Fine-needle Aspiration Samples Research Articles

High Diagnostic Accuracy of Epigenetic Imprinting Biomarkers in Thyroid Nodules.

To explore the novel diagnostic value of epigenetic imprinting biomarkers in thyroid nodules. A total of 550 patients with fine-needle aspiration (FNA)-evaluated and histopathologically confirmed thyroid nodules were consecutively recruited from eight medical centers. Quantitative chromogenic imprinted gene in situ hybridization (QCIGISH) was used to assess the allelic expression of imprinted genes SNRPN and HM13, on the basis of which a diagnostic grading model for thyroid nodules was developed. The model was retrospectively trained on 124 postsurgical thyroid samples, optimized on 32 presurgical FNA samples, and prospectively validated on 394 presurgical FNA samples. Blinded central review-based cytopathologic and histopathologic diagnoses were used as the reference standard. For thyroid malignancy, the QCIGISH test achieved an overall diagnostic sensitivity of 100% (277/277), a specificity of 91.5% (107/117; 95% CI, 86.4 to 96.5), a positive predictive value (PPV) of 96.5% (95% CI, 94.4 to 98.6), and a negative predictive value (NPV) of 100% in the prospective validation, with a diagnostic accuracy of 97.5% (384/394; 95% CI, 95.9 to 99.0). QCIGISH demonstrated a PPV of 97.8% (95% CI, 94.7 to 100) and NPV of 100%, with a diagnostic accuracy of 98.2% (111/113; 95% CI, 95.8 to 100), for indeterminate Bethesda III-V thyroid nodules. QCIGISH demonstrated a PPV of 96.6% (95% CI, 91.9 to 100) and a NPV of 100%, with a diagnostic accuracy of 97.5% (79/81; 95% CI, 94.2 to 100), for Bethesda III-IV. For Bethesda VI, QCIGISH showed a 100% (184/184) accuracy. This imprinting biomarker-based test can effectively distinguish malignant from benign thyroid nodules. The high PPV and NPV make the test both an excellent rule-in and rule-out diagnostic tool. With such a diagnostic performance and its technical simplicity, this novel thyroid molecular test is clinically widely applicable.

FNA as a Powerful Modality to Detect Granular Cell Tumor in Male Breast: A Report and Review of Literature

Abstract Introduction/Objective Granular cell tumor (GCT) is a benign mesenchymal tumor of Schwannian origin. Granular cell tumor is an uncommon tumor of the breast with a prevalence of 1:1000 to 6.7:1000 of all breast tumors. Most cases are diagnosed on surgical resection. When reported that are most commonly in middle-aged premenopausal women. Diagnosis of GCT on fine needle aspiration (FNA) in male patients is, however, extremely rare. We present here an unusual case of GCT of the breast in a male patient clinically mimicking malignancy. Methods/Case Report A 34-year-old male patient presented with a palpable lump in the left breast. An imaging study showed a 0.6 cm hypoechoic mass with microlobulated borders and no internal vascularity suspicious for breast carcinoma (BIRAD-4). Fine needle aspiration (FNA) of the mass was performed. Rapid onsite specimen evaluation was performed and additional passes were performed to collect sample for cell block to perform ancillary studies. FNA revealed clusters of polygonal cells with abundant eosinophilic granular cytoplasm and poorly defined borders. Those cells had round to ovoid nuclei with vesicular chromatin pattern and prominent nucleoli. A differential diagnosis of GCT, apocrine metaplasia, and lesions rich in histiocytes such as granulomatous inflammation, fat necrosis and duct ectasia were considered. Immunohistochemical staining with S100 and CD68 was performed which stained granular cytoplasm in tumor cells of GCT. A review of literature from the past 5 years shows 23 cases of GCT of the breast are reported in the literature and of these only 3 (13%) are reported in males. These tumors ranged from 20-68 years with tumor size ranging from 5- 28 mm. Only 1(4%) cases were preoperatively diagnosed using FNA. No cases of GCT were preoperatively diagnosed on FNA samples from the breast. Results (if a Case Study enter NA) N/A. Conclusion We show here that a powerful relatively noninvasive and rapid modality like FNA can be effectively utilized to detect very rare benign tumor like GCT in the male breast where imaging studies suspect breast cancer.

