INTRODUCTION TO GENE Therapy/Gene TRANSFER

In gene therapy, or gene transfer, genetic material, DNA, is introduced into the cells of an organism to alter the expression of a gene and its associated protein. The introduced DNA may inactivate an existing gene in the cell or produce a required protein that would otherwise be absent or exist at inadequate levels in the organism. Alternatively, the introduced DNA may regulate the expression of other genes, either increasing or decreasing their expression to produce its effect. A newly expressed protein may be secreted into the bloodstream and affect other cells in the organism, producing a widespread effect.

Gene therapy in humans has been used to cure diseases and there are currently several commercially available therapies on the market. In most cases, this treatment is used in people that are born with genetic defects that result in a “missing” protein, which can be restored by delivering the gene that codes for that protein, in essence reversing the genetic defect. Examples of successes in human medicine include treatments for lipoprotein lipase deficiency (1), hemophilia B (2) Leber's congenital amaurosis (3) and spinal muscular atrophy (4). Currently there are thousands of human gene therapy clinical trials underway for a wide variety of conditions, including cancer (5), HIV (6), heart disease (7) and muscular dystrophy (8).

The most common method of delivering DNA into cells is using a virus that has been engineered for that purpose. Viruses are used because they have evolved to become highly efficient in infecting a wide variety of tissues. Viruses used in gene therapy trials include adenovirus, lentivirus, and adeno-associated virus (AAV) (9). These viral vectors deliver the therapeutic gene but viral genes involved in replication or those that have toxic effects on the cells have been deleted reducing the possibility of safety issues. Testing of these viral vectors always includes testing in an animal model, such as mice, dogs, or pigs where they have been shown to be safe and provide high levels of gene expression.

AAV is the most commonly used viral vector for gene therapy and has been shown to infect a variety of tissues, produce persistent expression and be nonpathogenic (10). Expression of genes delivered using AAV in animal models and human clinical trials have been observed to persist for at least a decade and suggest some alterations may be permanent (11). AAV vectors do not appear to infect germline cells, so these alterations are not passed onto subsequent generations.

GENE THERAPY RESEARCH TO STERILIZE CATS AND DOGS

In the case of animal contraception, gene therapy could be used to introduce genes that could suppress reproduction in a variety of ways, depending on the gene used. It could produce long-term or even permanent effects, eliminating the need for re-treatment.

Such a treatment holds the potential to become an alternative to surgery that is faster, not require a surgeon’s time, is less expensive, and comes without post-surgical complications.

The most promising studies at the moment focus on contraception of female cats using adeno-associated virus (AAV) to deliver relevant genes (14, 15). In June 2023, Nature Communications published research on delivery of a cat-specific version of AMH using a single intramuscular (IM) injection (15). At a high level, AMH inactivates the primordial follicles required for reproduction and blocks ovulation. Delivering this as a gene opens the door for potential lifetime expression of AMH. No adverse effects were observed at the site of the injection and blood chemistry parameters remained normal for all cats, indicating the treatment was safe (14, 15). In the Nature Communications study, cats were also monitored for over 2 years for physiological measures of transgene expression, anti-transgene antibodies, and reproductive hormones, plus mating behavior and pregnancy. The study concludes that the treatment prevents any “breeding induced ovulation, results in complete infertility, and may constitute a safe and durable strategy to control reproduction in the domestic cat.” 

Based on these results, this research is shifting from into those studies needed to for FDA regulatory approval in the United States.

Other research has targeted Gonadotropin-releasing hormone (GnRH), which is required for both gametogenesis and sex steroid production, and which controls fertility in both males and females. The approach was to deliver a gene that codes for an antibody against GnRH. Many studies of GnRH vaccines, which contain the GnRH protein and produce anti-GnRh antibodies after injection, have shown that if serum levels of anti-GnRH antibody can be produced (usually requiring several booster injections), then reproduction is suppressed. In this approach, instead of a protein, the gene that codes for the anti-GnRH antibody is delivered, which then could be expressed for a long period of time after a single injection.

A study in mice demonstrated that a gene encoding a anti-GnRH antibody (SMI41) expressed using a recombinant AAV vector was both safe and effective in producing long-term (1 year) infertility (16). To test this approach in cats, three female cats were given an intramuscular injection of AAV containing the gene encoding this antibody. No injection site reactions or adverse effects were observed in the cats. Relative to the mouse, only slight increases in SMI41 antibody titers were seen and in two of the three cats these levels rapidly returned to baseline (15). It is likely that the antibody was seen as foreign by the cats and inactivated due to the cats mounting an immune response. It is our understanding that the project has not continued.

Further studies and regulatory approval are needed before any gene therapy/transfer approach to fertility suppression is available for commercial use and cats or dogs. However, the research delivering AMH using an AAV vector (15), shows that the potential to deliver a “single shot” lifetime suppression of fertility is there.