Delivery and evaluation of recombinant adeno-associated viral vectors in the equine distal extremity for the treatment of laminitis.
Abstract: Our long-term aim is to develop a gene therapy approach for the prevention of laminitis in the contralateral foot of horses with major musculoskeletal injuries and non-weightbearing lameness. Objective: The goal of this study was to develop a practical method to efficiently deliver therapeutic proteins deep within the equine foot. Methods: Randomised in vivo experiment. Methods: We used recombinant adeno-associated viral vectors (rAAVs) to deliver marker genes using regional limb perfusion through the palmar digital artery of the horse. Results: Vector serotypes rAAV2/1, 2/8 and 2/9 all successfully transduced equine foot tissues and displayed similar levels and patterns of transduction. The regional distribution of transduction within the foot decreased with decreasing vector dose. The highest transduction values were seen in the sole and coronary regions and the lowest transduction values were detected in the dorsal hoof-wall region. The use of a surfactant-enriched vector diluent increased regional distribution of the vector and improved the transduction in the hoof-wall region. The hoof-wall region of the foot, which exhibited the lowest levels of transduction using saline as the vector diluent, displayed a dramatic increase in transduction when surfactant was included in the vector diluent (9- to 81-fold increase). In transduced tissues, no significant difference was observed between promoters (chicken β-actin vs. cytomegalovirus) for gene expression. All horses tested for vector-neutralising antibodies were positive for serotype-specific neutralising antibodies to rAAV2/5. Conclusions: The current experiments demonstrate that transgenes can be successfully delivered to the equine distal extremity using rAAV vectors and that serotypes 2/8, 2/9 and 2/1 can successfully transduce tissues of the equine foot. When the vector was diluted with surfactant-containing saline, the level of transduction increased dramatically. The increased level of transduction due to the addition of surfactant also improved the distribution pattern of transduction.
© 2015 EVJ Ltd.
Publication Date: 2016-01-16 PubMed ID: 26663470PubMed Central: PMC7764945DOI: 10.1111/evj.12547Google Scholar: Lookup
The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
- Clinical Trial
- Journal Article
Summary
This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.
The research study is focused on developing a gene therapy approach to prevent laminitis (a painful condition affecting horse’s feet) using gene-delivering viruses (recombinant adeno-associated viral vectors, rAAVs). These vectors were able to successfully introduce marker genes to equine foot tissues and the inclusion of a surfactant to the vector increased the effectiveness and distribution of the gene delivery.
Objective and Methods of Study
- The research aimed to create a method for delivering therapeutic proteins deep within the foot of a horse. This will contribute to the broader goal of preventing laminitis in horses suffering from major musculoskeletal injuries.
- The team utilized recombinant adeno-associated viral vectors (rAAVs) to convey marker genes into the horse’s foot. This was achieved through regional limb perfusion, a method of delivering medication to specific parts of the body, targeting the palmar digital artery of the horse.
Results of the Methods Used
- Three types of vector serotypes (styles of the virus designed to carry the therapy), rAAV2/1, 2/8, and 2/9, were used. All three successfully transduced (transferred) genes into equine foot tissues and had comparable levels and patterns of transduction.
- The researchers found variations in transduction levels within different regions of the horse’s foot. Maximum transduction was noted in the sole and the coronary regions of the foot, whereas minimum transduction was observed in the dorsal hoof-wall region.
- To enhance transduction distribution and effectiveness in the hoof-wall region, a surfactant-enhanced vector diluent was used. This resulted in increased transduction levels, between 9 and 81-fold, in an area that initially showed low levels when saline was used as the vector diluent.
- The study found no significant difference in gene expression between the two promoters used, chicken β-actin vs. cytomegalovirus.
Conclusion of Study
- The researchers concluded that the transgenes can indeed be effectively introduced into a horse’s distal extremity (lower part of the limbs) using rAAV vectors.
- The use of surfactant-enriched vector diluent significantly enhanced the transduction level and distribution within the foot.
Cite This Article
APA
Mason JB, Gurda BL, Van Wettere A, Engiles JB, Wilson JM, Richardson DW.
(2016).
Delivery and evaluation of recombinant adeno-associated viral vectors in the equine distal extremity for the treatment of laminitis.
Equine Vet J, 49(1), 79-86.
https://doi.org/10.1111/evj.12547 Publication
Researcher Affiliations
- Department of Clinical Studies, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Kennett Square, USA.
- Gene Therapy Program, Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, USA.
- Utah Veterinary Diagnostic Laboratory, School of Veterinary Medicine, Utah State University, Logan, USA.
- Department of Clinical Studies, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Kennett Square, USA.
