The New Nematicide Cyclobutrifluram Targets the Mitochondrial Succinate Dehydrogenase Complex in Bursaphelenchus xylophilus.

Bursaphelenchus xylophilus is a dangerous quarantine pest that causes extensive damage to pine ecosystems worldwide. Cyclobutrifluram, a succinate dehydrogenase inhibitor (SDHI), is a novel nematicide introduced by Syngenta in 2013. However, the nematocidal effect of cyclobutrifluram against plant-parasitic nematodes remains underexplored. Therefore, here, we aim to address this knowledge gap by evaluating the toxicity, effects, and mode of action of cyclobutrifluram on B. xylophilus. The result shows that cyclobutrifluram is the most effective agent, with an LC50 value of 0.1078 mg·L-1. At an LC20 dose, it significantly reduced the population size to 10.40 × 103 ± 737.56-approximately 1/23 that of the control group. This notable impact may stem from the agent's ability to diminish egg-laying and hatching rates, as well as to impede the nematodes' development. In addition, it has also performed well in the prevention of pine wilt disease, significantly reducing the incidence in greenhouses and in the field. SDH consists of a transmembrane assembly composed of four protein subunits (SDHA to SDHD). Four sdh genes were characterized and proved by RNAi to regulate the spawning capacity, locomotion ability, and body size of B. xylophilus. The mortality of nematodes treated with sdhc-dsRNA significantly decreased upon cyclobutrifluram application. Molecular docking further confirmed that SDHC, a cytochrome-binding protein, is the target. In conclusion, cyclobutrifluram has a good potential for trunk injection against B. xylophilus. This study provides valuable information for the screening and application of effective agents in controlling and preventing PWD in forests.

Characterisation of an Adult Zebrafish Model for SDHB-Associated Phaeochromocytomas and Paragangliomas.

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.

[The relationship between clinical characteristics and pathological features in patients with pheochromocytomas/paragangliomas].

Objective: To explore the correlation between clinical characteristics and pathological features in patients with pheochromocytoma/paraganglioma (PPGLs). Methods: A case series study. A retrospective analysis was conducted on patients with single and primary PPGLs after postoperative pathological diagnosis who were admitted to Peking Union Medical College Hospital between January 2019 and December 2022. The patients were divided into the Ki-67<3% group and the Ki-67≥3% group with Ki-67 proliferation index of 3% as the threshold. The relationship between clinical and pathological characteristics of PPGLs was analyzed. Results: A total of 399 PPGLs patients were included, with 177 males and 222 females, aged [M(Q1, Q3)] 45.0(35.5, 53.0) years. Among them, 226 (56.6%) cases originated from the adrenal gland, while 104 cases (26.1%) from the retroperitoneum. 20.9% (27/129) of the patients were found to harbor germline mutations of susceptibility genes, with SDHB mutations being the most common (10.1%, 13/129). The Ki-67 staining was performed on 302 cases, with a Ki-67 proliferation index [M(Q1, Q3)] of 2.0% (1.0%, 3.0%). There were 194 cases in Ki-67<3% group and 108 cases in Ki-67≥3% group. Compared with the patients in Ki-67<3% group, the age of onset in Ki-67≥3% group was younger (P=0.029). Compared with the patients with paragangliomas without SDHB or Cluster 1A-related gene mutations, positive 131I-meta-iodobenzylguanidine (131I-MIBG) imaging or negative O-6-methylguanine-DNA methyltransferase (MGMT) immunohistochemistry staining, those with SDHB or Cluster 1A-related gene mutations, negative 131I-MIBG imaging or positive MGMT immunohistochemistry staining had a higher Ki-67 index (all P<0.05). Compared with adrenal pheochromocytoma, retroperitoneal paragangliomas had a higher proportion of SDHB mutations and a higher proportion of normetanephrine (NMN) secretory types (all P<0.05). Compared with adrenal pheochromocytoma, the maximum diameter of head and neck paraganglioma tumors was smaller [3.0 (1.9, 3.8) cm vs 4.7 (3.4, 6.4) cm, P<0.001] and the proportion of Ki-67≥3% was higher (61.3% vs 33.8%, P=0.007). Conclusions: PPGLs patients with earlier onset age, SDHB or Cluster 1A-related gene mutations, negative 131I-MIBG imaging, or positive MGMT immunohistochemistry staining tend to have a higher Ki-67 index. Head and neck tumors, though smaller, exhibit a higher proliferation potential.

