Patents and technology transfer.

Although the discovery and transfer of technology from universities to industry has been taking place for many years, the surge of activity in areas related to biotechnology, over the past fifteen years, has been remarkable. As the very essence of university research requires rapid publication of results, it is particularly important that timely patenting activity take place if an orderly and profitable transfer of technology is to occur.

The aminoglycoside antibiotics.IV. Synthesis of aminodeoxy analogs of neamine.

An efficient synthesis of the key 3',4'-galacto epoxide intermediate 4 obtained from the known 5,6-O,O'-cyclohexylidene-N,N'-bis-(methoxycarbonyl)-4-O-[2,6-bis(methoxycarbony lamino)-alpha-D-glucopyranosyl]-2-deoxystreptamine (5) is described. Treatment of this epoxide with sodium azide, followed by reduction and acetylation, yielded the protected4'-amino-4'-deoxyneamine 18 (3',4'-diequatorial), whereas treatment with ammonia followed by acetylation yielded the protected 3'-amino-3'-deoxyneamine analog 19 with a diaxial configuration of its 3' and 4' positions. Reaction of the previously described protected 3',4'-allo epoxide 3 with sodium azide yielded separable mixtures of the protected 3'-amino-3'-deoxyneamine 14 and the protected diaxial 4'-amino-4'-deoxyneamine isomer 13, the ratios of products depending on the solvent temperature. Structural assignments for 13, 14, 18 and 19 were based on their PMR spectra. An additional 4'-amino-4'-deoxyneamine analog (24) with an axial configuration as its 4' position was also prepared by azide displacement of an approximately protected 4'-methanesulfonyl neamine intermediate 10. The five protected isomers were deblocked to yield a series of aminodeoxyneamine analogs (15, 16, 20, 21 and 25), all of which were less active in vitro than neamine against a group of Gram-positive and Gram-negative bacteria.

Serum protein binding alterations of selected cephalosporin antibiotics by fatty acids and their derivatives.

Saturated fatty acids containing 10--14 carbon atoms were more potent inhibitors of serum protein binding than those containing shorter or longer carbon chains. Introduction of unsaturation into chains containing 16 or 18 carbons increased their inhibitory potency. Triglycerides and fatty acid esters, chlorides, thiols, and amides had no inhibitory activity. When inhibition was observed, it was concentration dependent and occurred when the molar ratio of fatty acid to protein equaled or exceeded three. The change in percent serum protein binding in the presence of an effective inhibitor was the greatest with cephalosporins that were most highly bound in the absence of an inhibitor.

In vitro and in vivo laboratory studies on three hydroxyiminophenylacetyl cephalosporins with particular reference to SK&F 80303, an unusually long-acting cephalosporin.

Three cephalosporins with 7-(2-hydroxyiminophenylacetamido) side chains (SK&F 79433, 80000 and 80303), differing in their 3-substituents, exhibited similar broad-spectrum antibacterial activity in vitro against strains of Staphylococcus aureus, Streptococcus faecalis and various Gram-negative bacilli. All three were active in vivo (s.c., mouse) against S. aureus, Escherichia coli or Klebsiella pneumoniae, but they differed significantly in serum pharmacokinetic profiles. SK&F 80303 produced high and extremely prolonged serum levels and protected mice when administered up to 24 hours prior to challenge with beta-lactamase-producing S. aureus or K. pneumoniae. It was resistant to hydrolysis by beta-lactamases from S. aureus, and variably so to beta-lactamases from E. coli strains. SK&F 80303 was bacteriolytic to logarithmically growing S. aureus, E. coli, Proteus mirabilis, K. pneumoniae and Enterobacter cloacae (partially). SK&F 80303 illustrates further the effect of the 3-sulfoalkyltetrazole substituent on the pharmacokinetic properties of cephalosporins. Its combined biological properties make it a possible candidate for therapeutic and long-term prophylactic use.

Presumptive identification of aminoglycoside antibiotics by the pH susceptibility disc agar-diffusion method.

Using the pH (buffered) sensitivity discs for the agar-diffusion bioassay of aminoglycoside antibiotics, characteristic response curves were obtained. Since the nature of the activities observed are structure-related, this method can serve as a useful aid for primary identification of members of this class of antibiotics.

Semisynthetic cephalosporins with antifungal activity: laboratory studies on 2-pyridinethiol 1-oxide cephalosporins.

