex tumor fragments favors RCC. UCs also display larger nuclear size, higher nuclear size variation and nuclear-cytoplasmic ratio. These cytomorphological features with corroboration of clinical/radiological findings, can aid in raising a diagnosis of RCC.

Methodological and TNM Focus-Based Comparison of EGFR Mutation Status in Non-Small-Cell Lung Carcinomas.

Epidermal growth factor receptor (EGFR) mutations in non-small-cell lung carcinomas (NSCLC) are a frequent class of driver mutations, and tyrosine kinase inhibitor (TKI) therapy provides considerable clinical benefits. Using the most effective and also easiest method for EGFR analysis is cost-effective and time-saving. In this study, we aimed to determine which method could be more effective by comparing the incidences of EGFR mutations in cytological and histological samples which were obtained by different methods also, whether there was a difference in the incidences of EGFR mutations between the primary foci, mediastinal lymph nodes, and distant metastatic foci. We retrospectively reviewed 420 cases of cytological materials, small biopsies, and surgical samples reported as NSCLC underwent EGFR analysis in our department between 2016 and 2022. We collected the data and interpreted the results from two different perspectives. We identified 36 EGFR mutations in 362 biopsies (9.94%) and 17 in 58 cytology samples (29.31%). There is a significant difference between the two methods (P = 0.01*). We observed 38 EGFR mutations in 320 primary foci (11.87%), 7 EGFR mutations in 36 mediastinal or subcarinal lymph nodes (19.44%), and 8 EGFR mutations in 64 distant metastatic foci (12.50%). A significant difference was also observed in pleural samples (P = 0.005*). We observed more successful results with cell blocks obtained from liquid-based cytological specimens than with formalin-fixed, paraffin-embedded tissues obtained from resection or otherwise in our clinical routine. Our study results highlight the benefits of cytological specimens in molecular treatments and current therapy modalities.

The effect of carbomer versus noncarbomer lubricant on the adequacy of cervical cytology specimens

Cervical cytology remains a critical screening tool for cervical cancer. While various factors can influence cytology quality, the effect of lubricant type used during specimen collection has been previously studied with inconclusive results. This study aimed to evaluate the impact of surgical lubricant on cervical cytology results and elucidate risk factors associated with unsatisfactory results. We hypothesized that switching from a carbomer-containing lubricant to a noncarbomer, water-soluble lubricant would improve specimen adequacy in cervical cytology. A retrospective chart review was performed examining patient cytologic results from January to December 2017 at a single academic institution. After historical rates of unsatisfactory cytology were higher than acceptable standards, the practice changed lubricant formulation from a carbomer containing lubricant to a noncarbomer, water soluble lubricant. Demographic data and treatment characteristics were collected for eligible patients. Matched analysis was performed to examine factors associated with an unsatisfactory cytology result. After the change in lubricant, there was a significant decline in the rates of unsatisfactory cytology from 9.6% to 5.7%, P = 0.01. This decline was also observed when patients were matched based on menopausal status, personal history of gynecologic malignancy, pregnancy status, and cytology specimen type (10.0% to 4.8%, P = 0.001). Change in lubricant from a carbomer containing to noncarbomer, water soluble product was associated with a statistically significant decline in the rates of unsatisfactory cytology. Although prior data have had mixed results as to the etiology of unsatisfactory cytology, we feel that this directly contributed to the high rates observed at our institution.

Utility of an anchored multiplex polymerase chain reaction‐based fusion assay for diagnosis of soft tissue tumors in cytology

AbstractBackgroundFine‐needle aspiration specimens from soft tissue tumors are complicated by lack of tissue architecture and limited material for ancillary testing. There are little data on the feasibility of next‐generation sequencing techniques for fusion detection on soft tissue cytology specimens. This study explored the role of an anchored multiplex polymerase chain reaction (PCR)‐based gene fusion assay in aiding the diagnosis of mesenchymal neoplasms on cytology samples.MethodsThe laboratory information system was queried for cytology specimens that had undergone testing by anchored multiplex PCR. After exclusion of epithelial and hematolymphoid neoplasms, clinical and pathologic information was collected on the remaining cases.ResultsThere were 1609 cytology specimens tested with anchored multiplex PCR. Of these, 48 (3%) were cytology specimens from mesenchymal tumors. Anchored multiplex PCR was positive for a reportable fusion transcript in 14 of 48 cases (29%); there was no fusion detected in 32 cases (67%), and there was insufficient tissue for analysis in two cases (4%). The detectable fusion partners included ALK (n = 4), STAT6 (n = 4), EWSR1 (n = 3), and one each of SS18, YAP1, and PHF1. Of the cases in which a fusion partner was detected, eight of 14 were disease‐defining on cytology preparation, and six of 14 provided molecular confirmation of a metastatic focus of a previously diagnosed tumor.ConclusionsThe anchored, multiplex PCR‐based gene fusion assay is a powerful orthogonal tool in helping diagnose mesenchymal neoplasms on cytology specimens. The material obtained for cytologic analysis yields sufficient quality/quantity of tissue in the majority of cases tested.

