July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
Biocompatibility and stability of an AAV vector for choroideremia gene therapy following passage through its surgical device
Author Affiliations & Notes
  • Maria In?s Patrício
    NDCN, University of Oxford, Oxford, ENGLAND, United Kingdom
    Oxford Eye Hospital, Oxford University Hospitals NHS Trust, Oxford, United Kingdom
  • Cristina Martinez-Fernandez Dela Camara
    NDCN, University of Oxford, Oxford, ENGLAND, United Kingdom
  • Christopher I Cox
    Nightstar Therapeutics plc, London, United Kingdom
  • Clare Blue
    Nightstar Therapeutics plc, London, United Kingdom
  • Alun R Barnard
    NDCN, University of Oxford, Oxford, ENGLAND, United Kingdom
    Oxford Eye Hospital, Oxford University Hospitals NHS Trust, Oxford, United Kingdom
  • Robert E MacLaren
    NDCN, University of Oxford, Oxford, ENGLAND, United Kingdom
    Oxford Eye Hospital, Oxford University Hospitals NHS Trust, Oxford, United Kingdom
  • Footnotes
    Commercial Relationships   Maria Patrício, Nightstar Therapeutics plc (P), University of Oxford (E); Cristina Martinez-Fernandez Dela Camara, University of Oxford (E); Christopher Cox, Nightstar Therapeutics plc (E); Clare Blue, Nightstar Therapeutics plc (E); Alun Barnard, Nightstar Therapeutics plc (C), University of Oxford (E); Robert MacLaren, Choroideremia Research Foundation (F), Euretina (S), Nightstar Therapeutics plc (I), Nightstar Therapeutics plc (C), Nightstar Therapeutics plc (E), Nightstar Therapeutics plc (P), Nightstar Therapeutics plc (F), Spark Therapeutics Inc (C), University of Oxford (E), University of Oxford (P)
  • Footnotes
    Support  NIHR Oxford Biomedical Research Centre; Nightstar Therapeutics Ltd
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 4541. doi:
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      Maria In?s Patrício, Cristina Martinez-Fernandez Dela Camara, Christopher I Cox, Clare Blue, Alun R Barnard, Robert E MacLaren; Biocompatibility and stability of an AAV vector for choroideremia gene therapy following passage through its surgical device. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4541.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Gene therapy for choroideremia is developing rapidly, as an increasing number of sites recruit patients worldwide. The subretinal delivery of the therapeutic agent, AAV2-REP1, is an established procedure in vitreoretinal surgery. Concerns exist, however, that the contact of the vector solution with the standardised surgical device results in loss of physical titre and/or level of expression and activity. Here we assessed the biocompatibility and stability of AAV2-REP1 following passage through the injection device to mimic the clinical scenario.

Methods : Three separate surgical devices (loading syringe and delivery syringe with tip needle) were screened using two doses of research-grade AAV2-REP1: high (1E+12 DRP/mL) and low (1E+11 DRP/mL). The low dose was prepared using either formulation buffer (with PF-68 as surfactant agent) or balanced salt solution (BSS). Vector preparations were analysed at baseline and over time after loaded into the systems (+4oC for 6 hours, followed by 90 and 180 min at +20oC). Samples were collected from all devices at all time-points and their titre determined by qPCR. The levels of REP1 expression and activity were determined by WB and in vitro prenylation assay, respectively, for some of the time-points.

Results : The mean genomic titre (DRP/mL) analysis was run ensuring good precision between sample replicates. The high-dose samples did not show any significant losses at any of the time-points. The low-dose samples, however, showed significant losses in the genomic titre of samples diluted with BSS for all time-points: up to 75% drop when compared to baseline. These were therefore excluded from protein analysis. Low-dose samples prepared with formulation buffer showed no significant difference to baseline for any of the time-points. Analysis of REP1 expression corroborated the data obtained by qPCR: no significant differences were detected in REP1 protein levels. Similarly, the levels of biotinylated Rab substrate did not vary significantly from baseline.

Conclusions : Neither the genomic titre nor the protein stability of an AAV2-REP1-containing solution was affected following passage through the surgical device when PF-68 was present as a surfactant. We provide evidence that inclusion of this surfactant in the formulation buffer ensures biocompatibility and stability of AAV2-REP1 even at a lower dilution, over a period up to 10 hours.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

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