The PTPN2/PTPN1 inhibitor ABBV-CLS-484 unleashes potent anti-tumour immunity.

Immune checkpoint blockade is effective for some patients with cancer, but most are refractory to current immunotherapies and new approaches are needed to overcome resistance1,2. The protein tyrosine phosphatases PTPN2 and PTPN1 are central regulators of inflammation, and their genetic deletion in either tumour cells or immune cells promotes anti-tumour immunity3-6. However, phosphatases are challenging drug targets; in particular, the active site has been considered undruggable. Here we present the discovery and characterization of ABBV-CLS-484 (AC484), a first-in-class, orally bioavailable, potent PTPN2 and PTPN1 active-site inhibitor. AC484 treatment in vitro amplifies the response to interferon and promotes the activation and function of several immune cell subsets. In mouse models of cancer resistant to PD-1 blockade, AC484 monotherapy generates potent anti-tumour immunity. We show that AC484 inflames the tumour microenvironment and promotes natural killer cell and CD8+ T cell function by enhancing JAK-STAT signalling and reducing T cell dysfunction. Inhibitors of PTPN2 and PTPN1 offer a promising new strategy for cancer immunotherapy and are currently being evaluated in patients with advanced solid tumours (ClinicalTrials.gov identifier NCT04777994 ). More broadly, our study shows that small-molecule inhibitors of key intracellular immune regulators can achieve efficacy comparable to or exceeding that of antibody-based immune checkpoint blockade in preclinical models. Finally, to our knowledge, AC484 represents the first active-site phosphatase inhibitor to enter clinical evaluation for cancer immunotherapy and may pave the way for additional therapeutics that target this important class of enzymes.

Dual CCR2/CCR5 antagonist treatment attenuates adipose inflammation, but not microvascular complications in ob/ob mice.

Diabetic peripheral neuropathy (DPN) and diabetic kidney disease (DKD) are common diabetic complications with limited treatment options. Experimental studies show that targeting inflammation using chemokine receptor (CCR) antagonists ameliorates DKD, presumably by reducing macrophage accumulation or activation. As inflammation is implicated in DPN development, we assessed whether CCR2 and CCR5 antagonism could also benefit DPN. Five-week-old ob/ob mice were fed a diet containing MK-0812, a dual CCR2-CCR5 receptor antagonist, for 8 weeks; DPN, DKD and metabolic phenotyping were then performed to determine the effect of CCR inhibition. Although MK-0812 reduced macrophage accumulation in adipose tissue, the treatment had largely no effect on metabolic parameters, nerve function or kidney disease in ob/ob mice. These results conflict with published data that demonstrate a benefit of CCR antagonists for DKD and hyperglycaemia. We conclude that CCR signaling blockade is ineffective in ob/ob mice and suspect that this is explained by the severe hyperglycaemia found in this model. It remains to be determined whether MK-0812 treatment, alone or in combination with improved glycaemic control, is useful in preventing diabetic complications in alternate animal models.

Insulin Resistance Prevents AMPK-induced Tau Dephosphorylation through Akt-mediated Increase in AMPKSer-485 Phosphorylation.

Metabolic syndrome (MetS) is a cluster of cardiovascular risk factors including obesity, diabetes, and dyslipidemia, and insulin resistance (IR) is the central feature of MetS. Recent studies suggest that MetS is a risk factor for Alzheimer disease (AD). AMP-activated kinase (AMPK) is an evolutionarily conserved fuel-sensing enzyme and a key player in regulating energy metabolism. In this report, we examined the role of IR on the regulation of AMPK phosphorylation and AMPK-mediated Tau phosphorylation. We found that AMPK(Ser-485), but not AMPK(Thr-172), phosphorylation is increased in the cortex of db/db and high fat diet-fed obese mice, two mouse models of IR. In vitro, treatment of human cortical stem cell line (HK-5320) and primary mouse embryonic cortical neurons with the AMPK activator, 5-aminoimidazole-4-carboxamide 1-ß-D-ribofuranoside (AICAR), induced AMPK phosphorylation at both Thr-172 and Ser-485. AMPK activation also triggered Tau dephosphorylation. When IR was mimicked in vitro by chronically treating the cells with insulin, AICAR specifically induced AMPK(Ser-485), but not AMPK(Thr-172), hyperphosphorylation whereas AICAR-induced Tau dephosphorylation was inhibited. IR also resulted in the overactivation of Akt by AICAR treatment; however, preventing Akt overactivation during IR prevented AMPK(Ser-485) hyperphosphorylation and restored AMPK-mediated Tau dephosphorylation. Transfection of AMPK(S485A) mutant caused similar results. Therefore, our results suggest the following mechanism for the adverse effect of IR on AD pathology: IR → chronic overactivation of Akt → AMPK(Ser-485) hyperphosphorylation → inhibition of AMPK-mediated Tau dephosphorylation. Together, our results show for the first time a possible contribution of IR-induced AMPK(Ser-485) phosphorylation to the increased risk of AD in obesity and diabetes.

