A Therapeutic Anti–VEGF Antibody with Increased Potency Independent of Pharmacokinetic Half-life

Abstract

Bevacizumab [Avastin; anti-vascular endothelial growth factor (VEGF) antibody] is an antiangiogenic IgG approved for treating patients with certain types of colon, breast, and lung cancer. In these indications, bevacizumab is administered every 2 to 3 weeks, prompting us to study ways to reduce the frequency of administration. Increasing affinity to neonatal Fc receptor (FcRn) may extend the pharmacokinetic half-life of an antibody, but the quantitative effect of FcRn affinity on clearance has not been clearly elucidated. To gain further insight into this relationship, we engineered a series of anti-VEGF antibody variants with minimal amino acid substitutions and showed a range of half-life improvements in primates. These results suggest that, if proven clinically safe and effective, a modified version of bevacizumab could potentially provide clinical benefit to patients on long-term anti-VEGF therapy through less-frequent dosing and improved compliance with drug therapy. Moreover, despite having half-life similar to that of wild-type in mice due to the speciesspecific FcRn binding effects, the variant T307Q/N434A exhibited superior in vivo potency in slowing the growth of certain human tumor lines in mouse xenograft models. These results further suggest that FcRn variants may achieve increased potency through unidentified mechanisms in addition to increased systemic exposure. Cancer Res; 70(8); 3269-77. ©2010 AACR.

Impact of SPR biosensor assay configuration on antibody: Neonatal Fc receptor binding data

“…These values are generally in good agreement with those reported for human IgG1 binding to FcRn under similar experimental conditions. 13,23,24,37,38,39,40 In contrast to rhumAb1, the binding interactions between rhumAb2 and FcRn were highly dependent on the assay format used. In format 1, the average K D values of rhumAb2 binding to recombinant rat, cynomolgus monkey, and human FcRn were 1.2, 2.7, and 3.2 uM, respectively.…”

“…Therefore, the choice of assay formats seems to be mainly driven by a combination of practical considerations (e.g., reagent availability, ease of molecule comparison) and the preference of each individual laboratory. Some may choose format 1 13,20,23,39 because it most likely offers monovalent binding affinity, and some may prefer format 2 17,18,24,36 because it better mimics the rhumAb-FcRn interaction orientation in endosomes. 13,55 Others may opt for an indirect format 25,43 or measuring solution affinity based on FcRn active concentration determination using an antibodyimmobilized assay format.…”

“…13,17,23,24,26,35,36,37,38 This variability in K D values has also been seen in studies of FcRn proteins from other species. At least 15-fold differences in K D values were reported for rhumAbs binding to FcRn proteins derived from cynomolgus monkey and rat (104 nM to 2.4 mM for cynomolgus monkey, 17,37,39 and 35 nM to 567 nM for rat, 25,26,40 ). A number of factors may contribute to this broad range of reported binding activities, including the source and quality of the assay reagents (FcRn, rhumAbs), the experimental setup (e.g., assay format, ligand immobilization level, temperature, analyte concentrations), and the data evaluation models used.…”

Influence of improved FcRn binding on the subcutaneous bioavailability of monoclonal antibodies in cynomolgus monkeys

“…[8][9][10][11][12][13][14] This is likely due to the fact that the intravenous route eliminates the potentially complicating role of IgG formulation and subsequent absorption from an extravascular site on the systemic pharmacokinetics. To date, there is little information examining the ability of engineering the Fc-FcRn interaction to influence the subcutaneous bioavailability of mAbs.…”

“…[8][9][10][11][12][13][14] These reports have provided evidence that specific mutations (T250Q/M428L, M428L, M252Y/S254T/T256E, M428L/N434S, N434A) to a humanized IgG1, IgG4 or IgG2 molecule can result in ~2-to ~4-fold longer in vivo elimination phase half-life in either cynomolgus or rhesus monkeys. [8][9][10][11][12][13][14] All…”