A Tetravalent Biparatopic Antibody Causes Strong HER2 Internalization and Inhibits Cellular Proliferation

Abstract

The overexpression of tyrosine kinase HER2 in numerous cancers, connected with fierce signaling and uncontrolled proliferation, makes it a suitable target for immunotherapy. The acquisition of resistance to currently used compounds and the multiplicity of signaling pathways involved prompted research into the discovery of novel binders as well as treatment options with multiple targeting and multispecific agents. Here we constructed an anti-HER2 tetravalent and biparatopic symmetrical IgG-like molecule by combining the Fab of pertuzumab with a HER2-specific Fcab (Fc fragment with antigen binding), which recognizes an epitope overlapping with trastuzumab. In the strongly HER2-positive cell line SK-BR-3, the molecule induced a rapid and efficient reduction in surface HER2 levels. A potent anti-proliferative effect, specific for the HER2-positive cell line, was observed in vitro, following the induction of apoptosis, and this could not be achieved with treatment with the mixture of pertuzumab and the parental Fcab. The inhibitory cytotoxic effect of our antibody as a single agent makes it a promising contribution to the armory of anti-cancer molecules directed against HER2-addicted cells.

Engineering hydrophobicity and manufacturability for optimized biparatopic antibody–drug conjugates targeting c-MET

“… 42 and Lin and coworkers. 43 There is broad evidence that an optimal combination of paratopes can foster inter-molecular target binding and its degradation, 18 , 19 , 44 , 45 but, in addition to target removal, the forced degradation of antibodies or, if applied, ADCs can yield enhanced payload delivery and cytotoxic potency. 22 , 46 While optimal paratope combinations may be identified in large unbiased screens as well, 47 , 48 modeling-supported choice of paratope combination for preferential inter-target binding could save resources and shorten discovery times.…”

“… 15–17 One molecular mechanism for effective target degradation is crosslinking of more than two target molecules via dual binding of suitable combinations of paratopes and their optimal orientation in a biparatopic antibody to favor inter- over intra-molecular binding ( Figure 1a ). 18 , 19 The underlying modes of action are enhanced apparent affinity for cellular binding, increased internalization by cross-linking, degradation 20 and with it signal transduction inhibition, as well as higher potency cytotoxicity. Such enhanced target removal also provokes degradation of the therapeutic mAb or a corresponding antibody–drug conjugate (ADC), leading to enhanced intracellular payload delivery.…”

Biparatopic antibodies: therapeutic applications and prospects

“…For therapy of solid tumors, many bpAbs target the HER2, 11 , 19 , 23 , 36 , 39 , 53 , 54 , 57 , 65–70 which is a receptor tyrosine kinase that is involved in various cellular functions including cell proliferation. High levels of HER2 overexpression are observed in 20–25% of breast cancer patients.…”