Download Novel Polymyxins for the treatment of serious Gram

Northern
Antibiotics
Vaara, Marttia,b, and Vaara, Timoa
a Northern
Antibiotics Ltd, Helsinki, Finland, and
b Division of Clinical Microbiology,
Helsinki University Hospital, Helsinki, Finland
Background: The emerging very multiresistant Gram-negative bacteria cause
remarkable therapeutic challenges. There are no novel classes of agents in
clinical development for the treatment of Gram-negative infections. Polymyxins
(polymyxin B, colistin) were abandoned in the seventies but are now back in
the therapy as the last resort. Their toxicity to renal proximal tubuli may
complicate the therapy or even necessitate its discontinuation. Less toxic
polymyxin derivatives would be very welcome.
Purpose: We wanted to develop polymyxin derivatives that differ from
polymyxin B and colistin (polymyxin E) in possessing no more than three (3)
positive charges under physiological conditions. Such derivatives can be
expected to have toxicological and pharmacokinetic properties different from
those of the old polymyxins.
This is a review on our results described in
• Vaara et al., AAC 2008,52:3229
• Ali et al., JAC 2009,64:1067
• Vingsbo Lundberg et al., JAC 2010,65:981
• Vaara et al., JAC 2010,65:942
• Vaara et al., AAC 2010,54:3341
• Vaara and Vaara, Peptides 2010; doi:10.1016/j.peptides.2010.09.010
Colistin [5]
MHA/MOA -Dab+ -Thr -Dab+ -cy[Dab -Dab+ -DLeu -Leu -Dab+ -Dab+ -Thr]
Polymyxin B [5] MHA/MOA -Dab+ -Thr -Dab+ -cy[Dab -Dab+ -DPhe -Leu -Dab+ -Dab+ -Thr]
NAB739 [3]
OA
- -Thr -DSer -cy[Dab -Dab+ -DPhe -Leu -Dab+ -Dab+ -Thr]
NAB7061 [3]
NAB741 [3]
OA
Ac
-
-Thr -Abu -cy[Dab -Dab+ -DPhe -Leu -Dab+ -Dab+ -Thr]
-Thr -DSer -cy[Dab -Dab+ -DPhe -Leu -Dab+ -Dab+ -Thr]
a
Highlighted parts indicate locations where the compounds are not identical to polymyxin B. Abbreviations for the nontrivial amino acyl residues: Dab, diaminobutyryl; Abu, aminobutyryl. Other abbreviations: MHA/MOA, the mixture of
methyl octanoyl and methyl heptanoyl; DA, decanoyl; OA, octanoyl; Ac, acetyl; cy, the cyclic portion indicated with
brackets. The positive charge of the free α- and γ-amino group is also shown. The net positive charge of the compound
is shown in brackets after the compound´s name.
18
16
14
Number of strains
Novel derivatives of polymyxin B,
a review on the NAB compounds
Table 1. Structure of colistin (polymyxin E) , polymyxin B, NAB739, NAB7061, and NAB741a.
12
10
8
6
4
Polymyxin B
2
NAB739
NAB7061
0
In addition, these results have been recently reviewed in
• Vaara, Curr Opin Pharmacol 2009,9:571
• Vaara, Curr Opin Microbiol 2010,13:574
0.125
0.25
0.5
1
2
4
MIC (µg/ml)
8
16
32
Results: The novel polymyxin derivatives NAB739, NAB7061 and NAB741 have
their cyclic part identical to that of polymyxin B, but their side chain consists of
octanoyl-threonyl-D-serinyl, octanoyl-threonyl-aminobutyryl, and acetylthreonyl-Dserinyl, respectively (Table 1). Accordingly, their linear part lacks the
two (2) diaminobutyryl (Dab) residues present in the linear part of polymyxin B
and colistin.
Figure 1. Cumulative MIC values (µg/ml) of polymyxin B, NAB739 and NAB7061
for 17 strains of E. coli, as determined by the agar dilution method according to
CLSI. Note the marked difference between NAB739 and NAB7061, compounds
that are identical except that the second amino acyl residue in the linear peptide
part is DSer in NAB739 and Abu (aminobutyryl) in NAB7061.
The MICs of NAB739 and polymyxin B for seventeen (17) strains of E. coli are
shown in Figure 1. The MIC90 of both drugs is 1 µg/ml. The MIC90 of NAB739
and polymyxin B for other polymyxin-susceptible Enterobacteriaceae (n=12)
are 2 µg/ml and 1 µg/ml, respectively. The MIC range of NAB739 and
polymyxin B for the polymyxin-susceptible carbapenemase-producing strains of
E. coli and Klebsiella pneumoniae (n=9, including KPC, OXA-48, VIM, and IMPproducing strains) is 1-4 µg/ml and 1-2 µg/ml, respectively.
Table 2. Susceptibilities of Gram-negative bacteria to rifampin and
clarithromycin in the presence of NAB7061 [Vaara et al. AAC 52:3229].
The MIC for Acinetobacter baumannii (n=3) is 2-8 µg/ml. At the subinhibitory
concentration of 1 μg/ml, NAB739 decreases the MIC of clarithromycin,
rifampin, vancomycin, and fusidic acid for A. baumannii (n=2) by factors of 4885, 670-750, >256, and >256, respectively. For Pseudomonas aeruginosa (n=1),
the MIC is 8 µg/ml.
