Tall Cell Research Articles

Glomangiosarcoma-like Anaplastic Transformation in Papillary Thyroid Carcinoma: A Novel Form of Heterologous Differentiation and a Systematic Review of Heterologous Element Prevalence.

Anaplastic thyroid carcinoma (ATC) demonstrates a wide variety of morphologies and is characteristically associated with a differentiated thyroid carcinoma component. Heterologous differentiation is a rare, potentially challenging phenomenon in ATC, mostly observed as osteosarcomatous or chondrosarcomatous differentiation. We now describe a novel 'glomangiosarcoma-like' differentiation, review our archival experience from two institutions (UPMC, CC), and perform a systematic review for the prevalence of heterologous elements in ATC. The patient is a 57-year-old female who presented with 4.5cm left thyroid, and 3.4cm neck masses. Histologically, the thyroid demonstrated a differentiated high grade papillary thyroid carcinoma, tall cell and hobnail/micropapillary subtypes transitioning into an anaplastic component with spindled to ovoid cells with hemangiopericytoma-like vasculature showing CD34 positivity, variable muscle marker expression and pericellular lace-like type IV collagen deposition. The neck mass consisted solely of the latter morphology. Targeted next-generation sequencing was performed on high grade DTC and adjacent ATC from the thyroid as well as ATC from the neck metastasis. All three components shared BRAFV600E, TERT promoter, and PIK3CA mutations confirming a clonal origin. Archival (UPMC: n = 150, CC: n = 74) and literature review showed no prior examples. Systematic review and meta-analysis of prevalence showed a baseline pooled prevalence (generalized linear mixed model) of heterologous elements of any type to be 1.6% (95% confidence interval: 1.0-2.6%) for studies where this was specifically addressed. ATC with glomangiosarcoma-like heterologous differentiation is a rarity among an already rare morphologic category with unique diagnostic pitfalls.

Subcentimetric papillary thyroid carcinoma with extensive lymph node and brain metastasis: case report and review of literature.

We report a 61-year-old male patient without personal history of thyroid carcinoma or radiation exposure. In 2011, he presented with a cervical mass whose biopsy diagnosed a papillary thyroid carcinoma (PTC) in a lymph node metastasis (LNM). Total thyroidectomy with lymphadenectomy of central and ipsilateral compartment was performed. Histopathology identified a 2 mm follicular variant of PTC and LNM in 25/25 lymph nodes. The patient was treated with 150 mCi of radioactive iodine (RAI), followed by levothyroxine suppressive therapy. In 2016, a retrotracheal mass was diagnosed, suggesting local recurrence; patient was submitted to surgical excision and RAI therapy (120 mCi). Due to seizures, in 2019, a brain CT was performed that diagnosed brain metastases. The patient underwent debulking of the main lesion. Histopathology analysis confirmed a metastatic lesion with variated morphology: classical PTC and follicular pattern and hobnail and tall cell features. Molecular analysis revealed BRAFV600E in LNM at presentation and BRAFV600E and TERT promoter (TERTp) mutations in the recurrent LNM and brain metastasis. Based upon this experience we review the reported cases of subcentimetric PTC with brain metastases and discuss the molecular progression of the present case. Papillary microcarcinoma (PMCs) usually have very good prognosis with low impact on patient survival. PMCs presenting in elderly patients with LNM at diagnosis may carry a guarded outcome. Brain metastasis although rare indicate aggressive phenotypic features. Patient risk stratification of PMCs based on histopathological analysis and genetic testing may have a significant impact on prognosis providing therapeutic markers, that may predict disease progression and overall outcome.

Adjuvant External Beam Radiotherapy and Histopathologic Prognostic Factors in Tall Cell Papillary Thyroid Carcinoma

