Antibiotic Resistance Acquisition · Antibiotic Resistance Acquisition Paul M. Tulkens, MD, PhD....

14/06/2014 XXXth Rocourt Neonatology Meeting 1

Antibiotic Resistance Acquisition

Paul M. Tulkens, MD, PhD

Cellular and Molecular Pharmacology & Center for Clinical Pharmacy Louvain Drug Research Institute, Université catholique de Louvain

Brussels, Belgium

Saturday June 14th, 2014 - Cercle de Wallonie Liège - Esplanade du Val, Seraing


Disclosures

Financial support from

• the Belgian Fonds de la Recherche Scientifique for basic research on pharmacology antibiotics and related topics

• Université catholique de Louvain for past personal support

• Commercial Relationships:– AstraZeneca, GSK, Sanofi-Aventis, Bayer HealthCare, Cempra

Pharmaceuticals, The Medicines Company, Northern Antibiotics, RibX, Cubist, Galapagos, …

• Other relationships in relation to this talk– Belgian Antibiotic Policy Coordination Committee,

– European Medicines Agency (as expert for the agency and for Industry)

– European Commmittee for Antibiotic Susceptibility Testing (EUCAST)

Slides: http://www.facm.ucl.ac.be Lectures


Do we have a problem ?

This man discovered the mode of action of penicillin

and died from invasive pneumococcal infection …

http://www.cip.ulg.ac.be/newsite/pdf/jmghuysen.pdf

http://www.cip.ulg.ac.be/newsite/pdf/jmghuysen.pdf


But what about this patient ?

Gram stain of Staphylococcus aureus in pustular exudatehttp://www.tjclarkdirect.com/bacterial_diseases/staphylococcus.htm

Last accessed: 10/06/2014

Etok et al. Aetiology and antimicrobial studies of surgical wound infections in University of Uyo

Teaching Hospital (UUTH) Uyo, Akwa Ibom State, Nigeria. 1:341. doi:10.4172/scientificreports.341

http://www.tjclarkdirect.com/bacterial_diseases/staphylococcus.htm


Which problem ?

wild type


Which problem ?

resistant bacteria

elimination

active effluximpermeabilisation

wild type

attack

inactivation of the antibiotic

(biotransformation)

avoidance

modification of the target

way around

alternativetargetor target multiplication

no more active antibiotic ! "useless" antibiotic "overwhelmed" antibiotic

antibiotic absent or in insufficient concentration


But be aware ! Several mechanisms may coexist !elimination


wild type

attack


(biotransformation)

avoidance


way around


multiresistantbacteria


Which problem ?

resistant bacteria

elimination


wild type

attack


(biotransformation)

avoidance


way around





Attack: the example of aminoglycosides

Van Bambeke et al. Mechanisms of Action. In: Infectious Diseases (3d edition; J. Cohen,

W. Powderly & S. Opal, eds), chapter 130, pp 1288-1307, Elsevier/Mosby, 2010


Aminoglycoside resistance in Pseudomonas aeruginosa: an example in Belgium for patients with nososomial pneumonia

0.015

625

0.031

250.0

625

0.125 0.2

5 0.5 1 2 4 8 16 32 64 128

256

512

0

25

50

75

100

gentamicin

MIC (µg/mL)

cum

ulat

ive

perc

enta

ge (%

)

0.015

625

0.031

250.0

625

0.125 0.2

5 0.5 1 2 4 8 16 32 64 128

256

512

0

25

50

75

100amikacin

MIC (µg/mL)

cum

ulat

ive

perc

enta

ge (%

)

Data from Riou et al. Int J Antimicrob Agents. 2010 Dec;36(6):513-22

EUCAST ( > )

CLSI ( ≥

)R breakpoint


Aminoglycoside resistance in Pseudomonas aeruginosa: the situation may be worse elsewhere

European Centre for Disease Prevention and Control. Antimicrobial resistance surveillance in Europe 2012. Annual Report of the European Antimicrobial Resistance Surveillance Network (EARS-Net). Stockholm: ECDC; 2013.


