Chemotherapeutics are defined as certain specific chemicals that can be put into the human body: chemical agents, antibiotics, antibodies They must fulfill the following criteria: 1. they must destroy or prevent the activity of a parasite WITHOUT injury to the host cell or cause ONLY LIMITED damage to the host cell (SELECTIVE TOXICITY) 2. they must be able to come into contact with the parasite mainly by penetration through the tissues or cells 3. they must NOT interfere with the host's natural defenses Historical Overview Mercury compounds were used to treat syphilis during the Renaissance. In the 1880's an extract of the chinchona bark (quinine) was discovered as a treatment for malaria. The establishment of most modern day work began with Joubert, Tyndall and Pasteur. One of the first chemotherapeutic agent came from the Gram - bacterium Pseudomonas. It produces a water soluble green dye, known as Pyocyanous, that inhibits Gram + rods - unfortunately this compound also hemolyzes red blood cells. In 1899, Metchnikoff, a Russian microbiologist (who also noted phagocytosis of white blood cells), worked with the Gram + Lactobacillus acidophilus. The growth of this organism in the intestine will modify the intestinal pH. It changed the normally alkaline small intestine to acid. This in turn killed off large numbers of Gram - rods, such as the typhoid and Shigella organisms. To accomplish this the patients had to drink sour milk. Unfortunately there were a number of serious side effects. During the time period 1906-1910, the chemist Paul Ehrlich sought a cure for syphilis. After several years and many trials he discovered Salvarsan (the 606th compound tested). This arsenic compound (arspenamide) served as the definitive treatment of syphilis for over 40 years - until the availability of penicillin In 1924, Drs. Dath and Gratin from the University of Pennsylvania introduced Actinomycetin. They developed a technique which involved the filtrate of a lysed culture of a strain of actinomyces. It was active against Gram + cocci and spore forming rods but not effective against Gram - rods. Their technique became the basis for future work in extraction of antibiotics. In 1929, Alexander Fleming made his initial observations on the fungus Penicillium. This ultimately led to the isolation and commercial development of penicillin. During 1936-1937 Dr. Damagh discovered sulfanilamide. This represented a big breakthrough because it could be prepared commercially. The sulfa drugs provided most of the microbial control of wounds during World War II. In 1942, penicillin became more available and practical but its use was not widespread. After 1938, American microbiologists began to appreciate the importance of penicillin. Florey (1940) harvested experimental penicillin through the urine of treated patients and recrystalized it because there was so little available for use. In 1939, Rene' Dubois discovered Gramicidin - unfortunately it was too toxic for protracted human use. In 1944 two graduate students working for Selman Waksman of Rutgers University isolated and purified Streptomycin. This was the first of the broad spectrum antibiotics (works against both Gram + and Gram - organisms). Waksman received the Nobel Prize, Rutgers got the patent, and the graduate students got their degrees. The discovery of streptomycin, in conjunction with the avail- ability of penicillin, ushered in what is known as The Age of Antibiotics. Modes of Action (How and Where Chemotherapeutic Agents Work) see also - Major Spectrum of Chemotherapeutic Agents 1. Inhibition of cell wall synthesis - attacks an enzyme which is involved in cross linkage of NAM-NAG; penicillin is an analog of D-alanine and substitutes for it. 2. Combines with the cell membrane and destroys it perme- ability. Polymyxin-B works against some strains of Pseudomonas. Poly-B has a + charged region which binds to the - charged phospholipid in the cell membrane. The lipid-soluble portion of Poly-B reacts with the rest of the membrane making it porous. 3. Inhibitors of Protein Synthesis - most of the broad spec- trum antibiotics work this way. While most work only on the smaller bacterial ribosomes, some can also interfere with normal host cell protein synthesis. Long term use must be avoided for this reason. 4. Alteration of Nucleic Acids - reacts with guanine residue of DNA and links two strands together so that replication is impossible. This prevents the formation of mRNA since it acts on the site of synthesis of m-RNA. Major Spectrum of Some Chemotherapeutic Agents Compete with PABA (Para-AminoBenzoic Acid) Sulfonamides Para-aminosalicylic acid (PAS) Compete with Pyridoxine (Vitamin B6) Isonictotinic acid hydrazide (INH) Inhibit cell wall peptidoglycan synthesis Penicillins Cephalosporins Bacitracin Vancomycin Ristocetin Inhibit protein synthesis by binding to 50S subunit of ribosome Chloramphenicol Macrolide Antibiotics Erythromycin Oleandomycin Carbomycin Spiramycin Lincomycin Clindamycin Inhibit protein synthesis by binding to 30S subunit of ribosome Tetracyclines Chlortetracycline Minocycline Oxytetracycline Tetracycline Streptomycin Other aminoglycoside antibiotics Amikacin Gentamycin Kanamycin Neomycin Tobramycin Disruption of cell membranes Polymyxins Polyene antibiotics Nystatin Amphotericin B Inhibit DNA synthesis (prevent replication and transcription) Mitomycin Actinomycin Nalidixic acid Novobiocin Griseofulvin Inhibit RNA synthesis Rifampicins Inhibit purine synthesis (Adenine and Guanine) Trimethoprim Complications of Chemotherapy A serious problem encountered with many chemotherapeutic agents is an allergic reaction developed by many patients. This type of sensitivity, called HYPERSENSITIVITY, elicits various reactions characteristic of allergic conditions. Skin rashes and fever are the most common manifestations, but a number of deaths have been attributed directly to antibiotic hypersensitivity. Penicillin