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Reactions That Translate mRNA into Drug-like Molecules

Periodic Reporting for period 3 - TRIGGDRUG (Reactions That Translate mRNA into Drug-like Molecules)

Reporting period: 2019-01-01 to 2020-06-30

The increased life span achieved in the most societies correlates with an increased probability of an individual to be diagnosed cancer. The aging of the society leads to an increase of cancer prevalence which concomitantly increases the number of cancer deaths. Despite the impressive improvements of cancer treatments, we do require new concepts for personalized therapies with little side effects.
In cancer, gene expression is deregulated due to amplification, mutation and translocation of genes. Next generation RNA sequencing provides us with the opportunity to identify the number and identity of the gene products aberrantly expressed in a patient. We propose a method that could take advantage of the personalized sequence data. The idea is to use the RNA molecules produced in cancer cells as instructors for the chemical synthesis of drug-like molecules that cure the disease. Accordingly, drug-like molecules would only be formed in those cells that express the disease-specific RNA molecules. Such a personalized molecular therapy would eliminate side effects caused by unwanted perturbation of healthy cells.
The idea to use cellular RNA molecules as triggers for drug synthesis requires chemical methods that couple recognition of the “cancer RNA” with a change of chemical reactivity. Reactive molecules must be able to “read” and “translate” the sequence of a RNA molecule into a drug-like output. We will develop mRNA-triggered reactions that i) proceed with turnover in template to cope with low mRNA copy numbers and ii) allow the single-step synthesis of highly active drug-like molecules to address deregulated protein targets inside cancer cells. To achieve this aim, we will advance chemical acyl transfer and alkylidene transfer as well as photocatalytic cleavage reactions. The reactions will form peptides, peptidomimetics or small molecules which will bind and inhibit those proteins that allow the cancer cell to survive. Since the products will able to target both RNA (by virtue of the “read” step) and deregulated proteins (by virtue pf the “translate” step) synergy between the nucleic acid and protein worlds will be harnessed.
In a nutshell, we will develop a chemistry-based tool to hijack disease mRNA and jump-start the cell death program of cancer cells.
The research project comprises four subprojects.
Subproject I. We seek to develop reactions that are triggered by RNA and produce peptides that antagonize protein (Bcl-XL) which allows the cancer cells to grow and escape programmed cell death. So far, we have identified two suitable chemical reactions. We found a reaction that puts the synthesis of a 16 amino acid long peptide under the control of a specific RNA molecule. We have shown that the product of this reaction binds cell death inhibiting protein (Bcl-XL), kills cancer cells and we have also identified the conditions that allow the delivery of the reagents into the cell. The second reaction leads to a phosphopeptide that prevents cell growth. We now have evidence that the chemical reaction can indeed be triggered by specific RNA molecules inside live cells and perturbs a protein-protein interaction required to signal cell growth.
In subproject II, we developed chemical syntheses and a RNA-triggered reaction system leading to dual activity agents. One part of the product molecule was designed to antagonize a road block installed by the cancer cell to prevent the intrinsic pathway of programmed cell death. The other part is meant to restore the blocked extrinsic pathway. We demonstrated that none of the parts has sufficient power to affect difficult to kill cancer cell lines. Rather, the two parts of the molecules are required to effectively kill cancer cells presumably by simultaneously targeting two different cancer Achilles heels.
In subproject III, we chemically modified small-molecules that inhibit cell growth signaling cancer proteins (receptor tyrosine kinases, RTKs). We demonstrated their sufficient bioactivity. We developed the chemical synthesis of reactive conjugates that can bind RNA targets. We performed RNA templated reactions in test tubes and demonstrated that the formation of the RTK inhibitors can be put under the control of RNA.
Subproject 4 deals with a new photocatalytic cleavage reaction that would render a RNA-templated reaction to produce many product molecules per RNA molecule. The approach relies on fragmentation linkers FL which are incorporated into PNA hairpins. We designed/identified synthesis methods and new FLs for the preparation of molecules that recognize RNA and are susceptible to photocatalytic cleavage.
We have established four different acyl transfer reactions which are driven/controlled by synthetic RNA. The reaction systems lead to compounds that contain i) a peptide nucleic acid (PNA) part (to bind and “read” cancer-specific mRNA) and ii) a peptide, phosphopeptide, peptidomimetic or small molecule compound . We have first evidence that the starting materials can be delivered into live cells and that a chemical reaction can indeed be triggered by intracellular RNA. Our results suggest that the products of the RNA-triggered acyl transfer in live cells can interfere with protein-protein interactions required for signaling of cancer cell growth. This confirms the key working hypothesis of the TRIGGDRUG project. We have furthermore demonstrated evidence for a proposed synergy principle i.e. linking the nucleic acid and protein worlds in peptidomimetic enhanced antisense conjugates allows the simultaneous attack on both RNA and protein targets. At the end of the project we will have chemical reaction systems that allow product formation in and selective killing of those cells that express disease-specific mRNA. In a nutshell, the cancer will instruct its own destruction.