Search Results (16)

Pheochromocytomas (PCCs) are tumors arising from chromaffin cells in the adrenal medulla, and paragangliomas (PGLs) are tumors derived from extra-adrenal sympathetic or parasympathetic paraganglia; these tumors are collectively referred to as PPGL cancer. Treatment for PPGL primarily involves surgical removal of the tumor, and only limited options are available for treatment of the disease once it becomes metastatic. Human carriers of the heterozygous mutations in the succinate dehydrogenase subunit B (SDHB) gene are susceptible to the development of PPGL. A physiologically relevant PCC patient-derived cell line hPheo1 was developed, and SDHB_KD cells carrying a stable short hairpin knockdown of SDHB were derived from it. An untargeted metabolomic approach uncovered an overactive polyamine pathway in the SDHB_KD cells that was subsequently fully validated in a large set of human SDHB-mutant PPGL tumor samples. We previously reported that treatment with the polyamine metabolism inhibitor N¹,N¹¹-diethylnorspermine (DENSPM) drastically inhibited growth of these PCC-derived cells in culture as well as in xenograft mouse models. Here we explored the mechanisms underlying DENSPM action in hPheo1 and SDHB_KD cells. Specifically, by performing an RNAseq analysis, we have identified gene expression changes associated with DENSPM treatment that broadly interfere with all aspects of lipid metabolism, including fatty acid (FA) synthesis, desaturation, and import/uptake. Furthermore, by performing an untargeted lipidomic liquid chromatography–mass spectrometry (LC/MS)-based analysis we uncovered specific groups of lipids that are dramatically reduced as a result of DENSPM treatment. Specifically, the bulk of plasmanyl ether lipid species that have been recently reported as the major determinants of cancer cell fate are notably decreased. In summary, this work suggests an intersection between active polyamine and lipid pathways in PCC cells. Full article

Phaeochromocytomas and paragangliomas (PPGLs) are rare neuroendocrine tumours arising from chromaffin cells. Pathogenic variants in the gene succinate dehydrogenase subunit B (SDHB) are associated with malignancy and poor prognosis. When metastases arise, limited treatment options are available. The pathomechanism of SDHB-associated PPGL remains largely unknown, and the lack of suitable models hinders therapy development. Germline heterozygous SDHB pathogenic variants predispose to developing PPGLs with a life-long penetrance of around 50%. To mimic the human disease phenotype, we characterised adult heterozygous sdhb mutant zebrafish as a potential model to study SDHB-related PPGLs. Adult sdhb mutant zebrafish did not develop an obvious tumour phenotype and were anatomically and histologically like their wild-type siblings. However, sdhb mutants showed significantly increased succinate levels, a major hallmark of SDHB-related PPGLs. While basal activity was increased during day periods in mutants, mitochondrial complex activity and catecholamine metabolite levels were not significantly different. In conclusion, we characterised an adult in vivo zebrafish model, genetically resembling human carriers. Adult heterozygous sdhb mutants mimicked their human counterparts, showing systemic elevation of succinate levels despite the absence of a tumour phenotype. This model forms a promising basis for developing a full tumour phenotype and gaining knowledge of the pathomechanism behind SDHB-related PPGLs. Full article

Gastrointestinal stromal tumors (GISTs) are the most common mesenchymal tumors of the gastrointestinal tract, with proto-oncogene, receptor tyrosine kinase (c-kit), or PDGFRα mutations detected in around 85% of cases. GISTs without c-kit or platelet-derived growth factor receptor alpha (PDGFRα) mutations are considered wild-type (WT), and their diverse molecular alterations and biological behaviors remain uncertain. They are usually not sensitive to tyrosine kinase inhibitors (TKIs). Recently, some molecular alterations, including neurotrophic tyrosine receptor kinase (NTRK) fusions, have been reported in very few cases of WT GISTs. This novel finding opens the window for the use of tropomyosin receptor kinase (TRK) inhibitor therapy in these subtypes of GIST. Herein, we report a new case of NTRK-fused WT high-risk GIST in a female patient with a large pelvic mass (large dimension of 20 cm). The tumor was removed, and the histopathology displayed spindle-predominant morphology with focal epithelioid areas, myxoid stromal tissue, and notable lymphoid infiltration with tertiary lymphoid structures. Ten mitoses were quantified in 50 high-power fields without nuclear pleomorphism. DOG1 showed strong and diffuse positivity, and CD117 showed moderate positivity. Succinate dehydrogenase subunit B (SDHB) was retained, Pan-TRK was focal positive (nuclear pattern), and the proliferation index Ki-67 was 7%. Next-generation sequencing (NGS) detected an ETV6::NTRK3 fusion, and this finding was confirmed by fluorescence in situ hybridization (FISH), which showed NTRK3 rearrangement. In addition, an RB1 mutation was found by NGS. The follow-up CT scan revealed peritoneal nodules suggestive of peritoneal dissemination, and Entrectinib (a TRK inhibitor) was administered. After 3 months of follow-up, a new CT scan showed a complete response. Based on our results and the cases from the literature, GISTs with NTRK fusions are very uncommon so far; hence, further screening studies, including more WT GIST cases, may increase the possibility of finding additional cases. The present case may offer new insights into the potential introduction of TRK inhibitors as treatments for GISTs with NTRK fusions. Additionally, the presence of abundant lymphoid infiltration in the present case may prompt further research into immunotherapy as a possible additional therapeutic option. Full article