Cytopathology Practice in the Covid-19 Pandemic, During the Lockdown and Post-lockdown Period: A Tertiary-Care Center Experience

Background: Covid-19 pandemic has changed the healthcare delivery and cytopathology practice worldwide. We evaluated our cytopathology laboratory workload during the lockdown and post-lockdown period and compared it with the same period in 2019, to see the impact of the Covid-19 pandemic. Methods: The cytological reports issued during the lockdown (10 March - 31 May 2020) and the post-lockdown period (1 June -31 August 2020); and the corresponding periods in 2019 were retrospectively reviewed from the database. Sample type, sampling site, and diagnostic categories were recorded. Results: During the Covid-19 lockdown period, the total number of cytological specimens, was reduced from n=3197 to n=745, with a rate of 76,7%. The most reduction was in thyroid fine-needle aspiration (FNA) and cervicovaginal smears. Relative increases were observed for soft tissue, lung, and liver FNA samples (p<0.05) and cerebrospinal fluid, peritoneal and pleural fluid samples (p<0.05). In the post-lockdown period, the total number reduced from n=2461 to n=2032 with a rate of 17.4%. Significant reduction continued for thyroid FNAs, but other samples have nearly reached the pre-covid levels. During the total six months period, the rate of the malignant category increased while the negative for malignancy category decreased compared to 2019. Conclusion: During the Covid-19 lockdown period, the reduction was primarily observed in the samples taken for screening purposes, and high-risk oncological patients continued to receive healthcare services. In the initial phase of the post-lockdown period, health services and cytopathology practice have rapidly reached almost the levels of the pre-pandemic period.

ODP494 Mediastinal Ectopic Thyroid Papillary Carcinoma with NTRK3 Fusion, Presenting with Thoracic Spine Metastasis

Abstract Background Ectopic thyroid tissue may be encountered anywhere from the base of the skull to the lower neck. It is rarely seen in the mediastinum. Thyroid cancer in ectopic thyroid tissue is a very rare entity. We report a patient with papillary thyroid cancer arising from upper mediastinal ectopic thyroid tissue. The patient presented with thoracic spine metastasis with cord compression. Molecular testing showed SQSTM1-NTRK3chromosomal rearrangement. Case Presentation The patient is a 67 -year-old female, presented with upper back pain. She reported no weakness or sensory changes. MRI showed suspected metastatic disease to the second and third thoracic vertebrae (T2 and T3). Therewasepidural extension of tumor with cord compression and severe stenosis at T2. She underwent laminectomy and decompression surgeryat T1-T3 level. Final pathology report showed Metastatic thyroid carcinoma with papillary features. She underwent external beam radiation to the affected spines. CT chest abdomen and pelvis showed 3. 0×2.8×2.3 cm soft tissue mass in the left superior mediastinum extending into the supraclavicular region. FDG-PET scan showedHypermetabolic foci in the upper mediastinum with SUV max of 25. Fine needle Aspiration (FNA) of the upper mediastinal mass was consistent with papillary thyroid cancer. Molecular testing from the FNA sample using Thyroseq V3 showed SQSTM1-NTRK3 chromosomal rearrangement. Total thyroidectomy was performed; however, the upper mediastinal mass was not resected due to proximity to vascular and neuronal structures. Pathology of the resected thyroid was benign. Recombinant TSH stimulatedI-123 scan showed intense uptake in the neck area, most likely represents remnant thyroid tissue. No clear uptake in the mediastinal mass or spine metastasis. She underwent removal of the upper mediastinal mass with lymph node dissection at a referral center. Pathology showed soft tissue involved by papillary thyroid carcinoma with focal tall cell features, forming a 4×2.5×2.5 cm mass. Surgery was followed by ablation with 100 mci of radioactive iodine (I-131). Thyroglobulin level decreased from a baseline of 15272 ng/ml to 1857 ng/ml, post-surgery, external beam radiation and radioactive iodine ablation. Conclusion This case highlights the possibility of developing thyroid cancer in ectopic thyroid tissue and the unique challenges in the diagnosis and management of such rare disease. Our patient has very aggressive disease presentation that required multimodality treatment including thoracic spine decompression, total thyroidectomy, mediastinal ectopic thyroid cancer resection, external beam radiation to the affectedspine area, and high dose radioactive iodine treatment. The use of molecular diagnostics is very important to improve the outcome in such patient. The tumor has SQSTM1-NTRK3 chromosomal rearrangement. This rearrangement causesconstitutive activation of the NTRK3 in the Ras-Raf-MEK-ERK Pathway. This fusioncan be targeted by medications like Larotrectinib and Endtrectinib. Presentation: No date and time listed

ODP465 BRAF V600 and NRAS Q61 mutations in thyroid fine-needle aspiration (FNA) with indeterminate cytology in Argentina