BTBR ob/ob mice as a novel diabetic neuropathy model: Neurological characterization and gene expression analyses.

Given the lack of treatments for diabetic neuropathy (DN), a common diabetic complication, accurate disease models are necessary. Characterization of the leptin-deficient BTBR ob/ob mouse, a type 2 diabetes model, demonstrated that the mice develop robust diabetes coincident with severe neuropathic features, including nerve conduction deficits and intraepidermal nerve fiber loss by 9 and 13 weeks of age, respectively, supporting its use as a DN model. To gain insight into DN mechanisms, we performed microarray analysis on sciatic nerve from BTBR ob/ob mice, identifying 1503 and 642 differentially expressed genes associated with diabetes at 5 and 13 weeks, respectively. Further analyses identified overrepresentation of inflammation and immune-related functions in BTBR ob/ob mice, which interestingly were more highly represented at 5 weeks, an observation that may suggest a contributory role in DN onset. To complement the gene expression analysis, we demonstrated that protein levels of select cytokines were significantly upregulated at 13 weeks in BTBR ob/ob mouse sciatic nerve. Furthermore, we compared our array data to that from an established DN model, the C57BKS db/db mouse, which reflected a common dysregulation of inflammatory and immune-related pathways. Together, our data demonstrate that BTBR ob/ob mice develop rapid and robust DN associated with dysregulated inflammation and immune-related processes.

Lack of both bradykinin B1 and B2 receptors enhances nephropathy, neuropathy, and bone mineral loss in Akita diabetic mice.

An insertion polymorphism of the angiotensin-I converting enzyme gene (ACE) is common in humans and the higher expressing allele is associated with an increased risk of diabetic complications. The ACE polymorphism does not significantly affect blood pressure or angiotensin II levels, suggesting that the kallikrein-kinin system partly mediates the effects of the polymorphism. We have therefore explored the influence of lack of both bradykinin receptors (B1R and B2R) on diabetic nephropathy, neuropathy, and osteopathy in male mice heterozygous for the Akita diabetogenic mutation in the insulin 2 gene (Ins2). We find that all of the detrimental phenotypes observed in Akita diabetes are enhanced by lack of both B1R and B2R, including urinary albumin excretion, glomerulosclerosis, glomerular basement membrane thickening, mitochondrial DNA deletions, reduction of nerve conduction velocities and of heat sensation, and bone mineral loss. Absence of the bradykinin receptors also enhances the diabetes-associated increases in plasma thiobarbituric acid-reactive substances, mitochondrial DNA deletions, and renal expression of fibrogenic genes, including transforming growth factor beta1, connective tissue growth factor, and endothelin-1. Thus, lack of B1R and B2R exacerbates diabetic complications. The enhanced renal injury in diabetic mice caused by lack of B1R and B2R may be mediated by a combination of increases in oxidative stress, mitochondrial DNA damage and over expression of fibrogenic genes.

Cytoplasmic TDP43 Binds microRNAs: New Disease Targets in Amyotrophic Lateral Sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal, and incurable neurodegenerative disease. Recent studies suggest that dysregulation of gene expression by microRNAs (miRNAs) may play an important role in ALS pathogenesis. The reversible nature of this dysregulation makes miRNAs attractive pharmacological targets and a potential therapeutic avenue. Under physiological conditions, miRNA biogenesis, which begins in the nucleus and includes further maturation in the cytoplasm, involves trans-activation response element DNA/RNA-binding protein of 43 kDa (TDP43). However, TDP43 mutations or stress trigger TDP43 mislocalization and inclusion formation, a hallmark of most ALS cases, that may lead to aberrant protein/miRNA interactions in the cytoplasm. Herein, we demonstrated that TDP43 exhibits differential binding affinity for select miRNAs, which prompted us to profile miRNAs that preferentially bind cytoplasmic TDP43. Using cellular models expressing TDP43 variants and miRNA profiling analyses, we identified differential levels of 65 cytoplasmic TDP43-associated miRNAs. Of these, approximately 30% exhibited levels that differed by more than 3-fold in the cytoplasmic TDP43 models relative to our control model. The hits included both novel miRNAs and miRNAs previously associated with ALS that potentially regulate several predicted genes and pathways that may be important for pathogenesis. Accordingly, these findings highlight specific miRNAs that may shed light on relevant disease pathways and could represent potential biomarkers and reversible treatment targets for ALS.

