Background: Due to its overexpression in a variety of tumor types, the chemokine receptor 4 (CXCR4) represents a highly relevant diagnostic and therapeutic target in nuclear oncology. Recently, [(68)Ga]pentixafor has emerged as an excellent imaging agent for positron emission tomography (PET) of CXCR4 expression in vivo. In this study, the corresponding [(68)Ga]-1,4,7-triazacyclononane-triacetic acid (NOTA) analog was preclinically evaluated and compared to the 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) parent compound [(68)Ga]pentixafor.
Methods: NOTA-pentixafor was synthesized by combining solid and solution-phase peptide synthesis. The CXCR4 receptor affinities of [(68)Ga]pentixafor and [(68)Ga]NOTA-pentixafor were determined in competitive binding assays using the leukemic CXCR4-expressing Jurkat T-cell line and [(125)I]FC131 as the radioligand. Internalization and cell efflux assays were performed using CXCR4-transfected Chem-1 cells. Small-animal PET and biodistribution studies were carried out using Daudi-tumor bearing SCID mice.
Results: [(68)Ga]NOTA-pentixafor showed a 1.4-fold improved affinity towards CXCR4 (IC50). However, internalization efficiency into CXCR4(+)-Chem-1 cells was substantially decreased compared to [(68)Ga]pentixafor. Accordingly, small-animal PET imaging and biodistribution studies revealed a 9.5-fold decreased uptake of [(68)Ga]NOTA-pentixafor in Daudi lymphoma xenografts (1.7 ± 0.4 % vs 16.2 ± 3.8 % ID/g at 90 min p.i.) and higher levels of non-specific accumulation, primarily in the excretory organs such as the liver, intestines, and kidneys (2.3 ± 0.9 % vs 2.0 ± 0.3 % ID/g, 1.9 ± 0.8 % vs 0.7 ± 0.2 % ID/g, and 2.7 ± 1.1 % vs 1.7 ± 0.9 % ID/g, respectively).
Conclusions: Despite enhanced CXCR4-affinity in vitro, the [(68)Ga]NOTA-analog of pentixafor showed reduced CXCR4 targeting efficiency in vivo. In combination with enhanced background accumulation, this resulted in significantly inferior PET imaging contrast, and thus, [(68)Ga]NOTA-pentixafor offers no advantages over [(68)Ga]pentixafor.
Keywords: CXCR4; Cancer; GPCR; NOTA; PET; Pentapeptide; Radiopharmaceutical; Tracer; [68Ga]pentixafor.