Design, Synthesis, and Antifungal Activities of Novel Potent Fluoroalkenyl Succinate Dehydrogenase Inhibitors.

The development of new fungicide molecules is a crucial task for agricultural chemists to enhance the effectiveness of fungicides in agricultural production. In this study, a series of novel fluoroalkenyl modified succinate dehydrogenase inhibitors were synthesized and evaluated for their antifungal activities against eight fungi. The results from the in vitro antifungal assay demonstrated that compound 34 exhibited superior activity against Rhizoctonia solani with an EC50 value of 0.04 µM, outperforming commercial fluxapyroxad (EC50 = 0.18 µM) and boscalid (EC50 = 3.07 µM). Furthermore, compound 34 showed similar effects to fluxapyroxad on other pathogenic fungi such as Sclerotinia sclerotiorum (EC50 = 1.13 µM), Monilinia fructicola (EC50 = 1.61 µM), Botrytis cinerea (EC50 = 1.21 µM), and also demonstrated protective and curative efficacies in vivo on rapeseed leaves and tomato fruits. Enzyme activity experiments and protein-ligand interaction analysis by surface plasmon resonance revealed that compound 34 had a stronger inhibitory effect on succinate dehydrogenase compared to fluxapyroxad. Additionally, molecular docking and DFT calculation confirmed that the fluoroalkenyl unit in compound 34 could enhance its binding capacity with the target protein through p-π conjugation and hydrogen bond interactions.

Analysis of clinical and pathological characteristics of retroperitoneal paraganglioma and associated prognostic factors.

BACKGROUND AND OBJECTIVES: The aim of this study is to explore the long-term prognostic risk factors associated with patients diagnosed with retroperitoneal paraganglioma (RPGL) and examine their clinical and pathological characteristics. METHODS: Expressions of biomarkers were identified using immunohistochemistry (IHC) and case databases were retrospectively searched. Survival analysis was performed using Kaplan-Meier and Cox risk regression to identify the factors that influence the postoperative progression-free survival of patients with RPGL. RESULTS: A total of 105 patients, most of whom had tumors situated in the paraaortic region, and whose average tumor size was 8.6 cm, were enrolled in this study. The average follow-up duration was 51 months, with a mortality rate of 19% and a recurrence and metastasis rate of 41.9%. Tumors were assessed using the modified Grading system for Adrenal Pheochromocytoma and Paraganglioma (GAPP), and SDHB, S-100, and Ki-67 were stained using IHC in all cases. Out of the total cases examined, negative in SDHB expression were observed in 18.1% of cases, S-100 expression was negative in 36.2% of cases, and endovascular tumor enboluswas present in approximately 25.7% of cases. The results of the univariate analysis indicated that several factors significantly influenced the progression-free survival of patients with PGL as follow: maximum tumor diameter (>5.5 cm), tumor morphological features, tumor grading (modified GAPP score > 6), SDHB negative, S-100 negative, and expression of proliferation index Ki-67 (>3%) (X2 = 4.217-27.420, p < 0.05). The results of the multivariate analysis indicated that negative of S-100 (p = 0.021) and SDHB (p = 0.038), as well as intravascular tumor thrombus (p = 0.047) expression were independent risk factors for progression-free survival in patients. CONCLUSION: RPGL is characterized by diverse biological features and an elevated susceptibility to both recurrence and metastasis. Both SDHB and S-100 can be employed as traditional IHC indicators to predict the metastatic risk of PGL, whereas the tumor histomorphology-endovascular tumor enbolus assists in determining the metastasis risk of RPGL.

Case report: A novel somatic SDHB variant in a patient with bladder paraganglioma.

