title: A pharmacokinetic analysis of molecular cardiac surgery with recirculation mediated delivery of βARKct gene therapy: developing a quantitative definition of the therapeutic window.
abstract: BACKGROUND
  Two major problems for translating gene therapy for heart failure therapy are: safe and efficient delivery and the inability to establish a relationship between vector exposure and in vivo effects. We present a pharmacokinetics (PK) analysis of molecular cardiac surgery with recirculating delivery (MCARD) of scAAV6-βARKct. MCARD's stable cardiac specific delivery profile was exploited to determine vector exposure, half-life, and systemic clearance.
  
  
  RESULTS
  Five naive sheep underwent MCARD with 10(14) genome copies of scAAV6-βARKct. Blood samples were collected over the recirculation interval time of 20 minutes and evaluated with quantitative polymerase chain reaction (qPCR). C(t) curves were generated and expressed on a log scale. The exposure, half-life, and clearance curves were generated for analysis. qPCR and Western blots were used to determine biodistribution. Finally, all in vivo transduction data was plotted against MCARD's PK to determine if a relationship existed. Vector concentrations at each time point were (cardiac and systemic, respectively): 5 minutes: 9.16 ± 0.15 and 3.21 ± 0.38; 10 minutes: 8.81 ± 0.19 and 3.62 ± 0.37; 15 minutes: 8.75 ± 0.12 and 3.69 ± 0.31; and 20 minutes: 8.66 ± 0.22 and 3.95 ± 0.26; P < .00001. The half life of the vector was 2.66 ± 0.24 minutes. PK model data revealed that only 0.61 ± 0.43% of the original dose remained in the blood after delivery, and complete clearance from the system was achieved at 1 week. A PK transfer function revealed a positive correlation between exposure and in vivo transduction. Robust βARKct expression was found in all cardiac regions with none in the liver.
  
  
  CONCLUSIONS
  MCARD may offer a viable method to establish a relationship between vector exposure and in vivo transduction. Using this methodology, it may be possible to address a critical need for establishing an effective therapeutic window.
authors.affiliation: Department of Surgery, Division of Cardiovascular Surgery, University of Pennsylvania Medical Center, Philadelphia, Pennsylvania, USA.
authors.forename: Anthony S
authors.initials: AS
authors.lastname: Fargnoli
authors.forename: Michael G
authors.initials: MG
authors.lastname: Katz
authors.forename: Charles
authors.initials: C
authors.lastname: Yarnall
authors.forename: Marina V
authors.initials: MV
authors.lastname: Sumaroka
authors.forename: Hansell
authors.initials: H
authors.lastname: Stedman
authors.forename: Joseph J
authors.initials: JJ
authors.lastname: Rabinowitz
authors.forename: Walter J
authors.initials: WJ
authors.lastname: Koch
authors.forename: Charles R
authors.initials: CR
authors.lastname: Bridges
mesh_terms: D000818:Animals; D006348:Cardiac Surgical Procedures / Q000379:methods*; D003326:Coronary Circulation / Q000502:physiology*; D018014:Gene Transfer Techniques; D015316:Genetic Therapy / Q000379:methods*; D010455:Peptides / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics*; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000097:blood* / Q000493:pharmacokinetics*; D012756:Sheep; D014018:Tissue Distribution / Q000502:physiology
chemical_list: D010455:Peptides; D011994:Recombinant Proteins; C445259:beta-adrenergic receptor kinase inhibitory peptide