In Vitro Activity of the New Glycopeptide LY333328 against Multiply Resistant Gram-Positive Clinical Isolates

Abstract

The in vitro activity of LY333328 was compared with those of vancomycin and teicoplanin against 425 gram-positive clinical isolates, including a variety of multiply resistant strains. LY333328 at ≤4 μg/ml inhibited all microorganisms tested, including methicillin- and teicoplanin-resistant staphylococci, glycopeptide-resistant enterococci, penicillin- and multiply resistant pneumococci, and viridans and beta-hemolytic streptococci.

The incidence of severe nosocomial infections caused by gram-positive cocci has increased in the last decades (1). This increase has been accompanied by the resistance of these microorganisms to multiple antimicrobials (5). The spread of methicillin-resistant staphylococci and penicillin-resistant pneumococci has led to the use of vancomycin, which in turn may have contributed to the emergence of glycopeptide-resistant enterococci and staphylococci (3). The risk of acquisition of infections due to these pathogens as well as the potential for the spread of vancomycin resistance to other microorganisms emphasizes the need for new compounds as alternatives for therapy. The new glycopeptide antibiotic LY333328 is a semisynthetic N-alkyl derivative of an analog of vancomycin (9, 11). Previous studies have demonstrated that LY333328 is active against gram-positive bacteria, including glycopeptide-resistant enterococci (4, 6, 7, 12). In this study, we examine the in vitro activity of LY333328 against recently obtained gram-positive clinical isolates, including a variety of multiply resistant strains.

(This study was presented at the 37th Interscience Conference on Antimicrobial Agents and Chemotherapy, Toronto, Ontario, Canada, 28 September to 1 October 1997.)

We tested a total of 425 nonduplicate, recently obtained clinical isolates, including 62 (30 methicillin-resistant) Staphylococcus aureus strains, 73 (43 methicillin-resistant and 11 teicoplanin-resistant) coagulase-negative staphylococci, 46 (20 vancomycin-resistant) Enterococcus faecalis strains, 43 (15 vancomycin-resistant) Enterococcus faecium strains, 101 (40 penicillin-intermediate and 40 penicillin-resistant) Streptococcus pneumoniae strains, 60 (20 penicillin-intermediate and 20 penicillin-resistant) Streptococcus viridans group strains, 20 (15 erythromycin-resistant) Streptococcus pyogenes strains, and 20 (5 erythromycin-resistant) Streptococcus agalactiae strains. LY333328 and vancomycin were provided by Lilly Research Laboratories (Indianapolis, Ind.) and teicoplanin was provided by Hoeschst Marion Roussel, Inc. (Cincinnati, Ohio). LY333328 was solubilized in water and diluted in cation-adjusted Mueller-Hinton broth, as indicated by the instructions of the manufacturers. Susceptibility studies were performed by the broth microdilution method with cation-adjusted Mueller-Hinton broth supplemented with 5% lysed horse blood when necessary. The recommendations of the National Committee for Clinical Laboratory Standards were followed (8). Susceptibilities to penicillin, ampicillin, oxacillin, cefotaxime, and erythromycin were determined with commercial panels (Sensititre; AccuMed International Ltd., West Sussex, England) by following the recommendations given by the manufacturers. Vancomycin and teicoplanin MICs of ≥16 μg/ml defined resistance. A projected breakpoint of ≤4 μg/ml was used to define percentages of susceptibility for LY333328. S. aureus ATCC 29213, E. faecalis ATCC 29212, and Streptococcus pneumoniae ATCC 49619 were used as control strains. Isolates were stored at −70°C, with three subcultures being made before the organisms were tested. Following inoculation (final inoculum concentration, ca. 5 × 10⁵ CFU/ml), MIC trays were incubated at 37°C in ambient air for 24 h before examination.

LY333328’s activity against all the strains tested was enhanced compared with those of vancomycin and teicoplanin. Table 1 compares the in vitro activities of all the antimicrobials tested against the 425 isolates. LY333328 inhibited all strains tested at concentrations of ≤4 μg/ml. The MICs at which 90% of the isolates were inhibited (MIC₉₀s) of all three glycopeptides for methicillin-resistant S. aureus were the same (2 μg/ml). LY333328 was also very active against methicillin-resistant coagulase-negative staphylococci, including teicoplanin-resistant strains (MIC₉₀, 2 μg/ml), and inhibited 100% of staphylococci tested at ≤4 μg/ml. In comparison with vancomycin and teicoplanin, LY333328 showed the best activity against enterococci, including vancomycin-resistant strains (VanA and VanB): all enterococci were inhibited at ≤2 μg/ml. LY333328 was 16- to 32-fold more active than vancomycin but had the same activity as teicoplanin (MIC₉₀, ≤0.01 μg/ml) against pneumococci. With Streptococcus viridans isolates the figures were similar, LY333328 was 8- to 32-fold more active than vancomycin, 8- to 16-fold more active than teicoplanin, and the most active of all antimicrobials tested against these microorganisms. The activity of this new compound against Streptococcus pyogenes and Streptococcus agalactiae was 16-fold and 2- to 16-fold higher than those of vancomycin and teicoplanin, respectively (Table 1).