And co-resistance in Pseudomonas aeruginosa is frequent

European Centre for Disease Prevention and Control. Antimicrobial resistance surveillance in Europe 2012. Annual Report of the European Antimicrobial Resistance Surveillance Network (EARS-Net). Stockholm: ECDC; 2013.


Aminoglycosides: can we do something ?

OO

O

CH2H2N

NH2

OH

HNH3C

CH3OH

HO

H2NO

NH2

Plazomicin (ACHN-490): made from sisomicin

OO

O

CH2H2N

NH2

OH

HNH3C

CH3OH

HO

HNO

HN

HOHCNH2

OH

plazomicin

sisomicin

still susceptible to AAC(2’)

(but rare enzyme)


Plazomycin future ?

Potential to demonstrate a mortality benefit over currently available therapy in the treatment of life-threatening CRE infections.

"We have designed our pivotal Phase 3 trial for plazomicin as a superiority trial with a primary efficacy endpoint of all-cause mortality at 28 days. The trial will compare a plazomicin-based regimen versus a colistin-based regimen for the treatment of CRE bloodstream infections and pneumonia."

http://www.achaogen.com/plazomicin/Last visited: 10/06/2014

http://www.achaogen.com/plazomicin/


Attack: the example

of the -lactamases

(part 1)

Van Bambeke et al. Mechanisms of Action. In: Infectious Diseases (3d edition; J. Cohen, W. Powderly & S. Opal, eds), chapter 130, pp 1288-1307, Elsevier/Mosby, 2010


Attack: the example

of the -lactamases

(part 2)



-lactamases: why so many ?



-lactamases and PBPs may be very close


and -lactamases are often in mobile, higly transmissible genetic elements (together with other resistance genes)



Schematic representation of blaNDM - associated genetic structures identified among Gram-negative clinical isolates.

(a) Structure found in A. baumannii (part of the composite transposon Tn125).

(b) Structures found in Enterobacteriaceaeand P. aeruginosa where ISAba125 is presented as full or truncated element with bleMBL gene also beingpresent as full or truncated gene.


Which problem ?

resistant bacteria

elimination


wild type

attack


(biotransformation)

avoidance


way around





this is where we have a problem

amoxicilin vs.S. pneumoniae (n = 136)

3.9×10

-030.0

0781

30.0

1562

60.0

3125

0.062

50.1

25 0.25 0.5 1 2 4 8 16 32

0

25

50

75

100

MIC50

MIC90

MICs (mg/L)

% o

f str

ains

(cum

ulat

ive)

Amoxicllin and Streptococcus pneumoniae

EUCAST: European Committee on Antimicrobial Susceptibility Testing (http://www.eucast.org) MIC: minimum inhibitory concentrationCAP: community-acquired pneumoniaCOPD: chronic obstructive pulmonary disease

EUCAST wild type

population

• Belgian isolates collected between 2009 and 2012 from patients with confirmed cases of CAP

• the high MICs of amoxicillin is driven by isolates from patients with past COPD

Tulkens, unpublished

http://www.eucast.org/


But which breakpoints do we need to use ?

Good Evil

To be honest, I always wondered ...


amoxicllin vs.S. pneumoniae (n = 136)

3.9×10

-030.0

0781

30.0

1562

60.0

3125

0.062

50.1

25 0.25 0.5 1 2 4 8 16 32

0

25

50

75

100

MIC50

MIC90

MICs (mg/L)

% o

f str

ains

(cum

ulat

ive)

MIC distribution is a continuous variable…

EUCAST wild type

population

EU clinical breakpoints

S ≤

0.5 – R > 2 *

EUCAST: European Committee on Antimicrobial Susceptibility Testing (http://www.eucast.org) MIC: minimum inhibitory concentrationCAP: community-acquired pneumoniaCOPD: chronic obstructive pulmonary dosease

