Cavity-Enhanced Lab-on-Fiber Technology: Toward Advanced Biosensors and Nano-Opto-Mechanical Active Devices

The integration of nanostructures able to manipulate light at the optical fiber tip is bringing the optical fibers to a renewed dimension since their beginnings. The past decade has seen unprecedented advancements in the lab-on-fiber technology field, pushed by the effective exploitation of nanoscale optical physics and supported by the improvement of nanofabrication techniques. In this context, here we report on a cavity enhanced lab-on-fiber optrode, which dramatically boosts the performances and widens the range of applications of the current lab-on-fiber systems, setting a new fundamental milestone along the roadmap of this technology. The “lab” integrated onto the fiber tip consists of a tunable optical cavity incorporating smart materials as the active layer. Specifically, the swelling/collapsing mechanism of multiresponsive microgels sandwiched in between gold layers induces the interplay between plasmonic resonances and cavity modes, according to optical cavity size. The combination of the optically resonant effects and microgels endows the optrode with the unique ability to work as both a sensor for detecting small molecules and a nano-opto-mechanical actuator triggered by light. In the specific case study here presented, we show that the device is able to detect glucose in solution, with a sensitivity improved by more than 1 order of magnitude compared to other state-of-the-art values. Moreover, we demonstrate that, by combining thermoplasmonics effects triggered by light coupled into the fiber and the microgel thermo-responsivity, it is possible to tune and control the optical cavity features, opening new avenues toward active nano-opto-mechanical actuators directly realized onto the fiber tip

Qualitative and Quantitative Proteomic Profiling of Cripto^−/− Embryonic Stem Cells by Means of Accurate Mass LC−MS Analysis

Cripto is one of the key regulators of embryonic stem cells (ESCs) differentiation into cardiomyocites vs neuronal fate. Cripto−/− murine ESCs have been utilized to investigate the molecular mechanisms underlying early events of mammalian lineage differentiation. 2D/LC−MS/MS and a label-free LC−MS approaches were used to qualitatively and quantitatively profile the cripto−/− ESC proteome, providing an integral view of the alterations induced in stem cell functions by deleting the cripto gene

Solution Structure of Mouse Cripto CFC Domain and Its Inactive Variant Trp107Ala^†

We report here for the first time the solution structures at pH 3 and pH 6 of the synthetic CFC domain of mouse Cripto and of the point mutated variant W107A that is unable to bind to the Alk4 Cripto receptor. NMR data confirm that the CFC domain has a C1−C4, C2−C6, C3−C5 disulfide pattern and show that structures are rather flexible and globally extended, with three noncanonical antiparallel strands. His104 and Trp107 side chains protrude from a protein edge and are strongly exposed to solvent, supporting previous evidence of direct involvement in receptor binding. On the opposite molecule side, several nonpolar residues are gathered, forming a large hydrophobic patch that supposedly acts as interface with the cell membrane or the adjacent EGF-like domain. A second hydrophilic patch surrounding His104 and Trp107 is present only in the wild type variant, suggesting a possible involvement in modulating Alk4 recognition

Discovery of a new class of triazole based inhibitors of acetyl transferase KAT2A

We have recently developed a new synthetic methodology that provided both N-aryl-5-hydroxytriazoles and N-pyridine-4-alkyl triazoles. A selection of these products was carried through virtual screening towards targets that are contemporary and validated for drug discovery and development. This study determined a number of potential structure target dyads of which N-pyridinium-4-carboxylic-5-alkyl triazole displayed the highest score specificity towards KAT2A. Binding affinity tests of abovementioned triazole and related analogs towards KAT2A confirmed the predictions of the in-silico assay. Finally, we have run in vitro inhibition assays of selected triazoles towards KAT2A; the ensemble of binding and inhibition assays delivered pyridyl-triazoles carboxylates as the prototype of a new class of inhibitors of KAT2A. </div

Discovery of a new class of triazole based inhibitors of acetyl transferase KAT2A

We have recently developed a new synthetic methodology that provided both N-aryl-5-hydroxytriazoles and N-pyridine-4-alkyl triazoles. A selection of these products was carried through virtual screening towards targets that are contemporary and validated for drug discovery and development. This study determined a number of potential structure target dyads of which N-pyridinium-4-carboxylic-5-alkyl triazole displayed the highest score specificity towards KAT2A. Binding affinity tests of abovementioned triazole and related analogs towards KAT2A confirmed the predictions of the in-silico assay. Finally, we have run in vitro inhibition assays of selected triazoles towards KAT2A; the ensemble of binding and inhibition assays delivered pyridyl-triazoles carboxylates as the prototype of a new class of inhibitors of KAT2A.</p

Image_9_MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.jpeg

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides’ immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.</p

Image_1_MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.jpeg

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides’ immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.</p

Image_3_MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.jpeg

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides’ immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.</p

Image_8_MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.jpg

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides’ immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.</p

Image_5_MHC-Optimized Peptide Scaffold for Improved Antigen Presentation and Anti-Tumor Response.jpeg

Tumor Associated Antigens (TAAs) may suffer from an immunological tolerance due to expression on normal cells. In order to potentiate their immunogenicity, heteroclitic peptides (htcPep) were designed according to prediction algorithms. In particular, specific modifications were introduced in peptide residues facing to TCR. Moreover, a MHC-optimized scaffold was designed for improved antigen presentation to TCR by H-2Db allele. The efficacy of such htcPep was assessed in C57BL/6 mice injected with syngeneic melanoma B16F10 or lung TC1 tumor cell lines, in combination with metronomic chemotherapy and immune checkpoint inhibitors. The immunogenicity of htcPep was significantly stronger than the corresponding wt peptide and the modification involving both MHC and TCR binding residues scored the strongest. In particular, the H-2Db-specific scaffold significantly potentiated the peptides’ immunogenicity and control of tumor growth was comparable to wt peptide in a therapeutic setting. Overall, we demonstrated that modified TAAs show higher immunogenicity compared to wt peptide. In particular, the MHC-optimized scaffold can present different antigen sequences to TCR, retaining the conformational characteristics of the corresponding wt. Cross-reacting CD8+ T cells are elicited and efficiently kill tumor cells presenting the wild-type antigen. This novel approach can be of high clinical relevance in cancer vaccine development.</p