DRUG PROTEIN BINDING. A Key Factor in Pharmacokineticspptx

Drug-Protein Binding: A Key Factor in Pharmacokinetics

Drug-protein binding refers to the interaction between a drug and proteins in the bloodstream, primarily albumin. This binding affects the drug’s distribution, metabolism, and excretion, impacting its bioavailability and therapeutic effectiveness.

When a drug binds to plasma proteins, only the unbound (free) portion is available for pharmacological action. This balance between bound and free drug determines how the drug is delivered to target tissues, influences drug-drug interactions, and plays a critical role in the onset, duration, and intensity of drug effects. Understanding drug-protein binding is crucial for drug development, dosage design, and clinical treatment plans.

1. DRUG PROTEIN BINDING 1

2. Group Members Naveed 020 Muhammad Imran Ali 043 Samra Arif 044 Muhammad Nabeel Sultan 049 Eman Aslam 054 Humayun khan 069 Fatima Shayed Abbasi 075 Eman Afsar 079 2

3. CONTENTS INTRODUCTION MECHANISM OF DRUG PROTEIN BINDING PROTEIN BINDING IN PLASMA AND INTERSTEIAL FLUID APPARENT VOLUME OF DISTRIBUTION HALF LIFE FACTORS AFFECTING DRUG PROTEIN BINDING CLINICAL SIGNIFICANCE OF DRUG PROTEIN BINDING DISCUSSION OF PROTEIN BINDING STUDIES OF PHENYTOIN DISCUSSION OF PROTEIN BINDING STUDIES OF CEFAZOLIN 3

4. INTRODUCTION The phenomenon of complex formation of drug with protein is called as Drug-Protein binding. The proteins are particularly responsible for such an interaction. Pharmacologically inactive due to its pharmacokinetic and Pharmacodynamic inertness. Protein + drug = drug-protein complex Protein binding may be divided into: 1. Intracellular binding. 2.Extracellular binding. 4

5. MECHANISMS OF DRUG PROTEIN BINDING Binding of drugs to proteins is generally of reversible & irreversible. Reversible generally involves weak chemical bond such as: 1. Hydrogen bonds 2.Hydrophobic bonds 3.Ionic bonds 4. Van der waal's forces. Irreversible drug binding, though rare, arises as a result of covalent binding and is often a reason for the carcinogenicity or tissue toxicity of the drug 5

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7. Protein Binding in Plasma and Interstitial Fluid Drugs that are highly bound to plasma proteins and have a relatively small apparent volume of distribution (V) are significantly influenced by protein binding in both plasma and interstitial fluid. Examples include ibuprofen, which remains largely within the circulating volume due to its high plasma protein binding, and drugs like amiodarone that bind extensively to tissue proteins, resulting in a large apparent V. 7

8. Half-Life and Apparent Volume of Distribution (V): Half-Life (t₁/₂): Changes in the extent of binding in plasma and interstitial fluid can alter the half-life of a drug by affecting the free fraction available for metabolism and excretion. Apparent Volume of Distribution (V): For drugs with high plasma protein binding, a decrease in plasma binding increases V, indicating more extensive distribution into tissues. 8

9. Tissue Binding: Distribution: Drugs with a large apparent V tend to distribute more extensively into tissues, and changes in tissue binding significantly affect their pharmacokinetics. Effectiveness: The therapeutic and toxic effects of a drug are determined by its concentration at the site of action, which is often within tissues. 9

10. Factors Influencing Protein Binding Number of Available Binding Sites Protein concentration Drug concentration Association Constant (K) Drug Properties Lipophilicity pKa Resemblance to endogenous ligands Environmental Conditions pH Temperature Endogenous ligands 10

11. Factors Influencing Protein Binding Protein Concentration: Higher protein levels in plasma increase the number of available binding sites for drugs. Drug Concentration: Higher drug concentrations can saturate available binding sites. 11

