Aboagye, E. O. | bioRxiv Science

bioRxiv2023

Development and optimisation of cationic lipid nanoparticles for mRNA delivery

Messenger RNA (mRNA) has been proposed as a therapeutic agent for various diseases, including cancer. To ensure effective transfection of cancer cells, mRNA needs to be transported with a delivery system that protects its integrity and functionality. In this regard, cationic lipid nanoparticles composed of dioleoylphosphatidylethanolamine (DOPE) and 3{beta}-[N-(N,N-dimethylaminoethane)-carbamoyl] cholesterol (DC-Chol) have emerged as common vectors to deliver mRNA. In this project, we aim to use luciferase mRNA as a reporter to synthesise mRNA-loaded cationic lipid nanoparticles, and optimise their mRNA encapsulation and transfection efficiency in ovarian cancer cells. The optimisation process included: 1) adjusting the lipid formulation; 2) adjusting the input mRNA concentration before lipid nanoparticle extrusion; and 3) adjusting the extrusion methods. After optimisation, the encapsulation efficiency was optimised to 62%, thus achieving a relatively high transfection luminescence signal (9.4 times compared to baseline). The lipid nanoparticles also demonstrated stable physical characteristics and high biocompatibility (above 75% cell viability after treatment) within 24 hours. Overall, this project evaluated the synthesis of DOPE/DC-Chol cationic lipid nanoparticles, and optimised their mRNA encapsulation and transfection efficiency in ovarian cancer cell lines. The optimised lipid nanoparticles can be utilised as an ideal system for mRNA delivery, which could be further developed as a potential platform for the immunotherapy in ovarian cancer.

bioengineering↗

bioRxiv2020

NOVEL NUCLEAR ROLE OF HDAC6 IN PROGNOSIS AND THERAPEUTIC TARGET FOR COLORECTAL CANCER

Histone deacetylase 6 (HDAC6) inhibition is a potential treatment of a wide range of cancer types via the acetylation of diverse proteins in the cytoplasm. However, the regulation of histone acetylation and the maintenance of higher-order chromatin structure remains unidentified. Here, we investigated the effect of selective inhibition of HDAC6 by histone acetylation, chromatin relaxation assays, co-immunoprecipitation, acetylome peptide array and in vivo RNA microarray. Our data shows that nuclear HDAC6 physically interacts with the Histone 4 lysine 12 residue, and that HDAC6 inhibition increases acetylation specifically at this residue in several cancer types. Inhibition induces major chromatin structure modulation, but has no equivalent effect on knockout HDAC6-/- MEF cells. We identified several novel HDAC6-deacetylated substrates and high expression of HDAC6 in colorectal cancer (CRC) tissue association with reduced levels of H4K12ac and independent of the key CRC driver mutations, but positively associated with EGFR expression. Furthermore, in vivo HDAC6 inhibition induces significant tumor regression in a CRC xenograft mice model with significant changes in the expression of functional nuclear genes. We also demonstrated that a DNA damaging agent in combination with selective HDAC6 inhibition is effective and acts synergistically, inducing chromatin relaxation and increased cell death in CRC cells. CRC tissues (Normal versus tumor; n=58 matched pairs) together with TCGA data analysis of 467 CRC patients showed that high HDAC6 expression is associated with metastasis, overall and disease-free survival, and is an independent risk factor of CRC stage progression. Our findings designate a new role for nuclear HDAC6 both in cancer prognosis and as a new therapeutic target for CRC and other types of cancer. HighlightHistone deacetylases 6 activity; Chromatin relaxation; Histone modifications; Gene array; DOX: doxorubicin; OXA: oxaliplatin; 5-FU: fluorouracil; Ac: acetylation; MNase: Micrococal nuclease.

cancer biology↗