No abstract

Amines whose pK values lie between about 5 and 9 are lysosomotropic because lysosomes are acidic intracellular compartments. If such amines bear long hydrophobic chains, they become detergents upon protonation inside the lysosomes, rupturing the lysosomal membrane and killing the cell. Six types of lysosomotropic amines have been prepared that all behave in the expected manner. They are cytotoxic to all lysosome-bearing cells but not red blood cells, which lack lysosomes. Their mode of action, the effect of alkyl chain length on activity, and the fact that their cytotoxic action appears only above a threshhold intracellular concentration support the belief that they behave as lysosomotropic detergents. Among the potential applications is cancer chemotherapy.

Macrophages isolated from the peritoneal cavity of untreated mice and maintained in tissue culture synthesize and release prostaglandins when challenged with zymosan. These cells also selectively release lysosomal acid hydrolases under the same conditions. The major prostaglandins released into the media are found to be prostaglandins E1, E2 and 6-oxoprostaglandin F1a, whereas prostaglandin F2a is not detected. Macrophages isolated from mice that have received an intraperitoneal injection of thioglycollate broth are far less responsive to zymosan challenge. These cells require 300 microgram of zymosan to synthesize and release one-third the amount of prostaglandins released from non-stimulated macrophages exposed to 50 microgram of zymosan. In addition, thioglycollate-stimulated macrophages release less than 10% of their lysosomal acid hydrolases when exposed to 300 microgram of zymosan whereas non-stimulated cells release approximately 50% of these enzymes after treatment with 50 microgram of zymosan. The zymosan-stimulated synthesis and release of prostaglandins are completely inhibited by indomethacin, whereas the increased selective release of lysosomal acid hydrolases is not affected. Macrophages, unlike fibroblasts, do not synthesize and release prostaglandins when exposed to serum or to bradykinin.

Topical application of arachidonic acid on mouse ears induces the synthesis of prostaglandin E2 and leukotrienes C4 and D4. The increased tissue levels of these products are quantitated by radioimmunoassay. The identity of the leukotrienes was confirmed by immunoreactivity of reverse-phase high-performance liquid chromatography fractions corresponding to authentic standards. Synthesis of the arachidonic acid metabolites precedes or is coincident with increased vascular permeability resulting in an edematous response, as measured by accumulation of [125I]albumin in the ear after i.v. injection or by tissue wet weight. When applied topically, anti-inflammatory drugs such as BW755C (3-amino-1-(m-[trifluoromethyl]phenyl)2-pyrazoline, indomethacin, and nordihydroguaiaretic acid inhibit edema and modulate the appearance of the arachidonic acid products. The data suggest the coinvolvement of prostaglandin E2 and leukotrienes C4 and D4 as mediators of inflammation in this in vivo model.

Several laboratories, including our own have reported the synthesis and activity of certain low relative molecular mass inhibitors of mammalian serine proteases, especially human leukocyte elastase (HLE, EC 3.4.21.37), an enzyme whose degradative activity on lung elastin has been implicated as a major causative factor in the induction of pulmonary emphysema, and which is present in the azurophil granules of human polymorphonuclear leukocytes (PMN). Normally, these granules fuse with phagosomes containing engulfed foreign material (such as bacteria), and HLE, in combination with other lysosomal enzymes, catabolizes the particles. Under certain pathological conditions, however, PMN become attached to host protein (elastin fibres, basement membrane, connective tissue, immune complexes), and in response to this adherence, the granules may fuse with the PMN outer membrane and release their contents, including HLE, directly onto the tissue. Besides emphysema, HLE may also contribute to the pathogenesis of disease states such as adult respiratory distress syndrome, and its potential involvement in rheumatoid arthritis makes HLE inhibitors of considerable interest. It is known that cephalosporin antibiotics (for example, cephalothin (compound I, Table 2)) are acylating inhibitors of bacterial serine proteases which help synthesize the cell wall by performing a transpeptidation reaction on a peptidyl substrate bearing a D-Ala-D-Ala terminus. We now report that neutral cephalosporins (that is, compounds not bearing a free carboxyl at position C-4) can be modified to become potent time-dependent inhibitors of HLE.