Background: Paragangliomas (PGL) are rare neuroendocrine tumors derived from the autonomic nervous system paraganglia. Urinary bladder paragangliomas (UBPGL) originate from the sympathetic neurons of the urinary bladder wall and represent 0.7% of all paragangliomas and <0.05% of all bladder tumors. PGL and UBPGL can be associated with SDHB, SDHD, NF1, and VHL gene variants, with the most common germline alterations found in SDHB and VHL. Case report: We report a case of a 42-year-old woman who presented with menorrhagia/hematuria, uterine leiomyomas, as well as cardiac and bladder masses. The cardiac mass was favored to be a myxoma based on clinical findings, while the bladder mass was diagnosed as UBPGL. A novel SDHB mutation (c.642G>A, p Q214Q), detected in the UBPGL, was proven to be somatic. Although this variant was seemingly synonymous, it was predicted to have a loss of function due to the splice site effect, which was further supported by the immunohistochemical loss of SDHB. Conclusion: This case highlights the challenges of diagnosing an extremely rare entity, bladder paraganglioma, with an emphasis on the multidisciplinary approach to navigate various clinical and imaging findings that may initially be misleading. In addition, a novel loss of function SDHB variant that could have been overlooked as a synonymous variant is herein reported, while also illustrating the importance of both germline and somatic mutation testing.

A myzozoan-specific protein is an essential membrane-anchoring component of the succinate dehydrogenase complex in Toxoplasma parasites.

Succinate dehydrogenase (SDH) is a protein complex that functions in the tricarboxylic acid cycle and the electron transport chain of mitochondria. In most eukaryotes, SDH is highly conserved and comprises the following four subunits: SdhA and SdhB form the catalytic core of the complex, while SdhC and SdhD anchor the complex in the membrane. Toxoplasma gondii is an apicomplexan parasite that infects one-third of humans worldwide. The genome of T. gondii encodes homologues of the catalytic subunits SdhA and SdhB, although the physiological role of the SDH complex in the parasite and the identity of the membrane-anchoring subunits are poorly understood. Here, we show that the SDH complex contributes to optimal proliferation and O2 consumption in the disease-causing tachyzoite stage of the T. gondii life cycle. We characterize a small membrane-bound subunit of the SDH complex called mitochondrial protein ookinete developmental defect (MPODD), which is conserved among myzozoans, a phylogenetic grouping that incorporates apicomplexan parasites and their closest free-living relatives. We demonstrate that TgMPODD is essential for SDH activity and plays a key role in attaching the TgSdhA and TgSdhB proteins to the membrane anchor of the complex. Our findings highlight a unique and important feature of mitochondrial energy metabolism in apicomplexan parasites and their relatives.

Quercetin induces itaconic acid-mediated M1/M2 alveolar macrophages polarization in respiratory syncytial virus infection.

BACKGROUND: Quercetin has received extensive attention for its therapeutic potential treating respiratory syncytial virus (RSV) infection diseases. Recent studies have highlighted quercetin's ability of suppressing alveolar macrophages (AMs)-derived lung inflammation. However, the anti-inflammatory mechanism of quercetin against RSV infection still remains elusive. PURPOSE: This study aims to elucidate the mechanism about quercetin anti-inflammatory effect on RSV infection. METHODS: BALB/c mice were intranasally infected with RSV and received quercetin (30, 60, 120 mg/kg/d) orally for 3 days. Additionally, an in vitro infection model utilizing mouse alveolar macrophages (MH-S cells) was employed to validate the proposed mechanism. RESULTS: Quercetin exhibited a downregulatory effect on glycolysis and tricarboxylic acid (TCA) cycle metabolism in RSV-infected AMs. However, it increased itaconic acid production, a metabolite derived from citrate through activating immune responsive gene 1 (IRG1), and further inhibiting succinate dehydrogenase (SDH) activity. While the suppression of SDH activity orchestrated a cascading downregulation of Hif-1α/NLRP3 signaling, ultimately causing AMs polarization from M1 to M2 phenotypes. CONCLUSION: Our study demonstrated quercetin stimulated IRG1-mediated itaconic acid anabolism and further inhibited SDH/Hif-1α/NLRP3 signaling pathway, which led to M1 to M2 polarization of AMs so as to ameliorate RSV-induced lung inflammation.

Genetic predisposition to pheochromocytoma and paraganglioma: 21 years of experience in the field.

