Feasible diet and circadian interventions reduce in vivo progression of FLT3-ITD-positive acute myeloid leukemia.

BACKGROUND: Acute myeloid leukemia (AML) with an internal tandem duplication in the fms-like tyrosine kinase receptor 3 gene (FLT3-ITD) is associated with poor survival, and few studies have examined the impact of modifiable behaviors, such as nutrient quality and timing, in this subset of acute leukemia. METHODS: The influence of diet composition (low-sucrose and/or low-fat diets) and timing of diet were tested in tandem with anthracycline treatment in orthotopic xenograft mouse models. A pilot clinical study to test receptivity of pediatric leukemia patients to macronutrient matched foods was conducted. A role for the circadian protein, BMAL1 (brain and muscle ARNT-like 1), in effects of diet timing was studied by overexpression in FLT3-ITD-bearing AML cells. RESULTS: Reduced tumor burden in FLT3-ITD AML-bearing mice was observed with interventions utilizing low-sucrose and/or low-fat diets, or time-restricted feeding (TRF) compared to mice fed normal chow ad libitum. In a tasting study, macronutrient matched low-sucrose and low-fat meals were offered to pediatric acute leukemia patients who largely reported liking the meals. Expression of the circadian protein, BMAL1, was heightened with TRF and the low-sucrose diet. BMAL1 overexpression and treatment with a pharmacological inducer of BMAL1 was cytotoxic to FLT3-ITD AML cells. CONCLUSIONS: Mouse models for FLT3-ITD AML show that diet composition and timing slows progression of FLT3-ITD AML growth in vivo, potentially mediated by BMAL1. These interventions to enhance therapy efficacy show preliminary feasibility, as pediatric leukemia patients responded favorable to preparation of macronutrient matched meals.

Computational immune infiltration analysis of pediatric high-grade gliomas (pHGGs) reveals differences in immunosuppression and prognosis by tumor location.

Immunotherapy for cancer has moved from pre-clinical hypothesis to successful clinical application in the past 15 years. However, not all cancers have shown response rates in clinical trials for these new agents, high-grade gliomas in particular have proved exceedingly refractory to immunotherapy. In adult patients, there has been much investigation into these failures, and researchers have concluded that an immunosuppressive microenvironment combined with low mutational burden render adult glioblastomas "immune cold". Pediatric cancer patients develop gliomas at a higher rate per malignancy than adults, and their brain tumors bear even fewer mutations. These tumors can also develop in more diverse locations in the brain, beyond the cerebral hemispheres seen in adults, including in the brainstem where critical motor functions are controlled. While adult brain tumor immune infiltration has been extensively profiled from surgical resections, this is not possible for brainstem tumors which can only be sampled at autopsy. Given these limitations, there is a dearth of information on immune cells and their therapeutic and prognostic impact in pediatric high-grade gliomas (pHGGs), including hemispheric tumors in addition to brainstem. In this report we use computational methods to examine immune infiltrate in pHGGs and discover distinct immune patterns between hemispheric and brainstem tumors. In hemispheric tumors, we find positive prognostic associations for regulatory T-cells, memory B-cells, eosinophils, and dendritic cells, but not in brainstem tumors. These differences suggest that immunotherapeutic approaches must be cognizant of pHGG tumor location and tailored for optimum efficacy.

Feasibility and early experience of a novel multidisciplinary alcohol-associated liver disease clinic.

BACKGROUND: Alcohol cessation improves mortality in alcohol-associated liver disease (ALD), but access to treatment is limited. To address this gap, implementation and early feasibility and outcomes of a multidisciplinary ALD clinic are described. METHODS: The clinic comprised a hepatologist, psychiatrist, psychologist, nurse, and social worker. Patients included those with alcohol-associated cirrhosis or acute alcoholic hepatitis who were not in the transplant evaluation process, who had less than 6 months' sobriety and willingness to engage in alcohol use treatment. Psychosocial metrics in addition to routine hepatic function labs were collected. Treatment plans were tailored based on patient preferences and needs after multidisciplinary discussion. RESULTS: 89 patients were referred from both inpatient and outpatient settings, with 51 seen during the initial year. 38 remained active in clinic (4 died, 6 discharged, 3 moved to transplant clinic). 55% were women, 88% were white, 61% had private insurance. 49% had alcoholic hepatitis. 71% were decompensated. 80% had severe alcohol use disorder (AUD) and 84% had at least 1 comorbid psychiatric or substance use disorder. 63% chose one-on-one AUD treatment, 57% were prescribed relapse prevention medications. Mean MELD-Na score improved from baseline of 14 (SD 6.6) to 11.3 at 6 months (p=0.01). Hospital utilization significantly declined when comparing 6 months before to 6 months after initial visit (emergency department visits: 0.51 to 0.20 per person-month; inpatient admission: 0.34 to 0.14 per person-month; (ß= -0.89, 95% CI -1.18 to -0.60). CONCLUSIONS: A multidisciplinary ALD clinic was feasible with encouraging early outcomes. Further research should explore ways to expand this model and increase clinic capacity.

Cigarette Smoke Exposure in Mice using a Whole-Body Inhalation System.

Close to 14% of adults in the United States were reported to smoke cigarettes in 2018. The effects of cigarette smoke (CS) on lungs and cardiovascular diseases have been widely studied, however, the impact of CS in other tissues and organs such as blood and bone marrow remain incompletely defined. Finding the appropriate system to study the effects of CS in rodents can be prohibitively expensive and require the purchase of commercially available systems. Thus, we set out to build an affordable, reliable, and versatile system to study the pathologic effects of CS in mice. This whole-body inhalation exposure system (WBIS) set-up mimics the breathing and puffing of cigarettes by alternating exposure to CS and clean air. Here we show that this do-it-yourself (DIY) system induces airway inflammation and lung emphysema in mice after 4-months of cigarette smoke exposure. The effects of whole-body inhalation (WBI) of CS on hematopoietic stem and progenitor cells (HSPCs) in the bone marrow using this apparatus are also shown.