Differentiated thyroid cancer is a diverse disease, with histopathologic characteristics contributing to prognosis and treatment decisions. Tall cell variant of papillary thyroid cancer (TCV-PTC) carries a poor prognosis relative to other histologies, with first site of failure most often in the neck. Several small studies suggest there may be a disease control benefit to adjuvant external beam radiotherapy (EBRT) for certain high-risk DTC populations; however, data is limited. In this study we aim to shed light on both histopathologic prognostic factors and survival outcomes of patients with TCV-PTC carcinoma treated with surgery followed by EBRT and Radioactive Iodine (RAI) or RAI alone. We hypothesize that EBRT with RAI leads to better disease control than RAI alone. Patients with TCV-PTC diagnosed between 1997-2020 at a single institution were included in the analysis. TNM staging was standardized to AJCC 8th edition. Demographics, staging, and histologic characteristics were compared between the two treatment groups using T-tests and Fisher's exact tests. Kaplan-Meier survival analyses were performed between patients receiving EBRT vs. RAI alone for disease-free survival (DFS), cancer-specific survival (CSS) and overall survival (OS), along with Cox Proportional Hazards analysis. A total of 93 patients were included. Of these patients, 22 received adjuvant RAI and EBRT, and 71 received RAI alone. There was no statistical difference in race, gender, tall cell percentage in the pathology specimen, TNM stage, AJCC group stage, or disease recurrence in patients receiving EBRT vs. RAI alone. However, those receiving EBRT were significantly older (RAI mean age (SD): 50.6 (18.7), EBRT mean age (SD): 59.7 (14.4), p = 0.02) and length of follow-up was shorter for the EBRT cohort (RAI mean (SD): 8.80 (5.81) years, EBRT mean (SD): 4.53 (2.14) years, p < 0.01). Multifocal disease (HR: 3.04, 95% CI: 1.30 - 7.08, p = 0.01) and soft tissue invasion (HR: 2.93, 95% CI: 1.28 - 6.75, p = 0.01) were associated with decreased DFS, whilst age (HR 1.08, 1.01 - 1.15: p = 0.03) was associated with decreased CSS; tall cell percentage in the pathology specimen, extra-thyroidal extension, vascular invasion, and positive surgical margin were not significant for either metric. There was no statistical difference in DFS (p = 0.5), CSS (p = 0.3), or OS (p = 0.6) between patients that received EBRT and RAI vs. RAI alone. DFS, CSS, and OS were not different between patients who received EBRT compared to RAI only in this high-risk subgroup of patients. Short interval follow-up for the EBRT group and significant age differences between the EBRT and RAI cohorts may confound results. Given the significant association of advanced age with CSS, an expanded, age-matched cohort analysis is underway.

Bortezomib Is Effective in the Treatment of T Lymphoblastic Leukaemia by Inducing DNA Damage, WEE1 Downregulation, and Mitotic Catastrophe.

T lymphoblastic leukemia (T-ALL) is an aggressive haematolymphoid malignancy comprising 15% of acute lymphoblastic leukemia (ALL). Although its prognosis has improved with intensive chemotherapy, the relapse/refractory disease still carries a dismal prognosis. Thus, there is an urgent need to develop novel therapy for T-ALL. Bortezomib, a 26S proteasome inhibitor, is licensed to treat plasma cell myeloma and mantle cell lymphoma. Due to its favorable side effect profile, it is a novel agent of research interest in the treatment of ALL. Despite an increasing number of clinical trials of bortezomib in T-ALL, its detailed mechanistic study in terms of DNA damage, cell cycle, and mitotic catastrophe remains elusive. Moreover, WEE1, a protein kinase overexpressed in ALL and involved in cell-cycle regulation, has been known to be a novel therapeutic target in many cancers. But the role of bortezomib in modulating WEE1 expression in ALL still remains elusive. In this study, we demonstrate the therapeutic efficacy of bortezomib on T-ALL primary samples and cell lines. Our findings reveal that bortezomib treatment induces DNA damage and downregulates WEE1, leading to G2-M cell-cycle progression with damaged DNA. This abnormal mitotic entry induced by bortezomib leads to mitotic catastrophe in T-ALL. In conclusion, our findings dissect the mechanism of action of bortezomib and provide further insights into the use of bortezomib to treat T-ALL. Our findings suggest the possibility of novel combination therapy using proteasome inhibitors together with DNA-damaging agents in the future, which may fill the research gaps and unmet clinical needs in treating ALL.

The Role of Novel Immunohistochemical Markers for Special Types of Breast Carcinoma.

Some histologic special types of breast carcinoma harbor specific recurrent genetic alterations that are not seen in other types of breast carcinoma (no special type), namely adenoid cystic carcinoma, secretory carcinoma, and tall cell carcinoma with reversed polarity. These tumors have unique morphologic features, are triple-negative, that is, do not express hormone receptors or HER2, and are generally associated with a favorable prognosis. Adenoid cystic carcinoma, like its counterpart in other organs, shows a MYB-NFIB fusion gene that is the result of a recurrent t(6;9)(q22-23;p23-24) translocation. Other MYB alterations have been described that result in overexpression of MYB . Secretory carcinoma is characterized by an ETV6-NTRK3 gene fusion that is the result of recurrent (12;15);(p13;q25) translocation, which is also seen in mammary analog secretory carcinoma of the salivary gland. Tall cell carcinoma with reversed polarity shows IDH2 p.Arg172 hotspot mutations. Immunohistochemical antibodies have emerged that identify the underlying genetic alterations in these tumors and serve as useful diagnostic tools. This review will provide an update on the molecular features and diagnostic immunohistochemical markers that have become increasingly popular to aid in diagnosing these uncommon triple-negative breast tumors.

