During the past decade, a wide series of CCK/gastrin analogues have been produced and tested in vitro and in vivo. Here we compared the biodistribution of 12 CCK2/gastrin receptor targeting peptides in mice with CCK2R-expressing tumours. Ten of these peptides were gastrin analogues, and two peptides were CCK8 derivatives. The CCK8 analogues showed lower tumour uptake accompanied by a low kidney retention. The sulphated peptide sCCK8[Phe2(p-CH2SO3H),HPG3,6] (SA106) also showed affinity for the CCK1 receptor, resulting in specific accumulation in murine pancreas . Specific uptake in the murine pancreas was not observed for the DOTA-CCK8 peptide (G-CCK8).
From the gastrin analogues, 111In-labelled MG0 and 111In-labelled sargastrin showed the highest kidney accumulation. This uptake was blocked by the coinjection of agents such as albumin fragments or gelatin-based plasma expanders . In addition, Béhé et al. showed efficient blocking of the kidney uptake of 111In-MG0 by using poly-Glu chains of various lengths . In particular, oligo-Glu chains of more than five amino acids showed effective reduction in kidney accumulation. These data suggest that the high kidney retention is a result of the presence of five Glu residues. Indeed, reducing the number of Glu residues in MG0 resulted in a strong reduction of kidney uptake (MG11), but led to reduced tumour localization of only one-third of the uptake of MG0. It has been reported that MG11 is unstable in serum , indicating that the penta-Glu sequence may be responsible for in vivo stability and kidney uptake. To diminish kidney retention of the gastrin analogues, four peptides were developed with various N-terminal sequences consisting of D-Glu and/or D-Gln residues. Except for these residues, peptide sequences were similar to those of MG0. Remarkably, replacing the five L-Glu residues of MG0 by five D-Glu, the new analogue PP-F11 showed markedly reduced kidney retention (90% reduced) as compared to that of MG0, while maintaining high tumour uptake. Replacement by six D-Gln residues PP-F10 even further reduced kidney uptake, at the cost of uptake in the tumour. Tumour uptake was further reduced when using only three D-Gln residues.
To further investigate the potential of a shortened gastrin analogue, Mather et al. screened a series of peptides  of which the peptide with two N-terminal His residues was selected for further studies. This peptide, APH070, combined improved stability with moderate tumour uptake, and although kidney uptake was very low, tumour-to-kidney ratios were lower as compared to several other gastrin analogues (Fig. 2).
The divalent MGD5 displayed good tumour uptake and retention, but showed high background levels in all organs. Blood levels at 1 h p.i. were generally tenfold higher than those of the other peptides investigated. These high background levels, most likely caused by the relatively high blood levels, are in disagreement with previous data  where background levels were three- to fivefold lower. This could be due to the fact that the present study was carried out in BALB/c nude mice, while the peptide was previously tested in CD1 (SCID) mice.
Previously, von Guggenberg et al. reported on the development of two 99mTc-labelled cyclic minigastrin derivatives . Based on HYNIC-cyclo-MG1, the DOTA-conjugated form of this peptide was tested. This peptide combined high tumour accumulation with relatively low kidney retention, resulting in high tumour-to-blood ratios and good tumour-to-kidney ratios. Uptake in all other organs, including the stomach, was the lowest of all peptides studied. Whereas the 99mTc compound showed good tumour retention at 1 and 4 h p.i. , the 111In-labelled DOTA cyclo-MG1 showed a more than 40% decrease in tumour retention between 1 and 4 h p.i.. Except for cyclo-MG1 and PP-F10, all other peptides showed good tumour retention of the radioactivity. The methods discussed above aimed to improve stability and affinity of the peptides. As described in another paper (Aloj et al., this issue), the affinity of all these 12 peptides was in the same low nanomolar range.
In summary, the highest tumour uptake was obtained with cyclo-MG1 (9.2 ± 1.0%ID/g), PP-F11 (9.7 ± 1.8%ID/g), MGD5 (8.9 ± 2.3%ID/g), sargastrin (11.8 ± 3.1%ID/g) and MG0 (13.3 ± 4.9%ID/g). Of these compounds, sargastrin and MG0 suffered from high kidney uptake. Tumour retention of the cyclic peptide was lower than that of the other peptides. Tumour retention of the divalent peptide was stable between 1 and 4 h p.i.. In general, tumour-to-blood and tumour-to-kidney ratios of the CCK analogues were lower than those of the gastrin analogues.