The pyruvate kinase activator mitapivat reduces hemolysis and improves anemia in a \u3b2-thalassemia mouse model

Anemia in \u3b2-thalassemia is related to ineffective erythropoiesis and reduced red cell survival. Excess free heme and accumulation of unpaired \u3b1-globin chains impose substantial oxidative stress on \u3b2-thalassemic erythroblasts and erythrocytes, impacting cell metabolism. We hypothesized that increased pyruvate kinase activity induced by mitapivat (AG-348) in the Hbbth3/+ mouse model for \u3b2-thalassemia would reduce chronic hemolysis and ineffective erythropoiesis through stimulation of red cell glycolytic metabolism. Oral mitapivat administration ameliorated ineffective erythropoiesis and anemia in Hbbth3/+ mice. Increased ATP, reduced reactive oxygen species production, and reduced markers of mitochondrial dysfunction associated with improved mitochondrial clearance suggested enhanced metabolism following mitapivat administration in \u3b2-thalassemia. The amelioration of responsiveness to erythropoietin resulted in reduced soluble erythroferrone, increased liver Hamp expression, and diminished liver iron overload. Mitapivat reduced duodenal Dmt1 expression potentially by activating the pyruvate kinase M2-HIF2\u3b1 axis, representing a mechanism additional to Hamp in controlling iron absorption and preventing \u3b2-thalassemia\u2013related liver iron overload. In ex vivo studies on erythroid precursors from patients with \u3b2-thalassemia, mitapivat enhanced erythropoiesis, promoted erythroid maturation, and decreased apoptosis. Overall, pyruvate kinase activation as a treatment modality for \u3b2-thalassemia in preclinical model systems had multiple beneficial effects in the erythropoietic compartment and beyond, providing a strong scientific basis for further clinical trials

Systemic detoxification of plant secondary metabolites by ruminant herbivores

The effect of xenobiotic compounds on drug metabolising enzymes was assessed in ruminants; the development of an in vivo approach using a drug cocktail to measure specific enzyme families of phase I and II metabolism has been developed. Predictive clearances based on in vitro data from tolbutamide, phenacetin, chlorzoxazone, midazolam and oxazepam incubations were correlated with clearance determined in vivo (R2= 0.695). A drug cocktail approach was used to assess the effect of plant secondary compounds on phase I and II enzymatic activities. This approach showed that phase I enzymes, e.g. CYP3A, can be affected by plant compounds, such as α-pinene or pyrrolizidine alkaloids. Additionally, due to carry over from one treatment period to the next, this experiment showed that interaction between plant secondary compounds can alter the overall effect of a single compound. Subsequently, in vitro experiments with hepatocytes or rumen microorganisms from sheep or goats, showed that pyrrolizidine alkaloids were not completely degraded in rumen fluid after 48h. Dose- and time-dependent toxicity of pyrrolizidine alkaloids to hepatocytes were observed. The magnitude of toxicity measured in goat and sheep hepatocytes was compatible with in vivo observations. Interaction between plant compounds observed in vivo was confirmed by in vitro assays. This concept of interaction between plant compounds was used to decrease the toxicity of pyrrolizidine alkaloids to hepatocytes. Exposure to certain plant compounds prior to the potentially toxic compounds could decrease pyrrolizidine alkaloid toxicity by acting on phase I or II enzymes. The mechanism of protection appears to be different between sheep and goats.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Experimental study of post-crack vibrations in dynamic fracture

International audienceVibrations induced by crack propagation in a strip of bonded silicon wafers are studied. A new optical setup enables the fast recording of crack-originated acoustic waves, emitted both ahead and behind the crack front, in bonded and separated wafers, respectively. Three types of crack-induced vibrations are identified, corresponding to different excitations and responses of the system: (1) “pneumatic” vibrations involving inertia and gas expansion/compression, (2) standard flexural waves involving inertia and bending rigidity, and (3) post-crack vibrations involving inertia, bending rigidity, and coupling to gas pressure. We show that a standard “beam on elastic foundation” model can explain these latter vibrations that occur along crack edges and is consistent with the observed frequencies

Inhibition of D-2HG leads to upregulation of a proinflammatory gene signature in a novel HLA-A2/HLA-DR1 transgenic mouse model of IDH1R132H-expressing glioma

BACKGROUND: Long-term prognosis of WHO grade II, isocitrate dehydrogenase (IDH)-mutated low-grade glioma (LGG) is poor due to high risks of recurrence and malignant transformation into high-grade glioma. Immunotherapy strategies are attractive given the relatively intact immune system of patients with LGG and the slow tumor growth rate. However, accumulation of the oncometabolite D-2-hydroxyglutarate (D-2HG) in IDH-mutated gliomas leads to suppression of inflammatory pathways in the tumor microenvironment, thereby contributing to the 'cold' tumor phenotype. Inhibiting D-2HG production presents an opportunity to generate a robust antitumor response following tumor antigen vaccination and immune checkpoint blockade. METHODS: An IDH1R132H glioma model was created in syngeneic HLA-A2/HLA-DR1-transgenic mice, allowing us to evaluate the vaccination with the human leukocyte antigens (HLA)-DR1-restricted, IDH1R132H mutation-derived neoepitope. The effects of an orally available inhibitor of mutant IDH1 and IDH2, AG-881, were evaluated as monotherapy and in combination with the IDH1R132H peptide vaccination or anti-PD-1 immune checkpoint blockade. RESULTS: The HLA-A2/HLA-DR1-syngeneic IDH1R132H cell line expressed the IDH1 mutant protein and formed D-2HG producing orthotopic gliomas in vivo. Treatment of tumor-bearing mice with AG-881 resulted in a reduction of D-2HG levels in IDH1R132H glioma cells (10 fold) and tumor-associated myeloid cells, which demonstrated high levels of intracellular D-2HG in the IDH1R132H gliomas. AG-881 monotherapy suppressed the progression of IDH1R132H gliomas in a CD4+ and CD8+ cell-dependent manner, enhanced proinflammatory IFNγ-related gene expression, and increased the number of CD4+ tumor-infiltrating T-cells. Prophylactic vaccination with the HLA-DR1-restricted IDH1R132H peptide or tumor-associated HLA-A2-restricted peptides did not enhance survival of tumor-bearing animals; however, vaccination with both HLA-A2-IDH1R132H and DR1-IDH1R132H peptides in combination with the IDH inhibitor significantly prolonged survival. Finally, tumor-bearing mice treated with both AG-881 and a PD-1 blocking antibody demonstrated improved survival when compared with either treatment alone. CONCLUSION: The development of effective IDH1R132H-targeting vaccine may be enhanced by integration with HLA class I-restricted cytotoxic T cell epitopes and AG-881. Our HLA-A2/HLA-DR1-syngeneic IDH1R132H glioma model should allow us to evaluate key translational questions related to the development of novel strategies for patients with IDH-mutant glioma