The Warthin-like variant of papillary thyroid carcinomas: a clinicopathologic analysis report of two cases

Abstract Obejectives Papillary thyroid carcinoma (PTC) is the most common type of thyroid carcinoma, comparising various subtypes including classical PTC (accounting for approximately 55–65 % of cases) and several variants. The identified variants include papillary microcarcinoma, encapsulated variant, follicular variant, diffuse sclerosing variant, tall cell variant, columnar cell variant, cribriform-morular variant, and Warthin-like variant (WLV-PTC). WLV-PTC is a rare malignant neoplasm. Case presentation The present study introduced two cases of WLV-PTC from Affiliated Hospital of Chengde Medical College in retrospect. The macroscopic characteristics, histological feature, immunophenotyping, gene mutations, local invasion, and distant metastasis of the tumors were observed, and the relevant literature on WLV-PTC was reviewed. The tumor cells exhibited nuclear the characteristics similar to those of classic papillary thyroid carcinoma. Immunohistochemical analysis showed strong positivity for thyroid transcription factor 1, cytokeratin 19, Galectin-3, and CK, weak positivity for thyroglobulin, and the Ki-67 proliferation index ranging from 5 to 10 %. Gene mutation analysis revealed that Case 1 had a BRAF V600E mutation, whereas Case 2 had a wild-type mutation. WLV-PTC has histological features similar to those of Warthin tumor originating from salivary gland. Conclusions The prognosis of WLV-PTC is similar to that of conventional PTC of comparable size and stage.

The circadian clock circuitry modulates leukemia initiating cell activity in T-cell acute lymphoblastic leukemia

BackgroundT-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy, characterized by restricted cellular subsets with asymmetrically enriched leukemia initiating cell (LIC) activity. Nonetheless, it is still unclear which signaling programs promote LIC maintenance and progression.MethodsHere, we evaluated the role of the biological clock in the regulation of the molecular mechanisms and signaling pathways impacting the cellular dynamics in T-ALL through an integrated experimental approach including gene expression profiling of shRNA-modified T-ALL cell lines and Chromatin Immunoprecipitation Sequencing (ChIP-Seq) of leukemic cells. Patient-derived xenograft (PDXs) cell subsets were also genetically manipulated in order to assess the LIC activity modulated by the loss of biological clock in human T-ALL.ResultsWe report that the disruption of the circadian clock circuitry obtained through shRNA-mediated knockdown of CLOCK and BMAL1 genes negatively impacted the growth in vitro as well as the activity in vivo of LIC derived from PDXs after transplantation into immunodeficient recipient mice. Additionally, gene expression data integrated with ChIP-Seq profiles of leukemic cells revealed that the circadian clock directly promotes the expression of genes, such as IL20RB, crucially involved in JAK/STAT signaling, making the T-ALL cells more responsive to Interleukin 20 (IL20).ConclusionTaken together, our data support the concept that the biological clock drives the expression of IL20R prompting JAK/STAT signaling and promoting LIC activity in T-ALL and suggest that the selective targeting of circadian components could be therapeutically relevant for the treatment of T-ALL patients.

Reduction of Ca2+ Entry by a Specific Block of KCa3.1 Channels Optimizes Cytotoxic Activity of NK Cells against T-ALL Jurkat Cells.

Degranulation mediated killing mechanism by NK cells is dependent on store-operated Ca2+ entry (SOCE) and has optimum at moderate intracellular Ca2+ elevations so that partial block of SOCE optimizes the killing process. In this study, we tested the effect of the selective blocker of KCa3.1 channel NS6180 on SOCE and the killing efficiency of NK cells from healthy donors and NK-92 cells against T-ALL cell line Jurkat. Patch-clamp analysis showed that only one-quarter of resting NK cells functionally express KCa3.1 current, which increases 3-fold after activation by interleukins 15 and 2. Nevertheless, blockage of KCa3.1 significantly reduced SOCE and intracellular Ca2+ rise induced by IL-15 or target cell recognition. NS6180 (1 μM) decreased NK degranulation at zero time of coculture with Jurkat cells but already after 1 h, the degranulation reached the same level as in the control. Monitoring of target cell death by flow cytometry and confocal microscopy demonstrated that NS6180 significantly improved the killing ability of NK cells after 1 h in coculture with Jurkat cells and increased the Jurkat cell fraction with apoptotic and necrotic markers. Our data evidence a strong dependence of SOCE on KCa3.1 activity in NK cells and that KCa3.1 specific block can improve NK cytotoxicity.

