A new antibiotic kills
pathogens without
detectable resistance
Losee L. Ling, Tanja Schneider2, Aaron J. Peoples, Amy L. Spoering, Ina Engels, Brian P. Conlon, Anna Mueller,
Till F. Scha¨berle, Dallas E. Hughes, Slava Epstein, Michael Jones, Linos Lazarides, Victoria A. Steadman, Douglas R. Cohen,
Cintia R. Felix, K. Ashley Fetterman, William P. Millett, Anthony G. Nitti, Ashley M. Zullo, Chao Chen & Kim Lewis
Saman Amirzadegan & Krista Jastrzembski
BINF 704
Antibiotic Resistance
Overview
• A Short History of Antibiotics
• Development of Antibiotic Resistance
• Teixobactin
• Cultivation
• Identification
• Mechanism of Action
• Discussion
The Antibiotic Era
1909 – Paul Ehrlich et. al. discover a drug effective against syphilis
Discovery of Penicillin
September 3, 1928
Alexander Fleming returns to his lab after a lengthy vacation to
discover Penicillin
Antibiotic Mechanisms of Action
1940 Penicillin
introduced –
Resistant
bacteria
uncommon
1950’s
Penicillin
resistant S.
aureus
common in
hospital
settings
1961 –
Methicillin
introduced
1962 –
Methicillin
resistant A.
aureus strains
begin to
appear
Superbugs from Super-fast Evolution
2002 –
Vancomycin
resistant strain
of S. aureus
isolated from
patient in
Michigan
Bacteria’s
evolutionary
advantages
 Vertical Transmission
 Generation time
 Population Size
 Horizontal Transmission
 No need to wait for
random mutations
 Multiple resistance genes
Mechanisms of
Antibiotic
Resistance
• Modification of the
Antibiotic
• Removal from the cell
• Modification of the target
site
The costs of Antibiotic
resistance
• 2 million yearly infections in the US
• 23,000 deaths directly related to
antibiotic resistant infections
• $20 billion lost to excessive healthcare
costs
• $35 billion due to lost productivity
Antibiotic Discovery tapers after 1960
Difficulty of new discovery combined with increasing antibiotic resistance results in potential public health crisis
iChip device for growing uncultured
microorganisms
Eleftheria terrae
• New species of b-proteobacteria
• New genus related to the genus Aquabacteria
• 16S sequencing
• DNA/DNA hybridization
Teixobactin
• Molecular Mass: 1,242 Da
• Depsipeptide containing
enduracididine,
methylphenylalanine and four Damino acids.
Image source: http://www.medscape.com/viewarticle/715971_2
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Resistance
• Teixobactin is
effective against
many gram
positive*
pathogens,
some of which
are
unresponsive to
vancomycin
(last resort)
Gram
negative
exception
20
Fig 2c
Image source:
https://commons.wikimedia.org/wiki/File:Vir
al_entry_(Endocytosis_and_lysis).svg 21
Resistance
• No mutants (S.aureus or M.tuberculosis) observed when plated on teixobactin –
NO RESISTANCE
• No S.aureus mutants observed, even after 27 day repeated exposure to
teixobactin – STILL NO RESISTANCE 
• Indicates “non – specific” mechanism & toxicity
• BUT no toxicity observed in mammalian cells (highest possible dose)
22
Mechanism of Action
• Teixobactin is non – haemolytic* and doesn’t bind DNA
• How did the research team figure out specifically where teixobactin is
active?
• Rate of label* incorporation into S.aureus’ main biosynthetic pathways
• Teixobactin inhibits peptidoglycan* synthesis
• Teixobactin has no observed effect on label incorporation into DNA, RNA, or protein
23
Fig 3a
24
Mechanism of Action
• In vancomycin, the absence of resistance suggests that the mechanism target is not
a protein. Could the same be true for teixobactin?
• Vancomycin binds lipid II* – does teixobactin bind this also?
• S.aureus built-up a peptidoglycan precursor (UDP-MurNAc-pentapeptide), when
exposed to teixobactin in concentrations of 1x – 5x the MIC*
• Vancomycin control does the same; indicates inhibition of a peptidoglycan biosynthesis step
• In test tubes, teixobactin blocked peptidoglycan synthesis reactions: lipid I, lipid II,
or undecaprenyl - pyrophosphate
25
Mechanism of Action
• What we know so far: Teixobactin is specifically involved with
peptidoglycan precursors, as opposed to the other enzymes involved in its
biosynthetic pathway
• The follow – up question: What specific part of these precursors is the is
teixobactin targeting?
• How to investigate: Study direct involvement with peptidoglycan
precursors coupled in pairs
26
• Paired peptidoglycan
precursors:
• purified and incubated with
teixobactin at various molar
ratios
• extracted and analyzed by thin –
layer chromatography
• Teixobactin bound if amount
Image Source:
http://www.waters.com/waters/en_US/HPLC---HighPerformance-Liquid-Chromatography-Beginner'sGuide/nav.htm?cid=10048919&locale=en_US
of lipid intermediates were
reduced
27
Mechanism of Action
• What did teixobactin bind? All peptidoglycan precursors and wall teichoic
acid*
• Fun fact! Lipid I and lipid II form a stable complex impermeable to the
effects of teixobactin
• Teixobactin > Vancomycin??
28
Mechanism of Action
• WTA is not needed for the organism to survive, inhibition of wall teichoic
acid biosynthesis has lethal toxic intermediates (late stage)
• Teichoic acids bind autolysins*; complex prevents uncontrolled
peptidoglycan break down.
• Inhibition of teichoic acid synthesis by teixobactin releases autolysins;
lytic/killing activity of the bacteria increases
29
In vivo efficacy
Teixobactin stable in
serum; low toxicity in
mice
Extended Data;
Fig 8a
30
In vivo efficacy
• Teixobactin also observed to be effective in mice with:
• MRSA* septicemia* dose of 90% lethality, if administered with as little as 1mg/kg
intravenously within an hour
• In a follow up experiment the PD50* was observed to be 0.2 mg/kg; better than
vancomycin* (PD50 of 2.75 mg/kg)
• Thigh model of MRSA infection*
• Streptococcus pneumonie infections
31
Summary
• Antibiotics are an invaluable medical advancement that have greatly reduced
suffering and death
• Bacteria have evolutionary advantages when it comes to evading antibiotic activity
• New techniques are necessary to advance the discovery of novel antibiotics
• Teixobactin is a strong and promising antibiotic with efficacy better than that of the
last resort antibiotic - Vancomycin
• Further research must be done, in terms of clinical trials – in vitro human trials?
32
References
• Clin Orthop Relat Res. 2005 Oct;439:23-6. THE CLASSIC: penicillin as a chemotherapeutic agent. 1940. Chain
E, Florey HW, Gardner AD, Heatley NG, Jennings MA, Orr-Ewing J, Sanders AG.
• Cell. 2007 Sep 7;130(5):797-810. A common mechanism of cellular death induced by bactericidal antibiotics.
Kohanski MA1, Dwyer DJ, Hayete B, Lawrence CA, Collins JJ.
• Lancet Infect Dis. 2010 Dec;10(12):816-8. doi: 10.1016/S1473-3099(10)70259-0. VRSA-doomsday superbug or
damp squib? Gould IM.
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