Naked DNA expressing two isoforms of hepatocyte growth factor induces collateral artery augmentation in a rabbit model of limb ischemia

Abstract

Hepatocyte growth factor (HGF) has been shown to induce angiogenesis in vivo and has potential as a candidate gene for 'therapeutic angiogenesis'. In vivo, two isoforms of HGF, HGF 723 and HGF 728 , consisting of 723 and 728 amino acids, are generated through alternative splicing between exons 4 and 5, but the biological effects of their coexpression have not yet been elucidated. In this study, we generated a series of genomic-complementary DNA (cDNA) hybrids of the HGF gene by inserting various truncated intron 4 into the junction of exons 4 and 5 of HGF cDNA and analyzed the biological activities of these hybrid constructs. We showed that: (1) the hybrid called HGF-X7, which contained 1502 base pairs of intron 4, could drive a higher level of HGF expression than other hybrid constructs and cDNAs of each isoform alone; (2) the pCK vector was most efficient for the gene expression of HGF-X7; (3) coexpression of both isoforms of HGF could more efficiently induce the migration of human umbilical vein endothelial cell (HUVEC) and of the mouse myoblast cell line C 2 C 12 myoblasts than a single isoform of HGF and human vascular endothelial growth factor (VEGF) 165 at a given concentration; (4) intramuscular administration of pCK-HGF-X7 resulted in transient and localized HGF expression in the injected muscle without an increase in the HGF protein levels in other tissues including serum; and (5) intramuscular injection of pCK-HGF-X7 could more efficiently increase the number of angiographically recognizable collateral vessels, as well as improve an intra-arterial Doppler wiremeasured blood flow in the rabbit model of hindlimb ischemia when compared with the identical vector encoding VEGF 165 gene. These results showed that transfer of the genomiccDNA hybrid of the HGF gene could be used as a potential therapeutic approach to human vascular diseases.

Phase 1/2 Open-label Dose-escalation Study of Plasmid DNA Expressing Two Isoforms of Hepatocyte Growth Factor in Patients With Painful Diabetic Peripheral Neuropathy

“…The safety and efficacy of VM202 for cardiovascular diseases have been demonstrated previously in extensive preclinical studies [7][8][9][10][11][12][13] as well as three independent phase I trials. [14][15][16] VM202 is a plasmid DNA containing a therapeutic gene, HGF-X7, which encodes two isoforms of hepatocyte growth factor (HGF).…”

“…Our preclinical studies showed that coexpression of the two isoforms of HGF results in greater angiogenesis than single expression of HGF 728 or VEGF 165 (vascular endothelial growth factor) in various animal models of cardiovascular disease. 7,8 HGF is also neurotrophic for peripheral sensory, sympathetic, and motor neurons, and it promotes neuronal survival and axonal outgrowth both in vitro and in vivo. [23][24][25][26][27][28][29][30] The combined angiogenic and neurotrophic properties of HGF make it an ideal candidate for the treatment of PDPN.…”

Safety of a non-viral plasmid-encoding dual isoforms of hepatocyte growth factor in critical limb ischemia patients: a phase I study

“…12,15,31 The key feature of HGF-X7 is that it was designed by inserting the truncated intron 4 into the junction between exon 4 and exon 5 of hgf cDNA, so that both isoforms of HGF protein are expressed simultaneously and efficiently as in the human genome. 15 Because there is no change in the coding region of the hgf gene, HGF proteins generated from VM202 are identical to the wild-type human HGF proteins.…”

“…14 Moreover, in a rabbit hindlimb model, investigators found that co-expression of both isoforms of HGF could more efficiently induce the migration of HUVEC and C 2 C 12 myoblasts than VEGF 165 or single isoform of HGF. 15 However, the safety and efficacy of the dual isoforms of HGF have not yet been studied in humans.…”

Understanding and Augmenting Collateral Blood Flow During Ischemic Stroke

“…Hepatocyte growth factor (HGF) is another angiogenic growth factor which promotes the growth of endothelial cells. HGF has been shown to efficiently increase the number of collateral vessels and improved the blood flow after hindlimb ischemia in rabbits (Pyun et al, 2010). Perhaps the most promising data relating angiogenic compounds to improved stroke outcome relates to G-CSF and granulocyte monocyte colony stimulating factor (GM-CSF).…”