Rational ab initio Design of a Humanized Nanobody against KRAS using the PIA Method

Abstract The KRAS oncoprotein remains a therapeutic challenge due to the limitations of current covalent inhibitors. This work presents the rational design of PIAKRASv2- Nb, a 100 Molecular dynamics simulations, extended up to 10 ns, confirm the persistence of the binding pose and reveal a three-phase mechanism: rapid anchoring, interface maturation, and convergence to a stable equilibrium state with ~30 residue-residue contacts. These dynamic metrics are consistent with those observed in successful therapeutic nanobodies like Caplacizumab or VHH72, consolidating the profile of PIA-KRASv2-Nb (seed 72) as a pan-mutant therapeutic candidate against KRAS. This study demonstrates that the PIA method can generate therapeutically optimal nanobodies ab initio, combining high affinity, intrinsic humanization, and conformational reproducibility, with direct implications for tackling targets traditionally considered "undruggable."