CONTEXT: Pheochromocytoma and paraganglioma (PPGL) are rare neuroendocrine tumors with high heritability, justifying systematic genetic screening for a germline variant in one of the twenty predisposing genes described to date. PURPOSE: To describe the experience of one endocrine oncogenetic laboratory over a period of 21 years (2001-2022), from the beginning of PPGL genotyping with Sanger sequencing in 2001 to the implementation of next-generation sequencing (NGS). METHOD: The activity database of an academic oncogenetic laboratory was searched to extract patients/relatives identified with a pathogenic variant/likely pathogenic variant (PV/LPV) over a period of 21 years. Clinical and genetic data were compared. RESULTS: In total, 606 index cases with PPGL and 444 relatives were genotyped. Genotyping of index cases was performed by Sanger sequencing and gene deletion analysis in 327 cases and by NGS in 279. Germline PV/LPV spanning 10 genes was identified in 165 index cases (27.2%). Several recurrent PV/LPVs in SDHx were observed in non-related index cases, the most frequent being SDHD, c.170-1G>T (n=28). This subgroup showed great phenotypic variability both between and within families in terms of both tumor location and number. Four patients (1.1%) with PV/LPV in SDHx had 3PA (Pituitary Adenoma and pheochromocytoma/paraganglioma) syndrome. 258 relatives (58.1%) had inherited a PV/LPV in one driver gene. The rate of PV/LPV carriers who were symptomatic at first imaging evaluation was 32%, but varied between<20% in SDHB and SDHC and >50% in SDHD, VHL and MAX. CONCLUSION: Our experience confirmed previously established genotype-phenotype correlations, but also highlights atypical clinical presentations, even for the same genetic variant. These data must be taken into account for optimal patient follow-up and management.

Synthesis, fungicidal activity and molecular docking of novel pyrazole-carboxamides bearing a branched alkyl ether moiety.

Succinate dehydrogenase inhibitors are essential fungicides used in agriculture. To explore new pyrazole-carboxamides with high fungicidal activity, a series of N-substitutedphenyl-3-di/trifluoromethyl-1-methyl-1H-pyrazole-4-carboxamides bearing a branched alkyl ether moiety were designed and synthesized. The in vitro bioassay indicated that some target compounds displayed appreciable fungicidal activity. For example, compounds 5d and 5e showed high efficacy against S. sclerotiorum with EC50 values of 3.26 and 1.52 µg/mL respectively, and also exhibited excellent efficacy against R. solani with EC50 values of 0.27 and 0.06 µg/mL respectively, which were comparable or superior to penflufen. The further in vivo bioassay on cucumber leaves demonstrated that 5e provided strong protective activity of 94.3 % against S. sclerotiorum at 100 µg/mL, comparable to penflufen (99.1 %). Cytotoxicity assessment against human renal cell lines (239A cell) revealed that 5e had low cytotoxicity within the median effective concentrations. Docking study of 5e with succinate dehydrogenase illustrated that R-5e formed one hydrogen bond and two π-π stacking interactions with amino acid residues of target enzyme, while S-5e formed only one π-π stacking interaction with amino acid residue. This study provides a valuable reference for the design of new succinate dehydrogenase inhibitor.

Design, Synthesis, and Antifungal Evaluation of Diverse Heterocyclic Hydrazide Derivatives as Potential Succinate Dehydrogenase Inhibitors.

