Effective gene therapy for haemophilic mice with pathogenic factor IX antibodies

Abstract

Formation of pathogenic antibodies is a major problem in replacement therapies for inherited protein deficiencies. For example, antibodies to coagulation factors (‘inhibitors’) seriously complicate treatment of haemophilia. While immune tolerance induction (ITI) protocols have been developed, inhibitors against factor IX (FIX) are difficult to eradicate due to anaphylactic reactions and nephrotic syndrome and thus substantially elevate risks for morbidity and mortality. However, hepatic gene transfer with an adeno-associated virus (AAV) serotype 8 vector expressing FIX (at levels of ≥4% of normal) rapidly reversed pre-existing high-titre inhibitors in haemophilia B mice, eliminated antibody production by B cells, desensitized from anaphylaxis (even if protein therapy was resumed) and provided long-term correction. High levels of FIX protein suppressed memory B cells and increased Treg induction, indicating direct and indirect mechanisms of suppression of inhibitor formation. Persistent presence of Treg was required to prevent relapse of antibodies. Together, these data suggest that hepatic gene transfer-based ITI provides a safe and effective alternative to eradicate inhibitors. This strategy may be broadly applicable to reversal of antibodies in different genetic diseases.

Lessons learned from lung and liver in-vivo gene therapy: implications for the future

“…in hemophilia B) and where enzyme replacement treatment often induces inhibitory antibodies to the protein. It has been shown that strong hepatocyte-specific transgene expression, whilst circumventing either the transduction of, or expression in, APCs, can also prevent and even abrogate the formation of such inhibitors [77,78]. …”

Enhanced liver gene transfer and evasion of preexisting humoral immunity with exosome-enveloped AAV vectors

“…At low vector doses (1 3 10 9 vg per mouse), 1 animal treated with standard AAV8-hF.IX developed an anti-hF.IX transgene antibody (supplemental Figure 6B), whereas none of the animals treated with exo-AAV8 vectors developed antitransgene antibodies (supplemental Figure 6). Accordingly, as previous studies showed that high expression levels promote better transgene-specific immune tolerance, 27,28 we evaluated the frequency of CD4 1 CD25 1 FoxP3 1 regulatory T cells (Tregs) in animals treated with exo-or standard AAV-hF.IX vectors at a dose of 1 3 10 9 vg per mouse. At this dose, no differences were noted in the spleens, however, a significant enhancement in the frequency of Tregs was noticeable in the lymph nodes of exo-AAV-treated animals.…”

Toward optimal therapy for inhibitors in hemophilia

“…Support for this therapeutic approach has been shown in the murine hemophilia B model in which liver-directed gene transfer using either adenoassociated viral vectors or lentiviral vectors can reverse preexisting anti-fIX high-titer inhibitors via T-regulatory cell induction. 56,57 In a canine hemophilia A model, liver-directed gene therapy led to tolerance in 3 of 4 dogs. 58 In addition, transplantation of hematopoietic stem cells engineered to express fVIII has been shown to induce fVIII-specific tolerance and eradicated fVIII inhibitors in mouse models.…”