A Clinical Inflammatory Syndrome Attributable to Aerosolized Lipid–DNA Administration in Cystic Fibrosis

Abstract

Immunologic reactivity to lipid-DNA conjugates has traditionally been viewed as less of an issue than with viral vectors. We performed a dose escalation safety trial of aerosolized cystic fibrosis transmembrane conductance regulator (CFTR) cDNA to the lower airways of eight adult cystic fibrosis patients, and monitored expression by RT-PCR. The cDNA was complexed to a cationic lipid amphiphile (GL-67) consisting of a cholesterol anchor linked to a spermine head group. CFTR transgene was detected in three patients at 2-7 days after gene administration. Four of the eight patients developed a pronounced clinical syndrome of fever (maximum of 103.3EF), myalgias, and arthralgia beginning within 6 hr of gene administration. Serum IL-6 but not levels of IL-8, IL-1, TNF-alpha, or IFN-gamma became elevated within 1-3 hr of gene administration. No antibodies to the cationic liposome or plasmid DNA were detected. We found that plasmid DNA by itself elicited minimal proliferation of peripheral blood mononuclear cells taken from study patients, but led to brisk immune cell proliferation when complexed to a cationic lipid. Lipid and DNA were synergistic in causing this response. Cellular proliferation was also seen with eukaryotic DNA, suggesting that at least part of the immunologic response to lipid-DNA conjugates is independent of unmethylated (E. coli-derived) CpG sequences that have previously been associated with innate inflammatory changes in the lung.

Gene Therapy Progress and Prospects: Cystic fibrosis

“…In a dose-escalating safety trial, liposome DNA complexes were aerosolised into the lungs of CF subjects, half of which, had a transient fever, muscle and joint pain shortly after liposome/DNA administration. 2 This was attributed to an immunological response against the liposome/DNA complexes; similar results were previously reported in a lung trial, using the same liposome/DNA formulation. Interestingly, Hyde et al demonstrated for the first time, that liposome/DNA complexes could be successfully readministered to the nose of CF patients.…”

“…36 Although, it has been demonstrated that repeated administration of liposome/DNA complexes to the nose of CF patients is possible, 3 there have been concerns raised regarding the inflammatory components of bacterial DNA. 2 The abundance of unmethylated CpG motifs in the bacterial plasmid DNA may at least in part be responsible for the inflammatory response. Several strategies are currently being explored to decrease these unwanted properties, such as (1) methylation of CpG sequences; (2) reduction of the CpG frequency by eliminating non-essential regions or by site-directed mutagenesis; and (3) the use of specific inhibitors of the CpG signalling pathway, such as chloroquine or quinacrine.…”

Pre-clinical evaluation of three non-viral gene transfer agents for cystic fibrosis after aerosol delivery to the ovine lung

“…Three GTAs that fulfilled the criteria for selection were cationic lipid 67 (GL67A), compacted DNA nanoparticles (NP) and 25 kDa-branched polyethyleneimine (PEI). In vivo gene transfer following aerosol delivery to the lung has been demonstrated previously with GL67A in both rodents and humans [12][13][14] and with PEI in rodents and sheep. 9,10,15,16 Evidence for in vivo gene transfer following administration of NP to the lungs of mice 17,18 and to CF patients has been reported 19 and although not delivered by aerosol in these studies, maintenance of the physical properties and biological function of NP collected post-aerosolisation has been described.…”

Shear stress increases cytotoxicity and reduces transfection efficiency of liposomal gene delivery to CHO-S cells

“…This also suggests that the increased toxicity of lipoplex in presence of shear stress is a result of cellular interaction with lipoplex in the presence of shear stress and likely not due to other longterm cellular changes upon exposure to shear stress. Lipoplexes with low toxicity in vitro, when injected into the blood stream where they are subjected to shear stress, have resulted in significant toxicity to blood cells in the form of transient leukopenia and neutropenia both in animal models and human clinical trials (Freimark et al 1998;Li et al 1999;Ruiz et al 2001;Tousignant et al 2000;Zhang et al 2005), though the cause of this is not clear. We did not observe a similar effect of higher toxicity in presence of shear stress in K562 cells.…”