Intraspinal cord delivery of IGF-I mediated by adeno-associated virus 2 is neuroprotective in a rat model of familial ALS.

BACKGROUND: Amyotrophic lateral sclerosis (ALS) is a devastating disease that is characterized by the progressive loss of motor neurons. Patients with ALS usually die from respiratory failure due to respiratory muscle paralysis. Consequently, therapies aimed at preserving segmental function of the respiratory motor neurons could extend life for these patients. Insulin-like growth factor-I (IGF-I) is known to be a potent survival factor for motor neurons. In this study we induced high levels of IGF-I expression in the cervical spinal cord of hSOD1(G93A) rats with intraspinal cord (ISC) injections of an adeno-associated virus serotype 2 vector (CERE-130). This approach reduced the extent of motor neuron loss in the treated segments of the spinal cord. However, a corresponding preservation of motor function was observed in male, but not female, hSOD1(G93A) rats. We conclude that ISC injection of CERE-130 has the potential to protect motor neurons and preserve neuromuscular function in ALS.

Mitochondrial uncoupling has no effect on microvascular complications in type 2 diabetes.

Diabetic peripheral neuropathy (DPN), diabetic kidney disease (DKD), and diabetic retinopathy (DR) contribute to significant morbidity and mortality in diabetes patients. The incidence of these complications is increasing with the diabetes epidemic, and current therapies minimally impact their pathogenesis in type 2 diabetes (T2D). Improved mechanistic understanding of each of the diabetic complications is needed in order to develop disease-modifying treatments for patients. We recently identified fundamental differences in mitochondrial responses of peripheral nerve, kidney, and retinal tissues to T2D in BKS-db/db mice. However, whether these mitochondrial adaptations are the cause or consequence of tissue dysfunction remains unclear. In the current study BKS-db/db mice were treated with the mitochondrial uncoupler, niclosamide ethanolamine (NEN), to determine the effects of mitochondrial uncoupling therapy on T2D, and the pathogenesis of DPN, DKD and DR. Here we report that NEN treatment from 6-24 wk of age had little effect on the development of T2D and diabetic complications. Our data suggest that globally targeting mitochondria with an uncoupling agent is unlikely to provide therapeutic benefit for DPN, DKD, or DR in T2D. These data also highlight the need for further insights into the role of tissue-specific metabolic reprogramming in the pathogenesis of diabetic complications.

Expressing TrkC from the TrkA locus causes a subset of dorsal root ganglia neurons to switch fate.

Tactile information is perceived by a heterogeneous population of specialized neurons. Neurotrophin receptors (the receptor tyrosine kinases, Trks) mark the major classes of these sensory neurons: TrkA is expressed in neurons that sense temperature and noxious stimuli, and TrkC is expressed in proprioceptive neurons that sense body position. Neurotrophin signaling through these receptors is required for cell survival. To test whether neurotrophins have an instructive role in sensory specification, we expressed rat TrkC from the TrkA (also known as Ntrk1) locus in mice. The surviving presumptive TrkA-expressing neurons adopted a proprioceptive phenotype, indicating that neurotrophin signaling can specify sensory neuron subtypes.

Juvenile murine models of prediabetes and type 2 diabetes develop neuropathy.

Peripheral neuropathy (neuropathy) is a common complication of obesity and type 2 diabetes in children and adolescents. To model this complication in mice, 5-week-old male C57BL/6J mice were fed a high-fat diet to induce diet-induced obesity (DIO), a model of prediabetes, and a cohort of these animals was injected with low-dose streptozotocin (STZ) at 12 weeks of age to induce hyperglycemia and type 2 diabetes. Neuropathy assessments at 16, 24 and 36 weeks demonstrated that DIO and DIO-STZ mice displayed decreased motor and sensory nerve conduction velocities as early as 16 weeks, hypoalgesia by 24 weeks and cutaneous nerve fiber loss by 36 weeks, relative to control mice fed a standard diet. Interestingly, neuropathy severity was similar in DIO and DIO-STZ mice at all time points despite significantly higher fasting glucose levels in the DIO-STZ mice. These mouse models provide critical tools to better understand the underlying pathogenesis of prediabetic and diabetic neuropathy from youth to adulthood, and support the idea that hyperglycemia alone does not drive early neuropathy.This article has an associated First Person interview with the first author of the paper.

