Citations

Abstract High-risk neuroblastoma remains therapeutically challenging to treat, and the mechanisms promoting disease aggression are poorly understood. Here, we show that elevated expression of dihydrolipoamide S-succinyltransferase (DLST) predicts poor treatment outcome and aggressive disease in patients with neuroblastoma. DLST is an E2 component of the α-ketoglutarate (αKG) dehydrogenase complex, which governs the entry of glutamine into the tricarboxylic acid cycle (TCA) for oxidative decarboxylation. During this irreversible step, αKG is converted into succinyl-CoA, producing NADH for oxidative phosphorylation (OXPHOS). Utilizing a zebrafish model of MYCN-driven neuroblastoma, we demonstrate that even modest increases in DLST expression promote tumor aggression, while monoallelic dlst loss impedes disease initiation and progression. DLST depletion in human MYCN-amplified neuroblastoma cells minimally affected glutamine anaplerosis and did not alter TCA cycle metabolites other than αKG. However, DLST loss significantly suppressed NADH production and impaired OXPHOS, leading to growth arrest and apoptosis of neuroblastoma cells. In addition, multiple inhibitors targeting the electron transport chain, including the potent IACS-010759 that is currently in clinical testing for other cancers, efficiently reduced neuroblastoma proliferation in vitro. IACS-010759 also suppressed tumor growth in zebrafish and mouse xenograft models of high-risk neuroblastoma. Together, these results demonstrate that DLST promotes neuroblastoma aggression and unveils OXPHOS as an essential contributor to high-risk neuroblastoma. Significance: These findings demonstrate a novel role for DLST in neuroblastoma aggression and identify the OXPHOS inhibitor IACS-010759 as a potential therapeutic strategy for this deadly disease.

Background While venous congestion in the peripheral vasculature has been described and accepted, intracranial venous congestion remains poorly understood. The characteristics, pathophysiology, and management of cerebral venous stasis, venous hypertension and venous congestion remain controversial, and a unifying conceptual schema is absent. The cerebral venous and lymphatic systems are part of a complex and dynamic interaction between the intracranial compartments, with interplay between the parenchyma, veins, arteries, cerebrospinal fluid, and recently characterized lymphatic-like systems in the brain. Each component contributes towards intracranial pressure, occupying space within the fixed calvarial volume. This article proposes a framework to consider conditions resulting in brain and neck venous congestion, and seeks to expedite further study of cerebral venous diagnoses, mechanisms, symptomatology, and treatments. Methods A multi-institution retrospective review was performed to identify unique patient cases, complemented with a published case series to assess a spectrum of disease states with components of venous congestion affecting the brain. These diseases were organized according to anatomical location and purported mechanisms. Outcomes of treatments were also analyzed. Illustrative cases were identified in the venous treatment databases of the authors. Conclusion This framework is the first clinically structured description of venous pathologies resulting in intracranial venous and cerebrospinal fluid hypertension. Our proposed system highlights unique clinical symptoms and features critical for appropriate diagnostic work-up and potential treatment. This novel schema allows clinicians effectively to approach cases of intracranial hypertension secondary to venous etiologies, and furthermore provides a framework by which researchers can better understand this developing area of cerebrovascular disease.

Introduction: Morbidity and mortality attributable to hypertension are higher in African American (AAEH) compared to Caucasian essential hypertensive (EH) patients, possibly related to a differential effect on vascular injury and repair. While circulating endothelial progenitor cells (EPC) preserve endothelial integrity IEC detach from sites of injury and represent markers of vascular damage. We hypothesized that plasma levels of IEC and inflammatory markers would be higher in AAEH compared to EH patients. METHODS: Inferior vena cava levels of CD34+/KDR+ (EPC) and VAP-1+ (IEC) cells were measured by FACS in EH and AAEH under fixed sodium intake (150 mEq/d) and blockade of the renin-angiotensin-system, and compared to systemic levels in normotensive control subjects (n=19 each). Systemic levels of inflammatory cytokines and EPC homing factors were measured by Luminex. Results: Blood pressure, serum creatinine, lipids, antihypertensive medications, and EPC levels did not differ between EH and AAEH patients. Circulating IEC were higher in AAEH, and inversely correlated with EPC levels (Figure). Systemic levels of inflammatory cytokines and EPC homing factors were higher in AAEH compared to EH patients (Table), and correlated directly with IEC. Conclusion: Despite preserved kidney function and controlled BP, circulating inflammatory markers were elevated in AAEH and correlated with increased IEC and decreased EPC levels. Increased release of cytokines and IEC in AAEH may impair EPC reparative capacity and predispose to hypertensive vascular injury. This process may aggravate vascular damage and accelerate hypertension-related morbidity/mortality rates in AAEH.