Plant pathogenic fungi pose a significant threat to agricultural production, necessitating the development of new and more effective fungicides. The ring replacement strategy has emerged as a highly successful approach in molecular design. In this study, we employed the ring replacement strategy to successfully design and synthesize 32 novel hydrazide derivatives containing diverse heterocycles, such as thiazole, isoxazole, pyrazole, thiadiazole, 1,3,4-oxadiazole, 1,2,4-oxadiazole, thiophene, pyridine, and pyrazine. Their antifungal activities were evaluated in vitro and in vivo. Bioassay results revealed that most of the title compounds displayed remarkable antifungal activities in vitro against four tested phytopathogenic fungi, including Fusarium graminearum, Botrytis cinerea, Sclerotinia sclerotiorum, and Rhizoctonia solani. Especially, compound 5aa displayed a broad spectrum of antifungal activity against F. graminearum, B. cinerea, S. sclerotiorum, and R. solani, with the corresponding EC50 values of 0.12, 4.48, 0.33, and 0.15 µg/mL, respectively. In the antifungal growth assay, compound 5aa displayed a protection efficacy of 75.5% against Fusarium head blight (FHB) at a concentration of 200 µg/mL. In another in vivo antifungal activity evaluation, compound 5aa exhibited a noteworthy protective efficacy of 92.0% against rape Sclerotinia rot (RSR) at a concentration of 100 µg/mL, which was comparable to the positive control tebuconazole (97.5%). The existing results suggest that compound 5aa has a broad-spectrum antifungal activity. Electron microscopy observations showed that compound 5aa might cause mycelial abnormalities and organelle damage in F. graminearum. Moreover, in the in vitro enzyme assay, we found that the target compounds 5aa, 5ab, and 5ca displayed significant inhibitory effects toward succinate dehydrogenase, with the corresponding IC50 values of 1.62, 1.74, and 1.96 µM, respectively, which were superior to that of boscalid (IC50 = 2.38 µM). Additionally, molecular docking and molecular dynamics simulation results revealed that compounds 5aa, 5ab, and 5ca have the capacity to bind in the active pocket of succinate dehydrogenase (SDH), establishing hydrogen-bonding interactions with neighboring amino acid residues.

Two sisters diagnosed with familial paraganglioma syndrome type 1 (FPGL1) and multiple endocrine neoplasia type 2A (MEN2A).

BACKGROUND: In clinical practice, genetic testing has become standard for many cancerous diseases. While a diagnosis of a single hereditary syndrome is not uncommon, the coexistence of two genetic diseases, even with partially common symptoms, remains unusual. Therefore, targeted next-generation sequencing (NGS), along with genetic consultation and imaging studies, is essential for every patient with confirmed paraganglioma. In this report, we present two sisters diagnosed with multiple endocrine neoplasia type 2 (MEN2A) and familial paraganglioma syndrome type 1 (FPGL1). CASE PRESENTATION: After presenting to the clinic with neck tumors persisting for several months, both patients underwent tumor removal procedures following imaging and laboratory studies. Pathological reports confirmed the diagnosis of paragangliomas. Subsequently, genetic testing, including NGS, revealed a mutation in the rearranged during transfection (RET) gene: the heterozygous change (c.2410G > A), (p.Val804Met), and a variant of the succinate dehydrogenase complex subunit D (SDHD) gene: (c.64 C > T), (p.Arg22Ter). Subsequently, thyroidectomy procedures were scheduled in both cases. CONCLUSION: To the best of our knowledge, this is the first report presenting these two mutations in two related patients, resulting in distinctive genetic syndromes with similar manifestations. This underscores that although infrequent, multiple hereditary disorders may co-occur in the same individual.

Uterine Leiomyomas with Specific Histology Features of Two Fumarate Hydratase/Succinate Dehydrogenase-Deficient Tumors: A Double Case Report.

Background and Objectives: Mutations in succinate dehydrogenase (SDH) and fumarate hydratase (FH) give rise to various familial cancer syndromes, with these alterations being characteristic of certain types of histomorphologically specific leiomyomas that hold significant predictive value. Materials and Methods: This study presents two cases of uterine leiomyomas exhibiting rare histomorphological and genetic characteristics, which are crucial for prognosis and further treatment. Results: Distinct histopathological features such as marked nuclear atypia, intracellular eosinophilic globules, and abnormal intratumoral vessels raise suspicion for specific leiomyoma subtypes, which carry predictive significance for additional hereditary cancer syndromes. Immunohistochemical analysis confirmed FH/SDH deficiency in both patients, who underwent careful follow-up. Conclusions: This study describes two cases involving unusual leiomyomas, the histopathological characteristics of which may easily go unrecognized. These features hold predictive significance because their specific mutations point to additional hereditary cancer syndromes, highlighting the need for further examinations.

Hesperidin, vanillic acid, and sinapic acid attenuate atorvastatin-induced mitochondrial dysfunction via inhibition of mitochondrial swelling and maintenance of mitochondrial function in pancreas isolated mitochondria.

