Tetracyclines glycicyclines macrolides and ketolides

1. PHAR 532
LECTURE 7
TETRACYCLINES, GLYCICYCLINES,
MACROLIDES, AND KETOLIDES
Antimicrobial agents and
Immunization

2. Tetracyclines
Tetracyclines are broad-spectrum agents, with
activity against both gram-positive and gram-
negative bacteria and intracellular chlamydiae,
mycoplasmas, and rickettsiae.
They show ability to treat parasitic infections such as
infection by Plasmodium falciparum, Giardia,
Trichomonas, and Toxoplasma.
Also used for the treatment of acne.
Treatment of protozoan infections and non-infecting
conditions such as acne could induce development of
resistance

3. Chemical name Generic name Trade name Yr of discovery Status Therapeutic
administration
7-Chlortetracycline Chlortetracycline Aureomycin 1948 Marketed Oral
5-Hydroxytetracycline Oxytetracycline Terramycin 1948 Marketed Oral and parenteral
Tetracycline Tetracycline Achromycin 1953 Marketed Oral
6-Demethyl-7-
chlortetracycline
Demethylchlortet
racycline
Declomycin 1957 Marketed Oral
2-N-
Pyrrolidinomethyltetracyclin
e
Rolitetracycline Reverin 1958 Marketed Oral
2-N-
Lysinomethyltetracycline
Limecycline Tetralysal 1961 Marketed Oral and parenteral
N-Methylol-7-
chlortetracycline
Clomocycline Megaclor 1963 Marketed Oral
6-Methylene-5-
hydroxytetracycline
Methacycline Rondomycin 1965 Marketed Oral
6-Deoxy-5-
hydroxytetracycline
Doxycycline Vibramycin 1967 Marketed Oral and parenteral
7-Dimethylamino-6-
demethyl-6-
deoxytetracycline
Minocycline Minocin 1972 Marketed Oral and parenteral
9-(t-butylglycylamido)-
minocycline
Tertiary-
butylglycylamido
minocycline
Tigilcycline 1993 Phase II clinical
trials

4. Structure of 6-deoxy-6-demethyltetracycline, the minimum tetracycline pharmacophore.
Ian Chopra, and Marilyn Roberts Microbiol. Mol. Biol. Rev.
2001;65:232-260

5. Tetracyclines mode of action
Tetracycline reversibly inhibits bacterial protein
synthesis by binding to the ribosomal complex,
preventing the association of aminoacyl-tRNA with
the bacterial ribosome.
 Each of the rings in the linear fused tetracyclic
nucleus must be six membered and purely
carbocyclic for the molecules to retain antibacterial
activity.

6. Resistance
Genes
Efflux
tet(A),tet(B), tet(C), tet(D),tet(E), tet(G), tet(H),tet(I), tet(J), tet (Z),tet(30) tet(31)
tet(K),tet(L)
otr(B),tcr3c
tetP(A)
tet(V)
tet(Y)d
Ribosomal protection
tet(M), tet(O),tet(S), tet(W)
tet(Q),tet(T)
otr(A),tetP(B),e
te tc
Emzymatic,tet(X)
Unknownf
tet(U),otr(C)

7. Resistance – efflux proteins
The efflux proteins are the best studied of the Tet
proteins. The genes encoding them belong to the
major facilitator superfamily (MFS), whose products
include over 300 individual proteins.
Export of tetracycline reduces the intracellular drug
concentration and thus protects the ribosomes
within the cell. Efflux genes are found in both gram-
positive and gram-negative species.

8. Resistance – efflux proteins
Each of the efflux genes codes for an approximately
46-kDa membrane-bound efflux protein. These
proteins have been divided into six groups based on
amino acid sequence identity.
Group 1 contains Tet(A), Tet(B), Tet(C), Tet(D),
Tet(E), Tet(G), Tet(H), Tet(Z), and probably Tet(I),
Tet(J), and Tet(30)

9. Efflux proteins

10. Resistance - ribosomal protection
Nine ribosomal protection cytoplasmic proteins
protect the ribosomes from the action of tetracycline
and confer resistance to doxycycline and
minocycline.
They confer a wider spectrum of resistance to
tetracyclines than is seen with bacteria that carry
tetracycline efflux proteins, with the exception of
Tet(B). The ribosomal protection proteins have
homology to elongation factors EF-Tu and EF-G.

11. Resistance - ribosomal protection
The Tet(M), Tet(O), and OtrA proteins reduce the
susceptibility of ribosomes to the action of
tetracyclines. The Streptomyces Otr(A) protein has
greatest overall amino acid similarity to elongation
factors.

12. The pathway of Tet(O)-mediated tetracycline release is illustrated by cryo-EM reconstructions
of ribosomes in various functional states (2, 45).
Sean R. Connell et al. Antimicrob. Agents Chemother.
2003;47:3675-3681

13. Tetracycline resistance
In the case of human pathogens, tetracycline
resistance is typically acquired via horizontal gene
transfer and occurs almost exclusively by ribosomal
protection or antibiotic efflux.
Both of these resistance mechanisms have their
evolutionary origins in the environment, but are now
found widely distributed in many commensal and
pathogenic bacteria.

14. TetX
Enzyme with activity that leads to destruction of
tetracycline.
Rare in clinical isolates but identified in samples in
2013 from Sierra Leone.
Possibly more prevalent in the environment than
previously thought.

15. Glycylcyclines
The glycylcyclines are synthetic analogues of the
tetracyclines.
The only antibiotic approved for clinical use is
tigecycline, a derivative of minocycline.
The glycylcyclines inhibit protein synthesis in a
manner similar to the tetracyclines but they
demonstrate more avid binding to the ribosome.
Tigecycline is active against a broad range of gram-
positive and gram-negative bacteria, including
strains resistant to the typical tetracyclines.