Skin cancer is the most common malignant disease worldwide and, therefore, also poses a challenge from a pharmacotherapeutic perspective. Derivatives of indirubin are an interesting option in this context. In the present study, the effects of 3-[3′-oxo-benzo[b]thiophen-2′-(Z)-ylidene]-1-(β-d-glucopyranosyl)-oxindole (KD87), a thia-analogous indirubin N-glycoside, on the viability and mitochondrial properties of melanoma (A375) and squamous cell carcinoma cells (A431) of the skin were investigated. In both cell lines, KD87 caused decreased viability, the activation of caspases-3 and -7, and the inhibition of colony formation. At the mitochondrial level, a concentration-dependent decrease in both the basal and ATP-linked oxygen consumption rate and in the reserve capacity of oxidative respiration were registered in the presence of KD87. These changes were accompanied by morphological alterations in the mitochondria, a release of mitochondrial cytochrome c into the cytosol and significant reductions in succinate dehydrogenase complex subunit B (SDHB, subunit of complex II) in A375 and A431 cells and NADH:ubiquinone oxidoreductase subunit B8 (NDUFB8, subunit of complex I) in A375 cells. The effect of KD87 was accompanied by a significant upregulation of the enzyme heme oxygenase-1, whose inhibition led to a partial but significant reduction in the metabolic-activity-reducing effect of KD87. In summary, our data show a mitochondria-targeting effect of KD87 as part of the cytotoxic effect of this compound on skin cancer cells, which should be considered in future studies with this class of compounds. Full article

The oriental river prawn (Macrobrachium nipponense) is a commercially important species in Asia. A previous study showed that the succinate dehydrogenase complex iron sulfur subunit B (SDHB) gene participates in testes development in this species through its effect on the expression of the insulin-like androgenic gland hormone gene. This study knocked-down the Mn-SDHB genes in M. nipponense using RNAi. A transcriptome analysis of the androgenic gland and testes was then performed to discover the male sex-related genes regulated by SDHB and investigate the mechanism of male sexual development in this species. More than 16,623 unigenes were discovered in each sample generated. In the androgenic gland, most of the differentially expressed genes were enriched in the hypertrophic cardiomyopathy pathway, while in the testes, they were enriched in the citrate cycle pathway. In addition, after Mn-SDHB knockdown, five genes were found to be downregulated in the androgenic gland in a series of biological processes associated with phosphorylated carbohydrate synthesis and transformations in the glycolysis/gluconeogenesis pathway. Moreover, a total of nine male sex-related genes were identified including Pro-resilin, insulin-like androgenic gland hormone, Protein mono-ADP-ribosyltransferase PAPR11, DNAJC2, C-type Lectin-1, Tyrosine-protein kinase Yes, Vigilin, and Sperm motility kinase Y-like, demonstrating the regulatory effects of Mn-SDHB, and providing a reference for the further study of the mechanisms of male development in M. nipponense. Full article

Pheochromocytoma (PHEO) and paraganglioma (PGL) are rare neuroendocrine tumors derived from neural crest cells. Germline variants in approximately 20 PHEO/PGL susceptibility genes are found in about 40% of patients, half of which are found in the genes that encode succinate dehydrogenase (SDH). Patients with SDH subunit B (SDHB)-mutated PHEO/PGL exhibit a higher likelihood of developing metastatic disease, which can be partially explained by the metabolic cell reprogramming and redox imbalance caused by the mutation. Reactive oxygen species (ROS) are highly reactive molecules involved in a multitude of important signaling pathways. A moderate level of ROS production can help regulate cellular physiology; however, an excessive level of oxidative stress can lead to tumorigenic processes including stimulation of growth factor-dependent pathways and the induction of genetic instability. Tumor cells effectively exploit antioxidant enzymes in order to protect themselves against harmful intracellular ROS accumulation, which highlights the essential balance between ROS production and scavenging. Exploiting ROS accumulation can be used as a possible therapeutic strategy in ROS-scavenging tumor cells. Here, we focus on the role of ROS production in PHEO and PGL, predominantly in SDHB-mutated cases. We discuss potential strategies and approaches to anticancer therapies by enhancing ROS production in these difficult-to-treat tumors. Full article

