Airway gene transfer in a non-human primate: Lentiviral gene expression in marmoset lungs

Abstract

Genetic therapies for cystic fibrosis (CF) must be assessed for safety and efficacy, so testing in a non-human primate (NHP) model is invaluable. In this pilot study we determined if the conducting airways of marmosets (n = 2) could be transduced using an airway pre-treatment followed by an intratracheal bolus dose of a VSV-G pseudotyped HIV-1 based lentiviral (LV) vector (LacZ reporter). LacZ gene expression (X-gal) was assessed after 7 days and found primarily in conducting airway epithelia as well as in alveolar regions. The LacZ gene was not detected in liver or spleen via qPCR. Vector p24 protein bio-distribution into blood was transient. Dosing was well tolerated. This preliminary study confirmed the transducibility of CF-relevant airway cell types. The marmoset is a promising NHP model for testing and translating genetic treatments for CF airway disease towards clinical trials.

Barriers to inhaled gene therapy of obstructive lung diseases: A review

“…However, lentiviruses have limited tissue tropism and, thus, capsid engineering is required to enable their use in gene therapy applications [124]. Initial preclinical studies have demonstrated efficient, persistent pseudotyped lentivirus-mediated gene expression in the lung [125–131]. For example, a recent study using pseudotyped lentivirus delivered intranasally showed gene expression in the lungs of mice that lasted up to 22 months after initial dosing without signs of toxicity or insertional mutagenesis (Figs.…”

“…They were able to transduce via the apical surface of primary HAE isolated from a ΔF508/ΔF508 homozygous CF patient, resulting in ~30% restoration of CFTR-mediated chloride currents [112]. Initial inhaled gene transfer studies with lentivirus focused on vectors pseudotyped with vesicular stomatitis virus G glycoprotein (VSV-G) [125–129]. However, this approach was generally developed to broaden the tropism of lentivirus rather than to specifically design a system for inhaled gene therapy applications [124].…”

“…However, this approach was generally developed to broaden the tropism of lentivirus rather than to specifically design a system for inhaled gene therapy applications [124]. It was later found for VSV-G-pseudotyped lentivirus that pre-treatment with compounds disrupting tight junction is required for gene transfer to the airway epithelium, suggesting that the transduction was achieved through the basolateral surface [125, 129]. To bypass the need for adjuvant treatments, hybrid lentiviral vectors were engineered by incorporating envelope proteins from SeV or baculovirus that have demonstrated tropism for the apical surface of airway epithelium [9].…”

“…Transduction by AdV was greatly enhanced in vivo in mouse nasal and airway epithelium pre-treated with PDOC [326] and C10 [327]. Likewise, pre-treatment with LPC has been shown to enhance VSV-G-pseudotyped lentivirus transduction in vitro in polarized HAE culture [273] and in vivo in marmoset [125] and ferret [129] airways. Calcium-chelating agents, such as ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA), can also be used as adjuvants to reduce the intracellular concentration of calcium ions, thereby disrupting calcium-dependent formation of tight junction protein complexes [328].…”

“…However, this study revealed that EGTA increased cell counts and the levels of pro-inflammatory cytokines in BALF, while C10 and C12 altered airway responsiveness after methacholine stimulation in vivo in the lungs of mice [330]. LPC has been used most extensively as an adjuvant to VSV-G-pseudotyped lentivirus-mediated inhaled gene therapy to enhance gene transfer with minimal adverse effects reported [125, 129]. …”