Comparison Study of Two Differently Clicked 18F-Folates-Lipophilicity Plays a Key Role

Abstract

Within the last decade, several folate-based radiopharmaceuticals for Single Photon Emission Computed Tomography (SPECT) and Positron Emission Tomography (PET) have been evaluated; however, there is still a lack of suitable ¹⁸F-folates for clinical PET imaging. Herein, we report the synthesis and evaluation of two novel ¹⁸F-folates employing strain-promoted and copper-catalyzed click chemistry. Furthermore, the influence of both click-methods on lipophilicity and pharmacokinetics of the ¹⁸F-folates was investigated. ¹⁸F-Ala-folate and ¹⁸F-DBCO-folate were both stable in human serum albumin. In vitro studies proved their high affinity to the folate receptor (FR). The lipophilic character of the strain-promoted clicked ¹⁸F-DBCO-folate (logD = 0.6) contributed to a higher non-specific binding in cell internalization studies. In the following in vivo PET imaging studies, FR-positive tumors could not be visualized in a maximum intensity projection images. Compared with ¹⁸F-DBCO-folate, ¹⁸F-Ala-folate (logD = -1.4), synthesized by the copper-catalyzed click reaction, exhibited reduced lipophilicity, and as a result an improved in vivo performance and a clear-cut visualization of FR-positive tumors. In view of high radiochemical yield, radiochemical purity and favorable pharmacokinetics, ¹⁸F-Ala-folate is expected to be a promising candidate for FR-PET imaging.

Keywords: 18F-folates; PET; click chemistry; copper-catalyzed click; folate receptor; folic acid; strain promoted click.

Scheme 1

Synthesis of both reference compounds ¹⁹F-Ala-folate and ¹⁹F-DBCO-folate starting with folate-azide.

Scheme 2

Synthesis of ¹⁸F-Alakyne and subsequent conversion to ¹⁸F-Ala-folate.

Figure 1

Displacement assay: % specific bound [³H]folic acid against ¹⁹F-DBCO-folate and ¹⁹F-Ala-folate.

Figure 2

Internalization of ¹⁸F-DBCO-folate (a) and ¹⁸F-Ala-folate (b) in KB and OC316 cells at 4 °C and 37 °C for 1 h including blocking with native folic acid.

Figure 3

Results of biodistribution of ¹⁸F-DBCO-folate (a) and ¹⁸F-Ala-folate (b) at 60 min p.i.

Figure 4

¹⁸F-DBCO-folate PET images of a KB-tumor bearing mouse (50–60 min p.i.). (a) representative transverse, coronal and sagittal plane with a green crosshair pointing at KB-tumor and (b) Maximum intensity projection. Tu = KB-tumor, Li = liver, Ki = kidney, Int = intestines, Bl = bladder.

Figure 5

¹⁸F-Ala-folate. PET images of a KB-tumor bearing mouse. Static scan over 10 min 50 min p.i. (a) representative transverse, coronal and sagittal plane with a green crosshair pointing at KB-tumor and (b) Maximum intensity projection. Tu = KB-tumor, Gb = gallbladder, Li = liver, Ki = kidney, Int = intestines, Bl = bladder.