Creating therapeutic Disulfide-Rich Peptides targeting transmembrane proteins including ion channels, transporters, and GPCRs

Veneno Technologies was established in July 2020 to accelerate the research and development of therapeutic Disulfide-Rich Peptides (DRP) and contribute to advanced and sustainable medicine.

Introduction

Transmembrane proteins, such as ion channels and GPCRs, have long been regarded as one of the most important drug targets.

In particular, ion channels play fundamental roles in cells and tissues and are important drug targets for many neurological, cardiovascular, and metabolic diseases, as well as for cancer and immune regulation.

Following the initial registration of ion channel drugs, known as “classics,” the number of ion channel-targeting drugs increased rapidly in the early 1950s and the early 1980s. However, most of these are “Me-too” drugs around the original chemical structure and incrementally property-adjusted drugs.

This tremendous and long-standing R&D effort across the entire pharmaceutical industry should have resulted in the creation of innovative ion channel medicines. However, these efforts have been focused predominantly on small molecules, and, unfortunately, very few registrations of new drugs have resulted from these efforts.

In contrast, the feasibility of targeting ion channels with biologics, such as peptides, has already been amply demonstrated in nature. We have finally begun to understand that venomous animals have developed and evolved a myriad of highly potent and selective peptides that can both inhibit and activate ion channels.

We will approach the ion channel drug discovery with the next generation modality “DRP” and the cutting-edge technology PERISS™!
(PERISS: Intra Periplasm Secretion and Selection)

Summary of our technologies.

We have developed three core technologies that enable the practical application of DRP molecules:
a) Huge DRP genotype library;
b) Novel DRP high-throughput screening technology, PERISS™;
c) DRP mass production technology.

What is DRP?

Molecules comprising DRPs are significant components of the venom. DRPs have a unique “knot” structure with multiple disulfide bonds in the molecule. Owing to the rigidity of such a framework, DRPs are highly stable to pH, heat, and enzymes, especially in comparison with linear peptides. Similar to venoms, DRPs are bioactive peptides that affect a wide range of neuro-receptors and ion channels, with high potency and selectivity. Due to these attractive properties, DRPs are considered to be a next-generation modality for drug discovery and development.

PERISS™ Technology

PERISS™ is a new screening method based on evolutionary molecular engineering. We have already established the overwhelming size of the DRP library, which consists of plasmid DNAs encoding one billion DRP variants.
The advantage of our library is not only its size. We created this library from a natural DRP used as a template to take advantage of the excellent biological activity of DRPs, which have evolved naturally as a significant component of venom against transmembrane proteins, such as ion channels. Therefore, a much higher probability of finding bioactive molecules can be expected than that of a conventional random-sequence library.

In the PERISS™ cycle, the DRP genetic variants undergo molecular selection by phenotype, and by repeating this cycle, the target DRP candidate can be efficiently obtained within a short time.
Using the nature of plasmid incompatibility, in the PERISS™ method, one E. coli acts as one reaction field, and one DRP variant translated from one plasmid is screened for affinity for the target membrane protein. Therefore, the PERISS™ method enables rapid and easy screening of up to one billion libraries by the HTS method in a single flask containing billions of E. coli.

Although mRNA display and phage display are the most commonly used protein screening methods, these are not suitable for screening DRP molecules.
Our PERISS™ method is a unique technology utilizing the periplasmic space of E. coli that provides an optimal condition for generating correctly structured DRPs, and can simultaneously express full-length target membrane proteins on the inner membrane of E. coli. PERISS™ method is an ideal solution for screening the DRP targeting ion channels.

Our technology is not specialized for ion channels. We have already confirmed that various types of human transmembrane proteins can be expressed in our system, and are now ready to target various transmembrane proteins, including GPCRs.

DRP mass production system

The production of bioactive DRPs requires the correct construction of multiple SS bonds in the molecule and the proper folding of the peptide. However, mass production using standard chemical synthesis is difficult and expensive, and it has become one of the major hurdles for R&D in DRP drug discovery.
Instead, we are developing a new mass production method using E. coli to overcome this critical issue.

The advantages of using E. coli are:

1. Extensive experience of this system in drug manufacturing (e.g., recombinant insulin, human growth hormone, interferon, cytokines, and antibody drugs)
2. Compared with animal cells and yeast, E. coli has a faster growth rate, can be grown in an inexpensive medium, and has excellent productivity.
3. By using the E. coli periplasmic space, peptides with complex structures, such as DRP, can be expressed with the correct structure.

In addition, we have developed a method to secrete the peptide outside the E. coli membrane (culture supernatant) with a major chemical company.

Our business models

As a business of peptide drugs targeting membrane proteins (such as various channels/transporters/GPCRs), we will develop two business models:

1. Drug Discovery Partnering: Providing tailor-made peptides using our screening platform technology;
2. In-house pipeline: Promote R&D as an in-house pipeline, with the aim of licensing to a pharmaceutical company.

Reference

Screening Techniques Using the Periplasmic Expression of Peptide Libraries and Target Molecules, Kimura, T., Journal of Bioanalysis & Biomedicine, Vol. 9(5):263-269 (2017)

High Proteolytic Resistance of Spider-Derived Inhibitor Cystine Knots, Kikuchi, K. Sugiura, M. Kimura, T., International Journal of Peptides, Vol. 2015: Article ID 537508 (2015)

The application of the Escherichia coli giant spheroplast for drug screening with automated planar patch clamp system, Kikuchi, K. Sugiura, M. Nishizawa-Harada, C. Kimura, T. Biotechnolgy Reports. Vol. 7:17-23 (2015)

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