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1. Click Conjugation of Boron Dipyrromethene (BODIPY) Fluorophores to EGFR-Targeting Linear and Cyclic Peptides

   https://doi.org/10.3390/molecules26030593  

   Williams, Tyrslai M.; Kaufman, Nichole E.; Zhou, Zehua; Singh, Sitanshu S.; Jois, Seetharama D.; Vicente, Maria da (February 2021, Molecules)

   null (Ed.)

   Through a simple 1,3-cycloaddition reaction, three BODIPY-peptide conjugates that target the extracellular domain of the epidermal growth factor receptor (EGFR) were prepared and their ability for binding to EGFR was investigated. The peptide ligands K(N3)LARLLT and its cyclic analog cyclo(K(N3)larllt, previously shown to have high affinity for binding to the extracellular domain of EGFR, were conjugated to alkynyl-functionalized BODIPY dyes 1 and 2 via a copper-catalyzed click reaction. This reaction produced conjugates 3, 4, and 5 in high yields (70–82%). In vitro studies using human carcinoma HEp2 cells that overexpress EGFR demonstrated high cellular uptake, particularly for the cyclic peptide conjugate 5, and low cytotoxicity in light (~1 J·cm−2) and darkness. Surface plasmon resonance (SPR) results show binding affinity of the three BODIPY-peptide conjugates for EGFR, particularly for 5 bearing the cyclic peptide. Competitive binding studies using three cell lines with different expressions of EGFR show that 5 binds specifically to EGFR-overexpressing colon cancer cells. Among the three conjugates, 5 bearing the cyclic peptide exhibited the highest affinity for binding to the EGFR protein. 

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2. The Synthesis of BODIPY-TKI Conjugates and Investigation of Their Ability to Target the Epidermal Growth Factor Receptor

   https://doi.org/10.3390/targets1010005  

   Dhingra, Simran; Shrestha, Prajesh; Chowdhury, Arpan; Zhou, Zehua; Jois, Seetharama D; Vicente, Maria_da_Graça H (September 2023, Targets)

   A near-IR BODIPY was covalently conjugated via its isothiocyanate groups to one or two Erlotinib molecules, a known tyrosine kinase inhibitor (TKI), via triethylene glycol spacers, to produce two novel BODIPY-monoTKI and BODIPY-diTKI conjugates. The ability of these conjugates to target the intracellular domain of the epidermal growth factor receptor (EGFR) was investigated using molecular modeling, surface plasma resonance (SPR), EGFR kinase binding assay, time-dependent cellular uptake, and fluorescence microscopy. While both the BODIPY-monoTKI and the BODIPY-diTKI conjugates were shown to bind to the EGFR kinase by SPR and accumulated more efficiently within human HEp2 cells that over-express EGFR than BODIPY alone, only the BODIPY-monoTKI exhibited kinase inhibition activity. This is due to the high hydrophobic character and aggregation behavior of the BODIPY-diTKI in aqueous solutions, as shown by fluorescence quenching. Furthermore, the competition of the two Erlotinibs in the diTKI conjugate for the active site of the kinase, as suggested by computational modeling, might lead to a decrease in binding relative to the monoTKI conjugate. Nevertheless, the efficient cellular uptake and intracellular localization of both conjugates with no observed cytotoxicity suggest that both could be used as near-IR fluorescent markers for cells that over-express EGFR. 

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3. Targeting EGFR Overexpression at the Surface of Colorectal Cancer Cells by Exploiting Amidated BODIPY‐Peptide Conjugates

   https://doi.org/10.1111/php.13234  

   Williams, Tyrslai M.; Zhou, Zehua; Singh, Sitanshu S.; Sibrian‐Vazquez, Martha; Jois, Seetharama D.; Henriques Vicente, Maria da Graça (April 2020, Photochemistry and Photobiology)