Background: Insulin-like growth factor binding protein7 (IGFBP-7) and tissue inhibitor of metalloproteinase-2 (TIMP-2) reflect G1-cell cycle arrest and are used as biomarkers for AKI. Recent studies show that these biomarkers rise in ischemic conditions and suggest that they actually may limit the severity of AKI. We tested the hypothesis that renal vein [IGFBP-7]*[TIMP-2] correlate with reductions in renal blood flow (RBF) and post-stent single kidney (SK)-GFR changes in patients with RVD undergoing contrast-based imaging and stent revascularization Methods: Inpatient studies were performed during 150 mEq Na+ intake and ACE/ARB Rx in patients with hemodynamically severe RVD (n=29, Doppler velocity = 318 ± 100cm/sec, and eGFR= 34.7 ± 11.7 mL/ min) scheduled for renal artery stenting, and compared to essential hypertensive (EH) healthy controls (n=32). Cortical and medullary RBFs (by multidetector CT) and renal vein levels of IGFBP-7 and TIMP-2 were measured before renal artery stenting and 3 months later Results: Pre-stenting IGFBP-7 and TIMP-2 levels were elevated in RVD compared to EH (18.5±2. vs 15.7±1.5 and 97.4±23.1vs 62.7±9.2 ng/mL respectively, P <0.0001). Baseline renal vein levels of IGFBP-7 * TIMP-2 correlated inversely with pre-stent RBF (r= - 0.53, P=0.01) and directly with the change (%) in SK-GFR observed 3 months after stenting Conclusion: IGFBP-7 and TIMP-2 levels are elevated in patients with chronic RVD as a function of the baseline reduction in RBF, and associate with subsequent improvement in SK-GFR 3 months after revascularization. These data are consistent with renovascular occlusion inducing cell-cycle arrest that serves to protect kidney function.

Percutaneous transluminal renal angioplasty and stenting (PTRS) for RVH may not revert the secondary cardiac dysfunction and remodeling. In experimental RVH, infusion of the tetrapeptide Bendavia (that inhibits mitochondrial permeability transition pore opening) during PTRS attenuated stenotic kidney inflammatory injury and improved its function. We hypothesized that adjunct Bendavia during PTRS would also attenuate cardiac remodeling after reversal of RVH. RVH was induced in 18 pigs, of which 6 were sham treated (RVH) and 12 underwent PTRS plus a 4h IV infusion of Bendavia (0.05 mg/kg) or vehicle (n=6, each); 6 normal pigs served as controls. Cardiac oxygenation and function were assessed 4 wks later by blood oxygen level‐dependent (BOLD)‐MRI and CT, and tissue studies performed. PTRS normalized blood pressure (Table). Systolic function was unchanged. Apoptotic activity (caspase‐3), hypoxia (BOLD‐R2*), inflammation (TNF‐α, IL‐6), and fibrosis (trichrome) remained elevated and diastolic function (E/A) impaired in PTRS+Vehicle, but all improved in PTRS+Bendavia (Figure). Thus, novel cardioprotective properties of Bendavia enhance cardiac recovery after reversal of experimental RVH, and may provide a novel therapeutic option.

Abstract Objectives To demonstrate clinicopathologic features and evaluate the clonality of double PIK3CA alterations in colorectal cancers (CRCs). Methods Clonality was examined in 13 CRCs with double PIK3CA alterations (1.7% of CRCs or 9.6% of PIK3CA-mutated CRCs). Multiregional analyses were performed to confirm subclonal PIK3CA alterations. Results PIK3CA alterations were detected within exon 9 (51%), exon 20 (23%), exon 1 (15%), and exon 7 (6.0%). CRCs with exon 7 alterations showed a significantly higher incidence of double PIK3CA alterations. Most double PIK3CA alterations consisted of a hotpsot alteration and an uncommon alteration; they were often clonal and present within a single tumor population. Multiregional analyses of CRCs with predicted subclonal double-alterations revealed multiclonal CRCs with divergent PIK3CA variant status originating from a common APC- and KRAS-mutated founder lineage of adenoma. Conclusions The findings supported multiclonal CRCs resulting from parallel evolution during the progression from adenoma to adenocarcinoma within the mitogen-activated protein kinase pathway, as previously demonstrated, or the mammalian target of rapamycin pathway. Further studies are warranted to elucidate clinical significance and potential targeted therapy for CRC patients with double PIK3CA alterations and impacts on clinical decision-making in patients with multiclonal CRCs harboring divergent PIK3CA mutational status.