Development of constitutively synergistic nanoformulations to enhance chemosensitivity in T-cell leukemia

Advances in multiagent chemotherapy have led to recent improvements in survival for patients with acute lymphoblastic leukemia (ALL); however, a significant fraction do not respond to frontline chemotherapy or later relapse with recurrent disease, after which long-term survival rates remain low. To develop new, effective treatment options for these patients, we conducted a series of high-throughput combination drug screens to identify chemotherapies that synergize in a lineage-specific manner with MRX-2843, a small molecule dual MERTK and FLT3 kinase inhibitor currently in clinical testing for treatment of relapsed/refractory leukemias and solid tumors. Using experimental and computational approaches, we found that MRX-2843 synergized strongly—and in a ratio-dependent manner—with vincristine to inhibit both B-ALL and T-ALL cell line expansion. Based on these findings, we developed multiagent lipid nanoparticle formulations of these drugs that not only delivered defined drug ratios intracellularly in T-ALL, but also improved anti-leukemia activity following drug encapsulation. Synergistic and additive interactions were recapitulated in primary T-ALL patient samples treated with MRX-2843 and vincristine nanoparticle formulations, suggesting their clinical relevance. Moreover, the nanoparticle formulations reduced disease burden and prolonged survival in an orthotopic murine xenograft model of early thymic precursor T-ALL (ETP-ALL), with both agents contributing to therapeutic activity in a dose-dependent manner. In contrast, nanoparticles containing MRX-2843 alone were ineffective in this model. Thus, MRX-2843 increased the sensitivity of ETP-ALL cells to vincristine in vivo. In this context, the additive particles, containing a higher dose of MRX-2843, provided more effective disease control than the synergistic particles. In contrast, particles containing an even higher, antagonistic ratio of MRX-2843 and vincristine were less effective. Thus, both the drug dose and the ratio-dependent interaction between MRX-2843 and vincristine significantly impacted therapeutic activity in vivo. Together, these findings present a systematic approach to high-throughput combination drug screening and multiagent drug delivery that maximizes the therapeutic potential of combined MRX-2843 and vincristine in T-ALL and describe a novel translational agent that could be used to enhance therapeutic responses to vincristine in patients with T-ALL. This broadly generalizable approach could also be applied to develop other constitutively synergistic combination products for the treatment of cancer and other diseases.

Impact of BRAFV600E Mutation on Event-Free Survival in Patients with Papillary Thyroid Carcinoma: A Retrospective Study in a Romanian Population.

We aimed to evaluate the prognostic value of BRAFV600E mutation in a series of 127 papillary thyroid carcinoma (PTC) cases as a single factor, and in synergic interaction with other standard risk factors. BRAFV600E mutation was assessed by real-time PCR. Event-free survival (EFS) was calculated between the date of the first evaluation and the date of occurrence of an adverse event or the date of the last known status. The prevalence of BRAFV600E mutation was 57.2%. The Kaplan-Meier analysis showed a significant reduction of EFS among cases harboring BRAFV600E mutation compared to non-mutated cases (p = 0.010). In addition, BRAFV600E mutation was found to better predict adverse outcomes when associated with the following risk factors: age ≥ 55 years old (p < 0.001), male gender (p < 0.001), conventional (p = 0.005) and tall cell (p = 0.014) histology, tumor size > 40 mm (p = 0.001), extrathyroidal extension (p = 0.001), multifocality (p = 0.001) and lymph node metastasis (p < 0.001). In univariate analysis, a 3.74-fold increased risk for a reduced EFS (p = 0.018) was found for BRAFV600E-mutated cases, but no increased risk was further confirmed by multivariate analysis. Our results highlight that BRAFV600E mutation cannot be used alone as an independent predictive factor in PTC patients, but is prognostically valuable if integrated in the context of other clinicopathological risk factors.

Classification of Thyroid Tumors Based on DNA Methylation Patterns.

