Discovery of an antagonist nanobody targeting the oxytocin receptor with tocolytic effect in mice.

Preclinical study involving in silico design, in vitro screening, and in vivo animal experiments

“In this study, we leverage AI-driven protein design to enable nanobody screening and identify several nanobodies with antagonist activity against the oxytocin receptor, a key target for preterm birth therapeutics.”

Synthetic nanobody library, human oxytocin receptor, human uterine smooth muscle cells, and pregnant C57BL/6J mice

“We conducted nanobody screening using a yeast display library of synthetic nanobodies.”

38 to 42 pregnant mice per experimental group in the in vivo preterm birth model

“n = 38, 42, 38, and 39 biologically independent samples for the Control, LPS, Atosiban, and Nb groups, respectively.”

In vivo monitoring occurred from embryonic day 15.5 until delivery or term

“Pregnant mice were administered LPS (30 µg/kg, i.p.) on embryonic day 15.5 (E15.5), a dose optimized to induce PTB in approximately 70% of animals within 24 h.”

Four nanobodies were identified with antagonist activity against the oxytocin receptor, with Nb2G5 showing potency comparable to Atosiban

“Functional screening revealed that four nanobodies exhibit antagonist activities; these candidates (named Nb2B1, Nb2B11, Nb2G5, Nb2H4) were selected for in-depth characterization.”

Nb2G5 demonstrated high selectivity for the oxytocin receptor with no inhibitory potency on the V1a receptor

“Conversely, Nb2G5 demonstrated no inhibitory potency on V1aR, clearly indicating its specificity for OTR.”

In a mouse model, Nb2G5 delayed preterm labor and improved fetal outcomes with efficacy comparable to Atosiban

“Treatment with Nb2G5 via intravenous injection significantly delayed preterm labor in a dose-dependent manner.”

Nb2G5 exhibits reduced affinity for the mouse oxytocin receptor compared to the human receptor, and safety profiles require validation in non-human primates

“However, while these results are promising, the species-selective nature of Nb2G5 means that mouse models have their limits.”