It has been reported that lipophilic statins such as atorvastatin can more readily penetrate into ß-cells and reach the mitochondria, resulting in mitochondrial dysfunction, oxidative stress, decrease in insulin release. Many studies have shown that natural products can protect mitochondrial dysfunction induced by drug in different tissue. We aimed to explore mitochondrial protection potency of hesperidin, vanillic acid, and sinapic acid as natural compounds against mitochondrial dysfunction induced by atorvastatin in pancreas isolated mitochondria. Mitochondria were isolated form rat pancreas and directly treated with toxic concentration of atorvastatin (500 µM) in presence of various concentrations hesperidin, vanillic acid, and sinapic acid (1, 10, and 100 µM) separately. Mitochondrial toxicity parameters such as the reactive oxygen species (ROS) formation, succinate dehydrogenases (SDH) activity, mitochondrial swelling, depletion of glutathione (GSH), mitochondrial membrane potential (MMP) collapse, and malondialdehyde (MDA) production were measured. Our findings demonstrated that atorvastatin directly induced mitochondrial toxicity at concentration of 500 µM and higher in pancreatic mitochondria. Except MDA, atorvastatin caused significantly reduction in SDH activity, mitochondrial swelling, ROS formation, depletion of GSH, and collapse of MMP. While, our data showed that all three protective compounds at low concentrations ameliorated atorvastatin-induced mitochondrial dysfunction with the increase of SDH activity, improvement of mitochondrial swelling, MMP collapse and mitochondrial GSH, and reduction of ROS formation. We can conclude that hesperidin, vanillic acid, and sinapic acid can directly reverse the toxic of atorvastatin in rat pancreas isolated mitochondria, which may be beneficial for protection against diabetogenic-induced mitochondrial dysfunction in pancreatic ß-cells.

3D-QSAR-Directed Synthesis of Halogenated Coumarin-3-Hydrazide Derivatives: Unveiling Their Potential as SDHI Antifungal Agents.

The pursuit of new succinate dehydrogenase (SDH) inhibitors is a leading edge in fungicide research and development. The use of 3D quantitative structure-activity relationship (3D-QSAR) models significantly enhances the development of compounds with potent antifungal properties. In this study, we leveraged the natural product coumarin as a molecular scaffold to synthesize 74 novel 3-coumarin hydrazide derivatives. Notably, compounds 4ap (0.28 µg/mL), 6ae (0.32 µg/mL), and 6ah (0.48 µg/mL) exhibited exceptional in vitro effectiveness against Rhizoctonia solani, outperforming the commonly used fungicide boscalid (0.52 µg/mL). Furthermore, compounds 4ak (0.88 µg/mL), 6ae (0.61 µg/mL), 6ah (0.65 µg/mL), and 6ak (1.11 µg/mL) showed significant activity against Colletotrichum orbiculare, surpassing both the SDHI fungicide boscalid (43.45 µg/mL) and the broad-spectrum fungicide carbendazim (2.15 µg/mL). Molecular docking studies and SDH enzyme assays indicate that compound 4ah may serve as a promising SDHI fungicide. Our ongoing research aims to refine this 3D-QSAR model further, enhance molecular design, and conduct additional bioactivity assays.

Geldanamycin confers fungicidal properties to azole by triggering the activation of succinate dehydrogenase.

AIMS: Azoles have been widely employed for the treatment of invasive fungal diseases; however, their efficacy is diminished as pathogenic fungi tolerate them due to their fungistatic properties. Geldanamycin (GdA) can render azoles fungicidal by inhibiting the ATPase and molecular chaperone activities of heat shock protein 90 (Hsp90). Nonetheless, the clinical applicability of GdA is restricted due to its cytotoxic ansamycin scaffold structure, its induction of cytoprotective heat shock responses, and the conservative nature of Hsp90. Hence, it is imperative to elucidate the mechanism of action of GdA to confer fungicidal properties to azoles and mitigate the toxic adverse effects associated with GdA. MATERIALS AND METHODS: Through various experimental methods, including the construction of gene-deleted Candida albicans mutants, in vitro drug sensitivity experiments, Western blot analysis, reactive oxygen species (ROS) assays, and succinate dehydrogenase activity assays, we identified Hsp90 client proteins associated with the tolerance of C. albicans to azoles. KEY FINDINGS: It was observed that GdA effectively hindered the entry of Hsp90 into mitochondria, resulting in the alleviation of inhibitory effect of Hsp90 on succinate dehydrogenase. Consequently, the activation of succinate dehydrogenase led to an increased production of ROS. within the mitochondria, thereby facilitating the antifungal effects of azoles against C. albicans. SIGNIFICANCE: This research presents a novel approach for conferring fungicidal properties to azoles, which involves specifically disrupting the interaction of between Hsp90 and succinate dehydrogenase rather than employing a non-specific inhibition of ATPase activity of Hsp90.