Cephalosporins are chemotherapeutic agents whose spectrum and use are limited to antibacterial activity. Therefore, it was of interest to find several new semisynthetic cephalosporins which possess in vitro antifungal activity against Trichophyton mentagrophytes, Candida albicans and certain other yeasts. Five new cephalosporins containing the 2-pyridinethiol 1-oxide grouping were examined. Four with this heterocyclic moiety in the 3-position were found to have activity. One with the grouping in the 7-acyl-side chain was inactive. The degree of the antifungal activity was influenced by the substituent at the 7-position. The cephalosporin with a p-hydroxyphenylglycyl side chain was the most potent against the fungal strains studied. Despite this in vitro antifungal activity, none of the compounds protected mice against a systemic Candida albicans infection. All of these cephalosporins had broad spectrum in vitro and in vivo antibacterial activity.

SK&F 75073, new parenteral broad-spectrum cephalosporin with high and prolonged serum levels.

SK&F 75073, a new parenteral cephalosporin, was found to have broad in vitro and in vivo antibacterial activity including isolates usually resistant to cephalothin and cefazolin. This activity included indole-positive Proteus and Enterobacter species and some Serratia isolates. Proteus mirabilis strains were particularly susceptible, as were Haemophilus influenzae and Neisseria species. The activity of SK&F 75073 against gram-positive bacteria was poorer than that of the control cephalosporins. This cephalosporin is highly bound to serum proteins, and a loss in in vitro activity was observed in the presence of serum. Parenteral administration of SK&F 75073 to experimental animals (mice, dogs, squirrel monkeys) resulted in high and prolonged serum levels when compared with cefazolin and other injectable cephalosporins. This favorable serum profile was reflected in the excellent protection observed in mice infected with pathogenic bacteria.

Semiautomated turbidimetric microbiological assay for cefazaflur.

A quantitative semi-automated turbidimetric bioassay for cefazaflur, using Streptococcus faecium as the indicator, is described. Assays were run at pH 6.5 approximately 7 for 3.75 hours at 37 degrees C using 2 approximately 12 microgram cefazaflur per ml assay broth for standards. The dose response line was plotted point to point using the natural log of the absorbance vs natural log of the concentration. This assay is both accurate and precise and is more rapid than traditional plate assays for antibiotics.

Biological and chemotherapeutic studies on three semisynthetic cephamycins.

Three semisynthetic cephamycin antibiotics (7alpha-methoxy-cephalosporins), SK&F 73678, SK&F 83088 (CS-1170) and cefoxitin, have been found to possess favorable biological and chemotherapeutic properties. All three cephamycins are active in vitro against strains of Staphylococcus aureus and a variety of gram-negative bacilli. Beta-lactamase producing organisms including indole-producing Proteus spp., Enterobacter spp. and Serratia strains as well as certain anaerobic bacteria were found to be susceptible to these antibiotics. SK&F 73768 showed somewhat better MIC values than cefoxitin against multiple strains of bacteria. Strains of Pseudomonas aeruginosa and group D streptococci are essentially insensitive to these compounds. Their binding to serum proteins is relatively low. In mice, cefoxitin showed the most favorable pharmacokinetics with respect to peak serum level, serum half-life and urinary recovery. These cephamycins protected mice experimentally infected with a variety of bacterial strains. All three compounds are rapidly bacteriolytic to the logarithmically growing Escherichia coli and belatedly so to Staphylococcus strains with complete sterilizing effect. SK&F 73678 and SK&F 83088 showed activity and potency comparable to or better than cefoxitin and thus can be considered candidates for clinical study.

The aminoglycoside antibiotics. I. Synthesis and biological evaluation of an analog of gentamicin.

The synthesis of 2-deoxy-4-O-(2,6-diamino-2,3,4,6-tetradeoxy-alpha-D-erythrohexopyranosyl)-6-O-(3-deoxy-3-methylamino-alpha-D-xylopyranosyl)-D-streptamine (1), an analog of gentamicin A, from diideoxyneamine and methyl 3-methylamino-3-deoxy-beta-D-xylopyranoside is described. The product was characterized by its 13C nmr spectrum and was found to exhibit broad spectrum antibacterial activity.

In vitro studies with cefazaflur and other parenteral cephalosporins.