Zymoseptoria tritici (Zt) populations adapt under the selection pressure of fungicides applied for disease control. The primary objective of this study was to assess fungicide sensitivity in the Estonian Zt population. A total of 282 Zt isolates from 2019 and 2020 were tested for sensitivity to azoles (DMIs; prothioconazole-desthio, epoxiconazole, mefentrifluconazole) and succinate dehydrogenase inhibitors (SDHIs; boscalid, fluxapyroxad). The efficacy of the tested fungicides varied considerably between the Estonian counties, but the Zt population is mainly sensitive to DMIs. Additionally, the frequencies of CYP51 gene alterations varied; D134G, V136C, A379G, and S524T had increased, but V136A and I381V showed a moderate decrease in 2020 in comparison to 2019. Sensitivity to SDHIs was stable, but boscalid was less effective than fluxapyroxad. SdhC gene mutations C-T33N, C-T34N, and C-N86S were common, but not linked with SDHI fungicide sensitivity assay results. Otherwise, mutation B-N225I in the SdhB subunit occurred in isolates with reduced sensitivity to SDHIs. Sensitivity to strobilurins was evaluated by the mutation G143A in the CytB gene, which was present in nearly half of the population. The data presented confirm the ongoing evolution of fungicide sensitivity in the Zt population in Estonia and highlight the importance of knowledge-based decisions for optimizing anti-resistance strategies in the field. Full article

Members of the Tribbles (TRIB) family of pseudokinases are critical components of intracellular signal transduction pathways in physiological and pathological processes. TRIBs, including TRIB2, have been previously shown as signaling mediators and scaffolding proteins regulating numerous cellular events such as proliferation, differentiation and cell death through protein stability and activity. However, the signaling network associated with TRIB2 and its binding partners in granulosa cells during ovarian follicular development is not fully defined. We previously reported that TRIB2 is differentially expressed in growing dominant follicles while downregulated in ovulatory follicles following the luteinizing hormone (LH) surge or human chorionic gonadotropin (hCG) injection. In the present study, we used the yeast two-hybrid screening system and in vitro coimmunoprecipitation assays to identify and confirm TRIB2 interactions in granulosa cells (GCs) of dominant ovarian follicles (DFs), which yielded individual candidate binding partners including calmodulin 1 (CALM1), inhibin subunit beta A (INHBA), inositol polyphosphate phosphatase-like 1 (INPPL1), 5′-nucleotidase ecto (NT5E), stearoyl-CoA desaturase (SCD), succinate dehydrogenase complex iron sulfur subunit B (SDHB) and Ras-associated protein 14 (RAB14). Further analyses showed that all TRIB2 binding partners are expressed in GCs of dominant follicles but are differentially regulated throughout the different stages of follicular development. CRISPR/Cas9-driven inhibition along with pQE-driven overexpression of TRIB2 showed that TRIB2 differently regulates expression of binding partners, which reveals the importance of TRIB2 in the control of gene expression linked to various biological processes such as proliferation, differentiation, cell migration, apoptosis, calcium signaling and metabolism. These data provide a larger view of potential TRIB2-regulated signal transduction pathways in GCs and provide strong evidence that TRIB2 may act as a regulator of target genes during ovarian follicular development. Full article

Mitochondrial dysfunction and systemic inflammation are major factors in the development of sarcopenia, but the molecular determinants linking the two mechanisms are only partially understood. The study of extracellular vesicle (EV) trafficking may provide insights into this relationship. Circulating small EVs (sEVs) from serum of 11 older adults with physical frailty and sarcopenia (PF&S) and 10 controls were purified and characterized. Protein levels of three tetraspanins (CD9, CD63, and CD81) and selected mitochondrial markers, including adenosine triphosphate 5A (ATP5A), mitochondrial cytochrome C oxidase subunit I (MTCOI), nicotinamide adenine dinucleotide reduced form (NADH):ubiquinone oxidoreductase subunit B8 (NDUFB8), NADH:ubiquinone oxidoreductase subunit S3 (NDUFS3), succinate dehydrogenase complex iron sulfur subunit B (SDHB), and ubiquinol-cytochrome C reductase core protein 2 (UQCRC2) were quantified by Western immunoblotting. Participants with PF&S showed higher levels of circulating sEVs relative to controls. Protein levels of CD9 and CD63 were lower in the sEV fraction of PF&S older adults, while CD81 was unvaried between groups. In addition, circulating sEVs from PF&S participants had lower amounts of ATP5A, NDUFS3, and SDHB. No signal was detected for MTCOI, NDUFB8, or UQCRC2 in either participant group. Our findings indicate that, in spite of increased sEV secretion, lower amounts of mitochondrial components are discarded through EV in older adults with PF&S. In-depth analysis of EV trafficking might open new venues for biomarker discovery and treatment development for PF&S. Full article

