HK-001 ATTENUATE AMYOTROPHIC LATERAL SCLEROSIS SYMPTOMS THROUGH DOWN-REGULATION AUTOPHAGY

Kuo-Wei Hsueh, Shinn-Zong Lin and Horng-Jyh Harn

Bioinnovation Center, Tzu Chu Fundation, Hualien, Taiwan

Abstract

Background: Amyotrophic lateral sclerosis (ALS) is a lethal degenerating disease, characterized by progressive muscular atrophy without any effective treatment. Here, we demonstrated the efficacy of abrograting autophagy in motor neurons (MN) by treatment with HK-001 in ALS transgenic mice (SOD1G93A). In this study, we found HK-001 may pose as a therapeutic regimen for ALS and relevant neurodegenerative diseases.

Methods: Animals were randomly distributed into three groups at 60 days of age: (1) vehicle, (2) HK-001 (100)-treated, given 100 mg HK-001/Kg/qd body weight, (3) HK-001 (500)-treated, given 500 mg HK-001/Kg/qd body weight, and (4) 250 mg HK-001/Kg/bid body weight. HK-001 was dissolved in olive oil (HK-001; Lancaster Synthesis Ltd., Newgate, Morecambe, UK), and administered by oral gavages once daily beginning at 60 days of age. The end-point was determined as the inability of the mouse to roll over within 30 s after being placed on its side (Gurney et al., 1996).

Results: Pre-symptomatic oral administration of 250 mg/kg/bid HK-001 significantly prolonged the survival period (203.9 ± 18.3 days), improved motor function, and attenuated MN loss compared to vehicle control (126.4 ± 7.2 days). This prolonged survival of ALS mice is much more robust than that reported with riluzole (140 days), which is an approved clinical therapy for ALS. Regarding the molecular mechanism, we demonstrated that HK-001 could decrease both of LC3-II (a biomarker of autophagy) and caspase 3 (a biomarker of apoptosis) expression, altogether increasing MN survival. This result was also confirmed by double transgenic mice (SOD1G93A:LC3-GFP) which showed that oral administration of HK-001 reduced GFP density and decreased caspase-3 expression. In addition, electron microscopy revealed that HK-001 administration not only decreased autophagosome number but also reduced morphological dysfunction of mitochondria. In summary, these results indicate that down-regulation of autophagy activation via HK-001 may as potential compound for testing therapeutic effect in ALS symptoms.

Keywords: ALS, HK-001, autophagy, bid, two times a day, qd, one times a day.