Abstract
Background
Acute intermittent porphyria (AIP) is an autosomal dominant disorder caused by the half‐normal activity of hydroxymethylbilane synthase (HMB‐synthase). Affected individuals can experience episodic, life‐threatening, acute neurological attacks that are precipitated by various drugs, dieting, and hormonal changes. Intravenous hematin is used to treat the attacks, but a more effective, preventive therapy is needed, especially for patients with frequent attacks. Since the disease is a hepatic encephalopathy, efforts were focused towards evaluating four different combinations of liver‐specific enhancers and promoters for maximal hepatic HMB‐synthase expression.
Methods
Four different mammalian expression vectors, each carrying a unique combination of liver‐specific enhancers and promoters driving murine HMB‐synthase cDNA expression, were transiently transfected into HepG2 cells. The vectors included: HMBS‐1; human α1‐microglobulin enhancer/α1‐antityrpsin promoter (α1Me/α1ATp), HMBS‐2; α1Me/human serum albumin promoter (α1Me/SAp), HMBS‐3; human prothrombin enhancer/SAp (PTe/SAp), and HMBS‐4; (PTe/α1ATp). Each HMB‐synthase construct and a luciferase reporter construct were hydrodynamically coinjected into mice with HMB‐synthase deficiency and evaluated for hepatic expression 24 h post‐injection, the time‐point of peak hepatic HMB‐synthase expression.
Results
Following transient transfection into HepG2 cells, HMBS‐1 (α1Me/α1ATp) had the highest HMB‐synthase expression level, with an ∼8‐fold increase over endogenous cellular activities. Construct HMBS‐1 also had the highest hepatic HMB‐synthase activity following hydrodynamic delivery into HMB‐synthase deficient mice, with a ∼6‐fold increase over saline‐treated mice.
Conclusions
These studies support the use of a gene therapy vector containing the α1Me/α1ATp combination for preclinical studies of the efficacy and safety of liver‐targeted gene therapy for AIP. Copyright © 2007 John Wiley & Sons, Ltd.