Abstract 2123: Validation of novel histone deacetylases (HDAC) specific nanoparticles using HDACs overexpressed human embryonic kidney cells
Authors: Lohana Sahil , Ramakrishnan Yogaraj , Balraj Premanand , Khan Md Rakib Hasan , Sanku Mallik , Sathish Venkatachalem , Quadir Mohiuddin 
Published: March 2024

Introduction: Histone deacetylase (HDAC) is a major enzyme system that is upregulated in numerous disease conditions including cancer. Harnessing the upregulation of HDAC can be a viable tool to design targeted therapy for these diseases. Nanoparticle technology has emerged as a powerful workhorse to suppress the survival of diseased cells selectively. As such, we aim to design a nanoscale, drug delivery platform that responds to HDAC enzymes to initiate drug release. In this poster, we show the pharmacological evidence that HDAC-sensitive nanoparticles are interacting with HDAC enzymes in vitro in time and concentration dependent pattern.

Methods: We synthesized HDAC-responsive nanoparticles via developing a block copolymer composed of a poly (ethylene glycol) block and a poly (L-acetylated lysine) block. Abbreviated as PEG-acPLK, these copolymers self-assemble as nanoparticles with negatively charged surface. Using in vitro culture of HEK 293 cells, we showed that the formed nanoparticles interact and interfere with HDAC enzymes. Cells were grown to 80-90% confluency in DMEM media w/10% FBS prior to experiments. HEK-293 cells were overexpressed with HDAC 7 & HDAC 8 using Lentiviral vectors. Western blot was done to prove the successful overexpression of the HEK cells. Cytotoxicity assay was done by MTT to analyze the efficacy of HDAC specific nanoparticle on control and overexpressed HEK-293 cells.

Results: Our work showed that, PEG-acPLK systems form nanoparticles with a hydrodynamic diameter (RH) of 120-150 nm. The particles were negatively charged on their surface, as such did not produce any non-specific toxicity to cells, such as damage of cell membranes. HEK-293 cells were infected with lentivirus vectors for HDAC 7 and HDAC 8. The medium was changed 8 h after infection. Forty-eight hours after infection, the positive cells were selected with 5 µg/mL puromycin. The lentivirus stable transformants of HEK-293 cells were collected and significant overexpression were confirmed by western blot. For the cytotoxicity study, control and overexpressed cells were treated with HDAC-nanoparticle and PEG-acPLKC (5 µg/mL to 70 µg/mL) for 24hrs. Both the HDAC-nanoparticle and PEG-acPLC (free form) showed more than 60% of toxicity at the concentration of 20 µg/mL. Whereas the HDAC 7 and 8 overexpressed HEK-293 cells did not showed any effects with nanoparticle and Ae-peg-PLKC, suggesting compensating effect due to highly available HDAC system in place and confirm the effectiveness of the HDAC nanoparticle.

Summary and Conclusion: Overall, these results showed that HDAC specific nanoparticle exhibited effects in HEK-293 HDACs overexpressed cells. Further molecular mechanistic studies will help to delineate how the HDAC-responsive particles can be used for drug delivery.

Citation Format: Sahil Lohana, Yogaraj Ramakrishnan, Premanand Balraj, Md Rakib Hasan Khan, Mallik Sanku, Venkatachalem Sathish, Mohiuddin Quadir. Validation of novel histone deacetylases (HDAC) specific nanoparticles using HDACs overexpressed human embryonic kidney cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2123.