The variable presentations of anaplastic spindle cell squamous carcinoma associated with tall cell variant of papillary thyroid carcinoma.

BACKGROUND: In 1976, Hawk and Hazard described the tall cell variant (TCV) of papillary thyroid carcinoma (PTC). While the lesions they described had cytologic features of papillary carcinoma, they showed more aggressive behavior with a greater propensity for extrathyroid extension and lymphovascular invasion than classic PTC. In 1991, Bronner and LiVolsi described a series of patients with TCV that progressed to spindle cell squamous carcinoma (SCSC), a unique form of anaplastic thyroid carcinoma. This study describes the variable clinical and pathologic presentations in 31 patients with anaplastic SCSC arising in association with TCV. METHODS: The surgical pathology archives as well as the personal consultation files of one of the authors (V.A.L.) were reviewed to retrieve cases of SCSC arising in association with TCV. The available clinical as well as pathologic information on all patients was reviewed. RESULTS: A total of 31 patients with SCSC arising in association with TCV were retrieved from our files. The average age at primary presentation was 67 (range 32-92) with a female-to-male ratio of 2:1. Three clinical scenarios for SCSC associated with TCV were identified. These were type I, consisting of TCV with SCSC at the time of presentation (18 patients); type II, consisting of SCSC arising as a recurrence in patients with a known history of TCV (5 patients); and type III, consisting of SCSC presenting as a primary laryngeal squamous cell carcinoma in a patient with or without a known history of TCV (8 patients). The type III cases were of most concern since they often were confused with primary laryngeal squamous cell carcinoma and most often were diagnosed after laryngectomy. CONCLUSIONS: SCSC of the thyroid is almost exclusively associated with TCV and can have variable clinical presentations. SCSC is most commonly seen associated with a primary diagnosis of TCV. SCSC may be seen, however, in patients with recurrent PTC and most importantly may present in a fashion similar to primary laryngeal SCSC. Therefore, caution should be exercised in evaluating laryngeal squamous lesions in patients with known history of TCV and without known risks factors for head and neck carcinogenesis

Mechanisms Responsible for omega-Pore Currents in Ca-v Calcium Channel Voltage-Sensing Domains

Mutations of positively charged amino acids in the S4 transmembrane segment of a voltage-gated ion channel form ion-conducting pathways through the voltage-sensing domain, named ω-current. Here, we used structure modeling and MD simulations to predict pathogenic ω-currents in CaV1.1 and CaV1.3 Ca2+ channels bearing several S4 charge mutations. Our modeling predicts that mutations of CaV1.1-R1 (R528H/G, R897S) or CaV1.1-R2 (R900S, R1239H) linked to hypokalemic periodic paralysis type 1 and of CaV1.3-R3 (R990H) identified in aldosterone-producing adenomas conducts ω-currents in resting state, but not during voltage-sensing domain activation. The mechanism responsible for the ω-current and its amplitude depend on the number of charges in S4, the position of the mutated S4 charge and countercharges, and the nature of the replacing amino acid. Functional characterization validates the modeling prediction showing that CaV1.3-R990H channels conduct ω-currents at hyperpolarizing potentials, but not upon membrane depolarization compared with wild-type channels

MCT1 in Invasive Ductal Carcinoma: Monocarboxylate Metabolism and Aggressive Breast Cancer.

