Targeting oncogenic KRAS with molecular brush-conjugated antisense oligonucleotides

Abstract

The mutant form of the guanosine triphosphatase (GTPase) KRAS is a key driver in human tumors but remains a challenging therapeutic target, making KRAS MUT cancers a highly unmet clinical need. Here, we report a class of bottlebrush polyethylene glycol (PEG)–conjugated antisense oligonucleotides (ASOs) for potent in vivo KRAS depletion. Owing to their highly branched architecture, these molecular nanoconstructs suppress nearly all side effects associated with DNA–protein interactions and substantially enhance the pharmacological properties of the ASO, such as plasma pharmacokinetics and tumor uptake. Systemic delivery to mice bearing human non–small-cell lung carcinoma xenografts results in a significant reduction in both KRAS levels and tumor growth, and the antitumor performance well exceeds that of current popular ASO paradigms, such as chemically modified oligonucleotides and PEGylation using linear or slightly branched PEG. Importantly, these conjugates relax the requirement on the ASO chemistry, allowing unmodified, natural phosphodiester ASOs to achieve efficacy comparable to that of chemically modified ones. Both the bottlebrush polymer and its ASO conjugates appear to be safe and well tolerated in mice. Together, these data indicate that the molecular brush–ASO conjugate is a promising therapeutic platform for the treatment of KRAS -driven human cancers and warrant further preclinical and clinical development.

A Long‐Circulating Vector for Aptamers Based upon Polyphosphodiester‐Backboned Molecular Brushes

“…Termed pacDNA (polymer‐assisted compaction of DNA), these nanoscopic bioconjugates produce a novel biodistribution profile, elevate blood circulation times, and augment tissue retention (up to 15 weeks post intravenous injection). These improvements result in massively boosted antisense activities in vivo, with 1–2 orders of magnitude reduction in dosage requirement in certain cancer xenograft models [11] . However, our initial pacDNA system may not be ideal for aptamers due to their tendency to undergo endocytosis, which we speculate stems from the hydrophobic polynorbornene (PN) backbone of the bottlebrush polymer [12] .…”

A mechanistic study on the cellular uptake, intracellular trafficking, and antisense gene regulation of bottlebrush polymer-conjugated oligonucleotides

“…27 In addition, the treatment was free of common deleterious effects such as acute toxicity, inflammation, coagulopathy, and anti-PEG immunity. 27 When tested in vitro, the pacDNA exhibits moderate cellular uptake and can regulate mRNA expression with reasonable efficiency if equipped with an appropriate ASO or siRNA. However, to date, the mechanism for the cellular uptake of the pacDNA and how it regulates protein expression at the molecular level is not well understood.…”

A Mechanistic Study on the Cellular Uptake, Intracellular Trafficking, and Antisense Gene Regulation of Bottlebrush Polymer-Conjugated Oligonucleotides

“…13 In addition, the treatment was free of common deleterious effects such as acute toxicity, inflammation, coagulopathy, and anti-PEG immunity. 13…”

“…12 In one example where we compared pacDNA with AZD4785, a clinical ASO targeting wild-type KRAS mRNA, pacDNA achieved more pronounced tumor suppression levels than AZD4785 at a fraction (2.5%) of the dosage and with greatly reduced dosing frequency in non-small cell lung carcinoma (NSCLC) mouse models. 13 In addition, the treatment was free of common deleterious effects such as acute toxicity, inflammation, coagulopathy, and anti-PEG immunity. 13 When tested in vitro, the pacDNA exhibits moderate cellular uptake and can regulate mRNA expression with reasonable efficiency if equipped with an appropriate ASO or siRNA.…”

“…13 In addition, the treatment was free of common deleterious effects such as acute toxicity, inflammation, coagulopathy, and anti-PEG immunity. 13 When tested in vitro, the pacDNA exhibits moderate cellular uptake and can regulate mRNA expression with reasonable efficiency if equipped with an appropriate ASO or siRNA. However, to date, the mechanism for the cellular uptake of the pacDNA and how it regulates protein expression at the molecular level is not well understood.…”