12. Factors Influencing Protein Binding Association Constant (K): The affinity between the drug and the binding site determines the extent of protein binding. 12

13. Factors Influencing Protein Binding Lipophilicity : Lipid-soluble drugs tend to bind more to plasma proteins. pKa : The ionization state of the drug influences its binding affinity. Resemblance to Endogenous Ligands: Drugs structurally similar to endogenous ligands may compete for the same binding sites. 13

14. Factors Influencing Protein Binding pH: Changes in pH affect the solubility and binding affinity of drugs. Temperature: Typically, body temperature remains within a narrow range, but significant deviations can affect binding dynamics. Endogenous Ligands: Competition with endogenous substances like bilirubin can affect drug binding. 14

15. Proteins and Blood Components that Bind Drugs Albumin : The primary binding protein for most drugs, with six binding sites. α-1 Acid Glycoprotein: Binds basic drugs and steroid molecules. Lipoproteins: Bind lipid-soluble drugs, e.g., cannabinoids and cyclosporin . Globulins: Bind fat-soluble vitamins (A, D, E, K). Hemoglobin: Binds drugs like pentobarbital and phenytoin. Red Cell Membrane: Binds drugs such as chlorpromazine and imipramine. Carbonic Anhydrase: Binds drugs like acetazolamide and chlorthalidone . Specific Transport Proteins: Bind drugs intended for transport, e.g., thyroxin-binding globulin. 15

16. Clinical Significance of Protein Binding Volume of Distribution (VOD): Smaller for drugs highly bound to plasma proteins. Larger for drugs extensively bound to tissue proteins. Metabolism and Clearance: Only the free fraction is typically available for renal elimination and hepatic metabolism. Exceptions exist where drugs are actively transported or metabolized despite high protein binding. 16

17. Clinical Significance of Protein Binding Measured Drug Levels: Total drug levels may not reflect the biologically active free fraction. Hypoalbuminemia can increase the free fraction, affecting clinical outcomes. Displacement by Other Drugs: Competition for binding sites can lead to displacement and increased free fraction of the displaced drug, potentially causing toxicity. 17

18. Discussion of Protein Binding Studies of Phenytoin 18

19. Protein Binding Properties of Phenytoin High Protein Binding : Phenytoin is approximately 90% bound to plasma albumins, meaning that a significant majority of the drug in the bloodstream is attached to plasma proteins rather than existing in a free form. Narrow Therapeutic Index : Phenytoin’s therapeutic range is narrow, indicating that small variations in drug concentration can lead to subtherapeutic effects or toxicity. Non-linear Pharmacokinetics : The drug exhibits Michaelis-Menten kinetics, meaning that its metabolism can become saturated, leading to non-linear relationships between dose and blood levels. 19

20. Methodology Ultrafiltration Technique: The protein binding of phenytoin was assessed using a centrifugal ultrafiltration method, which separates free (unbound) drug from the protein-bound fraction. Concentration Ranges: Three different concentrations of phenytoin were studied (25, 50, and 100 µg/mL), with the analysis performed in triplicate for accuracy. 20

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22. Results The study investigated the effect of tizoxanide (TZX) on the protein binding of phenytoin (PHT). The results were derived from two phases: Phase 1: Phenytoin Alone: At 100 µg/mL: Fraction unbound = 0.17 (±0.004) At 50 µg/mL: Fraction unbound = 0.13 (±0.001) At 25 µg/mL: Fraction unbound = 0.11 (±0.001) Mean free fraction of phenytoin: 0.14 22

23. Phase 2: Phenytoin with TZX: At 100 µg/mL: Fraction unbound = 0.47 (±0.005) At 50 µg/mL: Fraction unbound = 0.48 (±0.007) At 25 µg/mL: Fraction unbound = 0.48 (±0.005) Tizoxanide increased unbound phenytoin concentrations by 4.4, 3.7, and 2.8 times for the respective concentrations. 23

