Periodic Reporting for period 1 - MGLycan (Targeting hMGL-GalNAc interactions to reverse immune suppression in cancer)
Reporting period: 2019-06-01 to 2021-05-31
Cancer cells communicate with immune cells to persuade them to preclude an immune response. More in depth, the malignant cells mimic healthy cells by displaying a kind of molecules known as glycans. Besides, immune cells such as Dendritic Cells (DC) present a surface populated by lectins, lectins are proteins that interact with glycans. One of these lectins is MGL that is over expressed in immature Dendritic Cells (iDC). iDC are related with the regulation of the immune response. Thus, MGL is an appropriate target to block the immune response. Cancer cells employ their glycans to interact with MGL present in DC aborting the immune reaction and, facilitating the cancer evasion and progression. This key communication happens more frequently with iDCs due to the MGL up regulation. Therefore, it is needed a strategy to foster the maturation of DCs that would help to our natural defenses to trigger an effective immune response.
In this line, we propose:
A) Glycomimetics library able to bind tightly to the MGL to block cancer-DCs glycan mediated communication.
B) Build up a nanoparticles to take advantage of the multivalency to strength the binding.
Conclusions.
New synthetic approach to accelerate the glycomimetic obtaining has been developed.
New glycomimetic library has been synthesized.
In this line, we propose:
A) Glycomimetics library able to bind tightly to the MGL to block cancer-DCs glycan mediated communication.
B) Build up a nanoparticles to take advantage of the multivalency to strength the binding.
Conclusions.
New synthetic approach to accelerate the glycomimetic obtaining has been developed.
New glycomimetic library has been synthesized.
In this project, a chemoenzymatic synthesis of glycomimetics has been performed. The first approach focused on G2 derivatives these compounds featured azide-containing glycans (GalNAz) to perform click reaction in order to expand the library. The first approach did not yield any potential candidate, therefore we decided to synthesize asymmetrical compounds and also linear compounds. In addition, we included azidofucose moiety to perform the click reaction. Besides, the preparation of the glycan comprised the linker installation, to this end, we developed a new reaction that make the most of the amino group that is hanging on the anomeric carbon. After obtaining the glycan derivatives, they were printed onto the microarray slides, and afterwards the click reaction was performed to obtain the glycomimetics library that was tested against two different MGL lectins, the MGL-extracellular domain (MGL-ECD) and the carbohydrate recognition domain of the MGL (MGL-CRD). Both MGLs were previously tagged with a fluorescent tag. Preliminary results have been disseminated in departmental meetings and also in the European Chemical Biology Symposium, ECBS (May 2021, virtual conference).
In parallel, two computational works were carried out in collaboration with different groups from several Dutch Universities. In one of the projects, molecular dynamics (MD) simulations were applied to a Glycan-Lectin system to evaluate the binding of different glycans. In the second project, the MD simulations assisted in the identification of the molecular components required in the auxin analogs for their binding to TIR1.
In parallel, two computational works were carried out in collaboration with different groups from several Dutch Universities. In one of the projects, molecular dynamics (MD) simulations were applied to a Glycan-Lectin system to evaluate the binding of different glycans. In the second project, the MD simulations assisted in the identification of the molecular components required in the auxin analogs for their binding to TIR1.
Glycan libraries are mainly populated by natural molecules obtained by natural products extraction or chemoenzymatic synthesis. We have developed a methodology that incorporates non-natural carbohydrate moieties both in biantennary and linear glycans. These new derivatives display an azido functional group that open a via for further modifications. In addition, controlling the enzymatic extension reaction conditions we obtained asymmetrical biantennary glycomimetics.
Besides, the modification of N-glycans after natural product extraction is constrained to keep the asparagine, in order to use the amino group in the peptide backbone for further chemical modifications. However, both the conformation of the peptide bond and the length of the amino acid could have an effect on binding studies with target proteins. Another strategy relays in cleaving the peptide bond between the peptide and the glycan and perform a long chemical reaction to re-install the amino group in the anomeric position. In this regard, we have optimize the enzymatic peptide cleavage by controlling the reaction conditions without touching the native amino group on the anomeric position and we have installed a linker straightaway after peptide removal. This optimized methodology facilitates and reduce the time required in the modifications of natural products extracted glycans.
Besides, the modification of N-glycans after natural product extraction is constrained to keep the asparagine, in order to use the amino group in the peptide backbone for further chemical modifications. However, both the conformation of the peptide bond and the length of the amino acid could have an effect on binding studies with target proteins. Another strategy relays in cleaving the peptide bond between the peptide and the glycan and perform a long chemical reaction to re-install the amino group in the anomeric position. In this regard, we have optimize the enzymatic peptide cleavage by controlling the reaction conditions without touching the native amino group on the anomeric position and we have installed a linker straightaway after peptide removal. This optimized methodology facilitates and reduce the time required in the modifications of natural products extracted glycans.