chemotherapy of microbial diseases 殷明 2011.11. shanghai jiao tong university 1940...

Chemotherapy of microbial diseases

殷明2011.11

Shanghai Jiao Tong University

1940 年前，小儿肺炎、痢疾、伤寒、斑疹伤寒、鼠疫、霍乱、白喉、疟疾——不治之症

1930-1949 上海传染病死亡率 565/10 万

1990 0.52/10 万

1949 年，中国人平均寿命 39 岁

2004 年，中国人平均寿命 72岁（日本 82岁）

一些事实数据


GENERAL PRINCIPLES OF ANTIMICROBIAL THERAPY


Introduction

Mechanism of action

Antibiotic Resistance

General principles of clinical use

Shanghai Jiao Tong UniversityDefinition

In the strictest sense, antibiotics are antibacterial substances produced by various species of microorganisms (bacteria, fungi, and actinomycetes) that suppress the growth of other microorganisms.

Common usage often extends the term antibiotics to include synthetic antimicrobial agents, such as sulfonamides and quinolones.

Shanghai Jiao Tong Universityclassification

according to the actions against the type of organism

Antibacterial

Antiviral

Antifungal


1928: Alexander Fleming discovered the first antibiotic.

He observed that Penicillium fungus made an antibiotic, penicillin, that killed S. aureus.

Two Oxford University specialists Dr Howard W Florey and his German associate Dr Ernest Boris Chain spent month after month attempting to extract the essential substance from Fleming's mould.

On 12 February, 1941, the first dose of isolated penicillin was injected into a human being

Nobel Prize for Medicine in 1945

The discovery of Penicillin

Shanghai Jiao Tong University Antibiotics

Most significant discovery of modern medicine

Save millions of lives


Academic terms

Antimicrobial spectrum

Bactericidal-MBC, bacteriostatic-MIC

Therapeutic index: LD50/ED50

Post antibiotic effect


Concept 1: antibiotic spectrum

Different antibiotics target different kinds of bacteriaAntimicrobial spectrum

Antimicrobial spectrum of a drug means the species of microorganisms that the drug can inhibit or kill.Narrow spectrum

The agents act against a single or limited group of microorganisms (isoniazid: mycobacteria ) Extended spectrum

The agent that are effective against gram-positive organisms and also against a number of gram-negative bacteria (ampicillin).Broad spectrum

The agent affect a wide variety of microbial species (tetracycline)


Concept 2: Bacteriostatic vs. Bactericide

Antibiotics differ by mode of action

Bacteriostatic: arrest the growth or replication of the microorgani

sm. Sulfanilamide

Bactericide: The agents which can kill the microorganisms are call

ed bactericidal drugs; a drug may be bacteriostatic for one organism but bactericidal for another.

Fluoroquinolones, Penicillins, …

Shanghai Jiao Tong UniversityConcept 3:

Bactericidal-MBC, bacteriostatic-MIC

Chemotherapy index (CI) To evaluate the safety of chemotherapeutic drugs, the value is LD50/ED50

Minimal inhibitory concentration (MIC) MIC is the lowest concentration of antimicrobial agents that prevents visible growth in 18-24 hours incubation.


Concept 4: Post antibiotic effect

The antibiotic inhibits microbial multiplication beyond the time when the MIC is reached in plasma.• Aminoglycosides• Macrolides

Allows reduction in dosing schedule


Mechanism of action


Inhibitors of cell wall synthesisAgents that interfere with cell membrane permeabilityInhibitors of nucleic acid synthesis or activityInhibitors of protein synthesisInhibitors of bacterial metabolism…

CLASSES OF ANTIBIOTICS

Shanghai Jiao Tong University Antimicrobial site of action

Shanghai Jiao Tong UniversityCell Wall Synthesis Inhibitors

Penicillins 青霉素Cephalosporins

头孢菌素Vancomycin 万古霉素 Bacitracin 杆菌肽

β-lactam antibiotics


Peptidoglycan （肽聚糖） is a 3D molecule

Cross links are both horizontal and vertical between glycan chains stacked atop one another.


