The beta-lactam family of antibiotics includes many of the most heavily used antibacterials in clinical medicine. They are important, both historically and currently, because of their effectiveness and generally low toxicity. The beta-lactam structure is being exploited by many drug development groups in the search for new drugs with improved efficacy against resistant strains of bacteria.
The majority of the clinically useful beta-lactams belong to either the penicillin (penam) or cephalosporin (cephem) group. Moxalactam is an antibacterial that belongs to the oxacephem group which is closely related to the cephalosporins. For convenience, it is common to include it as a member of the cephalosporins because its pharmacology is so closely related to the so-called third generation cephalosporins. The beta-lactams also include the carbapenems (e.g., imipenem), the monobactams, e.g., aztreonam, and the beta-lactamase inhibitors, (e.g., clavulanic acid).
All members of the family have a beta-lactam ring and a carboxyl group resulting in similarities in the pharmacokinetics and mechanism of action of the group members.
The beta-lactams have carboxyl groups making them weak acids. As a result of ion trapping, at steady state they will have higher concentrations in the more alkaline (e.g., plasma) of two media with obvious ramifications for intracellular, cerebrospinal fluid, and intramammary drug concentrations. Their ionic character results in both high water solubility of the sodium and potassium salts of the drugs as well as slow, but useful diffusion through membranes.
Family members are substrates for the organic anion transport system in the renal tubule and choroid plexus. Thus, a major means of elimination for most members is by tubular secretion into the urine. Choroid plexus transport removes beta-lactams from cerebrospinal fluid. Because these transport systems are very active, members of the family tend to have short elimination half-lives (e.g., 30 minutes to 2 hours) and to produce low concentrations within the central nervous system. Therapeutic CNS concentrations can be achieved with many members of the family as a result of increased penetration allowed by inflammed menninges combined with massive doses. As a carrier mediated process, the organic acid transport system can be saturated. Therefore, other organic acids (e.g., probenecid) and high concentrations of the drugsthemselves can slow elimination into the urine and from the CSF. This will be discussed in more detail in the section on the penicillins.
A short summary of the mechanism of action of the beta-lactam antibacterials could be stated as follows. A slightly more detailed explanation will be presented next. Beta-lactam antibiotics inhibit bacterial cell wall synthesis. The drugs cause nicks in the peptidoglycan net of the cell wall that allow the bacterial protoplasm to "flow" from its protective net into the surrounding hypotonic medium. Fluid accumulates in the naked protoplast, as the cell now devoid of its wall is called, and it bursts resulting in death of the organism.
An overview of the major steps in synthesis of bacterial cell wall may aid one in understanding how many antibacterials act (See the accompanying figure). Bacteria have a cytoplasmic membrane much like that of eukaryotes. Surrounding this membrane is a periplasmic space which is in turn enclosed by a peptidoglycan layer and finally the outer membrane. The peptidoglycan layer is a cross-linked polymer that forms a net-like structure that helps provide structural rigidity to the organism and allows it to survive in mediums to which it may be strongly hypertonic. Bacterial cytoplasmic osmolality is usually much higher (5-20 atmospheres) than that of eukaryotes (7.6 atmospheres). Without the cell wall and its net-like peptidoglycan layer the bacterial protoplast (cytoplasm plus cell membrane) would swell and burst. The complexity of the cell wall of gram negative organisms is quite different from that of gram positive organisms. The composition of the cell wall also varies with species, but a complete discussion of cell wall composition is beyond the scope of this lecture. The mechanism of action for beta-lactams described below represents the simplest explanation.