Receptor for advanced glycation end products activation injures primary sensory neurons via oxidative stress.

The receptor for advanced glycation end products (RAGE) may promote diabetic vascular and renal disease through the activation of intracellular signaling pathways that promote oxidative stress. Oxidative stress is a mediator of hyperglycemia-induced cell injury and a unifying theme for all mechanisms of diabetic complications, but there are few studies on the expression and potential contribution of RAGE in diabetic neuropathy. The current study demonstrates that dorsal root ganglia neurons express functional RAGE and respond to the RAGE ligand S100 with similar downstream signaling, oxidative stress, and cellular injury as other diabetic complication-prone tissues. RAGE-induced phosphatidylinositol-3 kinase activity is associated with formation of reactive oxygen species, caspase-3 activation, and nuclear DNA degradation. These events are prevented by treatment with the antioxidant alpha-lipoic acid. Our data indicate that therapies aimed at decreasing RAGE ligands, blocking RAGE signaling, or preventing oxidative stress could significantly decrease the development of neuropathy in diabetic patients.

Human neural stem cell transplantation into the corpus callosum of Alzheimer's mice.

The hippocampus has been the target of stem cell transplantations in preclinical studies focused on Alzheimer's disease, with results showing improvements in histological and behavioral outcomes. The corpus callosum is another structure that is affected early in Alzheimer's disease. Therefore, we hypothesize that this structure is a novel target for human neural stem cell transplantation in transgenic Alzheimer's disease mouse models. This study demonstrates the feasibility of targeting the corpus callosum and identifies an effective immunosuppression regimen for transplanted neural stem cell survival. These results support further preclinical development of the corpus callosum as a therapeutic target in Alzheimer's disease.

Dietary reversal of neuropathy in a murine model of prediabetes and metabolic syndrome.

Patients with metabolic syndrome, which is defined as obesity, dyslipidemia, hypertension and impaired glucose tolerance (IGT), can develop the same macro- and microvascular complications as patients with type 2 diabetes, including peripheral neuropathy. In type 2 diabetes, glycemic control has little effect on the development and progression of peripheral neuropathy, suggesting that other metabolic syndrome components may contribute to the presence of neuropathy. A parallel phenomenon is observed in patients with prediabetes and metabolic syndrome, where improvement in weight and dyslipidemia more closely correlates with restoration of nerve function than improvement in glycemic status. The goal of the current study was to develop a murine model that resembles the human condition. We examined longitudinal parameters of metabolic syndrome and neuropathy development in six mouse strains/genotypes (BKS-wt, BKS-Leprdb/+ , B6-wt, B6-Leprdb/+ , BTBR-wt, and BTBR-Lepob/+ ) fed a 54% high-fat diet (HFD; from lard). All mice fed a HFD developed large-fiber neuropathy and IGT. Changes appeared early and consistently in B6-wt mice, and paralleled the onset of neuropathy. At 36 weeks, B6-wt mice displayed all components of the metabolic syndrome, including obesity, IGT, hyperinsulinemia, dyslipidemia and oxidized low density lipoproteins (oxLDLs). Dietary reversal, whereby B6-wt mice fed a HFD from 4-20 weeks of age were switched to standard chow for 4 weeks, completely normalized neuropathy, promoted weight loss, improved insulin sensitivity, and restored LDL cholesterol and oxLDL by 50% compared with levels in HFD control mice. This dietary reversal model provides the basis for mechanistic studies investigating peripheral nerve damage in the setting of metabolic syndrome, and ultimately the development of mechanism-based therapies for neuropathy.

Short-term hyperglycemia produces oxidative damage and apoptosis in neurons.

