A carregar...
Publicação
Development of traceless strategies for the selective release of BET inhibitors in cancer cells
| Nome: | Descrição: | Tamanho: | Formato: | |
|---|---|---|---|---|
| 31.46 MB | Adobe PDF |
Orientador(es)
Resumo(s)
Epigenome-targeting drugs are an emerging class of therapeutic agents against a variety of human cancers. This results from the increasingly clear understanding of histone post-translational modifications and their role in tumorigenesis and tumor progression. Proteins of the bromodomain and extraterminal (BET) family, in particular, are epigenetic regulators of great interest as biological targets. These family of proteins is comprised of BRD2, BRD3, BRD4 and BRDT, which bind to chromatin through recognition of acetylated lysine residues (KAc) on histone tails, leading to the recruitment and co-activation of master regulatory transcription factors including the oncogene MYC. Preclinical modulation of BET protein function in malignant models through small molecule inhibition has therefore resulted in marked phenotypic changes and promising therapeutic benefits, driving multiple effectors into clinical evaluation. However, early clinical trials have had modest results with only a few, short-lived, responses in both hematologic and solid tumors. Relevant toxicities were observed in many patients, including severe thrombocytopenia, fatigue, gastrointestinal (GI)-side effects, nausea, vomiting, and which significantly limited compliance to treatment.
The targeted delivery of BET inhibitors to cancer cells can, on the other hand, markedly improve their therapeutic index and overall efficacy. Through the conjugation of the therapeutic agent to a tumor-cellspecific ligand via a cleavable linker, it is possible to enhance the accumulation of drug at the tumor site while sparing healthy cells and the associated collateral toxicity. This project describes the engineering of a conditionally stable construct for the efficient, targeted and traceless release of a BETbromodomain protein effector in cancer cells.
Based on published evidence and organic chemistry methodologies, a BET inhibitor was designed to retain high binding efficacy to target proteins and potent antitumor activity, while enabling targeted delivery approaches. The derivatised compound, featuring a tertiary amine suitable to accommodate a self-immolative release mechanism, engaged BET protein targets with great affinity as determined by different biophysical assays. Crystallography studies and molecular modeling of compounds in complex with target proteins provided further mechanistic insights into their mode of binding, revealing unprecedented bond formation and establishing a rationale for the exquisite binding observed.
At the cellular level the tertiary-amine containing BET inhibitor exhibited potent antitumor activity against several human cancer cell lines from different tissues. Particularly in the context of prostate cancer, cellular target engagement strategies have shown successful binding of compounds to BET bromodomain proteins, while flow cytometry analysis displayed potent perturbations of cell cycle progression. Concomitantly, a substantial suppression of Myc protein levels was observed following compound treatment in a dose- and time-dependent manner, further corroborated by transcriptome analysis data showing a significant down-regulation of MYC and MYC-related gene signatures. Additional transcriptional perturbations were found in several growth-promoting gene signatures and known downstream effectors of BET bromodomain protein activity, further supporting the antitumoral efficacy and therapeutic potential of the tertiary-amine containing BET inhibitor.
Finally, the derivatised BET bromodomain effector was assembled in a unique ligand-targeted conjugate for its selective delivery in prostate cancer cells. The tertiary-amine functionality of the drug was used to tether a self-immolating motif, which allows for intracellular release upon an enzymatic trigger, further conjugated to a ligand that specifically targets prostate cancer cells. The data observed confirmed the labile nature of the release mechanism and selective liberation of the tertiary-amine derivative in the presence of a triggering event. Furthermore, in a mice model of prostate cancer it is shown that the unique ligand-linker-BET inhibitor conjugate has a remarkably superior therapeutic activity than that of the BET inhibitor alone, thus suggesting a therapeutic advantage over the nontargeted counterparts.
Descrição
Palavras-chave
Terapêutica cancerígena epigenética inibidores de proteínas BET cancro da próstata entrega seletiva de fármacos fármacos conjugados Epigenetic cancer therapy BET bromodomain inhibitors prostate cancer drug delivery drug conjugates