is the antibiotic most frequently administered. Therefore, it is not surprising that it is responsible for more side reactions than any other drug. Paradoxically, penicillin is one of the least toxic of the antibiotics. It can be taken by most people in enormous quantities with no undesirable results. The chart below gives some indication of the range of reactions associated with various antibiotics. Adverse Reactions and Major Contraindications for the Use of Various Antibiotics ----------------------------------------------------------------- Antibiotic Most Common Reaction or Contraindication Penicillins Hypersensitivity shown by about 5 % of Americans Cephalosporins similar to penicillin Chloramphenicol Irreversible aplastic anemia Erythromycin Relatively nontoxic; jaundice in about 0.4% of cases when used for over 10 days Lincomycin & Diarrhea, severe colitis Clindamycin Tetracyclines Permanent staining of teeth and bones if given during last half of pregnancy up to 8 years of age; increased photosensi- tivity in some adults; gastrointestinal irritation Streptomycin & 8th nerve damage (may be irreversible), Aminoglycosides skin eruptions; dizziness Polymyxins Toxic to kidneys Nalidixic Acid Gastrointestinal upset; rash; headache; photosensitivity Trimethoprim Rash; fever; kidney and liver damage; very rarely - cases of aplastic anemia Sulfonamides Similar to Trimethoprim Antibiotic Resistance (Why Some Organisms Become Resistant) 1. the sensitive target structure may be missing in the resistant form (cell wall, enzyme, ribosome) 2. the cellular structure that is the target of the antibiotic may undergo an alteration so that it no longer binds the antibiotic but can still carry out its normal function 3. the resultant organisms may be impermeable to the anti- biotic (e.g. it may have developed a capsule) 4. the organism may be able to modify the antibiotic to an inactive form; e.g. certain organisms produce the enzyme penicillinase which inactivates penicillin - PPNG (Penicillinase Producing Neisseria gonorrhea) (see also: "Antibiotics that Resist Resistance," Science, Vol.270, pp.724-727, November 3, 1995) No one drug is effective against all pathogens, so that care must be taken to select the appropriate drug. Additional Reasons for Ineffectiveness of Certain Chemicals 1. Many chemicals are inhibitory to pathogens in culture (IN VITRO) BUT are ineffective against the same pathogen in an infected host (IN VIVO) a. the chemical may be inactivated or destroyed by the host b. the chemical may be poorly absorbed or rapidly excreted c. a high concentration must be maintained at the site of the infection d. the pathogen may be alive in the host at some site in the body where the drug cannot penetrate (e.g. dead tissue) 2. In some cases, treatment of the infectious disease requires more than the use of drugs. 3. Drugs may have toxic side effects or cause allergic reactions. a. the pathogen may become drug resistant b. the drug may reduce or destroy the body's normal flora c. drugs may permit SUPERINFECTION - a natural pathogen, normally held in check by the normal flora, is able to flourish once the normal flora is gone' e.g. Candida d. the ability of the body to develop immunity to the patho- gen may be reduced if the pathogen is rapidly eliminated by drug treatment e. drug interaction - two or more drugs taken together may create a dangerous condition or inactivate one of the drugs - consult your pharmacist or AccuFays computer In very few infections is the drug alone responsible for a cure. Most of the defense and immunity systems of the body are essential to bring about a cure, even when a highly effective drug is used. Drugs may also be used to prevent future infections in people who are unusually susceptible to them - chemoprophylaxis - e.g. surgical patients; penicillin to prevent streptococcal sore throats in rheumatic fever patients. Antibiotic - defined as a chemical substance produced by a living organism that is capable of killing or inhibiting the growth of microorganisms. Three groups of microorganisms are responsible for the production of most of the antibiotics used in medicine. 1. fungi - especially the genus Penicillium (penicillin and griseofulvin); provides molecular backbone for semisynthetic penicillins 2. bacteria of the genus Bacillus (bacitracin, polymyxin) 3. actinomycetes (filamentous bacteria) of the genus Streptomyces (streptomycin, chloramphenicol, erythromycin, tetracycline); most antibiotics come from this group Antibiotics capable of effective action on both Gram + AND Gram - bacteria are known as BROAD SPECTRUM antibiotics. Testing of Antibiotics activity is measured by determining the smallest amount of the agent needed to inhibit the growth of a test organism 1. (MIC) Minimum Inhibitory Concentration - often called the tube dilution technique. It does not provide an absolute constant for a given agent since it is affected by: the kind of test organism used, inoculum size, composition of culture medium, and incubation conditions such as temperature, pH, aeration. With rigorous standardization, comparisons are possible. This procedure does not distinguish between CIDAL or STATIC agents since the chemical agent is present throughout the growth period. 2. Agar Diffusion or Disk Diffusion Method - most widely used method. The standardized version is known as the KIRBY-BAUER test. A zone of no growth called a ZONE OF INHIBITION occurs if the organisms are killed by the concentration of antibiotic in the paper disk. The size of the zone is affected by: the sensitivity of the test organism, type of culture medium, incubation conditions, rate of diffusion of the agent within the medium, and concentration of the antibiotic. In order to interpret the results, comparison with standardized tables is required. These tables provide information on the size of the zone (in vitro) required to be effective within the body (in vivo).