Abstract 17327: Ischemic Skeletal Muscle From Patients With Peripheral Arterial Disease Displays Marked Desialylation of Cell Surface Glycans: Implications for Ischemia-Targeted Gene Therapy

Abstract

Introduction: Ischemic skeletal muscle is preferentially targeted after IV injection of AAV9 in mouse models of hind-limb ischemia (HLI). Ischemia is known to induce desialylation of cell surface glycans, unmasking the cellular receptor for AAV9 and resulting in significantly higher levels of selective gene expression in ischemic vs non-ischemic muscle. To assess clinical relevance, we compared the differential distribution of sialylated and desialylated cell surface glycans in skeletal muscle biopsies from peripheral arterial disease (PAD) patients and normal controls. Methods: Gastrocnemius (GA) muscle samples from PAD patients and normals (n=29 each) were biopsied under IRB-approved protocols. Muscle sections were stained using the lectins MAL-I and ECL. MAL-I binds to α2,3-sialylated glycans whereas ECL binds to desialylated galactose residues on cell surface glycans. Lectins were labeled with Streptavidin-Alexa-488 (red) for MAL-I and SA-555 (green) for ECL, and visualized using fluorescence microscopy. Quantification of mean fluorescence signal intensity of lectin staining was performed on each section. Results: Visually, MAL-I staining was markedly higher and ECL staining lower in non-ischemic GA from normals when compared to ischemic GA from PAD patients (Figure). The ratio of MAL-I to ECL integrated fluorescence intensity in the ischemic muscles from PAD patients (1.13 ± 0.13) was significantly lower than the same ratio in non-ischemic muscles from normals (1.58 ± 0.14, p<0.02). Conclusions: Desialylation of cell surface glycans is significantly higher in ischemic skeletal muscle from PAD patients, consistent with previous work in a mouse model of HLI. This desialylation can potentially be exploited to treat human PAD with gene therapy, since these desialylated cell surface glycans serve as cell surface receptors for AAV9 and provide for efficient and selective gene transfer to ischemic skeletal muscle following systemic or local delivery.