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Peptidomimetics with photocontrolled biological activity

Periodic Reporting for period 2 - PELICO (Peptidomimetics with photocontrolled biological activity)

Reporting period: 2018-01-01 to 2019-12-31

One of the main problems in pharmacology is low success rate of drug candidates in clinical trials. Despite enormous resources spent on the research and discovery of investigational drugs, especially for treatment of cancer, they still have very low success rate. The drug candidates fail mostly because of their excess toxicity.

The project addressed the problem by discovering light-controllable drug candidates, the biological activity of which can be deactivated (“switched off”) or activated (“switched on”) by non-ionisable light irradiation. The light-controllable drugs can be administered in the inactive, non-toxic form, and then activated by light only when and where required for treatment. The activation by light can be done with very high spatiotemporal precision in the lesion site, leaving the rest of the patient body unaffected. Consequently, the light-controllable drugs will have favourable pharmacological characteristics, such as high selectivity of therapeutic action, reduced overall side-toxicity and significantly augmented safety that will increase their success rate in clinical trials. As the result, the healthcare industry will obtain a new treatment modality to address unmet medical needs in Europe and in the whole world.

The main objective of the Project was the design, synthesis, as well as detailed structural and functional studies of cyclic peptide analogues (peptidomimetics), whose biological activity could be effectively controlled by light. The idea behind the design of such compounds relies on the modification of the known biologically active cyclic peptides by using photoisomerizable (photoswitchable) building blocks to get photocontrolled peptidomimetics. The modified peptides can change their structure upon irradiation with light of different wavelengths (Fig. 1), so their properties can be controlled by light. Consortium members have explored and patented the design principles which allow obtaining practically applicable photocontrolled peptidomimetics.
The work was done along the following work packages (WP):
WP1 Pharmacokinetic characterization of the lead peptidomimetics;
WP2 Lead optimisation;
WP3 Discovery of novel photocontrolled peptidomimetics;
WP4 Novel photoswitches in peptidomimetics;
WP5 Management, communication and dissemination.

In the WP1, we evaluated the pharmacokinetic profile as well as toxicity (on mice) of the photocontrolled antimicrobial and anti-cancer peptidomimetics (leads) which were at hands of the participants before the Project started (Figure 2). This prepared the stage for the further lead optimisation in the WP2 by analysis of the obtained data. The main result from the studies of the WP1 is that the less active and toxic peptidomimetic GS-Sw(DPro)FP (closed) accumulated in the tumor tissue after administration and its concentration remained constant for at least 10 h, up to the point when it was cleared from the rest of the body. This fact is favourable for the therapy of cancer. However, the toxicities of both photoforms of the GS Sw(DPro)FP in vivo differ not much, justifying further optimisation studies (WP2).

The work on the WP2 - optimisation of the leads - were done by extensive variations of the lead structure, and subsequent screening of the compounds in vitro using cancer cell lines. Haemolytic activity against human erythrocytes was also measured. More that 40 analogues were tested; major improvement in terms of in vivo toxicity was achieved. We have demonstrated for the first time that photopharmacological therapeutic agents can indeed be safer comparing to their non-photocontrollable analogues. This might be regarded as one of the major accomplishment of the project. It justifies further efforts in the development of photocontrollable drugs and the potentially high costs of their application due to the need for specialized light sources and trained clinical personnel. It is our belief that these efforts will be rewarding in the long term, eventually resulting in the first photocontrollable drugs for human use.

WP3 was devoted to synthesis of novel photocontrolled peptidomimetics designed to interfere with protein-protein interactions (PPI) in living organisms. PPI control many cellular functions and are prospective drug targets. One of the most important PPI which is responsible for cancer is p53-MDM2 interaction. The tumor suppressor p53 protein is responsible for maintaining normal cellular function and replication. We focused in this WP on design and synthesis of photosensitive stapled peptides capable of inhibiting the p53-MDM2 interaction. The design included the use of the diarylethene-based photoswitchable linkers. We prepared the photocontrolled peptidomimetics with nanomolar binding affinity to MDM2. In addition, we explored if such a property of peptides as their ability to penetrate lipid bilayers can be controlled by light. We confirmed that cell-penetrating peptides can be made photoswitchable by modifying them with diarylethene fragments. Such peptides can act as vehicles to deliver therapeutic agents inside the cells. Finally, we also discovered powerful photoregulated inhibitors of an enzyme PIN1, DAE-containing analogues of neurotoxin mambalgin, photocontrollable analogues of cytotoxic depsipeptide didemnin, spiropyrane-containing analogues of a model amphipatic peptide. All these finding will potentially lead to development of novel research tools or therapeutic agents in the future.

WP4 was devoted to synthesis of novel photoswitches. We developed new synthetic approaches to photosensitive amino acids – spiropyrane derivatives. The building blocks suitable for SPPS were obtained.
The fundamental achievement of the project was the development of the design principles for the photocontrolled diarylethene-based peptidomimetics. We found the structural contexts where the diarylethene units can be efficiently used for the photocontrol. We confirmed a hypothesis that the DAE unit can serve as an enthalpy-entropy switch for biologically active peptodomimetic binding to biological targets.
The main practically useful achievement of the Project is the discovery of the optimised diarylethene-based antibiotics - analogues of the natural peptide gramicidin S. These antibiotics are expected to serve as drug candidates in preclinical and clinical development of the first-in-class photopharmacology therapeutics against cancer and bacterial infections. A start-up company LUMOBIOTICS GmbH, the spin-off of the two members of the Project (Enamine and Karlsruhe Institute of Technology) has been founded on July 2nd, 2017. The founders of the company are the inventors, the scientists who also participated in implementing the Project. LUMOBIOTICS GmbH is now at the stage of raising funds for the preclinical and clinical development and acquiring the IP rights. It aims at developing a safer and more efficient targeted therapy against solid tumors and antimicrobial multidrug-resistant infections (MDRI) to solve major problems in healthcare in Europe and worldwide. In the case of success, this Project outcome will have positive impact on one of the global EU/challenges - health, demographic change and wellbeing.
Figure 2. Gramicidin S and initial leads
Figure 1. Principle of the photocontrol of a peptidomimetic