Oral GALNS Peptides Suppress Antibody Responses to Therapeutic Protein in MPS IVA Mice
Pretreatment to reduce immune responses to therapeutic protein
In mice with mucopolysaccharidosis type IVA (MPS IVA), oral administration of human GALNS-derived peptides followed by adeno-associated virus serotype 9 (AAV9) gene therapy resulted in no detectable antibodies against the therapeutic protein and improved some bone pathology. The findings do not establish treatment efficacy or long-term durability in humans.
MPS IVA is a genetic disorder caused by deficiency of N-acetylgalactosamine-6-sulfatase (GALNS), which leads to accumulation of undegraded glycosaminoglycans. Their buildup in bone, cartilage, and other tissues causes impaired growth and progressive skeletal abnormalities. Gene therapy is designed to enable cells to produce the needed enzyme continuously, but an immune response to the enzyme produced could reduce treatment efficacy.
The researchers tested oral immune tolerance, an approach intended to induce immune nonresponsiveness to a specific antigen. The goal was to reduce responses to human GALNS rather than broadly suppress immune function.
No detectable antibodies and improvement in some bone pathology after peptide pretreatment
The study used newborn male mice deficient in GALNS, with 3–6 animals per group. Starting immediately after birth, the mice were fed every other day a mixture of three epitopes, or immune-recognition regions, from human GALNS. The peptide mixture was given at 2.5 μg or 5 μg per gram of body weight. At about 4 weeks of age, the mice received an intravenous AAV9 vector carrying the human GALNS gene. Comparison groups included mice fed human GALNS protein or buffer before receiving the same gene therapy.
Plasma anti-human GALNS immunoglobulin G (IgG) was not detected during the observation period in either peptide group. By contrast, antibody levels increased beginning 2 weeks after AAV9 administration in the GALNS-protein pretreatment and buffer pretreatment groups. The high-dose peptide group had the highest plasma enzyme activity, and high enzyme activity was also observed in the liver and heart of the peptide groups. These results show that oral pretreatment with short peptides and with the full-length protein produced different outcomes, even though both were derived from the same GALNS. However, equal mass doses contain different numbers of molecules bearing the relevant epitopes, so this comparison alone cannot establish that the peptides themselves are more effective at inducing immune tolerance.
Increased enzyme activity did not lead to complete recovery in every tissue. Both peptide groups had significantly reduced vacuolation in femoral articular cartilage compared with the untreated disease group. Vacuoles are a pathological finding associated with intracellular accumulation. Some bone-morphology measures returned to normal levels in the low-dose group, but the same result was not consistently observed in the high-dose group. In the heart, vacuolation was corrected at all assessed sites in both peptide groups. There was no significant difference between treatment groups, however, so an advantage from adding peptides to gene therapy cannot be established.
Human application and durability require further study
The findings support the possibility that pretreatment to reduce antibody responses to a therapeutic protein could aid enzyme supply and improve pathology in some tissues. They cannot be extended to claims that the immune response to the AAV vector itself was suppressed or that repeat administration was made possible.
The study was also limited by its small size and use of newborn male mice. The mice used do not fully reproduce the severe skeletal manifestations seen in humans, and individual differences in major histocompatibility complex (MHC) may mean that suitable peptides also differ. The researchers said reproducibility should be assessed in larger animal models and expanded samples. Whether the effect of a single gene therapy treatment can be maintained for longer remains a question for future study.
The methods and results are available in the full text of the paper in the International Journal of Molecular Sciences.
Mucopolysaccharidosis IVA (MPS IVA) is caused by the accumulation of undegraded glycosaminoglycans due to the deficiency of the N-acetylgalactosamine-6-sulfate sulfatase (GALNS) enzyme. MPS IVA manifests as progressive systemic skeletal dysplasia. Gene therapy (GT) is potentially a one-time treatment in which the enzyme is continuously produced, circulated, and delivered to target tissues. However, immune responses to gene products can diminish therapeutic efficacy. We hypothesized that oral delivery of tolerogenic peptides induces immune tolerance to human GALNS (hGALNS) in MPS IVA mice, enhancing therapeutic efficacy. Neonatal mice deficient in mouse GALNS (mGALNS) were treated orally with three T-cell/B-cell epitope peptides or hGALNS protein on alternate days from day 3 after birth to day 20 before intravenous injection with AAV9 vectors encoding human GALNS.
This provides preclinical evidence that MPS IVA gene therapy development should assess immune responses to the enzyme produced, alongside enzyme production levels. Follow-up studies could combine oral GALNS peptide pretreatment with AAV therapy and test whether the antibody-response profile, enzyme activity, and improvements in bone pathology are maintained together. However, undetectable antibodies do not mean that the entire skeleton has recovered. Human application would require peptide selection suited to MHC diversity, determination of the dose and schedule, and assessment of long-term safety. The current results do not justify claims that this approach can replace nonspecific immunosuppressants or guarantee gene therapy durability.