Background: Alterations in DNA methylation are stable epigenetic events that can serve as clinical biomarkers. The aim of this study was to analyze methylation patterns among various follicular cell-derived thyroid neoplasms to identify disease subtypes and help understand and classify thyroid tumors. Methods: We employed an unsupervised machine learning method for class discovery to search for distinct methylation patterns among various thyroid neoplasms. Our algorithm was not provided with any clinical or pathological information, relying exclusively on DNA methylation data to classify samples. We analyzed 810 thyroid samples (n = 256 for discovery and n = 554 for validation), including benign and malignant tumors, as well as normal thyroid tissue. Results: Our unsupervised algorithm identified that samples could be classified into three subtypes based solely on their methylation profile. These methylation subtypes were strongly associated with histological diagnosis (p < 0.001) and were therefore named normal-like, follicular-like, and papillary thyroid carcinoma (PTC)-like. Follicular adenomas, follicular carcinomas, oncocytic adenomas, and oncocytic carcinomas clustered together forming the follicular-like methylation subtype. Conversely, classic papillary thyroid carcinomas (cPTC) and tall cell PTC clustered together forming the PTC-like subtype. These methylation subtypes were also strongly associated with genomic drivers: 98.7% BRAFV600E-driven cancers were PTC like, whereas 96.0% RAS-driven cancers had a follicular-like methylation pattern. Interestingly, unlike other diagnoses, follicular variant PTC (FVPTC) samples were split into two methylation clusters (follicular like and PTC like), indicating a heterogeneous group likely to be formed by two distinct diseases. FVPTC samples with a follicular-like methylation pattern were enriched for RAS mutations (36.4% vs. 8.0%; p < 0.001), whereas FVPTC- with PTC-like methylation patterns were enriched for BRAFV600E mutations (52.0% vs. 0%, Fisher exact p = 0.004) and RET fusions (16.0% vs. 0%, Fisher exact p = 0.003). Conclusions: Our data provide novel insights into the epigenetic alterations of thyroid tumors. Since our classification method relies on a fully unsupervised machine learning approach for subtype discovery, our results offer a robust background to support the classification of thyroid neoplasms based on methylation patterns.

Effects of MYC Inhibitors on the Growth of Acute Leukaemia Cells.

MYC proto-oncogene bHLH transcription factor (MYC) proteins function as transcription factors by binding to MYC-associated factor X (MAX) proteins and are involved in various cancer growth, including leukaemia. This study aimed to examine the effects of synthetic MYC inhibitors, which block the MYC-MAX complex formation, in in vitro human acute leukaemia cell lines. Four cell lines, OCI/AML2 derived from acute myeloid leukaemia, NALM-6 from B-lymphoblastic leukaemia, and KOPT-K1 and Jurkat from notch receptor 1 (NOTCH1)-mutated T-lymphoblastic leukaemia (T-ALL), were treated with the small-molecule MYC inhibitors 10058-F4 and MYCi975. The expression of cell proliferation and signalling proteins was studied. These inhibitors suppressed the growth of leukaemia cell lines. Treatment with the two inhibitors down-regulated the protein expression of c-MYC, MAX, and activating enhancer-binding protein 4 (AP4) in all cell lines. Up-regulation of p27 and p21 was observed only in 10058-F4-treated OCI/AML2 cells and MYCi975-treated KOPT-K1 cells. These two inhibitors down-regulated the expression of NOTCH1, cleaved NOTCH1, and hes family bHLH transcription factor 1 (HES1) in both T-ALL cell lines. MYC inhibitors appear to be novel molecular-targeted drugs against acute leukaemia, including NOTCH1-mutated T-ALL. However, it is necessary to elucidate the precise molecular mechanisms of these effects before clinical use.

Augmenter of liver regeneration promotes drug resistance of acute lymphoblastic leukemia through the alteration of mitochondrial functions and the inhibition of the mitochondrial apoptosis pathway.

Acute T lymphoblastic leukemia (T-ALL) occurs in 25% of adults diagnosed with Acute lymphocytic leukemia (ALL), and drug resistance is still a clinical obstacle. Augmenter of liver regeneration (ALR) is important to ALL drug resistance and is involved in the regulation of mitochondrial function; we speculated that the high expression of ALR in T-ALL promotes drug resistance through the alteration of mitochondrial function and the inhibition of the mitochondrial apoptosis pathway. We silenced and overexpressed ALR in the T-ALL cell lines that were untreated or treated with dexamethasone (DXM) or methotrexate (MTX). Apoptosis, proliferation, reactive oxygen species and ATP productions, mitochondrial membrane potential, and mitochondrial respiratory chain complex expression in cells were examined. The data were collated to comprehensively evaluate the effects of ALR expression change on mitochondrial function and drug resistance in T-ALL cells. ALR knockdown led to the inhibition of proliferation, an increase in apoptosis, and the promotion of the cells' sensitivity to drugs. It also showed mitochondrial dysfunction. ALR knockdown actived the mitochondrial apoptosis pathway. The treatment of ALR knockdown T-ALL cells with MTX or DXM further altered the mitochondrial function of T-ALL cells and actived the mitochondrial apoptosis pathway. Overexpression of ALR promoted cell proliferation and drug resistance, reduced apoptosis, protected mitochondrial function, and inhibited the mitochondrial apoptosis pathway. T-ALL resistance caused by ALR through the alteration of mitochondrial function is associated with the inhibition of the mitochondrial apoptosis pathway.

Clinico-pathological initial outcome of a newly adopted novel surgical technique for nodal metastatic thyroid cancer at a large-volume centre in a high-income developing country.

