# 99mTc-Labeled Diarylpyrazoles for Single-Emission Computer Tomography Imaging of Neurotensin Receptor-Positive Tumors: A Comparative Preclinical Study

## TL;DR

This study develops and compares two new radioactive compounds for imaging tumors that overexpress a specific receptor, finding one to be more effective for potential clinical use.

## Contribution

The development of new 99mTc-labeled nonpeptide NTSR1 antagonists for SPECT imaging of NTSR-positive tumors is presented.

## Key findings

- The [99mTc]Tc-HYNIC complex ([99mTc]1) showed higher tumor accumulation and better tumor-to-background ratios in preclinical studies.
- The [99mTc]Tc-tricarbonyl complex ([99mTc]2) exhibited high intestinal accumulation, making it unsuitable for clinical translation.
- Both radioligands demonstrated stability in human serum and plasma with high receptor-specific uptake in HT-29 cells.

## Abstract

Background/Objectives: Neurotensin receptors (NTSRs), members of the G protein-coupled receptor (GPCR) family, have been found to be overexpressed in several types of human cancers, including breast, colon, lung, liver, prostate, and pancreatic cancer. In particular, NTSR1 is overexpressed in at least 75% of pancreatic ductal adenocarcinomas. The aim of the present study was the development and evaluation of new 99mTc-labeled nonpeptide NTSR1-antagonists for SPECT imaging of NTSR-positive tumors. Methods: Multistep syntheses of NTSR1 antagonist derivatives were performed following our previously described procedure. Two different chelating strategies were applied for 99mTc radiolabeling to provide the [99mTc]Tc-HYNIC complex [99mTc]1 and the [99mTc]Tc-tricarbonyl complex [99mTc]2. Receptor binding assays were performed using hNTSR1-expressing CHO cells. Radiochemical yields (RCYs) were determined by radio-HPLC. For [99mTc]1 and [99mTc]2, log D7.4, plasma protein binding, stability in human plasma and serum, and cellular uptake in HT-29 cells were determined. Biodistribution studies and small animal SPECT studies were performed in HT-29 tumor-bearing nude mice. Results: The radiosynthesis of [99mTc]1 (log D7.4 = −0.27) and [99mTc]2 (log D7.4 = 1.00) was successfully performed with RCYs of 94–96% (decay-corrected). Both radioligands were stable in human serum and plasma, showed plasma protein binding of 72% ([99mTc]1) and 82% ([99mTc]2), and exhibited high and specific uptake in HT-29 cells. Biodistribution studies in HT-29 tumor-bearing mice showed a higher tumor accumulation of [99mTc]1 compared to [99mTc]2 (8.8 ± 3.4 %ID/g vs. 2.7 ± 0.2 %ID/g at 2 h p.i.). [99mTc]2 showed exceptionally high intestinal accumulation (49 ± 22 %ID/g at 1 h p.i.) and was therefore considered unfavorable. In the SPECT/CT imaging of HT-29 tumor xenografts, [99mTc]1 showed a higher NTSR1-specific tumor uptake than [99mTc]2 at all time points after tracer injection, with 12 ± 2.8 %ID/g for [99mTc]1 vs. 3.1 ± 1.1 %ID/g for [99mTc]2 at 4 h p.i. and adequate tumor-to-background ratios. Conclusions: In particular, the [99mTc]Tc-HYNIC ligand ([99mTc]1) showed promising preclinical results, being a potential candidate for SPECT imaging and, therefore, appropriate for translation into the clinic.