Dorsal root ganglia neurons in culture die through programmed cell death when exposed to elevated glucose, providing an in vitro model system for the investigation of the mechanisms leading to diabetic neuropathy. This study examines the time course of programmed cell death induction, regulation of cellular antioxidant capacity, and the protective effects of antioxidants in neurons exposed to hyperglycemia. We demonstrate that the first 2 h of hyperglycemia are sufficient to induce oxidative stress and programmed cell death. Using fluorimetric analysis of reactive oxygen species (ROS) production, in vitro assays of antioxidant enzymes, and immunocytochemical assays of cell death, we demonstrate superoxide formation, inhibition of aconitase, and lipid peroxidation within 1 h of hyperglycemia. These are followed by caspase-3 activation and DNA fragmentation. Antioxidant potential increases by 3-6 h but is insufficient to protect these neurons. Application of the antioxidant alpha-lipoic acid potently prevents glucose-induced oxidative stress and cell death. This study identifies cellular therapeutic targets to prevent diabetic neuropathy. Since oxidative stress is a common feature of the micro- and macrovascular complications of diabetes, the present findings have broad application to the treatment of diabetic patients.

Increased ratio of circulating neutrophils to monocytes in amyotrophic lateral sclerosis.

OBJECTIVE: To elucidate amyotrophic lateral sclerosis (ALS) biomarkers and potential mechanisms of disease, we measured immune cell populations in whole blood from a large cohort of patients with ALS. METHODS: Leukocytes were isolated from the blood of 44 control patients and 90 patients with ALS. The percentages and total numbers of each cell population were analyzed using flow cytometry and matched with patient ALS Functional Rating Scale-Revised (ALSFRS-R) score to correlate leukocyte metrics with disease progression. RESULTS: We show a significant increase in the percentage of neutrophils and a significant decrease in the percentage of CD4 T cells and CD16(-) monocytes in the blood of patients with ALS compared to controls; however, only CD16(-) monocyte levels correlated with disease progression. We also examined the monocyte surface expression of CCRL2 and CCR3; CD16(-) monocytes displayed decreased percentages and total numbers expressing CCR3, but these numbers did not correlate with ALSFRS-R score. We found that combining multiple disease metrics yielded the most accurate predictor of disease progression: the ratio of neutrophils to CD16(-) monocytes (N:M ratio) is significantly increased in patients with ALS and better correlates with disease progression than any other single metric. CONCLUSIONS: These observations implicate neutrophils and monocytes as important factors in late disease progression.

Cell culture modeling to test therapies against hyperglycemia-mediated oxidative stress and injury.

The concept that oxidative stress is a key mediator of nerve injury in diabetes has led us to design therapies that target oxidative stress mechanisms. Using an in vitro model of glucose-treated dorsal root ganglion (DRG) neurons in culture, we can examine both free radical generation, using fluorimetric probes for reactive oxygen species, and cell death via the TUNEL assay. The cell culture system is scaled down to a 96-well plate format, and so is well suited to high-throughput screening. In the present study, we test the ability of three drugs, nicotinamide, allopurinol, and alpha-lipoic acid, alone and in combination to prevent DRG neuron oxidative stress and cell death. This combination of drugs is currently in clinical trial in type 1 diabetic patients. We demonstrate independent effects on oxidative stress and neuronal survival for the three drugs, and neuronal protection using the three drugs in combination. The data strengthen the rationale for the current clinical trial. In addition, we describe an effective tool for rapid preclinical testing of novel therapies against diabetic neuropathy.

Phosphatidylinositol 3-kinase and Akt effectors mediate insulin-like growth factor-I neuroprotection in dorsal root ganglia neurons.

Insulin-like growth factor-I (IGF-I) protects neurons of the peripheral nervous system from apoptosis, but the underlying signaling pathways are not well understood. We studied IGF-I mediated signaling in embryonic dorsal root ganglia (DRG) neurons. DRG neurons express IGF-I receptors (IGF-IR), and IGF-I activates the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. High glucose exposure induces apoptosis, which is inhibited by IGF-I through the PI3K/Akt pathway. IGF-I stimulation of the PI3K/Akt pathway phosphorylates three known Akt effectors: the survival transcription factor cyclic AMP response element binding protein (CREB) and the pro-apoptotic effector proteins glycogen synthase kinase-3beta (GSK-3beta) and forkhead (FKHR). IGF-I regulates survival at the nuclear level through accumulation of phospho-Akt in DRG neuronal nuclei, increased CREB-mediated transcription, and nuclear exclusion of FKHR. High glucose increases expression of the pro-apoptotic Bcl protein Bim (a transcriptional target of FKHR). However, IGF-I does not regulate Bim or anti-apoptotic Bcl-xL protein expression levels, which suggests that IGF-I neuroprotection is not through regulation of their expression. High glucose also induces loss of the initiator caspase-9 and increases caspase-3 cleavage, effects blocked by IGF-I. These data suggest that IGF-I prevents apoptosis in DRG neurons by regulating PI3K/Akt pathway effectors, including GSK-3beta, CREB, and FKHR, and by blocking caspase activation.