- Gene Therapy Program, Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, USA.
- Department of Clinical Studies, New Bolton Center, School of Veterinary Medicine, University of Pennsylvania, Kennett Square, USA.
MeSH Terms
- Adenoviridae / physiology
- Animals
- Extremities
- Foot Diseases / therapy
- Foot Diseases / veterinary
- Gene Expression Regulation, Enzymologic
- Gene Expression Regulation, Viral
- Genetic Therapy / methods
- Genetic Therapy / veterinary
- Genetic Vectors / metabolism
- Hoof and Claw / pathology
- Horse Diseases / therapy
- Horses
- Inflammation / therapy
- Inflammation / veterinary
- Transgenes
- beta-Galactosidase / genetics
- beta-Galactosidase / metabolism
Grant Funding
- P30 AR069619 / NIAMS NIH HHS
- P30 DK047757 / NIDDK NIH HHS
Conflict of Interest Statement
Authors’ declaration of interests. J.M. Wilson is an advisor to REGENXBIO, Dimension Therapeutics, Solid Gene Therapy, and Alexion, and is a founder of, holds equity in, and has a sponsored research agreement with REGENXBIO and Dimension Therapeutics; in addition, he is a consultant to several biopharmaceutical companies and is an inventor on patents licensed to various biopharmaceutical companies. All other authors declare that there are no conflicts of interest.
References
This article includes 25 references
- Mason JB, Vandenberghe LH, Xiao R, Wilson JM, Richardson DW. Influence of serotype, cell type, tissue composition, and time after inoculation on gene expression in recombinant adeno-associated viral vector-transduced equine joint tissues.. Am J Vet Res 2012 Aug;73(8):1178-85.
- Michelfelder S, Trepel M. Adeno-associated viral vectors and their redirection to cell-type specific receptors. .
- Xiao X, Li J, Samulski RJ. Production of high-titer recombinant adeno-associated virus vectors in the absence of helper adenovirus.. J Virol 1998 Mar;72(3):2224-32.
- Auricchio A, Hildinger M, O'Connor E, Gao GP, Wilson JM. Isolation of highly infectious and pure adeno-associated virus type 2 vectors with a single-step gravity-flow column.. Hum Gene Ther 2001 Jan 1;12(1):71-6.
- Hildinger M, Auricchio A, Gao G, Wang L, Chirmule N, Wilson JM. Hybrid vectors based on adeno-associated virus serotypes 2 and 5 for muscle-directed gene transfer.. J Virol 2001 Jul;75(13):6199-203.
- Gao GP, Alvira MR, Wang L, Calcedo R, Johnston J, Wilson JM. Novel adeno-associated viruses from rhesus monkeys as vectors for human gene therapy.. Proc Natl Acad Sci U S A 2002 Sep 3;99(18):11854-9.
- Mason JB, Vandenberghe LH, Wilson JM, Richardson DW. When selecting an adeno-associated viral vector serotype, cell monolayer transduction efficiency does not accurately predict tissue transduction efficiency in equine synovial tissues. FASEB J 23, 817.7.
- Bell P, Limberis M, Gao G, Wu D, Bove MS, Sanmiguel JC, Wilson JM. An optimized protocol for detection of E. coli beta-galactosidase in lung tissue following gene transfer.. Histochem Cell Biol 2005 Jul;124(1):77-85.
- Roessler BJ, Davidson BL. Genetic modification of synoviocytes in vivo using recombinant adenoviral vectors. Clin. Res 41, A171.
- Bell P, Moscioni AD, McCarter RJ, Wu D, Gao G, Hoang A, Sanmiguel JC, Sun X, Wivel NA, Raper SE, Furth EE, Batshaw ML, Wilson JM. Analysis of tumors arising in male B6C3F1 mice with and without AAV vector delivery to liver.. Mol Ther 2006 Jul;14(1):34-44.
- Calcedo R, Vandenberghe LH, Gao G, Lin J, Wilson JM. Worldwide epidemiology of neutralizing antibodies to adeno-associated viruses.. J Infect Dis 2009 Feb 1;199(3):381-90.
- Mason JB, Gurda BL, Engiles JB, Hankenson KD, Wilson JM, Richardson DW. Multiple recombinant adeno-associated viral vector serotypes display persistent in vivo gene expression in vector-transduced rat stifle joints.. Hum Gene Ther Methods 2013 Jun;24(3):185-94.
- Wang Z, Zhu T, Qiao C, Zhou L, Wang B, Zhang J, Chen C, Li J, Xiao X. Adeno-associated virus serotype 8 efficiently delivers genes to muscle and heart.. Nat Biotechnol 2005 Mar;23(3):321-8.
