Biological evaluation of integrin α3β1-targeted 68Ga-labeled HEVNPs in HCT 116 colorectal tumor-bearing mice

Abstract

<p>Integrins are cell surface receptors involved in multiple functions vital for cellular proliferation. Various tumor cells overexpress αβ-integrins, making them ideal biomarkers for diagnostic imaging and tumor-targeted drug delivery. LXY30 is a peptide that can specifically recognize and interact with the integrin α₃β₁, a molecule overexpressed in breast, ovarian and colorectal cancer. Hepatitis E virus nanoparticles (HEVNPs) are virus-like particles that have been investigated as drug delivery agents for the targeted delivery of nucleic acids and small proteins. HEVNPs can be a theranostic platform for monitoring and evaluating tumor-targeted therapies if tagged with a suitable diagnostic marker. Herein, we describe the radiolabeling and biological evaluation of integrin α₃β₁-targeted HEVNPs. HEVNPs were conjugated with DOTA and radiolabeled with gallium-68 (t_1/2 = 67.7 min), a short-lived positron emitter used in positron emission tomography (PET). The synthesized [⁶⁸Ga]Ga-DOTA-HEVNPs were used to evaluate the efficacy of conjugated LXY30 peptide to improve HEVNPs binding and internalization to integrin α₃β₁ expressing human colorectal HCT 116 cells. <em>In vivo</em> tumor accumulation of [⁶⁸Ga]Ga-DOTA-HEVNP-LXY30 was evaluated in HCT 116 colorectal tumor-bearing mice. [⁶⁸Ga]Ga-DOTA-HEVNP-LXY30 and non-targeted [⁶⁸Ga]Ga-DOTA-HEVNP were radiolabeled with radiochemical yields (RCY) of 67.9 ± 3.3% and 73.7 ± 9.8%, respectively. [⁶⁸Ga]Ga-DOTA-HEVNP-LXY30 exhibited significantly higher internalization in HCT 116 cells than the non-targeted [⁶⁸Ga]Ga-DOTA-HEVNPs (21.0 ± 0.7% vs. 10.5 ± 0.3% at 3 h, ****<em>P</em><0.0001). After intravenous administration to mice, accumulation of [⁶⁸Ga]Ga-DOTA-HEVNP-LXY30 to HCT 116 xenograft tumors was at its highest rate of 0.8 ± 0.4%ID/g at 60 min. [⁶⁸Ga]Ga-DOTA-HEVNP-LXY30 accumulated mainly in the liver and spleen (39.8 ± 13.0%%ID/g and 24.6 ± 24.1%ID/g, respectively). Despite the low targeting efficiency <em>in vivo</em>, we demonstrated that [⁶⁸Ga]Ga-DOTA-HEVNP is a promising diagnostic platform for quantitative analysis of HEVNP distribution <em>in vivo</em>. This nanosystem can be utilized in future studies assessing the success of further engineered HEVNP structures with optimized targeting efficiency in vivo.</p>