   Abstract Three BODIPY‐peptide conjugates designed to target the epidermal growth factor receptor (EGFR) at the extracellular domain were synthesized, and their specificity for binding to EGFR was investigated. Peptide sequences containing seven amino acids, GLARLLT (2)and KLARLLT (4), and 13 amino acids, GYHWYGYTPQNVI (3), were conjugated to carboxyl BODIPY dye (1) by amide bond formation in up to 73% yields. The BODIPY‐peptide conjugates and their “parent” peptides were determined to bind to EGFR experimentally using SPR analysis and were further investigated using computational methods (AutoDock). Results of SPR, competitive binding and docking studies propose that conjugate6including the GYHWYGYTPQNVI sequence binds to EGFR more effectively than conjugates5and7, bearing the smaller peptide sequences. Findings in human carcinoma HEp2 cells overexpressing EGFR showed nontoxic behavior in the presence of activated light (1.5 J cm⁻²) and in the absence of light for all BODIPYs. Furthermore, conjugate6showed about five‐fold higher accumulation within HEp2 cells compared with conjugates5and7, localizing preferentially in the cell ER and lysosomes. Our findings suggest that BODIPY‐peptide conjugate6is a promising contrast agent for detection of colorectal cancer and potentially other EGFR‐overexpressing cancers. 

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4. Gatekeeper mutations activate FGF receptor tyrosine kinases by destabilizing the autoinhibited state

   https://doi.org/10.1073/pnas.2213090120  

   Besch, Alida; Marsiglia, William M.; Mohammadi, Moosa; Zhang, Yingkai; Traaseth, Nathaniel J. (February 2023, Proceedings of the National Academy of Sciences)

   Many types of human cancers are being treated with small molecule ATP-competitive inhibitors targeting the kinase domain of receptor tyrosine kinases. Despite initial successful remission, long-term treatment almost inevitably leads to the emergence of drug resistance mutations at the gatekeeper residue hindering the access of the inhibitor to a hydrophobic pocket at the back of the ATP-binding cleft. In addition to reducing drug efficacy, gatekeeper mutations elevate the intrinsic activity of the tyrosine kinase domain leading to more aggressive types of cancer. However, the mechanism of gain-of-function by gatekeeper mutations is poorly understood. Here, we characterized fibroblast growth factor receptor (FGFR) tyrosine kinases harboring two distinct gatekeeper mutations using kinase activity assays, NMR spectroscopy, bioinformatic analyses, and MD simulations. Our data show that gatekeeper mutations destabilize the autoinhibitory conformation of the DFG motif locally and of the kinase globally, suggesting they impart gain-of-function by facilitating the kinase's ability to populate the active state. 

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5. Chirality-enhanced transport and drug delivery of graphene nanocarriers to tumor-like cellular spheroid

   https://doi.org/10.3389/fchem.2023.1207579  

   Jeon, Hyunsu; Zhu, Runyao; Kim, Gaeun; Wang, Yichun (August 2023, Frontiers in Chemistry)

   Chirality, defined as “a mirror image,” is a universal geometry of biological and nonbiological forms of matter. This geometry of molecules determines how they interact during their assembly and transport. With the development of nanotechnology, many nanoparticles with chiral geometry or chiroptical activity have emerged for biomedical research. The mechanisms by which chirality originates and the corresponding synthesis methods have been discussed and developed in the past decade. Inspired by the chiral selectivity in life, a comprehensive and in-depth study of interactions between chiral nanomaterials and biological systems has far-reaching significance in biomedicine. Here, we investigated the effect of the chirality of nanoscale drug carriers, graphene quantum dots (GQDs), on their transport in tumor-like cellular spheroids. Chirality of GQDs (L/D-GQDs) was achieved by the surface modification of GQDs withL/D-cysteines. As anin-vitrotissue model for drug testing, cellular spheroids were derived from a human hepatoma cell line (i.e., HepG2 cells) using the Hanging-drop method. Our results reveal that theL-GQDs had a 1.7-fold higher apparent diffusion coefficient than theD-GQDs, indicating that theL-GQDs can enhance their transport into tumor-like cellular spheroids. Moreover, when loaded with a common chemotherapy drug, Doxorubicin (DOX), via π-π stacking,L-GQDs are more effective as nanocarriers for drug delivery into solid tumor-like tissue, resulting in 25% higher efficacy for cancerous cellular spheroids than free DOX. Overall, our studies indicated that the chirality of nanocarriers is essential for the design of drug delivery vehicles to enhance the transport of drugs in a cancerous tumor. 

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