Inducing heat shock protein 70 expression provides a robust anti-thrombotic effect with minimal bleeding risk

Abstract

Antithrombotic medications target coagulation factors. Their use is associated with an increased bleeding risk. Safer drugs are needed. The heat shock protein 70 (Hsp70) exhibits antithrombotic properties that do not influence bleeding. By using murine models, we aimed to test the hypothesis that overexpressing Hsp70 with CM-695, a first in class dual inhibitor of HDAC6 and phosphodiesterase 9, protects against thrombosis while leaves bleeding tendency unaltered. CM-695 was used to induce Hsp70 overexpression. Hsp70 overexpressing mice were submitted to three thrombosis-triggering procedures. The ferric chloride carotid artery model was used to compare the antithrombotic role of CM-695 and rivaroxaban, a direct oral anticoagulant. The mouse tail transection model was used to compare the bleeding tendency upon CM-695 or rivaroxaban administration. Intraperitoneal (i. p.) 20 mg/kg CM-695 increased Hsp70 expression markedly in the murine aortic tissue. This treatment delayed thrombosis in the collagen/epinephrine [p=0.04 (Log-Rank test), n=10], Rose Bengal/laser [median vessel occlusion time (OT): 58.6 vs 39.0 minutes (min) in the control group (CG), p=0.008, n≥10] and ferric chloride (OT: 14.7 vs 9.2 min in the CG, p=0.032, n≥10) models. I.p. 80 mg/kg CM-695 (n≥9) and intravenous 3 mg/kg rivaroxaban (n≥8) significantly delayed thrombosis. CM-695 did not induce bleeding [median bleeding time (BT): 8.5 vs 7.5 min in the CG, n≥10]. However, BT was dramatically increased by rivaroxaban (30.0 vs 13.7 min in the CG, p=0.001, n=10). In conclusion, CM-695 is a new antithrombotic small molecule devoid of bleeding risk that may be envisioned as a useful clinical tool.

Taking Advantage of the Selectivity of Histone Deacetylases and Phosphodiesterase Inhibitors to Design Better Therapeutic Strategies to Treat Alzheimer’s Disease

“…HSPA1A/B was among the genes overexpressed in the Affymetrix microarray analysis and validated by qRT-PCR in the hippocampus of mice administered CM-695. Significantly, a similar induction was induced by CM-695 in a model of thrombosis (Allende et al, 2017). Hsp70 fulfills a neuroprotective role in AD by decreasing the oligomerization and production of toxic Aβ isoforms, and by increasing its degradation (Magrané et al, 2004; Muchowski and Wacker, 2005; Kumar et al, 2007).…”

Multitarget Approach for the Treatment of Alzheimer’s Disease: Inhibition of Phosphodiesterase 9 (PDE9) and Histone Deacetylases (HDACs) Covering Diverse Selectivity Profiles

“…Compound 31b showed moderate cytotoxicity in THLE-2 (8.4 μM and 10.4 μM after 72 and 24 h, respectively), with weaker proliferative effect on neurons and glial cells from wild-type mice (LC 50 = 16.7 μM at 72 h) and lower tolerance for peripheral blood mononuclear cells (PBMCs; LC 50 = 5 μM at 72 h). Compound 31b had minimal inhibition of five of the main cytochrome isoforms: <50% for 1A2, 2C19, and 2C9, 75% for 2D6, and 83% for 3A4 (10 μM) and good solubility (109.4 μg/mL) . Microsomal stability was moderate, with estimated half-lives of 37.1 and 55.9 min in mouse and human, respectively.…”

“…Compound 31b had minimal inhibition of five of the main cytochrome isoforms: <50% for 1A2, 2C19, and 2C9, 75% for 2D6, and 83% for 3A4 (10 μM) and good solubility (109.4 μg/mL). 54 Microsomal stability was moderate, with estimated half-lives of 37.1 and 55.9 min in mouse and human, respectively. Compound 31b had low plasma protein binding in human (18% of the unbound fraction).…”

VAMP3 and SNAP23 as Potential Targets for Preventing the Disturbed Flow-Accelerated Thrombus Formation

“…After incubation the artery was rinsed with saline. Thrombus formation was assessed by monitoring blood flow immediately after FeCl 3 application and for up to 40 min, if the blood flow did not stop after 40 min, the time of thrombus formation was recorded as 40 min (Allende et al, 2017 ). The time to occlusion was calculated as the time required for blood flow through the carotid artery was monitored until vessel occlusion reached 95% (Liang et al, 2015a ).…”