842. Repeated Administration of Sendai Virus into Lung

Abstract

Recombinant Sendai virus (SeV) is currently the most efficient vector for airway gene transfer. We have previously shown that after a second administration of transmission incompetent F protein-deficient DF/SeV to the lung gene expression was reduced by 60% when compared to a single administration, but was still significantly higher than in untreated mice. Here, we assessed, if this residual level is maintained after further virus administrations. Mice received three administrations one week apart of DF/SeVMtsHNtsPLmut, an F-deficient virus with mutations in the M, HN, P and L protein, that reduce cytotoxicity in vitro (week 1: SeV-GFP, week 2: SeV-GFP, week 3: SeV-lux). Because the immune response, might be depend on virus titre, individual cohorts of animals were treated with either 106, 107 or 108 pfu/mouse (n=20/dose). Luciferase (lux) expression was measured two days after SeV-lux administration. Importantly, repeat administration of DF/SeVMtsHNtsPLmut was well tolerated even at the highest titre. However, Lux expression after three administrations of SeV was significantly reduced compared to single administration, regardless of the virus titre used. A similar study in sheep has been instigated and will serve to indicate whether expression is similarly attenuated in a host in which wild type SeV infection is of no apparent significance. As a prelude to repeat administration the duration of expression after single administration was eastablished. We, therefore, aerosolised sheep with DF/SeV-lacZ (1010 pfu/sheep, n=3) using a Trudell AeroProbe catheter and compared bgal expression in bronchial biopsies (Bx, n=4/sheep from different lobes) and bronchial brushings (Br, n=4/sheep from different lobes) pre-and two days post-transfection. Despite the comparatively small amount of tissue sample bgal expression was significantly (p<0.05) increased post transfection (Bx pre: 2.1|[plusmn]|0.8, post: 253|[plusmn]|9.8, Br pre: 2.6|[plusmn]|1.2, post: 102.5|[plusmn]|72.6 LU/mg protein). We also assessed shedding of infective virus particles in oral and nasal swabs, broncho-alveolar lavage fluid (BALF) and lung tissue and importantly, did not detect any virus shedding. A second cohort of sheep was nebulised in a silimalar manner (1010 pfu/sheep, n=3) and Br, Bx, lavage fluid were collected and blood 2, 7, 14, 21 and 28 days after transfection to determine the dosing interval for repeat administration. At present only the safety data are available. DF/SeV transfection was well tolerated. There was no SeV-related increase in body temperature, or total and differential blood cell counts. However, serum haptoglobin, an inflammatory marker, increased 7 days after transfection, but had returned to baseline values by day 14 (pre: 0, day 7: 2.8|[plusmn]|0.2, day 14: 0.5|[plusmn]|0.3). Further histological and BALF analysis and bgal quantification to determine the time-course of SeV transfection in sheep is currently underway. These studies will inform repeat dosing intervals in the sheep.