Gram negative cell structure


Antimicrobials acting on the bacterial cell wall

Interfere with synthesis of peptidoglycan layer in cell wall

• eventually cause cell lysis

• bind to and inhibit activity of enzymes responsible for peptidoglycan synthesis

• “penicillin-binding proteins”(transpeptidase)

Shanghai Jiao Tong Universityβ-Lactams 内酰胺

β-lactams inhibit transpeptidase.Only effective against rapidly growing organisms that synthesize peptidoglycan. The size, charge and hydrophobicity of the molecule determines the extent of its antibacterial activity.


There is no molecule similar to peptidoglycan in humans, making drugs that target cell wall synthesis very selective in their toxicity against bacteria.


Antimicrobials acting on the cell membrane

amphotericin 两性霉素 : binding to the sterol-containing membranes of fungipolymyxins 多粘菌素 : acting like detergents and disrupt the Gram negative outer membrane.• Not used parenterally because of toxicity

to mammalian cell membranefluconazole and itraconazole: interfering with the biosynthesis of sterol in fungi


Bacteriostatic inhibitors of protein synthesis

Board Spectrum: tetracyclines 四环素 , chloramphenicol 氯霉素 Moderate: macrolides 大环内酯Narrow: clindamycin 克林霉素

peroperties

Suppress bacterial growth and replication but do not kill

Second-line agents


Antimicrobials acting on protein synthesis

Target the bacterial ribosome.• Bacterial – 70S (50S/30S)• Mammalian – 80S (60S/40

S)• High levels may interact with

mammalian ribosomes.Binding to 30s Subunit• Aminoglycosides 氨基糖苷

(bacteriocidal)• tetracyclines 四环素

Binding to the 50s subunit• chloramphenicol 氯霉素macrolides (erythromycin 红霉

素 , clarithromycin 克拉霉素 , azithromycin 阿齐霉素 )

30s subunit

50s subunit

mRNA

protein


Antimicrobials acting on nucleic acid synthesis

Inhibitors of DNA replication • Quinolones (e.g. ofloxacin ，氧氟沙星 )

inhibit DNA-gyrase 回旋酶• Orally active, broad spectrum

Damage to DNA• Metronidazole 甲硝唑 (anti-anaerobes)• nitrofurantoin 呋喃妥因 Inhibitors of

Transcription • rifampicin (key anti-TB drug) inhibits

bacterial RNA polymerase• flucytosine 氟胞嘧啶， incorporated into

yeast mRNA


Antimicrobials acting on Folic acid synthesis

Inhibitors of precursor synthesis• sulphonamides & trimethoprim are synthetic,

bacteriostatic agents• used in combination in co-trimoxazole （复方磺

胺甲噁唑） • Sulphonamides inhibit early stages of folate

synthesis （二氢叶酸合成酶）• dapsone( 氨苯砜 ), an anti-leprosy drug

• Trimethoprim inhibits final enzyme in pathway, dihydrofolate reductase （二氢叶酸合成酶）　

• Pyrimethamine （乙胺嘧啶）

Shanghai Jiao Tong University Mechanism of sulfonamides

蝶啶＋ → 二氢蝶酸→二氢叶酸→四氢叶酸

↑ ↑

二氢蝶酸合成酶二氢叶酸还原酶

sulfonamides TMP （甲氧苄啶）

Dapsone 氨苯砜 pyrimethamine

（乙胺嘧啶）

methotrexate


Antibiotic Resistance


2010 年英国媒体：南亚发现新型超级病菌，产生 NDM-1 （ New Delhi metallo-β-lactamase-1 ，新德里金属 -β- 内酰胺酶 -1 ），抗药性极强。 Superbug ：

Escherichia coli and

Klebsiella pneumoniae

克雷伯肺炎杆菌


日本科学家拍摄到的“超级细菌”照片


Bacteria

Antibiotic resistance

Definition: a microorganism is able to survive exposure to an antibiotic.1940’s, penicillin-resistant bacteria, 1%