Cefazaflur has a broad-spectrum of in vitro antibacterial activity equal to or greater than that of the commercially-available cephalosporins. In addition, cefazaflur has activity against isolates of Enterobacter, Citrobacter and indole-positive Proteus; however, this activity decreased markedly when the MIC determinations were carried out with a large inoculum size. A similar inoculum effect was observed with cefamandole, however, cefoxitin's activity was relatively unchanged at increased inoculum sizes. Human serum had a relatively small effect on the in vitro activity of cefazaflur.

Comparative stability of cephalothin and cefazolin in buffer or human serum.

A marked loss in potency was observed when cephalothin was incubated for 5 h in human serum at 37 degrees C. Cefazolin was stable under these conditions.

Semisynthetic cephalosporins. Synthesis and structure-activity relationships of analogues with 7-acyl groups derived from 2-(cyanomethylthio)acetic acid or 2-[(2,2,2-trifluoroethyl)thio]acetic acid and their sulfoxides and sulfones.

The synthesis and in vitro and in vivo activities of a series of cephalosporins having side chains derived from 2-[(2,2,2-trifluoroethyl)thio]acetic acid or 2-(cyanomethylthio)acetic acid and with acetoxymethyl or 3-heterocyclic thiomethyl substituents at the 3 position are described. In both series, increasing the oxidation state of the side-chain sulfur atom from sulfide to sulfoxide/sulfone decreased the in vitro gram-positive activity, but the effect on gram-negative activity was variable and less pronounced. The protective effectiveness in mice infected with Escherichia coli increased as the oxidation level of the side-chain sulfur was raised from sulfied to sulfoxide/sulfone. Replacement of the 3-acetoxymethyl by a 3-heterocyclic thiomethyl group resulted in overall improvement of activity both in vitro and in vivo for all oxidation states.

Semisynthetic cephalosporins. Synthesis and structure-activity relationships of 7-sulfonylacetamido-3-cephem-4-carboxylic acids.

The synthesis and in vitro and in vivo activities of a series of 7-sulfonylacetamido-3-cephem-4-carboxylic acids with acetoxymethyl or heterocyclic thiomethyl substituents at the 3 position are described. Lengthening the alkyl chain attached to the sulfonyl group increased gram-positive activity but the effect on gram-negative activity was variable. Other structural changes on the 7-acyl side chain resulted in only minor changes in vitro activity. the protective effectiveness in infected mice generally paralleled the in vitro activity, except that the butylsulfonyl derivatives were less protective than predicted by in vitro activity. replacement of the 3-acetoxymethyl by a 3-heterocyclic thiomethyl group resulted in an overall improvement of activity both in vitro and in vivo.

Cefatrizine (SK&F 60771), a new oral cephalosporin: serum levels and urinary recovery in humans after oral or intramuscular administration--comparative study with cephalexin and cefazolin.

Cefatrizine (SK&F 60771), a new broad-spectrum cephalosporin, was administered in a 0.5-g dose either orally or intramuscularly to volunteers in a crossover study. After oral administration, the average peak serum levels were 5.6 and 22.1 mug/ml for cefatrizine and cephalexin, respectively. The serum half-life of cefatrizine appeared to be more extended than that of cephalexin. Urinary recovery of cefatrizine (35%) was approximately half that of cephalexin (68%) after oral administration. After intramuscular injection of 0.5 g, the average peak serum level of cefatrizine (12.0 mug/ml) was approximately one-fourth that of cefazolin (44.0 mug/ml). The serum half-life after intramuscular injection was 86 min for cefatrizine and 118 min for cefazolin. Urinary recovery was 45% of the intramuscularly administered dose, as compared with cefazolin, which was 74%.

Relationships between in vitro susceptibility and efficacy in experimental mouse infection-protection assay of cefazolin and cephalothin using Escherichia coli strains.

The disc sensitivity and minimal inhibitory concentrations (MIC) of cefazolin and cephalothin were compared against a series of Escherichia coli isolates. These data were correlated with the mouse protective doses of the 2 cephalosporins in animals infected with E. coli strains selected according to their various degrees of in vitro sensitivity to the 2 cephalosporins. The overwhelming majority of E. coli strains showed a significantly higher degree of susceptibility and lower MIC values for cefazolin than for cephalothin. There has been found a good correlation between the inhibition zones and especially the MIC values and the ED50 results for both cephalosporins. Using E. coli clinical isolates, cefazolin was found to be superior to cephalothin not only in vitro experiments but also more potent in protecting the experimentally infected mice.