Thyroid cancer is the most common tumour in the endocrine system. In the past decade, the incidence of lymph node metastasis has increased, and so has the patient demand for a smaller scar. This research reports the surgical and patho-oncological short-term outcomes of a novel, minimally invasive neck dissection approach for thyroid carcinoma with lymph node metastases at the UAE's leading endocrine surgery centre. This study employed a prospectively maintained surgical database to retrospectively analyse relevant parameters such as surgical complications, including bleeding, hypocalcaemia nerve injury and lymphatic fistula, as well as oncological metrics such as the type of tumour and the ratio of lymph node metastasis to the number of harvested lymph nodes in 100 patients that underwent open minimally invasive selective neck dissection. The study comprised 50 patients with thyroidectomy, with bilateral central compartment neck dissection (BCCND; 50%); 34 with thyroidectomy, BCCND and selective bilateral lateral compartment neck dissection (BLCND; 34%); and 16 patients with selective unilateral central and lateral compartment neck dissection by recurrent nodal disease (ULCND; 16%). The female-to-male gender ratio was 78:22, with median ages of 36 and 42 years, respectively. Histopathology findings revealed that 92% of patients had papillary thyroid cancer (PTC) and 8% had medullary thyroid cancer. The mean total number of lymph nodes removed was 22 in the BLCND group, 17 in the ULCND group and 8 in the BCCND group (p = 0.001). Furthermore, the average lymph node metastasis was significantly higher in the BLCND group (p = 0.002). The percentage of temporary hypoparathyroidism was 29.8% and it persisted for 1.3%. The lateral compartment dissection morbidity was as follows: Four male patients with tall cell infiltrative PTC presented with pre-existing vocal cord paresis, necessitating nerve resection and anastomosis, and two more developed it after surgery (1.1% of the nerve at risk). Lymphatic fistulas occurred in four patients (4%) who were treated conservatively. Two patients were readmitted due to symptomatic neck collection. Horner syndrome was discovered in just one female patient. Male gender, aggressive histology and lateral compartment dissection independently increased surgical morbidity. In a high-volume endocrine centre, the adoption of minimally invasive selective neck dissections to treat nodal metastatic thyroid cancer did not increase specific cervical surgery complications.

A Shift in Molecular Drivers of Papillary Thyroid Carcinoma Following the 2017 World Health Organization Classification: Characterization of 554 Consecutive Tumors With Emphasis on BRAF-Negative Cases

Most studies for comprehensive molecular profiling of papillary thyroid carcinoma (PTC) have been performed before the 2017 World Health Organization (WHO) classification, in which the diagnostic criteria of follicular variants of PTC have been modified and noninvasive follicular thyroid neoplasm with papillary-like nuclear features has been introduced. This study aims to investigate the shift in the incidence of BRAF V600E mutations in PTCs following the 2017 WHO classification and to further characterize the histologic subtypes and molecular drivers in BRAF-negative cases. The study cohort consisted of 554 consecutive PTCs larger than 0.5 cm between January 2019 and May 2022. Immunohistochemistry for BRAF VE1 was performed for all cases. Compared with a historical cohort of 509 PTCs from November 2013 to April 2018, the incidence of BRAF V600E mutations was significantly higher in the study cohort (86.8% vs 78.8%, P = .0006). Targeted RNA-based next-generation sequencing using a FusionPlex Pan Solid Tumor v2 panel (ArcherDX) was performed for BRAF-negative PTCs from the study cohort. Eight cribriform-morular thyroid carcinomas and 3 cases with suboptimal RNA quality were excluded from next-generation sequencing. A total of 62 BRAF-negative PTCs were successfully sequenced, including 19 classic follicular predominant PTCs, 16 classic PTCs, 14 infiltrative follicular PTCs, 7 encapsulated follicular PTCs, 3 diffuse sclerosing PTCs, 1 tall cell PTC, 1 solid PTC, and 1 diffuse follicular PTC. Among them, RET fusions were identified in 25 cases, NTRK3 fusions in 13 cases, BRAF fusions in 5 cases including a novel TNS1::BRAF fusion, NRAS Q61R mutations in 3 cases, KRAS Q61K mutations in 2 cases, NTRK1 fusions in 2 cases, an ALK fusion in 1 case, an FGFR1 fusion in 1 case, and an HRAS Q61R mutation in 1 case. No genetic variants, from our commercially employed assay, were detected in the remaining 9 cases. In summary, the incidence of BRAF V600E mutations in PTCs significantly increased from 78.8% to 86.8% in our post–2017 WHO classification cohort. RAS mutations accounted for only 1.1% of the cases. Driver gene fusions were identified in 8.5% of PTCs and were clinically relevant given the emerging targeted kinase inhibitor therapy. Of the 1.6% of cases for which no driver alteration was detected, the specificity of drivers tested and tumor classification require further investigation.

