Mycograb (NeuTec Pharma plc) is a human genetically recombinant antibody against fungal heat shock protein 90 (HSP90). Antibody to HSP90 is closely associated with recovery in patients with invasive candidiasis who are receiving amphotericin B (AMB). Using in vitro assays developed for efficacy assessment of chemotherapeutic antifungal drugs, Mycograb showed activity against a wide range of yeast species (MICs against Candida albicans [fluconazole {FLC}-sensitive and FLC-resistant strains], Candida krusei, Candida tropicalis, Candida glabrata, and Candida parapsilosis, 128 to 256 g/ml). Mycograb (4 or 8 g/ml) showed synergy with AMB, the fractional inhibitory index being 0.09 to 0.31. Synergy was not evident with FLC, except for FLC-sensitive C. albicans. Murine kinetics showed that Mycograb at 2 mg/kg produced a maximum concentration of drug in serum of 4.7 g/ml, a half-life at alpha phase of 3.75 min, a half-life at beta phase of 2.34 h, and an area under the concentration-time curve from 0 to t h of 155 g ⅐ min/ml. Mycograb (2 mg/kg) alone produced significant improvement in murine candidiasis caused by each species: (i) a reduction (Scheffe's test, P < 0.05) in the mean organ colony count for the FLC-resistant strain of C. albicans (kidney, liver, and spleen), C. krusei (liver and spleen), C. glabrata (liver and spleen), C. tropicalis (kidney), and C. parapsilosis (kidney, liver, and spleen) and (ii) a statistically significant increase in the number of negative biopsy specimens (Fisher's exact test, P < 0.05) for C. glabrata (kidney), C. tropicalis (liver and spleen), and C. parapsilosis (liver). AMB (0.6 mg/kg) alone cleared the C. tropicalis infection but failed to clear infections caused by C. albicans, C. krusei, C. glabrata, or C. parapsilosis. Synergy with AMB, defined as an increase (Fisher's exact test, P < 0.05) in the number of negative biopsy specimens compared with those obtained using AMB alone, occurred with the

Heat-shock proteins (hsps) have been identified as molecular chaperones conserved between microbes and man and grouped by their molecular mass and high degree of amino acid homology. This article reviews the major hsps of Saccharomyces cerevisiae, their interactions with trehalose, the effect of fermentation and the role of the heat-shock factor. Information derived from this model, as well as from Neurospora crassa and Achlya ambisexualis, helps in understanding the importance of hsps in the pathogenic fungi, Candida albicans, Cryptococcus neoformans, Aspergillus spp., Histoplasma capsulatum, Paracoccidioides brasiliensis, Trichophyton rubrum, Phycomyces blakesleeanus, Fusarium oxysporum, Coccidioides immitis and Pneumocystis jiroveci. This has been matched with proteomic and genomic information examining hsp expression in response to noxious stimuli. Fungal hsp90 has been identified as a target for immunotherapy by a genetically recombinant antibody. The concept of combining this antibody fragment with an antifungal drug for treating life-threatening fungal infection and the potential interactions with human and microbial hsp90 and nitric oxide is discussed.

Seroconversion to hsp90 is associated with recovery from systemic candidiasis in humans, and a murine monoclonal antibody to this hsp90 antigen (LKVIRK epitope) was protective in mice. A human recombinant antibody to the same epitope was assessed in acute and chronic models of murine invasive candidiasis. Lethal intravenous challenge with fluconazole-susceptible (strain 4) or fluconazole-resistant (strain 019) Candida albicans, followed 2 h later by a single dose of recombinant antibody, was associated with a statistically significant drop in mortality of > or = 40% (two experiments in BALB/c mice given strain 4; one experiment in CD-1 mice given strain 019) or 23% (BALB/c mice, strain 019). In mice sublethally infected with strain 4, treatment with recombinant antibody was associated with improved renal clearance of infection. Antibody-mediated protection may involve neutralization of the protein-binding properties of circulating candidal hsp90, since LKVIRK strongly bound dexamethasone in vitro.

Immunoblotting sera from 26 patients with septicemia due to an epidemic strain of methicillin-resistant Staphylococcus aureus (EMRSA-15), 6 of whom died, revealed an immunodominant EMRSA-15 antigen at 61 kDa. There was a statistically significant correlate (P < 0.001) between survival and immunoglobulin G to the 61-kDa band. The antigen was identified by sequencing positive clones obtained by screening a genomic expression library of EMRSA-15 with pooled sera from patients taken after the septicemic episode. Eluted antibody reacted with the 61-kDa antigen on immunoblots. The amino terminus was obtained by searching the S. aureus NCTC 8325 and MRSA strain COL databases, and the whole protein was expressed in Escherichia coli TOP 10F. The derived amino acid sequence showed homology with ABC transporters, with paired Walker A and Walker B motifs and 73% homology to YkpA from Bacillus subtilis. Epitope mapping of the derived amino acid sequence with sera from patients who had recovered from EMRSA-15 septicemia delineated seven epitopes. Three of these epitopes, represented by peptides 1 (KIKVYVGNYDFWYQS), 2 (TVIVVSHDRHFLY NNV), and 3 (TETFLRGFLGRMLFS), were synthesized and used to isolate human recombinant antibodies from a phage antibody display library. Recombinant antibodies against peptides 1 and 2 gave logarithmic reductions in organ colony counts, compared with control groups, in a mouse model of the infection. This study suggests the potential role of an ABC transporter as a target for immunotherapy.

Recent case reports of vancomycin treatment failures in the United States, Japan, and France have prompted a retrospective analysis of 42 cases of septicemia caused by epidemic methicillin-resistant Staphylococcus aureus strain 15 (EMRSA-15), which is the most prevalent epidemic strain of methicillin-resistant S. aureus in the United Kingdom; all cases occurred in a teaching hospital in Manchester, United Kingdom, between 1994 and 1998. Mortality was lowest (4%) in patients with rifampin-susceptible isolates treated with vancomycin and rifampin. It rose to 38% in patients who were treated with both antibiotics but in whom the organism became resistant to rifampin during therapy, and it reached 78% in patients who had rifampin-resistant isolates or in whom rifampin was contraindicated (P<.0001; Fisher exact test, 2-tailed). All isolates were susceptible to vancomycin by conventional laboratory testing, but susceptibility was lost by growth in vancomycin in vitro, becoming resistant at a minimum inhibitory concentration of 8 mg/L. This was associated with accumulation of cell-wall material. The deoxyribonucleic acid fingerprint remained unchanged. This study suggests that rifampin played a key role in the prevention of deaths caused by an epidemic strain of methicillin-resistant S. aureus that readily gave rise to a subpopulation with reduced susceptibility to vancomycin.