21st century, 耐苯唑西林金黄色葡萄球菌（ MRSA: methicillin-resistant Staphylococcus aureus): 80-92%

耐万古霉素肠球菌（ VRE ），耐氨基糖苷类的高耐株（ high-level aminoglycoside resistant, HLAR ）： 0-8% ， 60-80%

2002.7: 美国 center for disease control (CDC) 公布第一株真正的耐万古霉素（ MIC>128 mg/L ）的葡萄球菌


19701960 20001990

金黄色葡萄球菌：获得性耐药株的新挑战 39thICAAC1999

1996

1980

耐青霉素的金黄色葡萄球菌1944

1961

1975

？

古典的MRSA

多重耐药MRSA

VRSA

VISA

VISA: Vancomycin-intermediate Staphylococcus aureus 万古霉素中介耐药葡萄球菌

VRSA: vancomycin-resistant Staphylococcus aureus


antibiotic era 抗生素时代 form 1940s: Penicillin was tested clinically an

d mass produced to 1975

Post-antibiotic era 后抗生素时代 multi-drug resistance 多药耐药


The kinds of antibiotic resistance

(1) Intrinsic resistance （天然 / 固有耐药）： genes in chromosome

(2) Acquired resistance （获得性耐药） : genes in plasma

transfer of antibiotic resistance: transformation 转化 conjugation 接合 transduction 转导


Mim

s C

et

al.

Med

ical

Mic

robi

olog

y. 1

998.

Shanghai Jiao Tong Universitymechanisms of resistance

Inactivating enzymes: hydrolases; synthases

Targets changes: protein structure change; production of protective protein

Decrease of antibiotics: increased efflux system; decreased permeability to antibiotics

Change of metabolic pathways; trapping mechanism; formation of biofilm

Shanghai Jiao Tong UniversityResistance to antibiotics

Shanghai Jiao Tong UniversityMechanisms of resistance

Modification or bypass of target

• by mutation or acquisition of extrinsic DNA

Shanghai Jiao Tong UniversityResistance Mechanisms

β-lactamase – hydrolyze the β-lactam ring.

Penicillinase – Staph

Alteration of penicillin-binding protein (PBP) affinity. (Strep. Pneumo., MRSA)


75% of antibiotics prescribed for respiratory tract infections

50% of antibiotic prescriptions are inappropriate

Most respiratory tract infections are caused by viruses

Antibiotics do not work against viruses

Antibiotics in theCommunity

Shanghai Jiao Tong UniversityAntimicrobial selection

Identify the infecting organismDrug sensitivity of the infecting organismHost factors – site of infection, status of patient, safety of the agent, cost therapy


Delaying the emergence of resistance

Prescribed only when neededNarrow-spectrum:

Cultures must be obtained prior to initiation of therapyAdequate dosage Limit use of newer drugsMinimize giving antibiotics to livestock

Shanghai Jiao Tong UniversityAntibiotic combinations

Severe infection

Mixed infections

Prevention of resistance – tuberculosis

Decreased toxicity

Enhanced antibacterial action

Shanghai Jiao Tong UniversityAppropriate prophylactic use

Before surgery

Prevent bacterial endocarditis

Protect fetus of an HIV-infected pregnant woman

Prevent TB


Quinolones, sulfonamides and other synthetic antibacterial drugs


X N

R1

COOH

O

R6

R71 2

34

5

67

8

Basic structure of quinolones

Shanghai Jiao Tong UniversityQuinolones （喹诺酮类）

1962 ， nalidixic acid （萘啶酸）

pipemidic acid （吡哌酸）

only used in urinary and intestinal infections caused by G － bacteria.

low efficacy; rapid resistance. removed from clinical uses.