Introduction: Monocarboxylate transporter 1 (MCT1) is an importer of monocarboxylates such as lactate and pyruvate and a marker of mitochondrial metabolism. MCT1 is highly expressed in a subgroup of cancer cells to allow for catabolite uptake from the tumor microenvironment to support mitochondrial metabolism. We studied the protein expression of MCT1 in a broad group of breast invasive ductal carcinoma specimens to determine its association with breast cancer subtypes and outcomes. Methods: MCT1 expression was evaluated by immunohistochemistry on tissue micro-arrays (TMA) obtained through our tumor bank. Two hundred and fifty-seven cases were analyzed: 180 cases were estrogen receptor and/or progesterone receptor positive (ER+ and/or PR+), 62 cases were human epidermal growth factor receptor 2 positive (HER2+), and 56 cases were triple negative breast cancers (TNBC). MCT1 expression was quantified by digital pathology with Aperio software. The intensity of the staining was measured on a continuous scale (0-black to 255-bright white) using a co-localization algorithm. Statistical analysis was performed using a linear mixed model. Results: High MCT1 expression was more commonly found in TNBC compared to ER+ and/or PR+ and compared to HER-2+ (p \u3c 0.001). Tumors with an in-situ component were less likely to stain strongly for MCT1 (p \u3c 0.05). High nuclear grade was associated with higher MCT1 staining (p \u3c 0.01). Higher T stage tumors were noted to have a higher expression of MCT1 (p \u3c 0.05). High MCT1 staining in cancer cells was associated with shorter progression free survival, increased risk of recurrence, and larger size independent of TNBC status (p \u3c 0.05). Conclusion: MCT1 expression, which is a marker of high catabolite uptake and mitochondrial metabolism, is associated with recurrence in breast invasive ductal carcinoma. MCT1 expression as quantified with digital image analysis may be useful as a prognostic biomarker and to design clinical trials using MCT1 inhibitors

Pyrimidine-2,4,6-triones are a new class of voltage-gated L-type Ca(2+) channel activators.

Cav1.2 and Cav1.3 are the main L-type Ca(2+) channel subtypes in the brain. Cav1.3 channels have recently been implicated in the pathogenesis of Parkinson’s disease. Therefore, Cav1.3-selective blockers are developed as promising neuroprotective drugs. We studied the pharmacological properties of a pyrimidine-2,4,6-trione derivative (1-(3-chlorophenethyl)-3-cyclopentylpyrimidine-2,4,6-(1H,3H,5H)-trione, Cp8) recently reported as the first highly selective Cav1.3 blocker. Here we show, in contrast to this previous study, that Cp8 reproducibly increases inward Ca(2+) currents of Cav1.3 and Cav1.2 channels expressed in tsA-201 cells by slowing activation, inactivation and enhancement of tail currents. Similar effects are also observed for native Cav1.3 and Cav1.2 channels in mouse chromaffin cells, while non-L-type currents are unaffected. Evidence for a weak and non-selective inhibition of Cav1.3 and Cav1.2 currents is only observed in a minority of cells using Ba(2+) as charge carrier. Therefore, our data identify pyrimidine-2,4,6-triones as Ca(2+) channel activators

Gating defects of disease-causing de novo mutations in Ca(v)1.3 Ca2+ channels

Recently, we and others identified somatic and germline de novo gain-of-function mutations in CACNA1D, the gene encoding the α1-subunit of voltage-gated Cav1.3 Ca2+-channels. While somatic mutations identified in aldosterone producing adenomas (APAs) underlie treatment-resistant hypertension, germline CACNA1D mutations are associated with a neurodevelopmental disorder characterized by a wide symptomatic spectrum, including autism spectrum disorder. The number of newly identified CACNA1D missense mutations is constantly growing, but their pathogenic potential is difficult to predict in silico, making functional studies indispensable to assess their contribution to disease risk. Here we report the functional characterization of previously identified CACNA1D APA mutations F747L and M1354I using whole-cell patch-clamp electrophysiology upon recombinant expression in tsA-201 cells. We also investigated if alternative splicing of Cav1.3 affects the aberrant gating of the previously characterized APA mutation R990H and two mutations associated with autism spectrum disorder (A479G and G407R). Splice-variant dependent gating changes are of particular interest for germline mutations, since the relative expression of Cav1.3 splice variants differs across different tissues and within brain regions and might therefore result in tissue-specific phenotypes. Our data revealed a complex gain-of-function phenotype for APA mutation F747L confirming its pathogenic role. Furthermore, we found splice-variant dependent gating changes in R990H, A749G and G407R. M1354I did not change channel function of Cav1.3 splice variants and should therefore be considered a rare non-pathogenic variant until further proof for its pathogenicity is obtained. Our new findings together with previously published data allow classification of pathogenic CACNA1D mutations into four categories based on prototypical functional changes