The peptidoglycan layer is composed of repeating units of N-acetylglucuronic acid (NAG) and N-acetylmuramic acid (NAMA). Cross links between the chains are provided by a glycine pentapeptide (in Staphylococci) which bridges the gap between D-alanyl-D-alanine peptides attached to NAMA moieties in the respective chains. D-alanine is not found in vertebrate tissues thus making it a substrate that is unique to bacteria. The cross linking reaction is catalyzed by a peptidoglycan transpeptidase located in the cell membrane. Because the beta-lactam ring of this group of antibacterials closely resembles the configuration of D-alanyl-D-alanine, intact beta-lactam antibiotics can serve as a substrate for the transpeptidase. Because the acylation of the enzyme is irreversible, the beta-lactams are said to be suicide inhibitors. Once they have combined with the transpeptidase enzymes, they remain bound regardless of the drug concentration in the medium. This irreversibility will be important during the discussion of post-antibiotic effect. There is growing evidence that mere inhibition of the transpeptidase reaction only inhibits bacterial growth. The cidal action results from some poorly understood sequence of events that lead to bacteriolysis and cell death. Nicks may be produced at the growth area of the cell wall. If these nicks are sufficiently severe, the protoplast may protrude into the medium and burst resulting in cell death. Under these conditions, the beta-lactams are cidal. If the bacterium were in a medium to which it is isotonic, death would not result and a naked protoplast would be formed. If the link between inhibition of cell wall synthesis and production of autolysins which cause the ninks is broken, bacteria are not killed. This may be one mechanism of resistance. Note the implication that the beta-lactams are effective in growing and dividing organisms.
Sub-cidal beta-lactam concentrations reveal that other mechanisms may also be involved. In this case, the drugs inhibit formation of the cross-wall which divides an elongating organism into two organisms. This results in abnormally elongated, grotesque, worm-like bacilli and the process is not completely understood.
Many antibacterials produce a post-antibiotic effect that is incompletely understood. This effect, of varying duration depending on the drug and degree of damage done, helps explain why one may allow the concentration of cidal drugs like the beta-lactams to drop below the MIC for a target organism for a short time during each dose interval.
The peptidoglycan transpeptidases may be thought of as one of the penicillin binding protein referred to in discussions of bacterial resistance elsewhere in this lecture. The beta-lactam must bind to these enzymes to be active. If the enzymes are modified in some way so that they do not bind the beta-lactam, the organism will be resistant to the drug.
Although the beta lactam antibiotics are similar in many respects, there are differences among them with respect to pharmacokinetics, toxicity, and antibacterial activity. The bases for differences in activity will be discussed next. Important differences in pharmacokinetics and toxicity will be discussed with each group.
One major pharmacokinetic difference should be mentioned here and that is acid stability. This is important because acid stable forms of beta-lactams may be given orally whereas the others must be given parenterally. Few, if any, beta-lactams are used topically.
The factors to be discussed next form the basis for why one cannot use one beta-lactam drug to predict the activity of the others. One must use a representative of each major category of penicillins in sensitivity testing. Depending on the reason for bacterial resistance, one may also rationally switch from a beta-lactam in one group to one in another group.
Obviously, a drug must reach its target site before it can act. The beta-lactams inhibit a bacterial transpeptidase that cross-links peptidoglycan chains. This enzyme is located in the outer surface of the cytoplasmic membrane, adjacent to the periplasmic space. Thus, drugs that act on it must penetrate the surrounding cell wall, the nature and complexity of which varies dramatically throughout the bacterial world. One can conceive of channels or pores in the wall through which drugs might be able to diffuse as being crypts and the ability of antibacterials to penetrate the wall as describing its crypticity. These channels are also called porins. Differences in lipophilicity and stereochemistry among the beta-lactams are reflected in their ability to enter the periplasmic space and thus, to demonstrate differences in effectiveness against various organisms.
A beta-lactam that closely resembles the conformation of D-alanyl-Dalanine and that simultaneously lacks groups that sterically hinder binding to the transpeptidase should have a high binding affinity for the enzyme. Addition of groups to retard action of other enzymes, e.g., beta-lactamases, may also decrease affinity for the transpeptidase. In light of this, it is no surprise that the simple penicillin G (benzylpenicillin) molecule is still one of the most effective beta-lactam antibacterials. Some drugs that are specifically designed to resist various beta-lactamases are actually less efficacious at the site of action than drugs like penicillin G, but may resist destruction or have better penetrability.
In some instances, the transpeptidase enzyme (and/or other enzymes that are less well characterized) may be altered in such a way that it fails to bind beta-lactam antibacterials. When this occurs, even the beta-lactamase resistant members will be ineffective and one should shift to another family of antibacterials. Methicillin resistant strains are thought to represent strains with enzymes (penicillin binding proteins) that have low affinity for members of the family.