Abstract It is known that thyroid cancer initiation and progression occurs because of gradual accumulation of genetic and epigenetic alterations at cell level. Many mutations associated with this disease have been reported to be present in genes encoding proteins in the mitogen-activated protein kinase (MAPK) signaling pathway, specifically BRAF and RAS genes, as well as in the PI3K/Akt signaling pathway, among others. In particular, molecular alterations in follicular cells in the BRAF or NRAS genes have been reported to be associated with the process of carcinogenesis. Previously we found BRAF and NRAS mutations are observed in a significant number of papillary and follicular thyroid carcinomas in our population, respectively. In this study, we tested the diagnostic performance of BRAF and NRAS mutation in thyroid carcinoma in fine-needle aspiration (FNA) specimens with indeterminate cytology. In total, residual material from 73 FNA samples was used (n= 16, Bethesda III; n=48, Bethesda IV and n=9 Bethesda V). BRAF codon 600 mutation and NRAS codon 61 mutations were investigated using qPCR and melting curve analysis was carried out. Diagnosis of malignancy was confirmed by histology on paired surgical specimen in 54 cases. Results In the cytologically indeterminate categories, 9.6% of specimens were BRAF V600 mutated: 1 of 16 were subcategorized as atypia of undetermined significance or follicular lesion of undetermined significance (6.3%); 2 of 48 as follicular neoplasm or suspicious for follicular neoplasm (4.1%); and 4 of 9 as suspicious for malignancy (44.4%). NRAS Q61 mutation was found in 4 patients (2 with Bethesda IV and 2 with cytology V). The BRAF V600 mutations were associated with carcinoma in 100% of cases (n = 7), whereas only 75% (n = 3) of the nodules with NRAS were associated with carcinoma. BRAF V600 and NRAS Q61 mutations had high specificity in predicting malignant carcinoma (96.8%); the sensitivity was 43.5% with a positive predictive value of 90.9% and a negative predictive value of 69.8%. In specific categories of indeterminate cytology, the pre-test risk of malignancy was 38%, 33% and 88% while the post molecular test risk of malignancy was 100%, 75% and 100% respectively. Conclusions BRAF V600 and NRAS Q61 molecular test in residual FNA samples improve diagnostic accuracy of the cytology analysis in our population and may help to better select patients for avoid inadequate or unnecessary surgeries. Presentation: No date and time listed

KRAS Mutation Testing on Endoscopic Ultrasound-Guided Fine-Needle Aspiration Samples Improves the Diagnostic Accuracy of Pancreatic Cancer.

Endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) cytology was one of the investigations for pancreatic masses. While the specificity approached 100%, its sensitivity remained low because of high rate of indeterminate and false-negative results. Meanwhile, KRAS gene was frequently mutated in up to 90% of pancreatic ductal adenocarcinoma and its precursor lesions. This study aimed to determine whether KRAS mutation analysis could improve the diagnostic sensitivity in EUS-FNA samples for pancreatic adenocarcinoma. The EUS-FNA samples from patients with a pancreatic mass obtained between January 2016 and December 2017 were reviewed retrospectively. The cytology results were classified as malignant, suspicious for malignancy, atypical, negative for malignancy, and nondiagnostic. KRAS mutation testing was performed using polymerase chain reaction followed by Sanger sequencing. A total of 126 EUS-FNA specimens were reviewed. The overall sensitivity and specificity by cytology alone were 29% and 100%, respectively. When KRAS mutation testing was performed in cases with indeterminate and negative cytology, the sensitivity increased to 74.2%, and the specificity remained at 100%. KRAS mutation analysis, especially when performed in cytologically indeterminate cases, improves the diagnostic accuracy for pancreatic ductal adenocarcinoma. This may reduce the need to repeat invasive EUS-FNA for diagnosis.

30P Molecular analysis of bone sarcomas by cytogenetic studies on fine needle aspiration samples

Generally tissue samples from histopathology or open biopsy are used for cytogenetic analysis and only few studies have used tissue samples from Fine Needle aspiration for chromosomal analysis. To start neoadjuvant chemotherapy in bone sarcoma patients, we need final diagnosis. Open biopsy has its own disadvantages. FNAC and cytological examination can give valuable information about the tumor. Apart from immunocytochemistry and electron microscopic examination of aspirated tissue, chromosomal analysis has also become a tool in diagnosing bone sarcoma.

Cobas HPV Genotyping of FNA Sample Supernatant and Frozen Section Scrapings of Suspected Head and Neck Cancer.