Systemic inflammation and mitochondrial dysfunction are involved in neurodegeneration in Parkinson’s disease (PD). Extracellular vesicle (EV) trafficking may link inflammation and mitochondrial dysfunction. In the present study, circulating small EVs (sEVs) from 16 older adults with PD and 12 non-PD controls were purified and characterized. A panel of serum inflammatory biomolecules was measured by multiplex immunoassay. Protein levels of three tetraspanins (CD9, CD63, and CD81) and selected mitochondrial markers (adenosine triphosphate 5A (ATP5A), mitochondrial cytochrome C oxidase subunit I (MTCOI), nicotinamide adenine dinucleotide reduced form (NADH):ubiquinone oxidoreductase subunit B8 (NDUFB8), NADH:ubiquinone oxidoreductase subunit S3 (NDUFS3), succinate dehydrogenase complex iron sulfur subunit B (SDHB), and ubiquinol-cytochrome C reductase core protein 2 (UQCRC2)) were quantified in purified sEVs by immunoblotting. Relative to controls, PD participants showed a greater amount of circulating sEVs. Levels of CD9 and CD63 were lower in the sEV fraction of PD participants, whereas those of CD81 were similar between groups. Lower levels of ATP5A, NDUFS3, and SDHB were detected in sEVs from PD participants. No signal was retrieved for UQCRC2, MTCOI, or NDUFB8 in either participant group. To identify a molecular signature in circulating sEVs in relationship to systemic inflammation, a low level-fused (multi-platform) partial least squares discriminant analysis was applied. The model correctly classified 94.2% ± 6.1% PD participants and 66.7% ± 5.4% controls, and identified seven biomolecules as relevant (CD9, NDUFS3, C-reactive protein, fibroblast growth factor 21, interleukin 9, macrophage inflammatory protein 1β, and tumor necrosis factor alpha). In conclusion, a mitochondrial signature was identified in circulating sEVs from older adults with PD, in association with a specific inflammatory profile. In-depth characterization of sEV trafficking may allow identifying new biomarkers for PD and possible targets for personalized interventions. Full article

Succinate dehydrogenase subunit B (SDHB) deficiency frequently occurs in cluster I pheochromocytomas and paragangliomas (PCPGs). SDHB-mutated PCPGs are characterized by alterations in the electron transport chain, metabolic reprogramming of the tricarboxylic cycle, and elevated levels of reactive oxygen species (ROS). We discovered that SDHB-deficient PCPG cells exhibit increased oxidative stress burden, which leads to elevated demands for glutathione metabolism. Mechanistically, nuclear factor erythroid 2-related factor 2 (NRF2)-guided glutathione de novo synthesis plays a key role in supporting cellular survival and the proliferation of SDHB-knockdown (SDHB^KD) cells. NRF2 blockade not only disrupted ROS homeostasis in SDHB-deficient cells but also caused severe cytotoxicity by the accumulation of DNA oxidative damage. Brusatol, a potent NRF2 inhibitor, showed a promising effect in suppressing SDHB^KD metastatic lesions in vivo, with prolonged overall survival in mice bearing PCPG allografts. Our findings highlight a novel therapeutic strategy of targeting the NRF2-driven glutathione metabolic pathway against SDHB-mutated PCPG. Full article

