Protein inhibitors of clostridium difficile toxin b

Inventors

Chen, Zhilei • Simeon, Rudo • Chamoun-Emanuelli, Ana M. • PENG, Zeyu • Feng, Hanping • Yu, Hua • ZHANG, Yongrong

Assignees

University of Maryland Baltimore • Texas A&M University

Abstract

In an embodiment, the present disclosure pertains to a method of treating or preventing C. difficile infections. In some embodiments, the method includes administering an antitoxin to a subject in need thereof. In some embodiments, the antitoxin includes a designed ankyrin repeat protein (DARPin). In an additional embodiments, the present disclosure pertains to a composition including an antitoxin for treating or preventing C. difficile infections. In some embodiments, the anti-toxin includes a DARPin. In some embodiments, the anti-toxin is a monomeric or dimeric DARPin for the neutralization of Clostridium difficile toxin B (TcdB).

Core Innovation

The present disclosure is directed to compositions and methods for the treatment or prevention of Clostridium difficile (C. difficile) infections by administering anti-toxins containing designed ankyrin repeat proteins (DARPins). These anti-toxins are specifically engineered to neutralize C. difficile toxin B (TcdB), and in particular comprise monomeric or dimeric DARPin proteins such as U3 (SEQ ID NO. 13) and 1.4E (SEQ ID NO. 2), which bind to a region on TcdB critical for toxin translocation into the host cytosol.

The main problem addressed is the inadequate efficacy of existing treatments for C. difficile infections, especially the high rate of recurrence even after the use of the FDA-approved monoclonal antibody bezlotoxumab. Current antibiotic therapies are becoming less effective due to the emergence of antibiotic-resistant and hypervirulent C. difficile strains, and the recurrence rate remains significant, driving the need for new, more potent, and cost-effective therapies.

This invention utilizes engineered DARPins, which are non-antibody binding scaffold proteins with high stability, resistance to proteases and denaturants, and low immunogenicity. The approach includes the creation of both monomeric and dimeric DARPins with very potent neutralization capabilities against TcdB, achieved through phage panning and high-throughput functional screening. Dimeric DARPins demonstrated synergistically enhanced toxin-neutralizing potency relative to the monomeric forms. These proteins can be efficiently expressed in E. coli, enabling large-scale, cost-effective production for use in intravenous, oral, or in situ therapeutic formulations.

Claims Coverage

There are three independent claims that establish the main inventive features of this invention concerning compositions and methods using DARPin-based anti-toxins for Clostridium difficile infections.

The inventive features center around the use of monomeric and dimeric designed ankyrin repeat proteins (DARPins), specifically including U3 and 1.4E sequences, which bind to and inhibit TcdB from C. difficile. These features encompass both compositions and methods of administration for treatment or prevention of infection.

Stated Advantages

Documented Applications