The effects of partial retro-inverso modifications of selected peptide bonds of the N-terminal tetrapeptide of gastrin have been studied. In some of the synthesized compounds, the phenylalanyl residue has been replaced by the (R,S)-2-benzylmalonyl, 3-phenylpropionyl, benzylcarbamoyl, or benzyloxycarbonyl moieties. All pseudopeptides showed affinity for the gastrin receptor, in vitro, with potencies varying from IC50 = 10(-7) to IC50 = 10(-4) M. These compounds exhibited little or no activity on acid secretion in the anesthetized rat but were able to antagonize the action of gastrin. Among the most potent were Boc-Trp-Leu-gAsp-CO-CH2CH2C6H5 (20) (ED50 = 0.15 microM/kg), Boc-Trp-Leu-gAsp-m(R,S)Phe-NH2 (3) (ED50 = 0.15 microM/kg), and Boc-Trp-gLeu-D-Asp-m(R,S)Phe-NH2 (7) (ED50 = 0.3 microM/kg).

ACE (angiotensin-converting enzyme; peptidyl dipeptidase A; EC 3.4.15.1), cleaves C-terminal dipeptides from active peptides containing a free C-terminus. We investigated the hydrolysis of cholecystokinin-8 [CCK-8; Asp-Tyr(SO3H)-Met-Gly-Trp-Met-Asp-Phe-NH2] and of various gastrin analogues by purified rabbit lung ACE. Although these peptides are amidated at their C-terminal end, they were metabolized by ACE to several peptide fragments. These fragments were analysed by h.p.l.c., isolated and identified by comparison with synthetic fragments, and by amino acid analysis. The initial and major site of hydrolysis was the penultimate peptide bond, which generated a major product, the C-terminal amidated dipeptide Asp-Phe-NH2. As a secondary cleavage, ACE subsequently released di- or tri-peptides from the C-terminal end of the remaining N-terminal fragments. The cleavage of CCK-8 and gastrin analogues was inhibited by ACE inhibitors (Captopril and EDTA), but not by other enzyme inhibitors (phosphoramidon, thiorphan, bestatin etc.). Hydrolysis of [Leu15]gastrin-(14-17)-peptide [Boc (t-butoxycarbonyl)-Trp-Leu-Asp-Phe-NH2] in the presence of ACE was found to be dependent on the chloride-ion concentration. Km values for the hydrolysis of CCK-8, [Leu15]gastrin-(11-17)-peptide and Boc-[Leu15]gastrin-(14-17)-peptide at an NaCl concentration of 300 mM were respectively 115, 420 and 3280 microM, and the catalytic constants were about 33, 115 and 885 min-1. The kcat/Km for the reactions at 37 degrees C was approx. 0.28 microM-1.min-1, which is approx. 35 times less than that reported for the cleavage of angiotensin I. These results suggest that ACE might be involved in the metabolism in vivo of CCK and gastrin short fragments.

estimated, but it appears that this event may be substantially more frequent with a cord blood source of stem cells. There are several potential explanations for leukemogenesis via this unusual route. [1][2][3][4] Perhaps, the most plausible was predicted for cord blood before the recent series of case reports. 5,6 Systematic screening of a large series of unselected cord blood samples revealed that B1% harboured putative pre-leukaemic clones with TEL-AML1 fusion or B0.2% with AML-ETO. 5 The frequency of such cells in positive cord bloods was of the order of 10 À3 -10 À4 , indicative of a clone size of B10 6 cells in the new born. It was suggested that the clinical use of such 'preleukaemic' samples as a source of stem cells might involve a risk of leukaemia in association with the proliferative stress consequent to transplantation. 5 Several inferences can be drawn from the data. First, if results with these particular fusion genes are typical of prenatal leukemia initiation in general, 7 then something like 5% of cord bloods may contain pre-leukemic clones. Second, the risk of the cord blood donor with such clones developing leukaemia him or herself is very low and of the order of 1%.These conclusions have implications for how one should respond to the cases of donor 'relapse'. The priority must be to establish the mechanism of donor cell leukaemia. If the above mechanism is at play, then the clear prediction is that cells bearing at least one of the molecular abnormalities found in the 'relapse' leukaemic blasts (i.e. the initiating mutation) would be present and detectable in archived donor cells. This would need to be assessed by carefully controlled reverse transcriptasepolymerase chain reaction plus combined immunophenotypefluorescence in situ hybridization technologies as described. 5 If this turns out to be the case, then there are other issues to be addressed, including additional ethical concerns. 8 Given the B1% risk of leukaemia in the donor, one would not expect to see leukaemia emerging in that individual and one can argue that it would be unethical to inform the donor or the donor's parents. Screening all cord bloods for potential 'contamination' with pre-leukaemic clones might be desirable, but would not be very practical given the wide variety of chromosomal and mutational events that could initiate acute leukaemia. Cord blood transplantation has been a very considerable success 9 and it is unlikely that donor cell leukaemia would seriously compromise its continued use. Nevertheless, there is now some urgency to establish the frequency of donor cell leukaemia by more systematic screening of 'relapse' cases in the EUROCORD project and elsewhere and identifying whether or not this does indeed reflect transfer of pre-leukaemic clones. MF Greaves