The major limit to efficacy of the beta-lactam antibacterials is destruction by a family of enzymes called beta-lactamases. Efficacy of these enzymes in hydrolyzing the beta-lactam ring, which is necessary for activity, varies widely and the enzymes could be conceived of as a family with a spectrum of activity. Those with strong proclivity for penicillins (6-aminopenicillanic acid derivatives) are called penicillinases. Those at the cephalosporin (7-aminocephalosporanic acid derivatives) end of the spectrum are cephalosporinases. There are also broad spectrum beta-lactamases that are active on both penicillins and cephalosporins.
It should be noted, however, that these terms are simplistic and that the family of beta-lactamases from gram-negative bacteria alone have been divided into five classes by Richmond and Sykes. Some understanding of these groups is clinically relevant, but not often appreciated because it is so hard to know what one is actually dealing with in practice.
CLASS CHROMOSOMAL INDUCIBLE / CLAVULANATE GROUP
CONSTITUTIVE SENSITIVE?
I Yes Inducible Nil Cephalosporins
(all 3
generations)
II Yes Constitutive Yes Penicillins
(rare)
III Plasmid Constitutive Yes Broad
IV Yes Constitutive Yes Broad
V Plasmid Constitutive Yes? Broad -- All
penicillins many
cephalosporins
For example, chromosomal cephalosporinases (Richmond & Sykes Class I) are inducible in such organisms as Enterobacter spp., Pseudomonas spp., and indole-positive Proteus spp. (Proteus vulgaris is "resistant" to beta-lactams; P. mirabilis is usually sensitive). These are the ONLY beta-lactamases NOT INHIBITED by clavulanic acid, a drug used clinically for this purpose. Class I beta-lactamases can produce resistance to all three generations of cephalosporin antibacterials.
Other Richmond & Sykes classes will be briefly mentioned to make the point that the enzymes are identifiable. Class II includes uncommonly encountered constitutive chromosomal penicillinases. Class III includes the commonly encountered plasmid-mediated broad spectrum beta-lactamases. This class is often found in ampicillin-resistant Haemophilus influenzae and penicillin-resistant Neisseria gonorrhoeae and many enterobacteriaceae. Class IV includes chromosomal broad-spectrum beta-lactamases such as the constitutively produced enzyme found in Klebsiella spp.. Finally, Class V includes plasmid mediated broad spectrum beta-lactamases that hydrolyze cephalosporins and ALL penicillins including the isoxazolyl derivatives (e.g., dicloxacillin). These constitutively produced enzymes are found in various entrerobacteriaceae and Pseudomonas aeruginosa. This plasmid is one of the reasons pseudomonas infections are so difficult to treat.
Staphylococci (gram postive) also produce a beta-lactamase that is highly active against many penicillins. Staphylococcal penicillinase is widespread and constititutes such a serious clinical problem that there is a whole category of penicillins to surmount it known as the antistaphylococcal penicillins.
Susceptibility/resistance to beta-lactamases is not absolute. Different plasmids produce lactamases with different specificities and turnover rates. Different drugs have different susceptibilities to the enzymes. For example, the TEM-1 plasmid beta-lactamase hydrolyzes no moxalactam, less than 1% of cefotaxime, and approximately 50% of cefoperazone within a specific time period.
Because one can identify specific problems that affect the clinical efficacy of drugs, one can also devise ways of overcoming those problems. One way is to systematically study structural modifications that confer certain characteristics, e.g., acid resistance or resistance to a particular beta-lactamase. You are invited to ponder the moxalactam structure in the following figure in which various sites are marked. The specific significance of each functional group is not required knowledge, but awareness of the significance that such information can be compiled and used by medicinal chemists to guide their creation of more effective medications is fundamental to understanding pharmacology.
The following comments refer to the moxalactam substituents noted in the figure.
Addition of these moieties to other beta-lactams often confers the same advantages. For example, addition of a p-hydroxy group to ampicillin yields amoxicillin which has improved activity over ampicillin, e.g., greater crypticity. Amoxicillin also has better pharmacokinetic characteristics, e.g., improved absorption from the gastrointestinal tract.