Cytological and Ultrasound Features of Thyroid Nodules Correlate With Histotypes and Variants of Thyroid Carcinoma.

Prognosis is excellent for papillary thyroid carcinoma (PTC), noninvasive follicular thyroid neoplasia with papillary-like nuclear features (NIFT-P), and follicular thyroid carcinoma (FTC) but is poor for poorly differentiated thyroid carcinoma (PDTC) and anaplastic thyroid carcinoma (ATC). Among PTCs, the prognosis is more favorable for follicular (FV-PTC) and classic (CV-PTC) than for tall cell (TCV-PTC), and solid (SV-PTC) variants. To associate histotypes and variants of thyroid carcinoma with ultrasound and cytological features. Histology of 1018 benign tumors and 514 PTC (249 CV, 167 FV, 49 TC, 34 SV, and 15 other variants), 52 NIFT-P, 50 FTC, 11 PDTC, and 3 ATC was correlated with fine-needle aspiration biopsy categories (Italian classification: TIR1, TIR2, TIR3A, TIR3B, TIR4, and TIR5) and ultrasound features at the Endocrinology Unit, University Hospital of Pisa. In total, 1117 patients with thyroid nodule(s) who underwent thyroidectomy were included. Of PTC, 36.3% had indeterminate cytology (TIR3A or TIR3B), 56.6% were suspicious for malignancy or malignant (TIR4 or TIR5); 84.0% FTC and 69.3% NIFT-P were TIR3A or TIR3B; 72.5% FV-PTC and 73.6% SV-PTC were TIR3A or TIR3B; 79.9% CV-PTC and 95.9% TCV-PTC were TIR4 or TIR5. The association of a hypoechoic pattern, irregular margins, and no microcalcifications was more frequent in TCV-PTC than in CV-PTC (P = .02, positive predictive value = 38.9%; negative predictive value = 85.5%). At cytology, most FTC, NIFT-P, FV-PTC, and SV-PTC were indeterminate, most CV-PTC and TCV-PTC were suspicious for malignancy or malignant. Ultrasound can be helpful in ruling out TCV-PTC.

Dissecting the tumor microenvironment of papillary thyroid carcinomas in primary tumor and metastatic lymph nodes at single-cell resolution.

e15182 Background: Although often well-differentiated and indolent, papillary thyroid carcinomas (PTCs) frequently metastasize to regional lymph nodes (LNs) especially in younger patients. It is unclear what factors of the local microenvironment promote LN metastases. To this end, we used single-cell RNA sequencing (scRNA-seq) to characterize the immune compartment of pair-matched primary tumor and metastatic LN samples from patients with PTC. Methods: In total, we processed 14 fresh surgical specimens from 7 patients undergoing primary total thyroidectomy and bilateral modified radical neck dissection between July 2018 and July 2019. CD45+ immune cells were isolated for library generation per 10X Genomics protocol for Single Cell 5’ v2 chemistry dual index. Data analysis was performed by Seurat and BBrowser. Differentially expressed genes were identified by Venice. Results: Pair-matched primary tumor and metastatic LN samples from 7 patients were included (mean [SD] age, 42.9 [17.4]). Histologic subtypes include classical PTC (n = 3), anaplastic (n = 2), tall cell variant (n = 1), and diffuse sclerosing variant (n = 1). The median number of positive metastatic LNs was 9 (range: 4-41) and the median number of examined LNs was 67 (range: 15-86). The majority of patients were pN1b (n = 6) or pN1a (n = 1) per AJCC 8th Edition. In total, we analyzed 26,659 individual cells by scRNA-seq, including 10,558 cells from the primary tumor and 16,101 cells from the metastatic LN. Unbiased clustering identified 6 major populations including B cells (CD79A/B, IGKC), T/natural killer (NK) cells (CD3E, CD247), myeloid cells (LYZ, FCER1G), thyrocytes (TG, EPCAM), endothelial cells (PECAM1, CD34), and fibroblasts (COL1A1/2, ACTA2). In a pooled analysis, we identified increased myeloid cells and thyrocytes within primary tumor samples. By contrast, there were increased B cells and T/NK cells within the metastatic LN samples. We found that IL7R is highly enriched by B and T/NK cells within metastatic LNs compared to primary tumor (B cells: log2 fold-change -1.26, P= 1.36x10-65; T/NK cells: log2 fold-change -1.23, P= 1.56x10-147). Further, CCL2 is significantly enriched by myeloid cells within the primary tumor compared to cells within the metastatic LN (log2 fold-change 2.16, P= 1.70x10-83). Lastly, TNFRSF12A (TWEAKR) is highly upregulated by thyrocytes within the metastatic LN as compared to cells from the primary tumor. Ongoing validation of these three targets (IL7R/FOXP3, CCL2/CD163, TNFRSF12A/PAX8) will be performed in an independent set of 25 patients with PTC with paired primary tumor-metastatic LNs. Conclusions: Collectively, our data indicate that there are increased immunosuppressive signals with the microenvironment that allow for PTC to colonize distant metastatic LNs. IL7R and TNFSRF12A may serve as a novel therapeutic target for patients with locally advanced PTC.

