HDAC11 deficiency disrupts oncogene-induced hematopoiesis in myeloproliferative neoplasms.

Protein acetylation is an important contributor to cancer initiation. Histone deacetylase 6 (HDAC6) controls JAK2 translation and protein stability and has been implicated in JAK2-driven diseases best exemplified by myeloproliferative neoplasms (MPNs). By using novel classes of highly selective HDAC inhibitors and genetically deficient mouse models, we discovered that HDAC11 rather than HDAC6 is necessary for the proliferation and survival of oncogenic JAK2-driven MPN cells and patient samples. Notably, HDAC11 is variably expressed in primitive stem cells and is expressed largely upon lineage commitment. Although Hdac11is dispensable for normal homeostatic hematopoietic stem and progenitor cell differentiation based on chimeric bone marrow reconstitution, Hdac11 deficiency significantly reduced the abnormal megakaryocyte population, improved splenic architecture, reduced fibrosis, and increased survival in the MPLW515L-MPN mouse model during primary and secondary transplantation. Therefore, inhibitors of HDAC11 are an attractive therapy for treating patients with MPN. Although JAK2 inhibitor therapy provides substantial clinical benefit in MPN patients, the identification of alternative therapeutic targets is needed to reverse MPN pathogenesis and control malignant hematopoiesis. This study establishes HDAC11 as a unique type of target molecule that has therapeutic potential in MPN.

Nutritional ketosis delays the onset of isoflurane induced anesthesia.

BACKGROUND: Ketogenic diet (KD) and exogenous ketone supplements can evoke sustained ketosis, which may modulate sleep and sleep-like effects. However, no studies have been published examining the effect of ketosis on the onset of general isoflurane induced anesthesia. Therefore, we investigated the effect of the KD and different exogenous ketogenic supplements on the onset of akinesia induced by inhalation of isoflurane. METHODS: We used a high fat, medium protein and low carbohydrate diet (KD) chronically (10 weeks) in the glucose transporter 1 (GLUT1) deficiency (G1D) syndrome mice model and sub-chronically (7 days) in Sprague-Dawley (SPD) rats. To investigate the effect of exogenous ketone supplements on anesthetic induction we also provided either 1) a standard rodent chow diet (SD) mixed with 20% ketone salt supplement (KS), or 2) SD mixed with 20% ketone ester supplement (KE; 1,3 butanediol-acetoacetate diester) to G1D mice for 10 weeks. Additionally, SPD rats and Wistar Albino Glaxo Rijswijk (WAG/Rij) rats were fed the SD, which was supplemented by oral gavage of KS or KE for 7 days (SPD rats: 5 g/kg body weight/day; WAG/Rij rats: 2.5 g/kg body weight/day). After these treatments (10 weeks for the mice, and 7 days for the rats) isoflurane (3%) was administered in an anesthesia chamber, and the time until anesthetic induction (time to immobility) was measured. Blood ketone levels were measured after anesthetic induction and correlation was calculated for blood beta-hydroxybutyrate (ßHB) and anesthesia latency. RESULTS: Both KD and exogenous ketone supplementation increased blood ketone levels and delayed the onset of isoflurane-induced immobility in all investigated rodent models, showing positive correlation between the two measurements. These results demonstrate that elevated blood ketone levels by either KD or exogenous ketones delayed the onset of isoflurane-induced anesthesia in these animal models. CONCLUSIONS: These findings suggest that ketone levels might affect surgical anesthetic needs, or could potentially decrease or delay effects of other narcotic gases.

Exogenous Ketone Supplements Improved Motor Performance in Preclinical Rodent Models.

Nutritional ketosis has been proven effective for neurometabolic conditions and disorders linked to metabolic dysregulation. While inducing nutritional ketosis, ketogenic diet (KD) can improve motor performance in the context of certain disease states, but it is unknown whether exogenous ketone supplements-alternatives to KDs-may have similar effects. Therefore, we investigated the effect of ketone supplements on motor performance, using accelerating rotarod test and on postexercise blood glucose and R-beta-hydroxybutyrate (R-ßHB) levels in rodent models with and without pathology. The effect of KD, butanediol (BD), ketone-ester (KE), ketone-salt (KS), and their combination (KE + KS: KEKS) or mixtures with medium chain triglyceride (MCT) (KE + MCT: KEMCT; KS + MCT: KSMCT) was tested in Sprague-Dawley (SPD) and WAG/Rij (WR) rats and in GLUT-1 Deficiency Syndrome (G1D) mice. Motor performance was enhanced by KEMCT acutely, KE and KS subchronically in SPD rats, by KEKS and KEMCT groups in WR rats, and by KE chronically in G1D mice. We demonstrated that exogenous ketone supplementation improved motor performance to various degrees in rodent models, while effectively elevated R-ßHB and in some cases offsets postexercise blood glucose elevations. Our results suggest that improvement of motor performance varies depending on the strain of rodents, specific ketone formulation, age, and exposure frequency.

Exogenous Ketones Lower Blood Glucose Level in Rested and Exercised Rodent Models.