Scaffolding LSD1 Inhibitors Impair NK Cell Metabolism and Cytotoxic Function Through Depletion of Glutathione.

Cell therapies such as chimeric-antigen receptor (CAR) T-cells and NK cells are cutting-edge methods for treating cancer and other diseases. There is high interest in optimizing drug treatment regimens to best work together with emerging cell therapies, such as targeting epigenetic enzymes to stimulate recognition of tumor cells by immune cells. Herein, we uncover new mechanisms of the histone demethylase LSD1, and various inhibitors targeting unique domains of LSD1, in the function of NK cells grown for cell therapy. Catalytic inhibitors (tranylcypromine and the structural derivatives GSK LSD1 and RN-1) can irreversibly block the demethylase activity of LSD1, while scaffolding inhibitors (SP-2509 and clinical successor SP-2577, also known as seclidemstat) disrupt epigenetic complexes that include LSD1. Relevant combinations of LSD1 inhibitors with cell therapy infusions and immune checkpoint blockade have shown efficacy in pre-clinical solid tumor models, reinforcing a need to understand how these drugs would impact T- and NK cells. We find that scaffolding LSD1 inhibitors potently reduce oxidative phosphorylation and glycolysis of NK cells, and higher doses induce mitochondrial reactive oxygen species and depletion of the antioxidant glutathione. These effects are unique to scaffolding inhibitors compared to catalytic, to NK cells compared to T-cells, and importantly, can fully ablate the lytic capacity of NK cells. Supplementation with biologically achievable levels of glutathione rescues NK cell cytolytic function but not NK cell metabolism. Our results suggest glutathione supplementation may reverse NK cell activity suppression in patients treated with seclidemstat.

Pharmacologic inhibition of lysine-specific demethylase 1 as a therapeutic and immune-sensitization strategy in pediatric high-grade glioma.

BACKGROUND: Diffuse midline gliomas (DMG), including brainstem diffuse intrinsic pontine glioma (DIPG), are incurable pediatric high-grade gliomas (pHGG). Mutations in the H3 histone tail (H3.1/3.3-K27M) are a feature of DIPG, rendering them therapeutically sensitive to small-molecule inhibition of chromatin modifiers. Pharmacological inhibition of lysine-specific demethylase 1 (LSD1) is clinically relevant but has not been carefully investigated in pHGG or DIPG. METHODS: Patient-derived DIPG cell lines, orthotopic mouse models, and pHGG datasets were used to evaluate effects of LSD1 inhibitors on cytotoxicity and immune gene expression. Immune cell cytotoxicity was assessed in DIPG cells pretreated with LSD1 inhibitors, and informatics platforms were used to determine immune infiltration of pHGG. RESULTS: Selective cytotoxicity and an immunogenic gene signature were established in DIPG cell lines using clinically relevant LSD1 inhibitors. Pediatric HGG patient sequencing data demonstrated survival benefit of this LSD1-dependent gene signature. Pretreatment of DIPG with these inhibitors increased lysis by natural killer (NK) cells. Catalytic LSD1 inhibitors induced tumor regression and augmented NK cell infusion in vivo to reduce tumor burden. CIBERSORT analysis of patient data confirmed NK infiltration is beneficial to patient survival, while CD8 T cells are negatively prognostic. Catalytic LSD1 inhibitors are nonperturbing to NK cells, while scaffolding LSD1 inhibitors are toxic to NK cells and do not induce the gene signature in DIPG cells. CONCLUSIONS: LSD1 inhibition using catalytic inhibitors is selectively cytotoxic and promotes an immune gene signature that increases NK cell killing in vitro and in vivo, representing a therapeutic opportunity for pHGG. KEY POINTS: 1. LSD1 inhibition using several clinically relevant compounds is selectively cytotoxic in DIPG and shows in vivo efficacy as a single agent.2. An LSD1-controlled gene signature predicts survival in pHGG patients and is seen in neural tissue from LSD1 inhibitor-treated mice.3. LSD1 inhibition enhances NK cell cytotoxicity against DIPG in vivo and in vitro with correlative genetic biomarkers.

Cutting Edge Therapeutic Insights Derived from Molecular Biology of Pediatric High-Grade Glioma and Diffuse Intrinsic Pontine Glioma (DIPG).

Pediatric high-grade glioma (pHGG) and brainstem gliomas are some of the most challenging cancers to treat in children, with no effective therapies and 5-year survival at ~2% for diffuse intrinsic pontine glioma (DIPG) patients. The standard of care for pHGG as a whole remains surgery and radiation combined with chemotherapy, while radiation alone is standard treatment for DIPG. Unfortunately, these therapies lack specificity for malignant glioma cells and have few to no reliable biomarkers of efficacy. Recent discoveries have revealed that epigenetic disruption by highly conserved mutations in DNA-packaging histone proteins in pHGG, especially DIPG, contribute to the aggressive nature of these cancers. In this review we pose unanswered questions and address unexplored mechanisms in pre-clinical models and clinical trial data from pHGG patients. Particular focus will be paid towards therapeutics targeting chromatin modifiers and other epigenetic vulnerabilities that can be exploited for pHGG therapy. Further delineation of rational therapeutic combinations has strong potential to drive development of safe and efficacious treatments for pHGG patients.