In vivo imaging of vesicular monoamine transporter 2 in pancreas using an (18)F epoxide derivative of tetrabenazine

Abstract

Objectives: Development of imaging agents for pancreatic beta cell mass may provide tools for studying insulin-secreting beta cells and their relationship with diabetes mellitus. In this paper, a new imaging agent, [(18)F](+)-2-oxiranyl-3-isobutyl-9-(3-fluoropropoxy)-10-methoxy-2,3,4,6,7,11b-hexahydro-1H-pyrido[2,1-a]isoquinoline [(18)F](+)4, which displays properties targeting vesicular monoamine transporter 2 (VMAT2) binding sites of beta cells in the pancreas, was evaluated as a positron emission tomography (PET) agent for estimating beta cell mass in vivo. The hydrolyzable epoxide group of (+)4 may provide a mechanism for shifting biodistribution from liver to kidney, thus reducing the background signal.

Methods: Both (18)F- and (19)F-labeled (+) and (-) isomers of 4 were synthesized and evaluated. Organ distribution was carried out in normal rats. Uptake of [(18)F](+)4 in pancreas of normal rats was measured and correlated with blocking studies using competing drugs, (+)dihydrotetrabenazine [(+)-DTBZ] or 9-fluoropropyl-(+)dihydro tetrabenazine [FP-(+)-DTBZ, (+)2].

Results: In vitro binding study of VMAT2 using rat brain striatum showed a K(i) value of 0.08 and 0.15 nM for the (+)4 and (+/-)4, respectively. The in vivo biodistribution of [(18)F](+)4 in rats showed the highest uptake in the pancreas (2.68 %ID/g at 60 min postinjection). In vivo competition experiments with cold FP-(+)-DTBZ, (+)2, (3.5 mg/kg, 5 min iv pretreatment) led to a significant reduction of pancreas uptake (85% blockade at 60 min). The inactive isomer [(18)F](-)4 showed significantly lower pancreas uptake (0.22 %ID/g at 30 min postinjection). Animal PET imaging studies of [(18)F](+)4 in normal rats demonstrated an avid pancreatic uptake in rats.

Conclusion: The preliminary results suggest that the epoxide, [(18)F](+)4, is highly selective in binding to VMAT2 and it has an excellent uptake in the pancreas of rats. The liver uptake was significantly reduced through the use of the epoxide group. Therefore, it may be potentially useful for imaging beta cell mass in the pancreas.

Fig. 1

Chemical structures of three ligands for VMAT2 binding: [¹¹C](+)-DTBZ, [¹⁸F](+)2 and [¹⁸F](+)4.

Fig. 2

HPLC chiral column separation profiles of racemic (±)4 and its corresponding active isomer (+)4. The profiles were obtained by using a AD chiral column (Regis Technology Co., http://www.registech.com/InfoPages/ChiralInfo/ChiralApplications.html) and eluted with 9:1 hexane and isopropanol.

Fig. 3

Comparison of pancreas to liver (P/L) and pancreas to kidney (P/K) ratios for [¹⁸F](+)4 and [¹⁸F](+)2 at different time points. The ratios were calculated based on %ID/g of each organ listed in Table 2. At 60 minutes after injection a significantly higher P/L ratio (4:1) for [¹⁸F](+)4 was obtained, while the previously reported [¹⁸F](+)2, showed a lower P/L ratio (2:1). It is important to note that the shifting the in vivo metabolism from liver to kidney for [¹⁸F](+)4, as compared to that of [¹⁸F](+)2, may contribute to this distinctive change.

Fig. 4

Comparison of in vivo PET images of normal male rats (29.6–44.4 MBq; 0.8–1.2 mCi, iv); A: [¹⁸F](+)2 (data acquired at 30–35 min) B: [¹⁸F](+)4, (data acquired at 30–33 min); C: [¹⁸F](+)4 (data acquired at 63–65 min). Both agents showed excellent pancreas uptake. The epoxide derivative, (+)4, showed a lower liver uptake (images B and C) as compared to the image A, suggesting a shift of the liver uptake and retention to the kidneys. The background activity (i.e. liver) is lower when using the epoxide derivative, [¹⁸F](+)4.

Fig. 5

A proposed hydrolysis of epoxide (+)4 to (+)4b in vivo and in vitro is shown in this figure. It is likely that the epoxide ring of (+)4 was opened leading to the production of di-hydroxyl derivative, (+)4b. When “cold” (+)4 was treated with sodium hydroxide, it produced (+)4b. 5A: [¹⁸F](+)4 and [¹⁸F](+)4b were prepared and equal amounts of these two tracers were then mixed together (~1:1) and co-injected along with a cold mixture of (+)4 and (+)4b (~1:1) (HPLC condition: Phenomenex C18, acetonitrile/50 mM ammonium formate buffer, pH 4), 1 mL/min). After an injection of [¹⁸F](+)4 (37 MBq, 1 mCi) a rat was sacrificed and radioactive material was isolated from plasma, liver and urine samples. To the extracted samples, 5 μL of “cold mixture” was added and then injected into HPLC. The samples were eluted with the same solvent and column system (5B: plasma; 5C: liver; 5D: urine).

Scheme 1

Scheme 2

Scheme 3

Scheme 4