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This 3D reconstruction depicts the viral capsid - the protein shell of a virus. The capsid encloses the genetic material of a virus, along with any THERAPEUTIC GENES added for therapy. Image Credit: Eric Zinn

Resurrecting ancient viruses to improve gene therapy

Researchers at Massachusetts Schepens Eye Research Institute have reconstructed an ancient virus to deliver gene therapy to the liver, muscle, or retina. A technique potentially more potent than currently available therapies.

"We believe our findings will teach us how complex biological structures, such as adeno-associated viruses (AAVs), are built. From this knowledge, we hope to design next-generation viruses to use as vectors for gene therapy."

Luk H. Vandenberghe PhD, Mass. Eye and Ear and Harvard Medical School Department of Ophthalmology, and study author.

A virus can be an ideal delivery system for gene therapy. To survive, an undetected virus infiltrates a host organism, transfers its genetic material into the host's cells where it proliferates and thrives. Taking advantage of viral stealth, researchers want to transfer therapeutic genes into viral capsules to treat damaged tissues instead of viral infections.

To date, AAVs (adeno-associated viruses) used in gene therapy are selected from those naturally circulating throughout our population. However, if a patient has been previously exposed to that particular virus, their body may identify it and mount an immune response, destroying it and its therapy genes before they can take affect. So, scientists want to engineer new and thoroughly benign viruses, unrecognizable to our immune system, in order to increase the possibilities of effective gene therapy.

However, efforts to engineer improved AAVs have been stymied by the intricate structure of viruses. Like pieces of a jigsaw puzzle, every protein in a viral shell must fit perfectly in order for the virus to function. Altering one part of the shell to achieve efficient gene transfer or reduce recognition by immune cells, can destroy the structural integrity of the entire virus.

To overcome this challenge, Vandenberghe and his colleagues at Harvard Medical School, Schepens Eye Research Institute, and Massachusetts Eye and Ear, have turned for guidance to evolutionary history.

Over time, AAV ancestors underwent a series of changes that kept their structural integrity, but slightly altered other viral functions. Researchers were therefore able to recreate an evolutionary timeline of these changes in the lab, and build nine synthetic ancestor viruses. Injected into mice, the most ancient ancestor virus recreated — Anc80 — successfully targeted liver, muscle, and retinal tissue without producing any bad side effects.

As research progresses, virus and host interplay will continue to be monitored to chart and locate any possible improved vector for use in clinical applications. Scientists will next examine the potential for virus Anc80 to deliver genes to treat liver disease and forms of retinal blindness.

This discovery process was published July 30 in Cell Reports

"The vectors developed and characterized in this study demonstrate unique and potent biology that justify their consideration for gene therapy applications."

Luk H. Vandenberghe PhD

Abstract
Highlights
•In silico ancestral sequence reconstruction leads to infectious viral particles
•Anc80, an ancestor of AAV1, 2, 8, and 9, is a potent in vivo gene therapy vector
•The putative evolutionary lineage of AAV is functionally restored
•Ancestral sequence reconstruction elucidates complex structure-function relations

Summary
Adeno-associated virus (AAV) vectors have emerged as a gene-delivery platform with demonstrated safety and efficacy in a handful of clinical trials for monogenic disorders. However, limitations of the current generation vectors often prevent broader application of AAV gene therapy. Efforts to engineer AAV vectors have been hampered by a limited understanding of the structure-function relationship of the complex multimeric icosahedral architecture of the particle. To develop additional reagents pertinent to further our insight into AAVs, we inferred evolutionary intermediates of the viral capsid using ancestral sequence reconstruction. In-silico-derived sequences were synthesized de novo and characterized for biological properties relevant to clinical applications. This effort led to the generation of nine functional putative ancestral AAVs and the identification of Anc80, the predicted ancestor of the widely studied AAV serotypes 1, 2, 8, and 9, as a highly potent in vivo gene therapy vector for targeting liver, muscle, and retina.

This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

About Massachusetts Eye and Ear
Mass. Eye and Ear clinicians and scientists are driven by a mission to find cures for blindness, deafness and diseases of the head and neck. Now united with Schepens Eye Research Institute, Mass. Eye and Ear is the world's largest vision and hearing research center, developing new treatments and cures through discovery and innovation. Mass. Eye and Ear is a Harvard Medical School teaching hospital and trains future medical leaders in ophthalmology and otolaryngology, through residency as well as clinical and research fellowships. Internationally acclaimed since its founding in 1824, Mass. Eye and Ear employs full-time, board-certified physicians who offer high-quality and affordable specialty care that ranges from the routine to the very complex. U.S. News & World Report's "Best Hospitals Survey" has consistently ranked the Mass. Eye and Ear Departments of Otolaryngology and Ophthalmology as top five in the nation. For more information about life-changing care and research, or to learn how you can help, please visit MassEyeAndEar.org.

About Harvard Medical School Department of Ophthalmology
The Harvard Medical School (HMS) Department of Ophthalmology (eye.hms.harvard.edu) is one of the leading and largest academic departments of ophthalmology in the nation. More than 350 full-time faculty and trainees work at nine HMS affiliate institutions, including Massachusetts Eye and Ear/Schepens Eye Research Institute, Massachusetts General Hospital, Brigham and Women's Hospital, Boston Children's Hospital, Beth Israel Deaconess Medical Center, Joslin Diabetes Center/Beetham Eye Institute, Veterans Affairs Boston Healthcare System, VA Maine Healthcare System, and Cambridge Health Alliance. Formally established in 1871, the department has been built upon a strong and rich foundation in medical education, research, and clinical care. Through the years, faculty and alumni have profoundly influenced ophthalmic science, medicine, and literature--helping to transform the field of ophthalmology from a branch of surgery into an independent medical specialty at the forefront of science.

This work was supported by the Harvard Medical School Department of Ophthalmology, Mass Eye and Ear, the Grousbeck Family Foundation, the Candyce Henwood Fund, the NIH Common Fund, Research to Prevent Blindness, and Foundation Fighting Blindness. Luk H. Vandenberghe (LHV) and Eric Zinn are inventors on a patent describing some of the methods and reagents described here. LHV is inventor on several patents on gene therapy technologies. LHV is co-founder, shareholder, member of the Scientific Advisory Board, and consultant of GenSight Biologics, an ophthalmology gene therapy company.

Cell Reports, Zinn et al. "In Silico Reconstruction of the Viral Evolutionary Lineage Yields a Potent Gene Therapy Vector"

Cell Reports, published by Cell Press, is a weekly open-access journal that publishes high-quality papers across the entire life sciences spectrum. The journal features reports, articles, and resources that provide new biological insights, are thought-provoking, and/or are examples of cutting-edge research. For more information, please visit http://www.cell.com/cell-reports. To receive media alerts for Cell Reports or other Cell Press journals, contactpress@cell.com.

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