Pancreatic ductal adenocarcinoma (PDAC) is amongst the most fatal malignancies and its development is highly associated with inflammatory processes such as chronic pancreatitis (CP). Since the succinate dehydrogenase subunit B (SDHB) is regarded as tumor suppressor that is lost during cancer development, this study investigated the impact of M1-macrophages as part of the inflammatory microenvironment on the expression as well as function of SDHB in benign and premalignant pancreatic ductal epithelial cells (PDECs). Immunohistochemical analyses on pancreatic tissue sections from CP patients and control individuals revealed a stronger SDHB expression in ducts of CP tissues being associated with a greater abundance of macrophages compared to ducts in control tissues. Accordingly, indirect co-culture with M1-macrophages led to clearly elevated SDHB expression and SDH activity in benign H6c7-pBp and premalignant H6c7-kras PDECs. While siRNA-mediated SDHB knockdown in these cells did not affect glucose and lactate uptake after co-culture, SDHB knockdown significantly promoted PDEC growth which was associated with increased proliferation and decreased effector caspase activity particularly in co-cultured PDECs. Overall, these data indicate that SDHB expression and SDH activity are increased in PDECs when exposed to pro-inflammatory macrophages as a counterregulatory mechanism to prevent excessive PDEC growth triggered by the inflammatory environment. Full article

Malignant pheochromocytomas (PCs) and paragangliomas (PGLs) are rare neuroendocrine neoplasms defined by the presence of distant metastases. There is currently a relatively paucity of data regarding the natural history of PCs/PGLs and the optimal approach to their treatment. We retrospectively analyzed the clinical, biochemical, imaging, genetic and histopathological characteristics of fourteen patients with metastatic PCs/PGLs diagnosed over 15 years, along with their response to treatment. Patients were followed-up for a median of six years (range: 1–14 years). Six patients had synchronous metastases and the remaining developed metastases after a median of four years (range 2–10 years). Genetic analysis of seven patients revealed that three harbored succinate dehydrogenase subunit B/D gene (SDHB/D) mutations. Hormonal hypersecretion occurred in 70% of patients; normetanephrine, either alone or with other concomitant hormones, was the most frequent secretory component. Patients were administered multiple first and subsequent treatments including surgery (n = 12), chemotherapy (n = 7), radionuclide therapy (n = 2) and radiopeptides (n = 5). Seven patients had stable disease, four had progressive disease and three died. Ectopic hormonal secretion is rare and commonly encountered in benign PCs. Ectopic secretion of interleukin-6 in one of our patients, prompted a literature review of ectopic hormonal secretion, particularly from metastatic PCs/PGLs. Only four cases of metastatic PC/PGLs with confirmed ectopic secretion of hormones or peptides have been described so far. Full article

Recently, genetic alterations in the genes encoding succinate dehydrogenase subunit B and D (SDHB and SDHD) were identified in pet dogs that presented with spontaneously arising pheochromocytomas (PCC) and paragangliomas (PGL; together PPGL), suggesting dogs might be an interesting comparative model for the study of human PPGL. To study whether canine PPGL resembled human PPGL, we investigated a series of 50 canine PPGLs by immunohistochemistry to determine the expression of synaptophysin (SYP), tyrosine hydroxylase (TH) and succinate dehydrogenase subunit A (SDHA) and B (SDHB). In parallel, 25 canine PPGLs were screened for mutations in SDHB and SDHD by Sanger sequencing. To detect large chromosomal alterations, single nucleotide polymorphism (SNP) arrays were performed for 11 PPGLs, including cases for which fresh frozen tissue was available. The immunohistochemical markers stained positive in the majority of canine PPGLs. Genetic screening of the canine tumors revealed the previously described variants in four cases; SDHB p.Arg38Gln (n = 1) and SDHD p.Lys122Arg (n = 3). Furthermore, the SNP arrays revealed large chromosomal alterations of which the loss of chromosome 5, partly homologous to human chromosome 1p and chromosome 11, was the most frequent finding (100% of the six cases with chromosomal alterations). In conclusion, canine and human PPGLs show similar genomic alterations, suggestive of common interspecies PPGL-related pathways. Full article

Germline mutations in succinate dehydrogenase subunit B and D (SDHB and SDHD) are predisposed to hereditary paraganglioma (PGL) and pheochromocytoma (PHEO). The phenotype of pathogenic variants varies according to the causative gene. In this retrospective study, we estimate the mortality of a nationwide cohort of SDHB variant carriers and that of a large cohort of SDHD variant carriers and compare it to the mortality of a matched cohort of the general Dutch population. A total of 192 SDHB variant carriers and 232 SDHD variant carriers were included in this study. The Standard Mortality Ratio (SMR) for SDHB mutation carriers was 1.89, increasing to 2.88 in carriers affected by PGL. For SDHD variant carriers the SMR was 0.93 and 1.06 in affected carriers. Compared to the general population, mortality seems to be increased in SDHB variant carriers, especially in those affected by PGL. In SDHD variant carriers, the mortality is comparable to that of the general Dutch population, even if they are affected by PGL. This insight emphasizes the significance of DNA-testing in all PGL and PHEO patients, since different clinical risks may warrant gene-specific management strategies. Full article