* non-meningitis






amoxicllin vs.S. pneumoniae (n = 136)

3.9×10

-030.0

0781

30.0

1562

60.0

3125

0.062

50.1

25 0.25 0.5 1 2 4 8 16 32

0

25

50

75

100

MIC50

MIC90

MICs (mg/L)

% o

f str

ains

(cum

ulat

ive)

MIC distribution is a continuous variable…

EUCAST wild type

population

EU clinical breakpoints

S ≤

0.5 – R > 2 *

CLSI clinical breakpoints

S ≤

2 – R ≥

8 *

CLSI: Clinical and Laboratory Standards Institute (http://clsi.org) EUCAST: European Committee on Antimicrobial Susceptibility Testing (http://www.eucast.org) MIC: minimum inhibitory concentrationCAP: community-acquired pneumoniaCOPD: chronic obstructive pulmonary disease

* non-meningitis* non-meningitis




http://clsi.org/



EUCAST calculations of target attainment rate for amoxicillin against S. pneumoniae

0.5 1 2 4 8 16 320

25

50

75

100

0.5 g 3x 1g 3x 2g 4x

MIC

targ

et a

ttai

nmen

t rat

e (%

)

By increasing the dose and multiplying the number of daily administration, you may cover bacteria with MIC up to 8 mg/L…

but the total daily dose will be very high…

* for f T >MIC = 40%

Graph prepared from data in http://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Rationale_documents/Amoxicillin_rationale_Nov2010_v_1.0.pdf

90 %

http://www.eucast.org/fileadmin/src/media/PDFs/EUCAST_files/Rationale_documents/Amoxicillin_rationale_Nov2010_v_1.0.pdf


And chlidren may harbour more resistant organisms

Diekema et al. Int. J. Antimicrob. Agents 2002; 412-418* longitudinal surveillance program designed to track antimicrobial resistance trends nationally

and internationally over a 5- to 10-year period and sponsored by Bristol-Myers Squibb


Staphylococcus aureushttp://www.microbewo rld.org/index.php?opti on=com_jlibrary&vie w=article&id=7611

• Nosocomial pneumonia involving hospital-acquired (HA) S. aureus is becoming increasingly frequent 1,2

• In parallel, pneumonia caused by community-acquired (CA) MRSA while remaining rare in Europe2 are becoming common in several other parts of the world including Asia 3

• As many strains (even MSSA) produce toxins, they cause major tissue damage, and, hence a high mortality 3,4,5

1. Jones, Clin Infect Dis. 2010;51(suppl 1):S81-72. Valour, et al Rev Pneumol Clin. 2013;69:368-82 3. Karampela, et al. Minerva Anestesiol. 2012 Aug;78(8):930-40

Kang & Song. Infect Chemother. 2013;45:22-314. Papazian & Donati. Nosocomial pneumonia. In Infectious Diseases, 3rd Edition,

Cohen, Powderly & Opal, eds. Elsevier (available on line at http://www.expertconsultbook.com ; last visisted: 4 April 2014)

5. Catena, et al Infez Med. 2012;20:205-10 /.MRSA methicillin-resistant Staphylococcus aureusMSSA methicillin-sensitive Staphylococcus aureus

http://www.microbeworld.org/index.php?option=com_jlibrary&view=article&id=7611

http://www.expertconsultbook.com/


S. aureushttp://www.microbewo rld.org/index.php?opti on=com_jlibrary&vie w=article&id=7611

• Nosocomial pneumonia involving hospital-acquired (HA) S. aureus is becoming increasingly frequent [1,2]

• In parallel, pneumonia caused by community-acquired (CA) MRSA while remaining rare in Europe [2] are becoming common in several other parts of the world including Asia [3]

• As many strains (even MSSA) produce toxins, they cause major tissue damage, and, hence a high mortality [3,4]

1. Jones Clin Infect Dis 2010;51(suppl 1):S81-72. Valour et al Rev Pneumol Clin. 2013;69:368-82 3. Karampela et al Minerva Anestesiol. 2012 Aug;78(8):930-40 / Kang & Song Infect Chemother 2013;45:22-314. Papazian & Donati Nosocomial pneumonia. In Infectious Diseases, 3rd Edition, Cohen, Powderly & Opal, eds. Elsevier (available on line at

http://www.expertconsultbook.com (Last visisted: 4 April 2014) / Catena et al Infez Med. 2012;20:205-10 /.