The human papillomavirus (HPV) status of squamous cell carcinomas (SCCs) of the head and neck is relevant for therapy planning, staging, and follow-up. Immunohistochemistry (IHC) for p16 is a surrogate marker of HPV status in oropharyngeal SCC, but not at other head and neck sites. We tested if the cobas HPV test was feasible and superior to p16-IHC on fine-needle aspiration (FNA) supernatants and frozen section (FS) scrapings of suspected SCC. A 500μL aliquots of postcentrifugation supernatant CytoRich Red media of FNA cellblock specimens and scrapings of FS suspended in SurePath media vials were tested with the cobas HPV test and compared with p16-IHC and/or HPV in situ hybridization (ISH) performed on cellblock and/or resections. Twenty-nine (n=29) FNAs were tested for a cobas HPV test, p16, and/or HPV-ISH. The mean collection to testing time was 6.3 days (range: 0 to 24d). Cobas yielded valid results in all cases; p16-IHC could not be interpreted in 4 (13%) cellblocks; correlation was performed on subsequent resections. Cohen κ correlation for cobas versus p16-IHC/HPV-ISH on FNA samples was 0.9, perfect agreement, sensitivity 100%, specificity 92.3%, positive predictive value 94.1%, negative predictive value 100%. Thirty-four (n=34) scrapings from FS were tested for cobas, p16, and/or HPV-ISH. The mean collection to testing time was 10.4 days (range: 1 to 28d). Cohen κ correlation for cobas versus p16-IHC/HPV-ISH on FS scrapings was 1, perfect agreement. Sensitivity, specificity, positive predictive value, and negative predictive value was 100%. Cobas genotype was HPV-16 in 87%, HPV-18 in 3%, and HPV-other in 10%. Cobas HPV test in FNA supernatant and FS scrapings outperformed or was equivalent to p16-IHC and provided genotyping information.

Follicular Neoplasm of Thyroid Revisited: Current Differential Diagnosis and the Impact of Molecular Testing.

The diagnosis of "follicular neoplasm" (FN) in thyroid cytopathology has a long history that originated not long after the practice of fine-needle aspiration (FNA) of thyroid nodules. From the outset, this interpretive category was intended to convey a set of differential diagnoses rather than a precise diagnosis, as key diagnostic features, such as capsular and vascular invasion, were not detectable on cytology preparations. Cytologic-histologic correlation studies over the past several decades have shown that FN interpretation can be applied to the spectrum of nonneoplastic tumors to carcinomas. Most tumors classified as FN include follicular adenoma, follicular carcinoma, noninvasive follicular thyroid tumor with papillary-like nuclear features, and follicular variant of papillary thyroid carcinoma. Less common entities that may be classified as FN on FNA include hyalinizing trabecular tumor (HTT), poorly differentiated thyroid carcinoma, medullary carcinoma, and nonthyroidal lesions such as parathyroid tissue, paraganglioma, and metastatic tumors. Advances in our ability to detect characteristic molecular alterations (eg, GLIS gene rearrangements for hyalinizing trabecular tumor) in FNA samples may assist in the identification of some of these entities. In this review, we summarize the pathophysiology, history, and evolution of the terminology and the current differential diagnosis according to the recently published 2022 World Health Organization classification, molecular testing, and management of nodules classified as FN.

Clinical Use of Raman Spectroscopy Improves Diagnostic Accuracy for Indeterminate Thyroid Nodules

Molecular analysis of thyroid fine-needle aspiration (FNA) specimens is believed to improve the management of indeterminate nodules. Raman spectroscopy (RS) can differentiate benign and malignant thyroid lesions in surgically removed tissues, generating distinctive structural profiles. Herein, the diagnostic performance of RS was tested on FNA biopsies of thyroid gland. Prospective, blinded, and single-center study. We enrolled 123 patients with indeterminate or more ominous cytologic diagnoses (TIR3A-low-risk indeterminate lesion, TIR3B-high-risk indeterminate lesion, TIR4-suspicious of malignancy, TIR5-malignant). All subjects were surgical candidates (defined by international guidelines) and submitted to FNA procedures for RS analysis. We compared RS data, cytologic findings, and final histologic assessments (as reference standard) using various statistical techniques. The distribution of our study population was as follows: TIR3A:37, TIR3B:32, TIR4:16, and TIR5:38. In 30.9% of patients, histologic diagnoses were benign. For predicting thyroid malignancy in FNA samples, the overall specificity of RS was 86.8%, with 86.5% specificity in indeterminate cytologic categories. In patients with high-risk ultrasound categories, the specificity of RS increased to 87.5% for TIR3A, reaching 100% for TIR3B. Benign histologic diagnoses accounted for 72.9% of patients classified as TIR3A and 31.3% of those classified as TIR3B. Based on positive RS testing, unnecessary surgery was reduced to 7.4% overall (TIR3A-33.3%, TIR3B-6.7%). This premier use of RS for thyroid cytology confirms its role as a valuable diagnostic tool and a valid alternative to molecular studies, capable of improving the management of indeterminate nodules and reducing unnecessary surgery.

