Infections
by intracellular pathogens are difficult to treat because
of the poor accessibility of antibiotics to the pathogens encased
by host cell membranes. As such, a strategy that can improve the membrane
permeability of antibiotics would significantly increase their efficiency
against the intracellular pathogens. Here, we report the design of
an adaptive, metaphilic cell-penetrating polypeptide (CPP)–antibiotic
conjugate (VPP-G) that can effectively eradicate the intracellular
bacteria both
in vitro
and
in vivo
. VPP-G was synthesized by attaching vancomycin to a highly membrane-penetrative
guanidinium-functionalized metaphilic CPP. VPP-G effectively kills
not only extracellular but also far more challenging intracellular
pathogens, such as
S. aureus
, methicillin-resistant
S. aureus
, and vancomycin-resistant
Enterococci
. VPP-G enters the host cell via a unique metaphilic membrane penetration
mechanism and kills intracellular bacteria through disruption of both
cell wall biosynthesis and membrane integrity. This dual antimicrobial
mechanism of VPP-G prevents bacteria from developing drug resistance
and could also potentially kill dormant intracellular bacteria. VPP-G
effectively eradicates MRSA
in vivo
, significantly
outperforming vancomycin, which represents one of the most effective
intracellular antibacterial agents reported so far. This strategy
can be easily adapted to develop other conjugates against different
intracellular pathogens by attaching different antibiotics to these
highly membrane-penetrative metaphilic CPPs.
Antibiotic
resistance is a daunting challenge in modern medicine,
and novel approaches that minimize the emergence of resistant pathogens
are desperately needed. Antimicrobial peptides are newer therapeutics
that attempt to do this; however, they fall short because of low to
moderate antimicrobial activity, low protease stability, susceptibility
to resistance development, and high cost of production. The recently
developed random peptide mixtures (RPMs) are promising alternatives.
RPMs are synthesized by incorporating a defined proportion of two
amino acids at each coupling step rather than just one, making them
highly variable but still defined in their overall composition, chain
length, and stereochemistry. Because RPMs have extreme diversity,
it is unlikely that bacteria would be capable of rapidly evolving
resistance. However, their efficacy against pathogens in animal models
of human infectious diseases remained uncharacterized. Here, we demonstrated
that RPMs have strong safety and pharmacokinetic profiles. RPMs rapidly
killed both Pseudomonas aeruginosa and Staphylococcus
aureus efficiently and disrupted preformed biofilms by both
pathogens. Importantly, RPMs were efficacious against both pathogens
in mouse models of bacteremia and acute pneumonia. Our results demonstrate
that RPMs are potent broad-spectrum therapeutics against antibiotic-resistant
pathogens.
Streptococcus pneumoniae (pneumococcus)
is a prevalent human pathogen that utilizes the competence regulon
quorum sensing circuitry to acquire antibiotic resistance and initiate
its attack on the human host. Therefore, targeting the competence
regulon can be applied as an anti-infective approach with minimal
pressure for resistance development. Herein, we report the construction
of a library of urea-bridged cyclic dominant-negative competence-stimulating
peptide (dnCSP) derivatives and their evaluation as competitive inhibitors
of the competence regulon. Our results reveal the first pneumococcus
dual-action CSPs that inhibit the group 1 pneumococcus competence
regulon while activating the group 2 pneumococcus competence regulon.
Structural analysis indicates that the urea-bridge cyclization stabilizes
the bioactive α-helix conformation, while in vivo studies using a mouse model of infection exhibit that the lead dual-action
dnCSP, CSP1-E1A-cyc(Dab6Dab10), attenuates group 1-mediated mortality
without significantly reducing the bacterial burden. Overall, our
results pave the way for developing novel therapeutics against this
notorious pathogen.
The
competence regulon of Streptococcus pneumoniae (pneumococcus)
is a quorum-sensing circuitry that regulates the
ability of this pathogen to acquire antibiotic resistance or perform
serotype switching, leading to vaccine-escape serotypes, via horizontal
gene transfer, as well as initiate virulence. Induction of the competence
regulon is centered on binding of the competence-stimulating peptide
(CSP) to its cognate receptor, ComD. We have recently synthesized
multiple dominant-negative peptide analogs capable of inhibiting competence
induction and virulence in S. pneumoniae. However,
the pharmacodynamics and safety profiles of these peptide drug leads
have not been characterized. Therefore, in this study, we compared
the biostability of cyanine-7.5-labeled wild-type CSPs versus dominant-negative
peptide analogs (dnCSPs) spatiotemporally by using an IVIS Spectrum in vivo imaging system. Moreover, in vitro cytotoxicity and in vivo toxicity were evaluated.
We conclude that our best peptide analog, CSP1-E1A-cyc(Dap6E10), is
an attractive therapeutic agent against pneumococcal infection with
superior safety and pharmacokinetics profiles.
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