ASO Visual Abstract: Diffuse Sclerosing Papillary Thyroid Carcinoma-Clinicopathological Characteristics and Prognostic Implications in Comparison with Classic and Tall Cell Papillary Thyroid Cancer.

ASO Visual Abstract: Diffuse Sclerosing Papillary Thyroid Carcinoma-Clinicopathological Characteristics and Prognostic Implications in Comparison with Classic and Tall Cell Papillary Thyroid Cancer.

Cumulative effects of weakly repressive regulatory regions in the 3’ UTR maintain PD-1 expression homeostasis in mammals

PD-1 has become a common target for cancer treatment. However, the molecular regulation of PD-1 expression homeostasis remains unclear. Here we report the PD-1 3’ UTR can dramatically repress gene expression via promoting mRNA decay. Deletion of the PD-1 3’ UTR inhibits T cell activity and promotes T-ALL cell proliferation. Interestingly, the robust repression is attributable to cumulative effects of many weak regulatory regions, which we show together are better able to maintain PD-1 expression homeostasis. We further identify several RNA binding proteins (RBPs) that modulate PD-1 expression via the 3’ UTR, including IGF2BP2, RBM38, SRSF7, and SRSF4. Moreover, despite rapid evolution, PD-1 3’ UTRs are functionally conserved and strongly repress gene expression through many common RBP binding sites. These findings reveal a previously unrecognized mechanism of maintaining PD-1 expression homeostasis and might represent a general model for how small regulatory effects play big roles in regulation of gene expression and biology.

Optimal Concentration and Duration of Endotracheal Tube Coating to Achieve Optimal Antimicrobial Efficacy and Safety Balance: An In Vitro Study

Ventilator-associated pneumonia (VAP) is a common and genuine complication in fundamentally sick patients accepting mechanical ventilation. Silver nitrate sol-gel (SN) has been proposed as a potential preventative measure against VAP. Be that as it may, the arrangement of SN with distinctive concentrations and pH values remains a basic factor influencing its effectiveness. Silver nitrate sol-gel was arranged with distinctive concentrations (0.1852%, 0.03496%, 0.1852%, and 0.01968%) and pH values (8.5, 7.0, 8.0, and 5.0) separately. The antimicrobial action of the silver nitrate and NaOH arrangements were assessed against Escherichia coli as a reference strain. The thickness and pH of the arrangements were measured, and biocompatibility tests were performed on the coating tube. The auxiliary changes in the endotracheal tube (ETT) tests after treatment were analyzed utilizing electron microscopy (SEM) and transmission electron microscopy (TEM). The pH estimations of the diverse arrangements showed that the pH values shifted depending on the test conditions, with pH values extending from 5.0 to 8.5. The consistency estimations of the arrangements showed that the thickness values expanded as the pH values drew closer to 7.5 and diminished when the pH values went over 7.5. The antimicrobial action of the silver nitrate and NaOH arrangements were successful against Escherichia coli, with microbial checks decreasing in concentration (0.03496%, 0.1852% (pH: 8), and 0.01968%). The biocompatibility tests revealed tall cell reasonability rates, demonstrating that the coating tube was secure for therapeutic utilization and did not hurt typical cells. The SEM and TEM investigation gave visual proof of the antibacterial impacts of the silver nitrate and NaOH arrangements on the bacterial surface or interior of the bacterial cells. Moreover, the investigation revealed that a concentration of 0.03496% was the foremost successful in hindering the development of ETT bacterial colonization at the nanoscale level. We propose that cautious control and alteration of the pH and thickness of the arrangements are essential to guaranteeing the reproducibility and quality of the sol-gel materials. The silver nitrate and NaOH arrangements may serve as a potential preventative degree against VAP in sick patients, with a concentration of 0.03496% appearing to show the most elevated viability. The coating tube may serve as a secure and viable preventative measure against VAP in sick patients. Further investigation is required to optimize the concentration and introduction time of the arrangements to maximize their adequacy in avoiding VAP in real-world clinical settings.