Development and evaluation of ^sup 99m^Tc-labeled LHRH peptide as potential tumor imaging agent

Objectives: Luteinizing hormone-releasing hormone (LHRH), is a decapeptide produced by the hypothalamus and released to the pituitary gland where it binds to specific receptors on the gonadotropes that regulate synthesis and secretion of gonadotropic hormones (luteinizing hormone (LH) and follicle-stimulating hormone (FSH)). Specific LHRH receptors are overexpressed in several tumors (breast, prostate, ovarian, etc). The aim of this work was to develop and optimize the radiolabeling procedure of LHRH peptide with 99mTc and explore whether it could target LHRH receptors in-vitro and evaluate its biodistribution in-vivo. Methods: HYNIC-GSG-LHRH was purchased from Siquimia Srl (Uruguay). 99mTc labeling was performed at 50°C in presence of different coligans including Tricine, ethylenediaminediacetic acid (EDDA), Tricine/EDDA and Tricine/nicotinic acid (NA). Radiolabeling conditions (i.e., pH, temperature, and reaction time) were optimized in order to standardize the procedure.Radiochemical purity was determined by HPLC and ITLC. Radiolabeled peptide was characterized with regard to lipophilicity and stability in serum and L-Cysteine up to 24 h. In-vitro studies were performed on human multiple myeloma (MM1S) cells up to 2 h. Biodistribution studies were performed in healthy Balb/c mice at 0.5, 1, 2 and 24 h. Results: HYNIC-GSG-LHRH could be radiolabeled with high specific activities using Tricine, Tricine/NA or Tricine/EDDA as coligands. From the explored combination of the described peptide, coligand and 99mTc, we found that [99mTc]tricine/HYNIC-GSG-LHRH and [99mTc]tricine-NA/HYNIC-GSG-LHRH revealed the highest labeling yields and in-vitro stability, accompanied by low lipophilicity with log P values of -2.59 ± 0.05 and -2.82 ± 0.04. In-vitro binding and competition assays confirmed that after its derivatization and radiolabelling, [99mTc]Tricine/HYNIC-GSG-LHRH retained its specificity of binding LHRH receptors.Biodistribution studies confirmed hydrophilicity of [99mTc]tricine-NA/HYNIC-GSG-LHRH by revealing greater kidney uptake and low blood retention. Conclusion: We were able to target LHRH receptors through [99mTc]tricine/HYNIC-GSG-LHRH. These encouraging results could be used to non-invasively monitor LHRH receptors expression in several tumors to be used as a potential cancer imaging agent as well as to evaluate the response to specific anti-LHRH drugs.