Background and Purpose: XO (xanthine oxidase) is a key enzyme of uric acid metabolism and is thought to contribute to oxidative pathways that promote atherosclerotic plaque progression, yet its role in plaque destabilization is not well elucidated. We hypothesized that XO is expressed in carotid plaque from symptomatic patients in association with cardiovascular risk factors. Methods: Patients were stratified by symptoms, defined as presentation with an ipsilateral cerebral ischemic event. Carotid atherosclerotic plaques were obtained from 44 patients with symptomatic plaque and 44 patients without ischemic cerebral events. Protein expression of XO was evaluated by immunohistochemical staining and the percentage of cells expressing XO and CD68 (macrophage marker) compared between the groups. Biochemical and demographic cardiometabolic risk factors of study participants also were measured. Results: Carotid atherosclerotic plaques from symptomatic patients were associated with significantly higher XO expression versus asymptomatic plaque (median [interquartile range]: 1.24 [2.09] versus 0.16 [0.34]; P <0.001) and with significantly higher circulating uric acid levels (mean±SD: 7.36±2.10 versus 5.37±1.79 mg/dL; P <0.001, respectively). In addition, XO expression in atherosclerotic carotid plaque was inversely associated with serum high-density lipoproteins cholesterol levels ( P =0.010, r =−0.30) and directly with circulating uric acid levels ( P <0.001, r =0.45). The average percentage of macrophages that expressed XO was significantly higher in symptomatic versus asymptomatic plaques (median [interquartile range]: 93.37% [25] versus 46.15% [21], respectively; P <0.001). Conclusions: XO overexpression in macrophages is associated with increased serum uric acid and low high-density lipoproteins cholesterol levels and may potentially have a mechanistic role in carotid plaque destabilization. The current study supports a potential role for uric acid synthesis pathway as a target for management of carotid atherosclerosis in humans.

3079 Background: RET fusions are targetable oncogenic drivers in multiple solid tumor types. ARROW study (NCT03037385) data supported the US FDA approval of pralsetinib, a once-daily (QD) oral highly potent and selective RET inhibitor, for RET-altered metastatic non-small cell lung cancer (NSCLC) and advanced/metastatic thyroid cancer. Here we provide an update on the clinical activity of pralsetinib in patients (pts) with advanced RET fusion-positive solid tumors other than NSCLC and thyroid cancer (“other” RET fusion–positive solid tumors). Methods: The global ongoing ARROW study (84 sites in 13 countries) includes phase 1 dose-escalation (30–600 mg [QD or twice daily]) and phase 2 expansion cohorts (400 mg QD) defined by tumor type and RET alteration status. Primary objectives are overall response rate (ORR; blinded independent central review per RECIST v1.1) and safety. Results: Updated analyses were completed as of Nov 6, 2020 (data cut-off) for 21 pts with other RET fusion–positive solid tumors enrolled by May 22, 2020 (enrollment cut-off) (lung other than NSCLC, n = 4; pancreatic, n = 3; colon, n = 3; cholangiocarcinoma, n = 3; unknown primary [UP], n = 2; other, n = 6). Overall, 11 (52%) pts received ≥2 prior lines of therapy for metastatic disease. The most common RET fusion partners were CCDC6 and KIF5B (24% each), NCOA4 (19%), other (10%), and unknown (24%). Two pts with colon cancer were excluded from efficacy analyses due to other driver mutations ( KRAS, PIK3CB). In 19 evaluable pts, ORR was 53% (95% CI, 29–76) with 2 (11%) complete responses (CR) and 8 (42%) partial responses (PR). Responses occurred across multiple tumor types including 3/3 pts with pancreatic cancer (including a CR ongoing at 20.8 months on treatment), 2/2 pts with UP, 2/3 pts with cholangiocarcinoma, and in pts with mesenchymal, salivary duct, and lung carcinoid tumors. Median duration of response was 19.0 months (95% CI, 5.5–not estimable). Clinical benefit rate (proportion with CR, PR, or stable disease persisting ≥16 weeks) was 68% (95% CI, 43–87). Tumor shrinkage was observed in 89% of 18 evaluable pts with post-baseline tumor assessment. In all pts enrolled in ARROW who received pralsetinib 400 mg QD irrespective of tumor type (n = 471) the most common (≥25%) treatment-related adverse events (TRAEs) were increased aspartate aminotransferase (39%), anemia (35%), increased alanine aminotransferase (28%), constipation (26%), and hypertension (25%). Overall, 6% of pts discontinued treatment due to TRAEs. Conclusions: Pralsetinib showed robust, durable antitumor activity in patients with multiple RET fusion‒positive, heavily pre-treated, advanced solid tumors, and was well tolerated. These data highlight the need for broad RET testing to identify candidates who could benefit from treatment with pralsetinib. Enrollment of patients with other RET fusion–positive solid tumors in ARROW is ongoing. Clinical trial information: NCT03037385.