Valour, et al Rev Pneumol Clin. 2013;69:368-82

"S. aureus accounts for 2 to 5% of the

etiologies of community-acquired

pneumonia"

"S. aureus represents 20 to 30% of cases of

hospital-acquired pneumonia, including ventilator-associated

pneumonia"




1. Jones Clin Infect Dis 2010;51(suppl 1):S81-72. Karampela et al Minerva Anestesiol. 2012 Aug;78(8):930-40 / Kang & Song Infect Chemother 2013;45:22-313. Papazian & Donati Nosocomial pneumonia. In Infectious Diseases, 3rd Edition, Cohen, Powderly & Opal, eds. Elsevier (available on line at

http://www.expertconsultbook.com (Last visisted: 4 April 2014) / Catena et al Infez Med. 2012;20:205-10 / Valour et al Rev Pneumol Clin. 2013;69:368-82.

S. aureushttp://www.microbewo rld.org/index.php?opti on=com_jlibrary&vie w=article&id=7611

• Nosocomial pneumonia involving hospital-acquired (HA) S. aureus is becoming increasingly frequent [1]

• In parallel, pneumonia caused by community-acquired (CA) MRSA while rare in Europe are becoming common in several other parts of the world including Asia [2]

• As many strains (even MSSA) produce toxins, they cause major tissue damage, and, hence high mortality [2,3]

Jones, et al. Clin Infect Dis. 2010;51(suppl 1):S81-7

HABP: hospital-acquired bacterial pneumoniaVABP: ventilator-associated bacterial pneumoniaMRSA: methicillin resistant Staphylococcus aureus




MRSA in AsiaPrevalence of methicillin resistance among S. aureus isolates.Some Asian countries have shown the highest prevalence rates of MRSA

< 1% 1-10% 20-25% 25-50% > 50%

Sri Lanka Singapore

Hong Kong

Taiwan

Japan

Korea

Kang & Song. Infect Chemother 2013;45:22-31MRSA methicillin restistant Staphylococcus aureus


Could ceftaroline (recently approved) be

a solution ?

An allosteric mechanism !

Otero et al. Proc Natl Acad Sci USA. 2013 Oct 15;110(42):16808-13.

Allosteric site


CEFTAROLINE: MICs

S. aureus (all; n = 240)

0.031

250.0

625

0.125 0.2

5 0.5 1 2 4 8

0

25

50

75

100

vancomycin linezolidceftaroline

MIC90

MIC50

MICs (mg/L)

N S

trai

ns (c

umul

ativ

epe

rcen

t)

MSSA (n = 83)

0.031

250.0

625

0.125 0.2

5 0.5 1 2 4 8

0

25

50

75

100 MIC90

MIC50

MICs (mg/L)

N S

trai

ns (c

umul

ativ

epe

rcen

t)

S.aureus MIC distributions

MRSA (n = 157)

0.031

250.0

625

0.125 0.2

5 0.5 1 2 4 8

0

25

50

75

100 MIC90

MIC50

MICs (mg/L)

N S

trai

ns (c

umul

ativ

epe

rcen

t)* isolates collected o, Belgium between 2011 and 2012 from patients suffering of wound infections in 3 hospitals (1 in South-East of Brussels; 1 in North of Brussels; 1 in Hainaut)

*

Tulkens et al. 26th ICC, 2013 and unpublished

EUCAST breakpoints of ceftaroline, vancomycin and linezolid


Which problem ?

resistant bacteria

elimination


wild type

attack


(biotransformation)

avoidance


way around





The VISA story…

• VISA stands for "Vancomycin Intermediate Staphylococcus Aureus" (but also termed GISA (Glycopeptide-Intermediate Staphylococcus Aureus) and denotes organisms with an increased MIC for vancomycin or teicoplanin

• First identified in Japan in 1997 but since then found in many other countries

• Resistance occurs by a tickening of the cell wall with increased amounts of free D-Ala-D-Ala termini that trap vancomycin (and glycopeptides).


