Drug Resistance Mechanisms

Drug Resistance

• Defined as the natural ability of a normal organism to remain unaffected by noxious agents in its environment
• A strain exhibits resistance to an antibiotic when its minimum inhibitory concentration (MIC) is considerably greater than average for the originally susceptible bacterial population
• Intrinsic resistance is a natural characteristic of certain bacterial species, while acquired resistance arises through genetic mutations or horizontal gene transfer, often as a response to antibiotic pressure.

Antimicrobial Resistance Factors

• Patients, the environment, healthcare providers, counterfeit medicine, food animals and humans

Drug Resistance Mechanisms

• Enzymatic degradation of antibacterial drugs
• Alteration of bacterial proteins that are antimicrobial targets
• Changes in membrane permeability to antibiotics
• Lack of entry due to decreased cell permeability
• Greater exit caused by active efflux
• Enzymatic inactivation of the antibiotic
• Altered target through modification of the drug receptor site
• Synthesis of a resistant metabolic pathway

Enzymatic Inhibition

• Beta-lactamases function by splitting the amide bond of the beta-lactam ring
• There are many types of enzymes inactivating antibiotics, characterized by amino acid and nucleotide sequencing
• Class A enzymes, which have a molecular weight (MWT) of approximately 29,000 daltons, play a crucial role in antibiotic resistance by hydrolyzing beta-lactam antibiotics, thus rendering them ineffective against targeted bacterial strains.
• Class A preferentially hydrolyze penicillins, e.g., TEM-1, prevalent in many gram-negative bacteria
• Class B enzymes are metalloenzymes and have a zinc-binding thiol group required for beta-lactamase activity
• Class C enzymes have a MWT of 39000 and are mainly cephalosporinases
• Blocking enzymes attach side chains to the antibiotic, inhibiting its function
• Examples of enzymes include Proteases, Carbohydrases, Nucleases and Cathelicidins

Alteration of Bacterial Membranes

• The outer membrane of Gram-negative bacteria acts as a barrier to antibiotics, especially hydrophobic ones
• The effectiveness of aminoglycosides relies heavily on their capacity to bind to specific transport proteins located in the bacterial cell membrane. These non-saturable anionic transporters facilitate the influx of the antibiotics, which is crucial for their therapeutic action against bacterial infections.
• They retain their positive charge and are pulled across the cytoplasmic membrane by the internal charge of the cell
• Promotion of antibiotic efflux is a major mechanism for tetracycline resistance in gram-negative bacteria, mediated by plasmid/chromosomal/transposon

Efflux/Influx Mechanism

• Restriction of influx is a physiological way to reduce toxicity to bacterial cells
• The AcrAB/TolC system in E. coli is a well-studied efflux system
• The E. coli system comprises an inner membrane protein Acr B, an outer membrane protein Tol C, and a periplasmic protein Acr A

Five Basic Mechanisms of Antibiotic Action Against Bacterial Cells

• Inhibition of cell wall synthesis, as seen with Beta-Lactams, Vancomycin, and Bacitracin
• Inhibition of protein synthesis (translation), as with Aminoglycosides, Tetracyclines, Chloramphenicol, Macrolides, and Clindamycin
• Alteration of cell membranes using Polymyxins and Bacitracin
• Inhibition of nucleic acid synthesis using Quinolones, Metronidazole, and Rifampin
• Antimetabolite activity, as seen with Sulfonamides, Trimethoprim, and Trimethoprim-Sulfamethoxazole

Mutations and Genetic Modifications

• Mutation is an alteration of the nucleotide sequence of the genome of an organism, virus, or extrachromosomal DNA or other genetic elements
• Mutations can occur due to errors during DNA replication (replication-dependent mutations)
• Mutations can also occur independently of DNA replication (replication-independent mutations)

Types of Mutations

• Point mutations, which affect a single nucleotide base in DNA sequences, can have varying effects on protein synthesis. Silent mutations result in no change in the amino acid sequence, while missense mutations lead to a different amino acid being incorporated, potentially altering protein function. Nonsense mutations create a premature stop codon, resulting in truncated, often nonfunctional proteins.
• Silent mutations occur when a base substitution results in a synonymous codon, leaving the amino acid sequence unchanged
• Missense mutations happen when a base substitution results in a codon that specifies a different amino acid, leading to a different polypeptide sequence
• The impact of the missense mutation can be conservative or nonconservative, depending on whether the substituted amino acid has similar biochemical properties to the original one, potentially preserving protein function or altering it significantly.
• Nonsense mutations occur when a single nucleotide change results in the formation of a premature stop codon within the protein-coding sequence. This truncation halts translation unexpectedly, leading to incomplete protein synthesis. Often, the resulting protein is nonfunctional or significantly impaired, which can disrupt essential cellular processes and lead to disease.