The Comparison of Real-Time PCR and Mutation-Specific Immunohistochemistry in EGFR Mutation Analysis of Non-Small Cell Lung Carcinomas

Objective: This study aims to identify activating mutations in the epidermal growth factor receptor (EGFR) gene in patients with non-small cell lung cancer (NSCLC) and to evaluate their correlation with responses to EGFR-tyrosine kinase inhibitors (TKI) treatment. This study aims to identify activating mutations in the epidermal growth factor receptor (EGFR) gene in patients with non-small cell lung cancer (NSCLC) and to evaluate their correlation with responses to EGFR-tyrosine kinase inhibitors (TKI) treatment. We conducted a comparative analysis of Real-Time PCR and immunohistochemistry to detect EGFR mutation status in non-small cell lung cancer patients, focusing on the sensitivity, specificity, and predictive values of immunohistochemistry. Material and Method: We evaluated 788 non-small cell lung cancer samples which were analyzed for EGFR mutation status by RT-PCR. We detected 126 EGFR mutated cases among these patients. We evaluated mutation-specific EGFR immunohistochemistry directed towards the exon 19 deletions (15 bp E746-A750) and exon 21 point mutation (L858R) to the 47 EGFR mutated patients histologic material and cell blocks of cytologic specimens. Results: 32 of the 47 cases (68%) had exon 19 deletion, 14 of them (30%) had point mutation in exon 21, and one of them (2%) showed exon 18 mutation. EGFR exon 19 (15 bp E746-A750 deletion) antibody showed a sensitivity of 100%, specificity of 40%, negative predictive value of 100%, and positive predictive value of 78%. The sensitivity of the exon 21 (L858R point mutation) antibody was 93%, specificity was 91%, negative predictive value was 97% and positive predictive value was 82%. Conclusion: Our investigation indicates that mutation-specific EGFR immunohistochemistry has demonstrated a notable sensitivity and specificity for exon 21. However, while sensitive, the exon 19 (15 bp E746-A750 deletion) antibody lacked specificity. While positive immunohistochemical staining may suggest the presence of an EGFR mutation, making the patient potentially eligible for TKI treatment, it should not be the sole determinant. If immunohistochemistry results are negative, it is essential to resort to molecular tests to ensure accurate diagnosis and appropriate therapeutic guidance. With evolving diagnostic landscapes, it is crucial to harness both IHC and molecular techniques judiciously for optimal patient care.

Prediction of the Aggressive Clinical Course of Papillary Thyroid Carcinoma Based on Fine Needle Aspiration Biopsy Molecular Testing.

Molecular genetic events are among the numerous factors affecting the clinical course of papillary thyroid carcinoma (PTC). Recent studies have demonstrated that aberrant expression of miRNA, as well as different thyroid-related genes, correlate with the aggressive clinical course of PTC and unfavorable treatment outcomes, which opens up new avenues for using them in the personalization of the treatment strategy for patients with PTC. In the present work, our goal was to assess the applicability of molecular markers in the preoperative diagnosis of aggressive variants of papillary thyroid cancer. The molecular genetic profile (expression levels of 34 different markers and BRAF mutations) was studied for 108 cytology specimens collected by fine-needle aspiration biopsy in patients with PTC having different clinical manifestations. Statistically significant differences with adjustment for multiple comparisons (p < 0.0015) for clinically aggressive variants of PTC were obtained for four markers: miRNA-146b, miRNA-221, fibronectin 1 (FN1), and cyclin-dependent kinase inhibitor 2A (CDKN2A) genes. A weak statistical correlation (0.0015 < p < 0.05) was observed for miRNA-31, -375, -551b, -148b, -125b, mtDNA, CITED1, TPO, HMGA2, CLU, NIS, SERPINA1, TFF3, and TMPRSS4. The recurrence risk of papillary thyroid carcinoma can be preoperatively predicted using miRNA-221, FN1, and CDKN2A genes.