The Metabolic Syndrome and Microvascular Complications in a Murine Model of Type 2 Diabetes.

To define the components of the metabolic syndrome that contribute to diabetic polyneuropathy (DPN) in type 2 diabetes mellitus (T2DM), we treated the BKS db/db mouse, an established murine model of T2DM and the metabolic syndrome, with the thiazolidinedione class drug pioglitazone. Pioglitazone treatment of BKS db/db mice produced a significant weight gain, restored glycemic control, and normalized measures of serum oxidative stress and triglycerides but had no effect on LDLs or total cholesterol. Moreover, although pioglitazone treatment normalized renal function, it had no effect on measures of large myelinated nerve fibers, specifically sural or sciatic nerve conduction velocities, but significantly improved measures of small unmyelinated nerve fiber architecture and function. Analyses of gene expression arrays of large myelinated sciatic nerves from pioglitazone-treated animals revealed an unanticipated increase in genes related to adipogenesis, adipokine signaling, and lipoprotein signaling, which likely contributed to the blunted therapeutic response. Similar analyses of dorsal root ganglion neurons revealed a salutary effect of pioglitazone on pathways related to defense and cytokine production. These data suggest differential susceptibility of small and large nerve fibers to specific metabolic impairments associated with T2DM and provide the basis for discussion of new treatment paradigms for individuals with T2DM and DPN.

Hyperglycemia-induced tau cleavage in vitro and in vivo: a possible link between diabetes and Alzheimer's disease.

Multiple lines of evidence link the incidence of diabetes to the development of Alzheimer's disease (AD). Patients with diabetes have a 50 to 75% increased risk of developing AD. In parallel, AD patients have a higher than normal tendency to develop type 2 diabetes or impaired fasting glucose. Tau is the major component of neurofibrillary tangles, one of the hallmarks of AD pathology. The current study examined the effect of hyperglycemia on tau modification. Glucose treatment of rat embryonic cortical neurons results in concentration-dependent apoptosis and caspase-3 activation. These changes are well correlated with glucose time- and concentration-dependent tau cleavage. Aß treatment induces tau cleavage and when added together with glucose, there is an additive effect on caspase activation, apoptosis, and tau cleavage. Tau cleavage is partially blocked by the caspase inhibitor, ZVAD. Cleaved tau displays a punctate staining along the neurites and colocalizes with cleaved caspase-3 in the cytoplasm. Both type 1 and type 2 diabetic mice display increased tau phosphorylation in the brain. In agreement with the effects of glucose on tau modifications in vitro, there is increased tau cleavage in the brains of ob/ob mice; however, tau cleavage is not observed in type 1 diabetic mouse brains. Our study demonstrates that hyperglycemia is one of major factors that induce tau modification in both in vitro and in vivo models of diabetes. We speculate that tau cleavage in diabetic conditions (especially in type 2 diabetes) may be a key link for the increased incidence of AD in diabetic patients.

Two dynamin-2 genes are required for normal zebrafish development.

Dynamin-2 (DNM2) is a large GTPase involved in clathrin-mediated endocytosis and related trafficking pathways. Mutations in human DNM2 cause two distinct neuromuscular disorders: centronuclear myopathy and Charcot-Marie-Tooth disease. Zebrafish have been shown to be an excellent animal model for many neurologic disorders, and this system has the potential to inform our understanding of DNM2-related disease. Currently, little is known about the endogenous zebrafish orthologs to human DNM2. In this study, we characterize two zebrafish dynamin-2 genes, dnm2 and dnm2-like. Both orthologs are structurally similar to human DNM2 at the gene and protein levels. They are expressed throughout early development and in all adult tissues examined. Knockdown of dnm2 and dnm2-like gene products resulted in extensive morphological abnormalities during development, and expression of human DNM2 RNA rescued these phenotypes. Our findings suggest that dnm2 and dnm2-like are orthologs to human DNM2, and that they are required for normal zebrafish development.