Irreversible Inhibtion of Cerebellar Gli Transcription Factors by Cobalt (III) Complexes

Hedgehog （Hh）信号通路的异常表达与肿瘤的形成和发育密切相关

Aberrant expression of the Hedgehog (Hh) signaling pathway is heavily implicated in the formation and proliferation of a subgroup of medulloblastoma (MB) tumors. MB is predominantly a pediatric brain cancer, with 70% of occurrences appearing in children under the age of 10. The SHH subgroup, driven by Hh signaling, is widely heterogeneous in genetic cause and histology, making effective treatment challenging and resulting in dismal outcomes. Current treatment strategies involve tumor resection, craniospinal irradiation, and chemotherapy, but suffer from short-term and long-term adverse effects. In efforts to inhibit the Hh pathway, research has targeted the protein Smoothened (Smo), but has been stunted by downstream mutations that lead to cancer recurrence with a much higher lethality. Thus, this project seeks to target Gli proteins, which belong to a family of zinc finger transcription factors (TFs) and are the final effectors of the Hh pathway. The Meade lab has developed a series of Cobalt (III)-Schiff base complexes (Co(III)-sb) coupled to TF consensus sequences that specifically and irreversibly inhibit zinc finger TFs of interest. This proposal seeks to reinvent the method for conjugating Co(III)-sb to the Gli consensus sequence to generate Co(III)-Gli, a highly specific and irreversible inhibitor of Gli proteins. This inhibitor will be conjugated to a Gd(III)-labeled gold nanoparticle (AuNP) delivery platform to evaluate and image the effectiveness of Gli inhibition both in vitro and in vivo. The first objective of this proposal is to redesign the synthetic route for coupling DNA to Co(III)-sb. The equatorial ligand scaffold of Co(III)-sb will be functionalized with an alkyne moiety to allow for clickable conjugation to an azide functionalized Gli consensus sequence, generating Co(III)-Gli. This is anticipated to significantly improve yields and scalability from the current conjugation method. The second and third objectives focus on evaluating the potency of Co(III)-Gli against Gli for the inhibition of tumor growth. Co(III)-Gli will be hybridized to a DNA capped AuNP to generate an optimized dehybridization sequence to release the agent at physiological temperature. Co(III)-Gli AuNPs will be labeled with Gd(III) magnetic resonance imaging (MRI) contrast agents to provide a platform for fate mapping the conjugates both in vitro and in vivo. The ability of Co(III)-Gli to inhibit exogenous Gli will be evaluated in vitro using human embryonic kidney cells and in vivo using a murine model that natively develops SHH subtype MB tumors. Co(III)-Gli is anticipated to inhibit Gli with high specificity, resulting in suppressed tumor growth. This project fully aligns with the mission of the NIH both in its short-term and long-term implications. It will further develop knowledge about inhibition of cerebellar Gli transcription factors in the short-term, and generate a platform for enhancing current treatment options for SHH subtype medulloblastoma long-term. The success of this project would present a unique opportunity for improving the survival rate and resulting quality of life for the most malignant childhood brain cancer.