Design, Synthesis, Antifungal Activity, and Action Mechanism of Pyrazole-4-carboxamide Derivatives Containing Oxime Ether Active Fragment As Succinate Dehydrogenase Inhibitors.

The dearomatization at the hydrophobic tail of the boscalid was carried out to construct a series of novel pyrazole-4-carboxamide derivatives containing an oxime ether fragment. By using fungicide-likeness analyses and virtual screening, 24 target compounds with theoretical strong inhibitory effects against fungal succinate dehydrogenase (SDH) were designed and synthesized. Antifungal bioassays showed that the target compound E1 could selectively inhibit the in vitro growth of R. solani, with the EC50 value of 1.1 µg/mL that was superior to that of the agricultural fungicide boscalid (2.2 µg/mL). The observations by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) demonstrated that E1 could reduce mycelial density and significantly increase the mitochondrial number in mycelia cytoplasm, which was similar to the phenomenon treated with boscalid. Enzyme activity assay showed that the E1 had the significant inhibitory effect against the SDH from R. solani, with the IC50 value of 3.3 µM that was superior to that of boscalid (7.9 µM). The mode of action of the target compound E1 with SDH was further analyzed by molecular docking and molecular dynamics simulation studies. Among them, the number of hydrogen bonds was significantly more in the SDH-E1 complex than that in the SDH-boscalid complex. This research on the dearomatization strategy of the benzene ring for constructing pyrazole-4-carboxamides containing an oxime ether fragment provides a unique thought to design new antifungal drugs targeting SDH.

[68Ga]DOTATOC PET-derived radiomics to predict genetic background of head and neck paragangliomas: a pilot investigation.

PURPOSE: To investigate the [68Ga]DOTATOC PET radiomic profile of head and neck paragangliomas (HNPGLs) and identify radiomic characteristics useful as predictors of succinate dehydrogenase genes (SDHx) pathogenic variants. METHODS: Sporadic and SDHx HNPGL patients, who underwent [68Ga]DOTATOC PET/CT, were retrospectively included. HNPGLs were analyzed using LIFEx software, and extracted features were harmonized to correct for batch effects and confronted testing for multiple comparison. Stepwise discriminant analysis was conducted to remove redundancy and identify best discriminating features. ROC analysis was used to define optimal cut-offs. Multivariate decision-tree analysis was performed using CHAID method. RESULTS: 34 patients harboring 60 HNPGLs (51 SDHx in 25 patients) were included. Three sporadic and nine SDHx HNPGLs were metastatic. At stepwise discriminant analysis, both GLSZM-Zone Size Non-Uniformity (ZSNU, reflecting tumor heterogeneity) and IB-TLSRE (total lesion somatostatin receptor expression) were independent predictors of genetic status, with 96.4% of lesions and 91.6% of patients correctly classified after cross validation (p < 0.001). Among non-metastatic patients, GLSZM-ZSNU and IB-TLSRE were significantly higher in sporadic than SDHx HNPGLs (p < 0.001). No differences were revealed in metastatic patients. Decision-tree analysis highlights multifocality and IB-TLSRE as useful variables, correctly identifying 6/9 sporadic and 24/25 SDHx patients. Model failed to classify one SDHA and three sporadic patients (2 metastatic). CONCLUSION: Radiomics features GLSZM-ZSNU and IB-TLSRE appear to reflect HNPGLs SDHx status and tumor behavior (metastatic vs. non-metastatic). If validated, especially IB-TLSRE might represent a simple and time-efficient radiomic index for SDHx variants early screening and prediction of tumor behavior in HNPGL cases.