- Mishra PC, Leach DH. Extrinsic and intrinsic veins of the equine hoof wall.. J Anat 1983 May;136(Pt 3):543-60.
- Bell P, Gao G, Haskins ME, Wang L, Sleeper M, Wang H, Calcedo R, Vandenberghe LH, Chen SJ, Weisse C, Withnall E, Wilson JM. Evaluation of adeno-associated viral vectors for liver-directed gene transfer in dogs.. Hum Gene Ther 2011 Aug;22(8):985-97.
- Wang L, Wang H, Bell P, McMenamin D, Wilson JM. Hepatic gene transfer in neonatal mice by adeno-associated virus serotype 8 vector.. Hum Gene Ther 2012 May;23(5):533-9.
- Raisler BJ, Berns KI, Grant MB, Beliaev D, Hauswirth WW. Adeno-associated virus type-2 expression of pigmented epithelium-derived factor or Kringles 1-3 of angiostatin reduce retinal neovascularization.. Proc Natl Acad Sci U S A 2002 Jun 25;99(13):8909-14.
- Yin L, Greenberg K, Hunter JJ, Dalkara D, Kolstad KD, Masella BD, Wolfe R, Visel M, Stone D, Libby RT, Diloreto D Jr, Schaffer D, Flannery J, Williams DR, Merigan WH. Intravitreal injection of AAV2 transduces macaque inner retina.. Invest Ophthalmol Vis Sci 2011 Apr 25;52(5):2775-83.
- Gray SJ, Foti SB, Schwartz JW, Bachaboina L, Taylor-Blake B, Coleman J, Ehlers MD, Zylka MJ, McCown TJ, Samulski RJ. Optimizing promoters for recombinant adeno-associated virus-mediated gene expression in the peripheral and central nervous system using self-complementary vectors.. Hum Gene Ther 2011 Sep;22(9):1143-53.
- Zhang FL, Jia SQ, Zheng SP, Ding W. Celastrol enhances AAV1-mediated gene expression in mice adipose tissues.. Gene Ther 2011 Feb;18(2):128-34.
- Davidoff AM, Gray JT, Ng CY, Zhang Y, Zhou J, Spence Y, Bakar Y, Nathwani AC. Comparison of the ability of adeno-associated viral vectors pseudotyped with serotype 2, 5, and 8 capsid proteins to mediate efficient transduction of the liver in murine and nonhuman primate models.. Mol Ther 2005 Jun;11(6):875-88.
- Hurlbut GD, Ziegler RJ, Nietupski JB, Foley JW, Woodworth LA, Meyers E, Bercury SD, Pande NN, Souza DW, Bree MP, Lukason MJ, Marshall J, Cheng SH, Scheule RK. Preexisting immunity and low expression in primates highlight translational challenges for liver-directed AAV8-mediated gene therapy.. Mol Ther 2010 Nov;18(11):1983-94.
- Calcedo R, Franco J, Qin Q, Richardson DW, Mason JB, Boyd S, Wilson JM. Preexisting Neutralizing Antibodies to Adeno-Associated Virus Capsids in Large Animals Other Than Monkeys May Confound In Vivo Gene Therapy Studies.. Hum Gene Ther Methods 2015 Jun;26(3):103-5.
- Nathwani AC, Gray JT, McIntosh J, Ng CY, Zhou J, Spence Y, Cochrane M, Gray E, Tuddenham EG, Davidoff AM. Safe and efficient transduction of the liver after peripheral vein infusion of self-complementary AAV vector results in stable therapeutic expression of human FIX in nonhuman primates.. Blood 2007 Feb 15;109(4):1414-21.
- Vandenberghe LH, Wang L, Somanathan S, Zhi Y, Figueredo J, Calcedo R, Sanmiguel J, Desai RA, Chen CS, Johnston J, Grant RL, Gao G, Wilson JM. Heparin binding directs activation of T cells against adeno-associated virus serotype 2 capsid.. Nat Med 2006 Aug;12(8):967-71.
Citations
This article has been cited 1 times.- Sanmiguel J, Gao G, Vandenberghe LH. Quantitative and Digital Droplet-Based AAV Genome Titration.. Methods Mol Biol 2019;1950:51-83.
Use Nutrition Calculator
Check if your horse's diet meets their nutrition requirements with our easy-to-use tool Check your horse's diet with our easy-to-use tool
Talk to a Nutritionist
Discuss your horse's feeding plan with our experts over a free phone consultation Discuss your horse's diet over a phone consultation
Submit Diet Evaluation
Get a customized feeding plan for your horse formulated by our equine nutritionists Get a custom feeding plan formulated by our nutritionists