Diagnosis of parathyroid incidentaloma detected on thyroid ultrasonography: the role of fine-needle aspiration cytology and washout parathyroid hormone measurements.

The aim of this study was to assess the diagnostic role of fine-needle aspiration cytology (FNAC) and analyze factors associated with false-negative FNAC results in patients with parathyroid incidentaloma who were referred for ultrasonography (US)-guided fine-needle aspiration (FNA) of thyroid nodules. In this study, 121 patients with suspected parathyroid lesions were enrolled. The patients underwent US-guided FNAC with measurements of washout parathyroid hormone (PTH) between January 2015 and May 2020. The diagnostic performance of FNAC for the diagnosis of parathyroid lesions was assessed using surgical results and elevated washout PTH as a reference standard. The clinical and radiologic features associated with false-negative results on FNAC for the diagnosis of parathyroid lesions were evaluated. Among the 121 nodules assessed, 38 were parathyroid lesions (31.4%), and 83 were non-parathyroid lesions (68.6%). The diagnostic performance of FNAC for parathyroid incidentaloma showed a sensitivity of 31.6% (12/38), specificity of 100% (83/83), positive predictive values of 100% (12/12), negative predictive values of 76.1% (83/109), and accuracy of 78.5% (95/121). The FNAC results of non-parathyroid lesions included thyroid nodules, lymph nodes, neurogenic tumors, and fat tissue. True-positive results on FNAC were significantly associated with performing FNA twice (58.3% vs. 23.1%, P=0.043). Considering the low sensitivity of FNAC, measuring washout PTH in addition to FNAC may help accurately diagnose parathyroid incidentaloma on thyroid US. Further, the falsenegative rate for FNAC can be reduced by obtaining two or more FNA samples.

Use of long non-coding RNAs for the molecular diagnosis of papillary thyroid cancer.

ObjectiveImproved molecular testing for common somatic mutations and the identification of mRNA and microRNA expression classifiers are promising approaches for the diagnosis of thyroid nodules. However, there is a need to improve the diagnostic accuracy of such tests for identifying thyroid cancer. Recent findings have revealed a crucial role of long non-coding RNAs (lncRNAs) in gene modulation. Thus, we aimed to evaluate the diagnostic value of selected lncRNAs from The Atlas of Noncoding RNAs in Cancer (TANRIC) thyroid cancer dataset.MethodsLncRNAs in TANRIC thyroid cancer dataset that have significantly increased or decreased expression in papillary thyroid cancer (PTC) tissues were selected as candidates for PTC diagnosis. Surgical specimens from patients who underwent thyroidectomy were used to determine the separation capability of candidate lncRNAs between malignant and benign nodules. Fine needle aspiration samples were obtained and screened for candidate lncRNAs to verify their diagnostic value.ResultsLRRC52-AS1, LINC02471, LINC02082, UNC5B-AS1, LINC02408, MPPED2-AS1, LNCNEF, LOC642484, ATP6V0E2-AS1, and LOC100129129 were selected as the candidate lncRNAs. LRRC52-AS1, LINC02082, UNC5B-AS1, MPPED2-AS1, LNCNEF, and LOC100129129 expression levels were significantly increased or decreased in malignant nodules compared to those in benign nodules and paired normal thyroid tissues. The combination of LRRC52-AS1, LINC02082, and UNC5B-AS1 showed favorable results for the diagnosis of PTC from fine needle aspirates, with 88.9% sensitivity and 100.0% specificity.ConclusionsLncRNA expression analysis is a promising approach for advancing the molecular diagnosis of PTC. Further studies are needed to identify lncRNAs of additional diagnostic value.

High-risk human papillomavirus testing in cytology aspiration samples from the head and neck part 1: a review of the literature on available testing options.