The VISA story…

• The MICs of these strains are above the susceptibility breakpoint of EUCAST (2 mg/L) !

Howden et al. Clin Microbiol Rev 2010;23:99–139.


Do you need to be afraid of VISA ?


VISA and co-resistance: the daptomycin problem


VISA and co-resistance: the daptomycin problem

daptomycin EUCAST breakpoint


Which problem ?

resistant bacteria

elimination


wild type

attack


(biotransformation)

avoidance


way around





An original observation with cancer cells…


Historical observations on tetracyclines …


Historical observations on tetracyclines …

McMurry et al., PNAS 1980; 77:3974-3977

Everted membranes

Whole bacteria


You said "antibiotic eflux"

1960 1970 1980 1990 2000 20100

250

500

750

1000

1250

1500

1750

Year

No.

of p

ublic

atio

ns /

2 ye

ars

No. of publications in PubMed with keywords: "antibiotic AND (efflux OR transporter)"


1960 1970 1980 1990 2000 20100

250

500

750

1000

1250

1500

1750

Year

No.

of p

ublic

atio

ns /

2 ye

ars

Historical landmarks …

Successive description of efflux- mediated resistance for major classes of antibiotics

tetracyclines

-lactams

fluoroquinolones

macrolides

linezolid

rifampin

aminoglycosides

daptomycin ** in eucaryotic cells

only (so far)


RoleRole of efflux of efflux pumpspumps in the in the clinicsclinics ……


Efflux in S. pneumoniae: is it important in the clinics ?

Suspected efflux based on phenotypic analysis (CIP MIC +/- reserpine)

CAP BPCO0

20

40

60

80

100 1 2 2

reserpine effect on MIC (x dilutions)

origin of isolates

% s

trai

ns

Lismond & Degives, unpublished

acute pathology

« one shot »antibiotic exposure

chronic pathology

repetitiveantibiotic exposures

183 strains 107 strains


Efflux in S. pneumoniae: is it important in the clinics ?

Identification of FQ transporters in clinical isolates

Lismond et al, ECCMID 2010

Inactivation of patA or patB as efficient as reserpine to

reduce MIC

• responsible for FQ efflux in clinical isolates

• work as heterodimers


Efflux in P. aeruginosa: is it important in the clinics ?

Prevalence of MexA and MexX overexpressers in 62 phylogentically-related pairs of P. aeruginosa isolated from ICU patients (VAP)

DAY x (%)

38.71%

22.58%

20.97%

17.74%

DAY 0 (%)

66.13%

12.90%

11.29%

9.68%

MexA-/MexX-

MexA+/MexX-

MexX+/MexA-

MexA+/MexX+

Riou et al, ECCMID 2010


amikacin (n=29)

D0 DL1

2

4

8

16

32

64

128

256

a

meropenem (n=28)

D0 DL0.125

0.25

0.5

1

2

4

8

16

32

64

128

256

*

piperacillin-tazobactam (n=31)

D0 DL

2

4

8

16

32

64

128

256

512

1024

*

cefepime (n=29)

D0 DL0.5

1

2

4

8

16

32

64

128

256

512

a

ciprofloxacin (n=11)

D0 DL0.015625

0.03125

0.0625

0.125

0.25

0.5

1

2

4

8

16

32

64

128

MIC

(mg/

L)

Emergence of resistance

during treatment

Riou, et al. Int J Antimicrob Agents. 2010;36:513-22

initial

isolate

Last

P. aeruginosa successive clonalisolates from the same patient(all patients treated with large doses of 1 to 3 antibiotics)

- D0: initial isolate DL: last isolate obtained

- individual values with geometric mean (95 % CI)

- S (lowest line) and R (highest line) EUCAST breakpoints

* p < 0.05 by paired t-test (two- tailed) and Wilcoxon non- parametric test

a p < 0.05 by Wilcoxon non- parametric test only

Note: stratification by time between D0 and DL gave no clue (too low numbers)


Is this all ?