TABLE 1.

In vitro activity of LY333328 compared to those of vancomycin, teicoplanin, and other antimicrobial agents against 425 gram-positive clinical isolates

Organism(s) and type (no. tested)	Antimicrobial agent	MIC (μg/ml)a			%b
		50%	90%	Range	Susc	Interm	Res	≤4 μg/ml
Staphylococcus aureus
Oxacillin susceptible (32)	LY333328	0.5	1	0.25–2				100
	Vancomycin	1	2	0.5–2	100
	Teicoplanin	0.5	2	0.12–4	100
Oxacillin resistant (30)	LY333328	1	2	0.12–4				100
	Vancomycin	1	2	0.5–4	100
	Teicoplanin	2	2	0.25–2	100
Coagulase-negative Staphylococcus spp.
Oxacillin susceptible (31)	LY333328	0.5	2	≤0.01–2				100
	Vancomycin	1	2	0.25–2	100
	Teicoplanin	0.5	2	0.03–4	100
Oxacillin resistant (42)c	LY333328	1	2	0.06–4				100
	Vancomycin	2	2	0.5–4	100
	Teicoplanin	2	64	0.06–64	74		26
Enterococcus faecalis
Vancomycin susceptible (25)	LY333328	0.5	1	0.06–1				100
	Vancomycin	1	2	0.5–4	100
	Teicoplanin	0.06	1	≤0.01–2	100
	Ampicillin	2	2	0.25–2	100
Vancomycin resistant (21)	LY333328	1	2	0.03–2				100
	Vancomycin	512	1,024	16–1,024			100
	Teicoplanin	2	64	≤0.03–64	24		76
	Ampicillin	1	2	1–2	100
Enterococcus faecium
Vancomycin susceptible (27)	LY333328	0.06	0.12	≤0.01–0.5				100
	Vancomycin	0.5	2	≤0.01–2	100
	Teicoplanin	0.25	0.5	≤0.01–1	100
	Ampicillin	>16	>16	0.25–>16	18		82
Vancomycin resistant (16)	LY333328	1	1	0.12–1				100
	Vancomycin	512	512	16–1,024			100
	Teicoplanin	64	128	0.01–128	19		81
	Ampicillin	128	256	1–256	25		75
Streptococcus pneumoniae
Penicillin susceptible (21)	LY333328	≤0.01	≤0.01	≤0.01				100
	Vancomycin	0.25	0.25	0.12–0.5	100
	Teicoplanin	≤0.01	≤0.01	≤0.01	100
	Erythromycin	≤0.25	≤0.25	≤0.25–>4	95		5
	Cefotaxime	≤0.06	≤0.06	≤0.06	100
	Penicillin	≤0.03	0.06	≤0.03–0.06	100
Penicillin intermediate (40)	LY333328	≤0.01	≤0.01	≤0.01–0.06				100
	Vancomycin	0.25	0.25	≤0.01–0.5	100
	Teicoplanin	≤0.01	≤0.01	≤0.01	100
	Erythromycin	4	>4	≤0.25–>4			100
	Cefotaxime	0.5	1	≤0.06–1	80	20
	Penicillin	0.5	1	0.12–1		100
Penicillin resistant (40)	LY333328	≤0.01	≤0.01	≤0.01–0.06				100
	Vancomycin	0.25	0.5	0.06–0.5	100
	Teicoplanin	≤0.01	≤0.01	≤0.01–0.12	100
	Erythromycin	>4	>4	≤0.25–>4			100
	Cefotaxime	1	2	0.5–2	12.5	55	32.5
	Penicillin	4	4	2–4			100
Viridans streptococci
Penicillin susceptible (20)	LY333328	≤0.01	≤0.01	≤0.01				100
	Vancomycin	0.5	0.5	0.12–1	100
	Teicoplanin	≤0.01	0.12	≤0.01–0.5	100
	Penicillin	≤0.03	0.12	≤0.03–0.12	100
	Erythromycin	≤0.25	2	≤0.25–>4	80	5	15
	Cefotaxime	≤0.06	0.12	≤0.06–0.12	100
Penicillin intermediate (20)	LY333328	≤0.01	≤0.01	≤0.01–0.25				100
	Vancomycin	0.25	0.5	0.12–0.5	100
	Teicoplanin	0.03	0.25	≤0.01–0.5	100
	Penicillin	0.5	1	0.5–2		100
	Erythromycin	2	>4	≤0.25–>4			100
	Cefotaxime	0.25	1	≤0.06–4	75	15	10
Penicillin resistant (20)	LY333328	≤0.01	0.06	0.01–0.06				100
	Vancomycin	0.25	0.5	0.25–1	100
	Teicoplanin	≤0.01	0.5	≤0.01–0.5	100
	Penicillin	4	>4	4–>4			100
	Erythromycin	>4	>4	≤0.25–>4	25		75
	Cefotaxime	4	>8	1–>8		15	85
Streptococcus spp.
Serogroup A (20)	LY333328	≤0.01	≤0.01	≤0.01–0.25				100
	Vancomycin	0.25	0.25	0.12–0.25	100
	Teicoplanin	≤0.01	0.25	≤0.01–0.25	100
	Penicillin	≤0.03	≤0.03	≤0.03	100
	Erythromycin	4	>4	≤0.25–>4	25		75
	Cefotaxime	≤0.06	≤0.06	≤0.06	100
Serogroup B (20)	LY333328	≤0.01	0.06	≤0.01–0.5				100
	Vancomycin	0.5	1	0.25–1	100
	Teicoplanin	≤0.01	0.12	≤0.01–0.25	100
	Penicillin	≤0.03	≤0.03	≤0.03	100
	Erythromycin	≤0.5	>4	≤0.5–>4	75		25
	Cefotaxime	≤0.06	≤0.06	≤0.06	100

