Fungicidal effect of three new synthetic cationic peptides against Candida albicans.
OBJECTIVE
Peptide antibiotics are considered a new class of antifungal agents. Of these, an alpha-helical, cationic peptide termed Dhvar 4, a relative of salivary histatin has been shown to be an antifungal of relatively high potency. Similarly, lactoferricin B (LFB) and a derivative thereof, LFB(17-30), disrupts the fungal cell membrane and acts against Candida albicans. As Dhvar 4 and LFB(17-30), exhibit almost identical amino acid sequences at their C-terminal, we hypothesized that laboratory synthesis of peptides with an alpha-helical structure and having similar amphipathic properties could lead to products with candidacidal activity. Hence, three such peptides - JH8194, JH8195 and JH 8944, were synthesized and their antifungal properties compared with recognized antifungals LFB, LFB(17-30), human lactoferricin (LFH), Histatin-5 and Dhvar 4, against two isolates of C. albicans.


METHODS
The antifungal agents were synthesized and their secondary structures evaluated according to a previously described protocol of Situ and Bobek (2000)Antimicrob Agents Chemother44: 1485-1493. The C. albicans strains were oral isolates from a human immunodeficiency virus-infected (isolate A2) and a healthy (A6) individual. A standard concentration of yeasts was exposed to a range of dilutions of the agents for a specific duration and the cell death (viability) in terms of the resultant colony forming units ml(-1) was quantified.


RESULTS
Dhvar 4, showed the most alpha-helical propensity, and was the least fungicidal while LFB and LFB(17-30) showed the highest antifungal potential, and demonstrated total kill of A6, and A2 at 5 and 10 microM concentrations, respectively whilst LFH killed both isolates at a l0 microM concentration. Of the three new synthetic peptides, JH 8194 was the most potent (total kill of A6/A2 strains at 1.25/2.5 microM), followed by JH 8195 (total kill of A6/A2 strains at 5/10 microM while JH 8944 was the least potent as a 25 microM concentration was required to kill either strain of Candida. On further analyses of the relationship between pI value of the peptides and their anticandicidal activity, a significant positive correlation was noted. In order to rule out a cytotoxic effect of the new synthetic peptides we compared the fungicidal and hemolytic activities under similar incubation conditions using freshly isolated erythrocytes and all three peptides exhibited no detectable hemolysis upto an concentration of 100 microM in contrast to the polyene antifungal amphotericin B that elicited significant initiation of hemolysis at a concentration of 5.0 microM.


CONCLUSIONS
Our data suggest that laboratory synthesis of agents with an alpha-helical structure and having amphipathic properties similar to known, natural antifungal agents may be a promising avenue to generate products with improved antifungal activity.
Nikawa
H
H
Department of Prosthetic Dentistry, Hiroshima University Faculty of Dentistry, Hiroshima, Japan. hirocky@hiroshima-u.ac.jp
Fukushima
H
H
Makihira
S
S
Hamada
T
T
Samaranayake
L P
LP
D000595:Amino Acid Sequence; D000935:Antifungal Agents / Q000138:chemical synthesis* / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000138:chemical synthesis* / Q000494:pharmacology*; D002176:Candida albicans / Q000187:drug effects*; D002942:Circular Dichroism; D015169:Colony Count, Microbial; D005260:Female; D006461:Hemolysis; D006801:Humans; D007526:Isoelectric Point; D008297:Male; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary
D000935:Antifungal Agents; D023181:Antimicrobial Cationic Peptides