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Evaluation of hit compounds in Caco-2 cells Caco-2 cells are derived from a human colon adenocarcinoma 49 ; . In culture, these cells spontaneously differentiate into polarized, columnar cells that are representative of the small intestine 50 ; . When grown on plastic, they form domes typical of normal and transporting epithelium 51, 52 ; . This cell line has been used extensively as a model for transport studies 50, 53 ; . Previous work has shown fatty acid uptake in Caco-2 cells is saturable, decreased by trypsin treatment of cells and can be competed by natural fatty acids 54 ; . These cells express higher level of both FATP2 and FATP4 than other FATP isoforms Black, Fraisl and DiRusso, unpublished data ; . To further evaluate hit compounds identified from our screening of yeast cells, we performed C1-BODIPY-C12 uptake inhibition experiments in 96-well format using Caco-2 cells. We initially tested the hit compounds in Caco-2 cells at 100 M and 10 M Table 7 ; . At 100M, most of these compounds inhibited Caco-2 uptake of C1-BODIPY-C12 to varying.

In response to the recent public concern over the issue of childhood cancer and Vitamin K given by injection, an Expert Committee met on 15 February, 1993 to discuss and issue a statement on the use of Vitamin K in infancy. A Joint Statement has been endorsed by both Councils of the Hong Kong College of Paediatricians and Hong Kong Paediatric Society. Enclosed herewith are the Joint Statement and a review of recent literature on this issue and megestrol. Substitutions in other recA' and recA + " mutants see Table I, not necessarily correlate with theDNA repair phenotype of a N-28, N-56, N-58, N-69, N-72, and Table 11, N-100, N-101, given mutant. N-102, N-104 ; , one might predict only a minimal effect on RecA Fourteen of the 29 coprt' mutants are red + , five of which function. However, N-116 shows a very low DNArepair activity. carry single amino acid substitutions. Mutants N-19 Ile'55 The coprotease activity is uneffected in this triple mutant. Met ; , N-29 G ~ u + Lys ; , and N-30 GIu"~ + Lys ; are ex'~~ N-31 Glu" * A different result that supports our observation of an unex- amples of strong coprt" mutants, whereas mutants pectedly high tolerance for mutation at most positions in this + Gln ; and N-33 GluI5' + Asn ; are examples of weak and V. region occurs in several triple mutants that carry combinations moderate coprt' mutants, respectively Table I ; The remainof hydrophobic substitutions. These include the following mu- ing 15 coprt' mutants show some decrease in DNA repair ac159, tivity, but this vanes widely from nearly r e d nearly r e d tants with the changes indicated at positions 156, 158, and respectively: N-78 Val, Leu, and Val ; , N-79 Val, Leu, and Leu ; , Table IV ; . Three of these mutants carry single substitutions, N-81 Gly, Val, and Val ; and N-115 Val, Phe, and Val ; . Whereas one being a weak coprt' mutant N-84, Ala153 + Glu ; and the the wild type RecA sequence in this region contains two nega- others showing moderate constitutive activity N-87, Gly'57 4 tive charges in stretch of 5 residues '5511e-Glu-Gly-Glu-Ile'5g ; , a Cys and N-88, Gly'57 + Asp ; . these four mutants create run of hydrophobic residues which, a No single substitution at positions 152, 154, 159 or any of the despite the dramatic chemical and stericdifferences from wild nontargeted residues resulted in a coprt" mutant. Of the nine type, support wild type-like DNA repair and coprotease single mutants obtained at Lys15' Tables I and 11 ; all showed functions. inducible coprotease activity. At G ~ all four of~the~single u ~ The results in the sections above regarding the effects of mutants have coprt phenotypes that correspond to the DNA mutations on DNA repair activity are summarized inFig. 2. repair phenotypes, three score as red-lcoprt- Table 111, N-121, Coprr Mutants-Of the 149 unique e d mutants character- N-122, and N-1231, and the other partially functional for both r is ized in this study found 29 that werecoprt'. The occurrence activities Table 11, N-85 ; . At position 159 all threesingle muwe of coprt' mutants as well as thelevel of constitutive activity do tants score as recA'lcoprt' Table I, N-34, N-35, and N-36.

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