16. Tigecycline

17. Tigecycline
The large substitution on the D ring seem to be
enough to increase the spectrum of activity and
overcome tetracycline resistance mechanisms.

18. Macrolides and lincosamides (MLS)

19. MLS
The binding sites for these antibiotics are on the 50S
ribosomal subunit.
Transpeptidation (catalyzed by peptidyl transferase)
is inhibited by blocking the binding of the aminoacyl
moiety of the charged transfer RNA (tRNA) molecule
to the acceptor site on the ribosome-messenger
(mRNA) complex. Thus, the peptide at the donor site
cannot be transferred to its amino acid acceptor.

20. Date of download: 11/2/2016 Copyright © 2016 McGraw-Hill Education. All rights reserved.
Steps in bacterial protein synthesis and targets of several antibiotics. Amino acids are shown as numbered circles. The 70S ribosomal mRNA complex is shown with its 50S and 30S subunits. In step 1, the charged tRNA unit carrying amino acid 6 binds to
the acceptor site on the 70S ribosome. The peptidyl tRNA at the donor site, with amino acids 1 through 5, then binds the growing amino acid chain to amino acid 6 (transpeptidation, step 2). The uncharged tRNA left at the donor site is released (step 3), and
the new 6-amino acid chain with its tRNA shifts to the peptidyl site (translocation, step 4). The antibiotic-binding sites are shown schematically as triangles. Chloramphenicol (C) and macrolides (M) bind to the 50S subunit and block transpeptidation (step 2).
The tetracyclines (T) bind to the 30S subunit and prevent binding of the incoming charged tRNA unit (step 1). (Reproduced, with permission, from Katzung BG, editor: Basic & Clinical Pharmacology, 12th ed. McGraw-Hill, 2012: Fig. 44–1.)
Legend:
From: Chloramphenicol, Tetracyclines, Macrolides, Clindamycin, Streptogramins, & Linezolid
Katzung & Trevor's Pharmacology: Examination & Board Review, 11e, 2015
From: Chloramphenicol, Tetracyclines, Macrolides, Clindamycin, Streptogramins, & Linezolid
Katzung & Trevor's Pharmacology: Examination & Board Review, 11e, 2015

21. MLS
Spectrum of activity limited to gram-positive cocci
(mainly staphylococci and streptococci) and bacilli,
to gram-negative cocci, and intracellular bacteria
(Chlamydia and Rickettsia species).
Gram negative bacilli are generally resistant, with
some important exceptions (i.e., Bordetella
pertussis, Campylobacter, Chlamydia, Helicobacter,
and Legionella species).

22. Erytromycin
Erythromycin usually is bacteriostatic but may be
bactericidal in high concentrations against
susceptible organisms.

23. Resistance
Bacteria resist macrolide and lincosamide antibiotics
in 3 ways:
 (1) through target-site modification by methylation or
mutation that prevents the binding of the antibiotic to its
ribosomal target
 (2) through efflux of the antibiotic
 (3) by drug inactivation.

24. Resistance – target site modification
Methylation of the ribosomal target of the antibiotics
leads to cross-resistance to macrolides, lincosamides,
and streptogramins B: MLSB phenotype.
MLSB phenotype encoded by a variety of erm
(erythromycin ribosome methylase) genes.
In pathogenic bacteria, Erm proteins dimethylate a
single adenine in nascent 23S rRNA, which is part of
the large (50S) ribosomal subunit.

25. erm genes
Nearly 40 erm genes have been reported so far.
Four major classes are detected in pathogenic
microorganisms: erm(A), erm(B), erm(C), and erm(F).
erm(A) and erm(C) typically are staphylococcal gene classes.
erm(B) class genes are mostly spread in streptococci and
enterococci
erm(F) class genes in Bacteroides species and other anaerobic
bacteria.
In addition to the erm(B) genes, the ermTR genes, which are
now considered a subset of the erm(A) class on the basis of
sequence homology, can be detected in b-hemolytic
streptococci .
That each class is relatively specific—but not strictly confined
—to a bacterial genus, reflects easy gene exchange

26. Resistance – enzymatic inactivation
There are a number of inactivating enzymes that act
on the MLS antibiotics.
The genes encode esterases, hydrolases, glycosylases,
phosphotransferases, nucleotidyltransferases, and
acetyltransferases and are found less frequently than
efflux and ribosome-modifying genes in clinical
isolates.

27. Resistance – efflux pumps
In gram-negative bacteria, chromosomally encoded
pumps contribute to intrinsic resistance to
hydrophobic compounds, such as macrolides.
In gram-positive organisms, acquisition of macrolide
resistance by active efflux is caused by 2 classes of
pumps, members of the ATP-binding-cassette (ABC)
transporter superfamily and of the major facilitator
superfamily (MFS).

28. Resistance – efflux pumps
To date, the only efflux proteins conferring acquired
macrolide resistance characterized in
Staphylococccus species are ABC transporters
encoded by plasmid borne msr(A) genes.
The efflux system appears to be multicomponent in
nature, involving msr(A) and chromosomal genes to
constitute a fully operational efflux pump that has
specificity for 14- and 15-membered macrolides and
type B streptogramins (the MSB phenotype)

29. Enzymatic inhibition
Esterases act on 14- (e.g., erythromycin) and 15-
(e.g., azithromycin) membered macrolides;
Hydrolases affect streptogramin B drugs
Acetyltransferases inactivate streptogramin A
antibiotics
Nucleotidyltransferases confer resistance to the
lincosamides (e.g., clindamycin).
Phosphotransferases modify 14-, 15-, and 16-
membered macrolides with varying specificities and
telithromycin.