Human papillomavirus (HPV)-associated oropharyngeal squamous cell carcinoma is increasing in incidence and is often first diagnosed on a cytology fine needle aspiration (FNA) specimen of metastatic nodal disease of the neck. In the setting of oropharyngeal squamous cell carcinoma, HPV status defines the disease with HPV-associated tumors having better overall prognosis than those that are HPV negative. Furthermore, metastatic squamous cell carcinoma of the neck of unknown origin requires testing for HPV as a positive result suggests an oropharyngeal primary. As a result, HPV testing in aspirate samples is increasingly important for the proper diagnosis and treatment of patients with head and neck squamous cell carcinoma. Although HPV testing in cervicovaginal cytology specimens is common and well-established, testing in head and neck FNA samples remains challenging. p16 immunohistochemistry is an excellent surrogate marker for HPV in tumors of known or suspected oropharyngeal origin, but the criteria used in histologic specimens may not be appropriate in cytology samples. FNA samples are more frequently hypocellular, and cytology cell blocks have variable fixation and processing steps, limiting the utility of p16 immunohistochemistry. Other potential testing options have been reported in the literature including staining of aspirate smears and molecular testing of liquid-based samples. The American Society of Cytopathology Clinical Practice Committee recently surveyed the American Society of Cytopathology membership to determine the current state of HPV testing in aspirate samples, and this review article is designed to provide a summary of the current literature on various testing options in FNA samples.

High-risk human papillomavirus testing in cytology aspiration samples from the head and neck part 2: a survey of the American Society of Cytopathology community.

High-risk human papillomavirus (HR-HPV) status is critical in the diagnosis of oropharyngeal squamous cell carcinoma, informing prognosis and choice of therapy. HR-HPV status additionally plays a key role in the evaluation of squamous cell carcinoma of unknown origin metastatic to cervical lymph nodes. Thus, HR-HPV testing of fine needle aspirate (FNA) specimens from the head and neck is invaluable for accurate diagnosis, prognostication, and treatment planning. American Society of Cytopathology members were surveyed to understand the current state of HR-HPV testing on FNA samples from the head and neck. The survey focused on 3 main topic areas: practice setting of respondents, methods of collection and processing of aspirate specimens for HR-HPV testing, and validation of HR-HPV testing methodologies on aspirate samples. The survey reveals that laboratories employ various methods to detect HR-HPV in FNA samples, most commonly p16 immunohistochemical staining of cell block sections. Although some laboratories have independently validated their HR-HPV detection method, such validation is not universal. Finally, not all respondents currently have HR-HPV testing available, but approximately half of those without a testing method desire to make HR-HPV testing of FNA samples available. Survey responses highlight that various testing modalities are utilized for HR-HPV detection in aspirate samples. However, internal laboratory validation of HR-HPV testing for FNA specimens is not ubiquitous despite professional society recommendations.

Performance of a Multigene Genomic Classifier in Thyroid Nodules with Suspicious for Malignancy Cytology.

Background: Molecular testing is increasingly used to refine the probability of cancer and assess recurrence risk in thyroid nodules with Bethesda III/IV fine needle aspiration (FNA) cytology. However, limited data exist for Bethesda V (suspicious for malignancy [SFM]) samples. This study evaluated the performance of ThyroSeq v3 (TSv3) in thyroid nodules with SFM cytology. Methods: In this single-institution retrospective cohort study, consecutive thyroid FNA samples diagnosed as SFM with TSv3 testing and known surgical outcome were identified. Clinical, pathology, and molecular findings were reviewed. The TSv3 Cancer Risk Classifier was used to determine molecular risk groups (MRGs). For test-negative cases diagnosed as cancer/noninvasive follicular thyroid neoplasm with papillary-like nuclear features, TSv3 was performed on the resected tumors. Results: Among 128 SFM samples studied, 100 (78.1%) were TSv3 positive, and 28 (21.9%) were negative. The cancer prevalence on surgery was 82.8%. Among test-positive samples, 95% were malignant and 5% benign. Among test-negative samples, 17 (60.7%) were benign and 11 (39.3%) malignant. Overall, TSv3 had a sensitivity of 89.6% (confidence interval; CI 82.4-94.1) and a specificity of 77.3% (CI 56.6-89.9). For a cancer prevalence of 50-75% expected in SFM cytology by the Bethesda system, the negative predictive value was expected to range from 71.2% to 88.1% and the positive predictive value from 79.8% to 92.2%. Among test-positive nodules, 20% were MRG-Low (mostly RAS-like alterations), 66% MRG-Intermediate (mostly BRAF-like alterations), and 14% MRG-High. Among patients with cancer, 65 (61.3%) were American Thyroid Association low risk, 25 (23.6%) intermediate risk, and 6 (5.7%) high risk. During the mean follow-up of 51.2 months (range: <1 to 470 months), 12 (13.0%) patients had disease recurrence, which was more common in MRG-High (54.6%) compared with MRG-Intermediate (9.5%) and MRG-Low (0%) cancers (p < 0.001). Upon reexamining tumors with false-negative results, half of evaluable cases had alterations likely missed due to limiting FNA sampling, and the remainder represented low-risk tumors. Potentially targetable alterations were identified in 10 samples. Conclusions: In this large series of SFM thyroid nodules, TSv3 further improved cancer prediction and detected RAS-like, BRAF-like, high-risk, and potentially targetable alterations, all of which may inform more optimal patient management. MRGs were associated with recurrence-free survival, offering potential preoperative cancer risk stratification.