Stromal Monocarboxylate Transporter MCT4 is a Poor Prognostic Factor in Squamous Cell Carcinoma

ABSTRACT Introduction: Monocarboxylate transporter 4 (MCT4) is the main exporter of lactate out of cells. It is also a critical component in the glycolytic metabolism of cancer cells. In this study, stromal MCT4 in oral SCC was correlated with risk of recurrence (ROR), extent of primary tumor (pT) and nodal metastasis (pN), perineural invasion (PNI), lymphovascular invasion (LVI), HPV status, extracapsular extension (ECE) and positive margin. Methods: Clinical data were collected for 86 consecutive patients with oral HNSCC. Tissue microarrays (TMA) were constructed from paraffin blocks of resection specimens and stained for MCT4. Immunohistochemistry (IHC) staining was assessed and quantified by digital image analysis with Aperio software. Using a co-localization algorithm we assessed the intensity of staining and the percentage of positive cells in the tumoral stromal cells. Correlations of MCT4 expression with clinicopathological features and survival were studied. Results: Increased IHC staining for MCT4 was strongly associated with an increased risk of recurrence, OR 1.96 (95%CI: 1.17-3.40), presence of PNI, OR 2.25 (95%CI: 1.33-3.95), higher pT, OR 1.68 (95%CI: 0.99-2.89), higher pN, OR 2.07 (95%CI: 1.25-3.57) and presence of LVI, OR 2.21 (95%CI: 1.11-4.67). We didn’t find any significant association between stromal MCT4 expression and HPV status, presence of ECE or positive margin. Conclusions: This study demonstrates that MCT4, a marker of glycolysis in cancer-associated stroma, is highly expressed in oral SCC. The IHC staining pattern of stromal MCT4 suggests that high MCT4 expression appears to be a useful marker for tumor progression and prognosis. We propose MCT4 serves as a new prognostic factor in oral SCC and can act as a potential therapeutic target marker considering pharmacological development of MCT4 inhibitors

Elevated APOBEC mutational signatures implicate chronic injury in etiology of an aggressive head-and-neck squamous cell carcinoma: a case report.

BACKGROUND: Aggressive squamous cell carcinomas (SCCs) present frequently in the context of chronic skin injury occurring in patients with the congenital blistering disease recessive dystrophic epidermolysis bullosa. Recently, these cancers were shown to harbor mutation signatures associated with endogenous deaminases of the active polynucleotide cytosine deaminase family, collectively termed APOBEC, and clock-like COSMIC [Catalogue of Somatic Mutations in Cancer] signatures, which are associated with normal aging and might result from cumulative DNA replication errors. We present a case of a nasal septal SCC arising in the context of recurrent injury, but also modest past tobacco use. Our genetic analysis of this tumor reveals unusually high APOBEC and clock-like but low tobacco-related COSMIC signatures, suggesting that chronic injury may have played a primary role in somatic mutation. This case report demonstrates how signature-based analyses may implicate key roles for certain mutagenic forces in individual malignancies such as head-and-neck SCC, with multiple etiological origins. CASE PRESENTATION: We report the case of a 43-year-old male former smoker who presented with congestion and swelling following a traumatic nasal fracture. During surgery, the mucosa surrounding the right nasal valve appeared abnormal, and biopsies revealed invasive keratinizing SCC. Frozen section biopsies revealed multiple areas to be positive for SCC. Gene sequencing showed loss of PTEN (exons 2-8), CDKN2A/B and TP53 (exons 8-9), MYC amplification, and BLM S338*. Exome sequencing data also revealed that 36% of mutations matched an APOBEC mutational signature (COSMIC signatures 2 and 13) and 53% of mutations matched the clock-like mutation signature (COSMIC signature 5). These proportions place this tumor in the 90th percentile bearing each signature, independently, in a reference data set combining cutaneous and The Cancer Genome Atlas (TCGA) head and neck SCC data. In contrast, few mutations harbored a tobacco-related COSMIC signature 4, representing about the 10th percentile in our reference SCC data set. The patient was treated with partial rhinectomy with local flap reconstruction, bilateral neck dissection, and adjuvant radiation therapy; the patient remains disease-free to date. CONCLUSION: Based on comparative mutational signature analysis, we propose that the history of tobacco use and traumatic injury may have collaborated in activating APOBEC enzymes and the clock-like mutational process, ultimately leading to cancer formation. Clinical awareness of the relationship between epithelial injury and tumorigenesis should enhance earlier detection of this particularly aggressive type of cancer