Baicalin attenuates neuronal damage associated with SDH activation and PDK2-PDH axis dysfunction in early reperfusion.

BACKGROUND: Energy deficiency and oxidative stress are interconnected during ischemia/reperfusion (I/R) and serve as potential targets for the treatment of cerebral ischemic stroke. Baicalin is a neuroprotective antioxidant, but the underlying mechanisms are not fully revealed. PURPOSE: This study explored whether and how baicalin rescued neurons against ischemia/reperfusion (I/R) attack by focusing on the regulation of neuronal pyruvate dehydrogenase kinase 2 (PDK2)-pyruvate dehydrogenase (PDH) axis implicated with succinate dehydrogenase (SDH)-mediated oxidative stress. STUDY DESIGN: The effect of the tested drug was explored in vitro and in vivo with the model of oxygen-glucose deprivation/reoxygenation (OGD/R) and middle cerebral artery occlusion/reperfusion (MCAO/R), respectively. METHODS: Neuronal damage was evaluated according to cell viability, infarct area, and Nissl staining. Protein levels were measured by western blotting and immunofluorescence. Gene expression was investigated by RT-qPCR. Mitochondrial status was also estimated by fluorescence probe labeling. RESULTS: SDH activation-induced excessive production of reactive oxygen species (ROS) changed the protein expression of Lon protease 1 (LonP1) and hypoxia-inducible factor-1ɑ (HIF-1ɑ) in the early stage of I/R, leading to an upregulation of PDK2 and a decrease in PDH activity in neurons and cerebral cortices. Treatment with baicalin prevented these alterations and ameliorated neuronal ATP production and survival. CONCLUSION: Baicalin improves the function of the neuronal PDK2-PDH axis via suppression of SDH-mediated oxidative stress, revealing a new signaling pathway as a promising target under I/R conditions and the potential role of baicalin in the treatment of acute ischemic stroke.

Effect of the mitochondrial transaminase (GOT2) on membrane potential-sensitive respiration in mitochondria of differentiated C2C12 muscle cells.

We previously showed that the transaminase inhibitor, aminooxyacetic acid, reduced respiration energized at complex II (succinate dehydrogenase, SDH) in mitochondria isolated from mouse hindlimb muscle. The effect required a reduction in membrane potential with resultant accumulation of oxaloacetate (OAA), a potent inhibitor of SDH. To specifically assess the effect of the mitochondrial transaminase, glutamic oxaloacetic transaminase (GOT2) on complex II respiration, and to determine the effect in intact cells as well as isolated mitochondria, we performed respiratory and metabolic studies in wildtype (WT) and CRISPR-generated GOT2 knockdown (KD) C2C12 myocytes. Intact cell respiration by GOT2KD cells versus WT was reduced by adding carbonyl cyanide-p-trifluoromethoxyphenylhydrazone (FCCP) to lower potential. In mitochondria of C2C12 KD cells, respiration at low potential generated by 1 µM FCCP and energized at complex II by 10 mM succinate + 0.5 mM glutamate (but not by complex I substrates) was reduced versus WT mitochondria. Although we could not detect OAA, metabolite data suggested that OAA inhibition of SDH may have contributed to the FCCP effect. C2C12 mitochondria differed from skeletal muscle mitochondria in that the effect of FCCP on complex II respiration was not evident with ADP addition. We also observed that C2C12 cells, unlike skeletal muscle, expressed glutamate dehydrogenase, which competes with GOT2 for glutamate metabolism. In summary, GOT2 KD reduced C2C12 respiration in intact cells at low potential. From differential substrate effects, this occurred largely at complex II. Moreover, C2C12 versus muscle mitochondria differ in complex II sensitivity to ADP and differ markedly in expression of glutamate dehydrogenase.NEW & NOTEWORTHY Impairment of the mitochondrial transaminase, GOT2, reduces complex II (succinate dehydrogenase, SDH)-energized respiration in C2C12 myocytes. This occurs only at low inner membrane potential and is consistent with inhibition of SDH. Incidentally, we observed that C2C12 mitochondria compared with muscle tissue mitochondria differ in sensitivity of complex II respiration to ADP and in the expression of glutamate dehydrogenase.