期刊
MOLECULAR PHARMACEUTICS
卷 17, 期 8, 页码 2831-2839出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.molpharmaceut.0c00231
关键词
transferrin receptor antibody; molecular Trojan horse; effectorless; Fc-effector function side-effects; pharmacokinetics; etythropoietin
资金
- National Institute of Health, NIA [R21AG055949, R01AG062840]
- Joseph H. Stahlberg Foundation
Erythropoietin (EPO), a hematopoietic growth factor and a promising therapy for Alzheimer's disease, has low permeability across the blood-brain barrier. The transferrin receptor antibody fused to EPO (TfRMAb-EPO) is a chimeric monoclonal antibody that ferries EPO into the brain via the transvascular route. However, TfRMAbs have Fc-effector function-related adverse effects including reticulocyte suppression. To overcome this, we recently developed an effectorless TERMAb-EPO fusion protein, designated TfRMAb-N292G-EPO, by eliminating the Fc N-linked glycosylation site at position 292 of the antibody heavy chain. The mutant fusion protein showed enhanced plasma clearance and dramatically reduced plasma concentrations compared with the wild-type (WT) nonmutant fusion protein. This increased dearance of the aglycosylated TfRMAb is expected to increase the injection dose of the mutant fusion protein. To provide a basis for future therapeutic uses of this IgG-neurotrophin fusion protein, the current study aimed to characterize the pharmacokinetic profile of this effectorless TfRMAb-N292G-EPO at different doses following different routes of administration in the mouse. Adult C57BL/6J male mice were injected with a single dose (3, 6, 9, or 20 mg/kg; n = 3-6 per dose) of TfRMAb-N292G-EPO through either the subcutaneous (SQ) or intraperitoneal (IP) route. TfRMAb-N292G-EPO plasma concentrations were determined using an enzyme-linked immunosorbent assay. Mice were sacrificed 24 h after injection, and terminal blood was used for a complete blood count. Brain concentrations in the WT- and mutant fusion protein-treated mice were compared. We observed stark differences in the plasma pharmacokinetics of TfRMAb-N292G-EPO between the IP and SQ routes of administration. Dose escalation from 3 to 20 mg/kg increased the plasma C-max only 3.5-fold for the SQroute, compared with a 35-fold increase for the IP route. The plasma C-max was 15.0 +/- 2.0, 21.3 +/- 4.1, 21.3 +/- 6.4, and 52.8 +/- 27.9 ng/mL following SQ injection and 288 +/- 47, 389 +/- 154, 633 +/- 194, and 10,066 +/- 7059 ng/mL following IP injection for 3, 6, 9, and 20 mg/kg doses, respectively. The plasma C-max following the SQ route was therefore 19- to 190-fold lower than that following the IP route. This finding is consistent with a 31-fold higher apparent clearance following the SQ route compared with the IP route at the highest dose administered. The brain concentrations in the mice treated with a 3 mg/kg dose of the mutant fusion protein were lower than those in the nonmutant WT-treated mice. No reticulocyte suppression was observed at the 3 mg/kg SQ dose of TfRMAb-N292G-EPO. However, reticulocyte suppression increased with an increase in dose and area under the plasma concentration-time curve (AUC) for both the IP and SQ routes. Overall, elimination of Pc N-linked glycosylation, to mitigate TfRMAb effector function side effects, has a profound effect on the plasma exposure of TfRMAb-N292GEPO at therapeutic as well as high doses (3-20 mg/kg). This effect is more pronounced following SQ injection. The low plasma concentrations of the mutant fusion protein following a 3 mg/kg dose resulted in negligible brain uptake. The beneficial rescue of reticulocyte reduction by the N292G mutation is a function of AUC and is negated at high doses of the N292G mutant.
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