Introduction: Subjects with renovascular hypertension (RVH) often manifest with metabolic syndrome (MetS), increasing risk for heart failure, yet the mechanisms underlying cardiac injury remain unknown. Reduced mitophagy and accumulation of impaired mitochondria have been linked to progression of heart failure. Hypothesis: We hypothesized that superimposition of MetS on RVH in swine inhibits myocardial mitophagy, aggravating mitochondrial damage, cardiac injury and dysfunction. Methods: Pigs were studied after 16 weeks of RVH (renal artery stenosis) with or without diet-induced MetS, and Lean controls (n=6 each). Cardiac function was assessed in vivo (multidetector-CT), and cardiac mitochondrial structure, function, mitophagy, and myocardial fibrosis ex vivo. Results: MetS groups developed obesity, whereas RVH groups developed hypertension (Table). Mitochondrial matrix density and ATP production were lower and H2O2 production higher in both RVH groups vs. Lean and MetS (Fig. AB). However, MetS+RVH failed to activate mitophagy and downregulate myocardial expression of mitophagy-related microRNAs achieved by Lean+RVH (Fig. CD). Myocardial expression of the mitophagy marker PTEN-induced kinase (PINK)-1 inversely correlated with myocardial fibrosis (R 2 =0.195, p=0.031). Cardiac fibrosis, hypertrophy, and diastolic dysfunction (decreased E/A ratio) were greater in MetS+RVH vs. Lean+RVH (Table, Fig. E). Conclusions: MetS aggravates myocardial mitochondrial damage and dysfunction in swine RVH. MetS+RVH failed to activate mitophagy, resulting in greater cardiac remodeling, fibrosis, and diastolic dysfunction. Mitochondrial injury and impaired mitophagy may constitute important mechanisms and potential therapeutic targets to ameliorate cardiac structural damage and dysfunction in patients with coexisting MetS and RVH.

Introduction: Bicuspid Aortic Valve (BAV), the most common congenital heart defect, is a major cause of aortic regurgitation or stenosis requiring valve replacement and thoracic aortic aneurysms predisposing to acute aortic dissections (TAD). The spectrum of BAV ranges from severe early onset valve and aortic complications to sporadic late onset disease. Hypothesis: Early onset BAV (EBAV) cases with valve or aortic complications that require intervention prior to age 30 are enriched for rare genetic variants that cause BAV and TAD. Methods: We performed whole exome sequencing of 147 EBAV cases in 141 families who were enrolled in the UTHealth Bicuspid Aortic Valve Research Registry. Candidate variants in the EBAV cohort (26% female, mean age 18, 44% with TAD) were compared to unselected controls from the Genome Aggregation Database (gnoMAD) and the Database of Genotypes and Phenotypes (dbGAP). We considered variants with minor allele frequencies (MAF) < 1%, Combined Annotation Dependent Depletion (CADD) scores > 25, and damaging (Polyphen-2) or deleterious (SIFT) functional prediction scores. Genomic copy number variants (CNVs) were detected using CoNIFER and prioritized when deletions involved genes with probability of loss intolerance (pLI) > 0.9. Variants were validated using quantitative PCR or Sanger sequencing. Results: We identified 6 rare variants of USP10 in 6 EBAV families (4% of cohort): 4 CNVs (2 duplications and 2 deletions) that are rare in dbGAP controls (4 in 15,414) and 2 deleterious rare missense variants (MAF<5x10 -5 in gnoMAD). Two of the 4 CNVs were de novo events in trios. In contrast, rare deleterious variants of the known causal BAV genes NOTCH1 (1), ROBO4 (1), GATA4 (1), GATA5 (1), and SMAD6 (4) were found in 7 total families. USP10 encodes a ubiquitin peptidase that is required for endothelial Notch signaling during vascular development. Conclusions: We identified rare and de novo variants of USP10 that implicate USP10 as a new candidate gene for BAV.