NAB7061 practically lacks intrinsic antibacterial activity (Figure 1) but strongly
synergizes the activity of antibiotics towards Gram-negative pathogens,
including antibiotics regarded as exclusively anti-Gram-positive drugs such as
rifampin, macrolides, fusidic acid and vancomycin. At 4 μg/ml, NAB7061
decreases the MIC of rifampin for E. coli (n=11), other polymyxin-susceptible
Enterobacteriaceae (n=12), and A. baumannii (n=3) by factors of 85-750 and
10-2000, and 25-125, respectively (Table 2). With clarithromycin, the
corresponding factors are 90->750, 10-1000, and 40-100, respectively. The
antibacterial properties of NAB741 are similar to those of NAB7061.
NAB739 is effective in treating experimental E. coli peritoneal infection in mice.
The single dose of 1 mg/kg per body weight decreased the bacterial count in 6
hours by three log10 from the initial level and prevented the appearance of
clinical symptoms. NAB7061 is effective when combined with erythromycin.
The affinity of the NAB compounds for isolated rat kidney brush border
membrane (BBM) is only approx. one-seventh (NAB739) or one-fifth
(NAB7061) that of polymyxin B and approx. one-third that of gentamicin
(Table 3).
The renal clearances of NAB741, NAB739, and NAB7061 are approx. 400, 50
and 30-fold higher than that of colistin (Table 4). This indicates that the
positively charged Dab residues in the side chain of the old polymyxins play an
important role in the reuptake of polymyxins by the proximal tubuli.
Conclusions: The MIC of NAB739 for Enterobacteriaceae and A. baumannii is
identical or very close to that of polymyxin B. NAB7061 and NAB741 lack direct
activity but strongly synergize the activity of several other antibiotics against
these bacteria. The observed differences in binding to BBM and in renal
handling suggest that NAB739 and the other tricationic NAB compounds are
less nephrotoxic than the pentacationic polymyxins. Several NAB compounds
are now in preclinical studies.
Materials and Methods: Polymyxin derivatives were synthesized by
conventional solid phase chemistry, using the standard Fmoc protection
strategy. For further information on this as well as for the other methods used,
see the individual publications listed above.
Acknowledgements: We thank our collaborators J. Fox (Univ. Birmingham,
United Kingdom), G. Loidl (Bachem AG, Bubendorf, Switzerland), J. Apajalahti
and O. Siikanen (Alimetrics Oy, Espoo, Finland), J. Nagai and M. Takano (Univ.
Hiroshima, Japan), F. Hansen, C. Vingsbo Lundberg and N. Frimodt-Møller
(Statens Serum Institute, Copenhagen, Denmark), and F.E. Ali, G. Cao, H. He, J.
Li, R.L. Nation, and A. Poudyal (Monash University, Melbourne, Australia).
Sensitization factorb to
Rifampin
Clarithromycin
85 - 750
90 - >750
10 - 500
10 - 170
680 - 2000
250 - 260
500 - 1500
85 - 1000
500 - 1000
250
24 - 125
40 - 100
2
<2
Speciesa
E. coli (11)
K. pneumoniae (5)
K. oxytoca (2)
E. cloacae (4)
C. freundii (1)
A. baumannii (3)
P. aeruginosa (1)
a
The number of strains tested shown in parentheses.
b
Ratio of the MIC in the absence of NAB7061 to that in the presence of 4 μg of NAB7061/ml.
Table 3. Affinity of polymyxin B, gentamicin, NAB739 and NAB7061 to the rat
brush border membrane (BBM) of the renal cortex [Vaara et al. AAC 52:3229].
Polymyxin B
Gentamicin
NAB739
NAB7061
a
ID50 in gentamicin
displacement assay
39 ± 6
90 ± 10
264 ± 91
187 ± 24
Relative
affinity to BBMa
1.00
0.43
0.15
0.21
Calculated from the ID50 values by setting the relative affinity of polymyxin as 1.00.
Table 4. Basic pharmacokinetic parameters of NAB739, NAB7061, and NAB741, as
compared to those of colistin (mean ± SD) [Ali et al. JAC 64:1067 and Vaara et al. AAC
54:3341].
Parameter
Half-life (min)
Volume of distribution (ml/kg)
Clearance (ml/min/kg)
Renal clearance (ml/min/kg)
Urinary recovery (% of dose)
Colistin
NAB739
NAB7061
NAB741
74.6 ± 13.2
496 ± 60
5.22 ± 0.41
0.010 ± 0.008
0.18 ± 0.14
69.0 ± 21.9
222 ± 20.5
2.63 ± 0.54
0.53 ± 0.30
19.4 ± 7.38
66.2 ± 12.3
339 ± 96
3.84 ± 0.75
0.28 ± 0.16
7.16 ± 3.70
32.7 ± 2.41
243 ± 24.0
7.39 ± 0.85
3.78 ± 1.11
50.9 ± 13.6
Contact: Professor Martti Vaara, M.D., Ph.D., CEO, CSO
Northern Antibiotics Ltd
Mailing address: P.O.Box 30, FI-00721, Helsinki, Finland
Visiting address: Eskolantie 1, FI-00720, Helsinki, Finland
Phone: +358-40-355 0822
Fax: +358-9-6842 0130
[email protected]
www.northernantibiotics.com