The Value of DPAS in the Detection of Breast Malignant Cells

Background: Fine-needle aspiration (FNA) has become an essential, critical test for breast masses. This study aimed to determine the value of diastase-resistant periodic acid-Schiff (DPAS) staining in the detection of malignant breast cells. Methods: This prospective cross-sectional study was conducted in Khartoum state (Sudan) among Sudanese women who suffered from breast lumps. FNA samples were collected from each patient, and the material was simultaneously smeared onto two labeled glass slides. The DPAS score and aspiration cytology (AC) grade are expressed as mean +/- SD, and the 95% confidence intervals of the means were calculated. Results: The findings revealed the following DPAS score frequencies among the studied women: negative (+/-) (28, 13.9%), one plus (+) (114, 56.7%), two plus (++) (27, 13.4%), and three-plus (+++) (32, 15.9%). Comparison of DPAS scores with the cytological categories (cytology results) revealed that DPAS positivity (++, +++) correlated best with malignancy. Of the 201 patients, the AC grades according to the International Academy of Cytology (IAC) system were: AC2 (30, 14.9%), AC3 (112, 55.7%), AC4 (27, 13.4%), and AC5 (32, 15.9%). Conclusions: DPAS positivity in atypical cells in FNA aspirates may assist in upgrading from a suspicious to a malignant diagnosis in women with breast lumps.

Diagnostic imaging findings in a referral population of dogs diagnosed with immune-mediated haemolytic anaemia: 50 cases (2015-2018).

To describe and characterise changes documented on thoracic and abdominal imaging of dogs with confirmed immune-mediated haemolytic anaemia. Medical records from a referral hospital were searched from 2015 to 2018 for all dogs diagnosed with immune-mediated haemolytic anaemia that underwent thoracic and abdominal imaging by radiography, ultrasound or computed tomography. Fifty dogs were included. Thoracic imaging revealed abnormalities in 10 dogs (20%) of which lymphadenopathy and cardiomegaly were documented in four dogs (8%) each, and pleural effusion and pleural thickening in one dog (2%) each. Abdominal imaging revealed abnormalities in 43 dogs (86%), in which hepatomegaly and peritoneal effusion were documented in 20 (40%) and 19 dogs (38%), gallbladder wall thickening and sludge in 16 (32%) and 14 dogs (28%) and splenic nodules and splenomegaly in 13 (26%) and seven dogs (14%), respectively. Hepatic and splenic abnormalities were further investigated via fine needle aspirates in 18 dogs and revealed extramedullary haematopoiesis in 12 hepatic (66.7%) and 14 splenic (77.8%) fine needle aspirate samples. Cholecystocentesis was performed in nine dogs with gallbladder abnormalities and revealed bactibilia in three samples (33.3%). In this population of dogs with immune-mediated haemolytic anaemia, thoracic imaging abnormalities were uncommon. Hepatomegaly, peritoneal effusion and gallbladder wall thickening were the most common abdominal imaging findings with bactibilia confirmed in one third of collected bile samples. Hepatosplenomegaly and abdominal lymphadenopathy were not associated with neoplasia in any of the dogs included in this study.