^a50% and 90%, MIC₅₀ and MIC₉₀, respectively. 

^bPercentages of isolates that were susceptible (Susc), intermediate (Interm), and/or resistant (Res) to the agent or inhibited with ≤4 μg of the agent per ml. 

^cIncludes 11 teicoplanin-resistant strains. 

LY333328 is a new semisynthetic glycopeptide with activity against methicillin-resistant S. aureus and vancomycin-resistant enterococci. Few studies have determined the activity of this compound against multiply resistant gram-positive emerging pathogens. Our study shows that LY333328’s ability to inhibit a variety of multiply resistant gram-positive clinical isolates is excellent. In our study, LY333328 showed an activity similar to those of vancomycin and teicoplanin against methicillin-resistant S. aureus and coagulase-negative staphylococci. Previous studies have reported various MIC results for staphylococci, with MIC₉₀s from 2 to 8 μg/ml and no significant differences in activities against methicillin-resistant and susceptible strains (2, 6). Other reports have shown MIC results similar to or slightly lower than ours (10). These small differences have been found only with staphylococci and may be explained by susceptibility testing conditions, physicochemical characteristics of the drug, and even strain-to-strain differences. LY333328 was active against all teicoplanin-resistant staphylococci tested. More-uniform results have been reported for enterococci, including vancomycin-resistant strains (6, 10). In our study all enterococci were inhibited with ≤2 μg of LY333328 per ml, this drug being the most active of the glycopeptides tested. Other gram-positive cocci were also tested in order to assess the general efficacy of the compound. LY333328 was very active against Streptococcus pyogenes and Streptococcus agalactiae, including erythromycin-resistant strains. The antipneumococcal activity of LY333328 has been previously reported (4). We tested the activity of LY333328 against multiply resistant pneumococci. In our study, all penicillin-, erythromycin-, and cefotaxime-resistant pneumococci were inhibited with ≤0.06 μg of LY333328 per ml, with LY333328 showing the same activity as teicoplanin and being 32-fold more active than vancomycin. This suggests a potential therapeutic option for the treatment of severe infections due to multiply resistant pneumococci, if results of toxicological and clinical studies point to development of this drug. This is, to our knowledge, the first study in which the activity of LY333328 has been evaluated against multiply resistant viridans streptococci, and this compound seems to be the most active of all tested antimicrobials, including the glycopeptides.

In summary, the glycopeptide LY333328 showed activity superior to those of vancomycin and teicoplanin. LY333328 is a promising new drug with high activity against many difficult-to-treat gram-positive multiply resistant microorganisms, including methicillin- and teicoplanin-resistant staphylococci, glycopeptide-resistant enterococci, penicillin- and multiply resistant pneumococci, and viridans streptococci.