Third and fourth generations: 　

1980’s: 氟喹诺酮类， norfloxacin （诺氟沙星） , ciprofloxacin （环丙沙星）， ofloxacin （氧氟沙星）， moxifloxacin （莫西沙星）， gatifloxacin （加替沙星）， sparfloxacin （司帕沙星）…

(me-too)

Shanghai Jiao Tong UniversityMechanism of action

Quinolones inhibit the bacterial DNA gyrase (topoisomerase Ⅱ) or topoisomerase Ⅳ ，拓朴异构酶Ⅳ ), thereby inhibiting DNA replication and transcription.


Gyrase 能在两条链上都产生切口，然后重新连结，并催化双链 DNA 形成双螺旋

Shanghai Jiao Tong UniversityResistance to quinolones

seen in numerous pathogens, eg. Staphylococccus aureus （金黄色葡萄球菌） , enterococci （肠球菌） and Streptococcus pyogenes（化脓性链球菌） , …

Mostly caused by inappropriate use (up to 99%)

Shanghai Jiao Tong UniversityResistance to quinolones

efflux pumps decrease intracellular quinolone concentration.

In gram-negative bacteria, plasmid-mediated resistance genes produce proteins that can bind to DNA gyrase, protecting it from the action of quinolones.

mutations at key sites in DNA gyrase or topoisomerase Ⅳ can decrease their binding affinity to quinolones, decreasing the drug's effectiveness.


Common properties of quinolones

Broad antibacterial spectrum: gram-negative and positive bacteria

Some effective to pseudomonas aeruginosa （铜绿假单胞菌，（从前：绿脓杆菌）， mycobacterium tuberculosis （结核分枝杆菌）， mycoplasma （支原体）， chlamydia （衣原体）， anaerobic bacteria （厌氧菌）Good pharmacokinetic features

Less and mild untoward effects, but arthropathy （关节病） often seen in children and phototoxicity


衣原体

厌氧菌

Shanghai Jiao Tong UniversityIndividual quinolones

Levofloxacin （左氧氟沙星） : chiral molecule, more powerful and less toxic

Sparfloxacin （司帕沙星）： wider antibacterial spectrum including M. tuboculosis

Moxifloxacin （莫西沙星）： high efficacy in anaerobic bacteria

Gatifloxacin （加替沙星）： wide antibacterial spectrum and low phototoxicity

Shanghai Jiao Tong Universitysulfonamides （磺胺类）

N SO2H

R2

H

R1

14

对氨基苯磺酰胺


Prontosil （百浪多息）

Shanghai Jiao Tong UniversityHistory

Gerhard Domagk ， German biochemist

Involved in WW I, experienced lots of infections

After MD, in dye company

Paid attention to effects of dyes on bacteria

Prontosil tested in mouse model of Streptococcus pyogenes infection

Used in his daughter suffered from Streptococcus pyogenes infection and succeded

Confirmed effective in President Roosevelt’s son

1939 Nobel prize in Medicine or Physiology


Antibacterial spectrum of sulfonamides

Antibacterial spectrum wide (bacteriostatic), but most of the bacteria resistant to these drugs now

Shanghai Jiao Tong University Mechanism of sulfonamides

蝶啶＋ → 二氢蝶酸→二氢叶酸→四氢叶酸

↑ ↑

二氢蝶酸合成酶二氢叶酸还原酶

sulfonamides TMP （甲氧苄啶）

Dapsone 氨苯砜 pyrimethamine

（乙胺嘧啶）

methotrexate

Shanghai Jiao Tong University Individual sulfonamides

Sulfamethoxazole （磺胺甲噁唑） +trimethoprim （甲氧苄啶）＝复方新诺明； used in mild urinary or respiratory infections

Sulfasalazine （柳氮磺吡啶）： ulcerative colitis （溃疡性结肠炎） Silver sulfadiazine （磺胺嘧啶银）： local use in burn wound

Shanghai Jiao Tong University nitrofurans （硝基呋喃类）

Nitrofurantoin （呋喃妥因）

urinary infections

Furazolidone （呋喃唑酮）

bacteroidal diarrhea

chemotherapy of microbial diseases 殷明 2011.11. shanghai jiao tong university 1940...

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