Diseases involving inflammation and oxidative stress can be exacerbated by high blood glucose levels. Due to tight metabolic regulation, safely reducing blood glucose can prove difficult. The ketogenic diet (KD) reduces absolute glucose and insulin, while increasing fatty acid oxidation, ketogenesis, and circulating levels of ß-hydroxybutyrate (ßHB), acetoacetate (AcAc), and acetone. Compliance to KD can be difficult, so alternative therapies that help reduce glucose levels are needed. Exogenous ketones provide an alternative method to elevate blood ketone levels without strict dietary requirements. In this study, we tested the changes in blood glucose and ketone (ßHB) levels in response to acute, sub-chronic, and chronic administration of various ketogenic compounds in either a post-exercise or rested state. WAG/Rij (WR) rats, a rodent model of human absence epilepsy, GLUT1 deficiency syndrome mice (GLUT1D), and wild type Sprague Dawley rats (SPD) were assessed. Non-pathological animals were also assessed across different age ranges. Experimental groups included KD, standard diet (SD) supplemented with water (Control, C) or with exogenous ketones: 1, 3-butanediol (BD), ßHB mineral salt (KS), KS with medium chain triglyceride/MCT (KSMCT), BD acetoacetate diester (KE), KE with MCT (KEMCT), and KE with KS (KEKS). In rested WR rats, the KE, KS, KSMCT groups had lower blood glucose level after 1 h of treatment, and in KE and KSMCT groups after 24 h. After exercise, the KE, KSMCT, KEKS, and KEMCT groups had lowered glucose levels after 1 h, and in the KEKS and KEMCT groups after 7 days, compared to control. In GLUT1D mice without exercise, only KE resulted in significantly lower glucose levels at week 2 and week 6 during a 10 weeks long chronic feeding study. In 4-month and 1-year-old SPD rats in the post-exercise trials, blood glucose was significantly lower in KD and KE, and in KEMCT groups, respectively. After seven days, the KSMCT group had the most significantly reduced blood glucose levels, compared to control. These results indicate that exogenous ketones were efficacious in reducing blood glucose levels within and outside the context of exercise in various rodent models of different ages, with and without pathology.

Corrigendum: Exogenous Ketone Supplements Reduce Anxiety-Related Behavior in Sprague-Dawley and Wistar Albino Glaxo/Rijswijk Rats.

[This corrects the article on p. 137 in vol. 9, PMID: 27999529.].

Efficacy of ALK5 inhibition in myelofibrosis.

Myelofibrosis (MF) is a bone marrow disorder characterized by clonal myeloproliferation, aberrant cytokine production, extramedullary hematopoiesis, and bone marrow fibrosis. Although somatic mutations in JAK2, MPL, and CALR have been identified in the pathogenesis of these diseases, inhibitors of the Jak2 pathway have not demonstrated efficacy in ameliorating MF in patients. TGF-ß family members are profibrotic cytokines and we observed significant TGF-ß1 isoform overexpression in a large cohort of primary MF patient samples. Significant overexpression of TGF-ß1 was also observed in murine clonal MPLW515L megakaryocytic cells. TGF-ß1 stimulated the deposition of excessive collagen by mesenchymal stromal cells (MSCs) by activating the TGF-ß receptor I kinase (ALK5)/Smad3 pathway. MSCs derived from MPLW515L mice demonstrated sustained overproduction of both collagen I and collagen III, effects that were abrogated by ALK5 inhibition in vitro and in vivo. Importantly, use of galunisertib, a clinically active ALK5 inhibitor, significantly improved MF in both MPLW515L and JAK2V617F mouse models. These data demonstrate the role of malignant hematopoietic stem cell (HSC)/TGF-ß/MSC axis in the pathogenesis of MF, and provide a preclinical rationale for ALK5 blockade as a therapeutic strategy in MF.

Exogenous Ketone Supplements Reduce Anxiety-Related Behavior in Sprague-Dawley and Wistar Albino Glaxo/Rijswijk Rats.

Nutritional ketosis has been proven effective for seizure disorders and other neurological disorders. The focus of this study was to determine the effects of ketone supplementation on anxiety-related behavior in Sprague-Dawley (SPD) and Wistar Albino Glaxo/Rijswijk (WAG/Rij) rats. We tested exogenous ketone supplements added to food and fed chronically for 83 days in SPD rats and administered sub-chronically for 7 days in both rat models by daily intragastric gavage bolus followed by assessment of anxiety measures on elevated plus maze (EPM). The groups included standard diet (SD) or SD + ketone supplementation. Low-dose ketone ester (LKE; 1,3-butanediol-acetoacetate diester, ~10 g/kg/day, LKE), high dose ketone ester (HKE; ~25 g/kg/day, HKE), beta-hydroxybutyrate-mineral salt (ßHB-S; ~25 g/kg/day, KS) and ßHB-S + medium chain triglyceride (MCT; ~25 g/kg/day, KSMCT) were used as ketone supplementation for chronic administration. To extend our results, exogenous ketone supplements were also tested sub-chronically on SPD rats (KE, KS and KSMCT; 5 g/kg/day) and on WAG/Rij rats (KE, KS and KSMCT; 2.5 g/kg/day). At the end of treatments behavioral data collection was conducted manually by a blinded observer and with a video-tracking system, after which blood ßHB and glucose levels were measured. Ketone supplementation reduced anxiety on EPM as measured by less entries to closed arms (sub-chronic KE and KS: SPD rats and KSMCT: WAG/Rij rats), more time spent in open arms (sub-chronic KE: SPD and KSMCT: WAG/Rij rats; chronic KSMCT: SPD rats), more distance traveled in open arms (chronic KS and KSMCT: SPD rats) and by delayed latency to entrance to closed arms (chronic KSMCT: SPD rats), when compared to control. Our data indicates that chronic and sub-chronic ketone supplementation not only elevated blood ßHB levels in both animal models, but reduced anxiety-related behavior. We conclude that ketone supplementation may represent a promising anxiolytic strategy through a novel means of inducing nutritional ketosis.