The priming effect of extracellular UTP on human neutrophils: Role of calcium released from thapsigargin-sensitive intracellular stores

P2Y2 receptors, which are equally responsive to ATP and UTP, can trigger intracellular signaling events, such as intracellular calcium mobilization and mitogen-activated protein (MAP) kinase phosphorylation in polymorphonuclear leukocytes (PMN). Moreover, extracellular nucleotides have been shown to prime chemoattractant-induced superoxide production. The aim of our study was to investigate the mechanism responsible for the priming effect of extracellular nucleotides on reactive oxygen species (ROS) production induced in human neutrophils by two different chemoattractants: formyl-methionyl-leucyl-phenylalanine (fMLP) and interleukin-8 (IL-8). Nucleotide-induced priming of ROS production was concentration- and time-dependent. When UTP was added to neutrophil suspensions prior to chemoattractant, the increase of the response reached the maximum at 1 min of pre-incubation with the nucleotide. UTP potentiated the phosphorylation of p44/42 and p38 MAP kinases induced by chemoattractants, however the P2 receptor-mediated potentiation of ROS production was still detectable in the presence of a SB203580 or U0126, supporting the view that MAP kinases do not play a major role in regulating the nucleotide-induced effect. In the presence of thapsigargin, an inhibitor of the ubiquitous sarco-endoplasmic reticulum Ca2+-ATPases in mammalian cells, the effect of fMLP was not affected, but UTP-induced priming was abolished, suggesting that the release of calcium from thapsigargin-sensitive intracellular stores is essential for nucleotide-induced priming in human neutrophils

Substance P Induces Rapid and Transient Membrane Blebbing in U373MG Cells in a p21-Activated Kinase-Dependent Manner

U373MG astrocytoma cells endogenously express the full-length neurokinin 1 receptor (NK1R). Substance P (SP), the natural ligand for NK1R, triggers rapid and transient membrane blebbing and we report that these morphological changes have different dynamics and intracellular signaling as compared to the changes that we have previously described in HEK293-NK1R cells. In both cell lines, the SP-induced morphological changes are Gq-independent, and they require the Rho, Rho-associated coiled-coil kinase (ROCK) signaling pathway. Using confocal microscopy we have demonstrated that tubulin is phosphorylated subsequent to cell stimulation with SP and that tubulin accumulates inside the blebs. Colchicine, a tubulin polymerization inhibitor, blocked SP-induced blebbing in U373MG but not in HEK293-NK1R cells. Although p21-activated kinase (PAK) is expressed in both cell lines, SP induced rapid phosphorylation of PAK in U373MG, but failed to phosphorylate PAK in HEK293-NK1R cells. The cell-permeable Rho inhibitor C3 transferase inhibited SP-induced PAK phosphorylation, but the ROCK inhibitor Y27632 had no effect on PAK phosphorylation, suggesting that Rho activates PAK in a ROCK-independent manner. Our study demonstrates that SP triggers rapid changes in cell morphology mediated by distinct intracellular signaling mechanisms in U373MG versus HEK293-NK1R cells