• Phenotypic "resistance"– small colony variants– dormant/persistent bacteria

http://www.facm.ucl.ac.be/intracellular_chemotherapy.htm Last visited: 10/06/2014

http://cmr.asm.org/content/15/2/167.figures-onlyRodney & Costerton Clin. Microbiol. Rev. 2002, 15(2):167.

http://infekt.ch/2006/10/small-colony-variants-von-staphylococcus- aureus-schwierig-zu-behandelnde-infektionen/Last visited 14/06/2014

http://www.facm.ucl.ac.be/intracellular_chemotherapy.htm

http://cmr.asm.org/content/15/2/167.figures-only

http://infekt.ch/2006/10/small-colony-variants-von-staphylococcus-aureus-schwierig-zu-behandelnde-infektionen/

http://infekt.ch/2006/10/small-colony-variants-von-staphylococcus-aureus-schwierig-zu-behandelnde-infektionen/


Intracellular infection and S. aureus

Mélard et al. J Antimicrob Chemother. 2013;68:648-58

-4 -3 -2 -1 0 1 2 3-6

-5

-4

-3

-2

-1

0

1

2

3

MIC = 0.25 mg/L (n=1)

MIC = 0.5 mg/L (n=3)

MIC = 1 mg/L (n=2)

Broth

MIC = 0.125 (n=1)MIC = 2 mg/L (n=4)

lo

g cf

u fr

om ti

me

0 (2

4 h)

-3 -2 -1 0 1 2 3-1

0

1

2

3

THP-1

-4 -3 -2 -1 0 1 2 3 4

THP-1Broth

-6

-5

-4

-3

-2

-1

0

1

2

3

log cfu from tim

e 0 (24 h)

log10 concentration (mg/L) log10 concentration (x MIC)

Activity of ceftaroline towards extracellular (broth) and intracellular forms of S. aureus with increasing MICs


Biofilms and S. pneumoniae

0

20

40

60

80

100

120

day 2day 11

0

20

40

60

80

100

120

naïve model

viability biomassATCC 49619

Perc

enta

ge o

f con

trol

val

ue

Log concentration (x MIC)

Vandevelde et al. Antimicrob Agents Chemother. 2014;58:1348-58.

0

20

40

60

80

100

120

LVX

LVXMXF

MXF

day 2

day 11

0

20

40

60

80

100

120

naïve modelPerc

enta

ge o

f con

trol

val

ue

v iability biomassATCC 49619

Log concentration (x MIC)


The question that I should have addressed…

• How does all that is acquired and spread ?

overexpression"let cook it first"and serve to those who can pay

existing genomic

informationhorizontal transfer "ready to eat"

for everyone

mutation selection "only the best will get it"

metabolic adaptation

stay in your niche


The question that I should have addressed…

overexpression"let cook it first"and serve to those who can pay

existing genomic

informationhorizontal transfer "ready to eat"

for everyone

mutation selection "only the best will get it"

metabolic adaptation

stay in your niche

but what can I do ?

hygieneisolation

reduce current antibiotic pressure

don't give a chance to selection

use "non-antibiotic"options


The real question …

What can do for him/her to have nice dreams … and to wake up in good health ?

Antibiotic Resistance Acquisition · Antibiotic Resistance Acquisition Paul M. Tulkens, MD, PhD....

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Transcript of Antibiotic Resistance Acquisition · Antibiotic Resistance Acquisition Paul M. Tulkens, MD, PhD....