Honey as a Cytological Fixative: A Comparative Study With 95% Alcohol

BackgroundNinety-five percent (95%) ethyl alcohol (ethanol) has been used as a standard cytological fixative but it is expensive, difficult to procure, and has addictive properties. Alternate substitutes like methanol, which give similar results to ethanol, have toxic potential. Honey, a known preservative, is an eco-friendly fixative and is of great value when ethanol is unavailable and economizing on cost is necessary. The present study was done to assess and compare the fixation property and cytomorphological features of smears fixed in 20% honey in comparison to 95% ethyl alcohol and to determine whether the former can be used as an alternative cytological fixative in routine practice.Material and methodsThe present prospective study was done in the cytology section of the Department of Pathology for one and a half years on 300 cytological samples comprising 100 samples each of various body fluids (peritoneal, pleural, bronchoalveolar lavage, and urine), cervical smears, and fine-needle aspiration samples. The smears from all the 300 cytological samples were fixed separately in 95% ethanol and 20% unprocessed natural honey for a minimum of 15 minutes and were then stained with Papanicolaou (Pap) stain. The cytomorphological parameters of both the smears were compared based on set criteria. Relevant statistical analysis was done using the student t-test, chi-square test, and test of agreement (kappa statistics).ResultsA comparable and good-quality staining pattern, preservation of morphology, and crisp nuclear and cytoplasmic staining were observed between the two fixatives for all three types of samples with a strong agreement between them (kappa value varying between 0.896 and 0.942) and a p-value of <0.05.ConclusionNatural honey is a readily available and non-toxic alternative to ethanol as a cytological fixative and can be used in routine practices, especially in resource-constrained settings.

Tumor-Derived Exosomal RNA From Fine-Needle Aspiration Supernatant as a Novel Liquid Biopsy for Molecular Diagnosis of Cancer.

Background: We hypothesized that the fine needle aspiration (FNA) supernatant from tumor might contain tumor-derived exosomes. The objective of this pilot study was to test if tumor-derived exosomal RNA could be found in FNA supernatants for molecular diagnosis of cancer. Methods: 10 FNA samples from pancreatic tumor were included. After the routine recuperation of cellular material by centrifugation, the cell-free Cytolyt liquid was collected instead of being discarded. 10 ml Cytolyt was used to isolate the exosomes. Transmission electronic microscopy (TEM) was used to examine the presence of exosomes. The exosomal marker CD63 was analyzed by flow cytometry. The exosomal RNA was extracted. RT-qPCR was performed to detect the GAPDH and the tumor marker of glypican 1 gene expression. Results: TEM confirmed the presence of exosomes from FNA supernatants. Flow cytometry showed a strong positive expression of exosome marker CD63. The concentration of exosomal RNA ranged from 18.81 to 354.75 ng/μl with an average of 81.76 ng/μl. The average exosomal RNA quantity was 1390.01 ng (range from 319.77 to 6030.75 ng) with an average 260/280 ratio of 2.12. GAPDH was detectable in all samples. Exosomal glypican 1 was detected in all samples of pancreatic ductal adenorcarcinomas (3/3) and absent from benign cystic samples (3/3). Furthermore, exosomal glypican 1 was positive in one sample with a non-contributive cytology and in one sample in which no malignant cell was found. Conclusion: This is the first report that the supernatants from FNA biopsy may contain tumor-derived exosomal RNA. These tumor-derived exosomes from FNA may provide a new liquid biopsy for the molecular diagnosis of cancer.

Clinicopathological features and outcomes of thyroid nodules with EIF1AX mutations.

EIF1AX gene mutations are reported in both benign and malignant thyroid tumors, with unclear outcomes when detected preoperatively. The aim of this study was to determine the features and outcomes of thyroid nodules with various types of mutation identified in cytologic (fine-needle aspiration) samples on preoperative ThyroSeq testing and with surgical outcomes. In this single-institution retrospective study of 31 consecutive patients, 77% were female and nodule size ranged from 1.5 to 9.4 cm with widely varying cytologic and TI-RADS ultrasound categorizations. Among two main mutational hotspots, 55% were located in exon 2 and 45% at the intron 5/exon 6 splice site. On histology, 45% of -positive nodules were cancer/noninvasive follicular thyroid neoplasm with papillary-like nuclear features (NIFTP) including 19% encapsulated follicular variant papillary thyroid carcinoma, 10% follicular carcinoma, 10% anaplastic carcinoma (ATC), and 7% NIFTP. Almost half (48%) of patients had one or more coexisting mutations, most frequently RAS. The prevalence of cancer/NIFTP was 80% for mutation with coexisting molecular alteration vs 13% with an isolated mutation (P = 0.0002). Cancer probability was associated with mutation type and was 64% for splice-site mutation and 29% for non-splice mutation (P = 0.075). All 3 nodules with EIF1AX+RAS+TERT+TP53 mutations were ATC. In summary, in this study, all nodules with an isolated non-splice mutation were benign, one-third of those with an isolated splice mutation were cancer, and most nodules with coexisting with RAS or other alterations were malignant. These findings suggest that clinical management decisions for patients with EIF1AX-mutant nodules should consider both the type of mutation and its co-occurrence with other genetic alterations.