24. Effect of Protein Binding of Phenytoin Impact of Tizoxanide on Phenytoin Binding Displacement Interaction : Tizoxanide significantly inhibited the protein binding of phenytoin. It increased the unbound fraction ( fu ) of phenytoin by 4.4, 3.7, and 2.8-fold at concentrations of 25, 50, and 100 µg/mL, respectively. This indicates a strong competitive displacement effect. Albumin Binding Sites : Both phenytoin and tizoxanide primarily bind to albumins. Tizoxanide binds specifically to Sudlow site I on albumin, suggesting that phenytoin may also bind to this site, leading to competitive displacement. 24

25. Impact on Pharmacokinetic (PK) Parameters Volume of Distribution ( Vd ) Increase in Vd : An increase in the unbound fraction of phenytoin can lead to a higher volume of distribution. Since Vd is influenced by the extent of protein binding, with less binding resulting in a larger volume, the displacement by tizoxanide will increase the Vd of phenytoin. Clearance (Cl) and Half-Life (t1/2) Clearance : Clearance may be affected due to changes in the free drug concentration. For phenytoin, which follows non-linear pharmacokinetics, the increase in the unbound fraction can lead to a disproportionate increase in the rate of metabolism until the metabolic pathways become saturated. After saturation, clearance will be reduced . 25

26. Half-Life : The half-life of phenytoin can be altered based on the changes in clearance and volume of distribution. Initially, as the unbound fraction increases and if the clearance is not saturated, the half-life may decrease. However, once metabolic saturation is reached, the half-life can increase due to reduced clearance capacity. 26

27. Protein Binding Studies of Cefazolin 27

28. Protein Binding Properties of Cefazolin High and Saturable Protein Binding : Cefazolin is a first-generation cephalosporin with high protein binding, typically quoted as 80–90%. The binding is saturable , meaning that as the concentration of cefazolin increases, the percentage bound to plasma proteins decreases. At low concentrations (e.g., 8.5 mg/L), the unbound fraction is about 9%, but at higher concentrations (e.g., 140 mg/L), it can rise to 51%. 28

29. Binding Sites : Cefazolin primarily binds to albumin at either the warfarin (site I) or bilirubin site. It can be displaced by endogenous substances like bilirubin and free fatty acids, and by other acidic drugs such as furosemide, clofibrate , and valproic acid. 29

30. Methodology Study Design : The study involved both between-patient and within-patient analyses. In the between-patient study, single random samples were taken from 31 patients treated with cefazolin. In the within-patient study, paired samples (trough and peak concentrations) were taken from 12 patients. Analytical Techniques : Total and unbound plasma concentrations were measured using HPLC. Ultracentrifugation was employed to separate unbound from bound drug fractions. 30

31. Results and Discussion Between-Patient Variability : Linear regression analysis indicated a significant relationship between the unbound percentage and unbound plasma concentrations (r² = 0.79). This supports the concept of saturable protein binding with increasing drug concentrations. Within-Patient Variability : Significant differences were observed between peak and trough concentrations within patients, confirming saturable protein binding. For instance, one patient had a trough concentration of 2.3 mg/L with 19.6% unbound cefazolin and a peak concentration of 101.6 mg/L with 21.1% unbound cefazolin. 31

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33. Impact on Pharmacokinetics: Saturable binding affects pharmacokinetic parameters such as volume of distribution ( Vd ) and clearance (Cl). As protein binding saturates and more drug remains unbound, Vd may increase. This change can lead to an apparent increase in the free drug available for activity. Clearance may also be impacted due to higher free drug concentrations, potentially altering the half-life (t½) of cefazolin. 33

34. Practical Implications Clinical Relevance : Saturable protein binding is particularly relevant when interpreting total drug concentrations against MIC values of infecting organisms. It is also significant in pharmacokinetic studies where dose adjustments might be needed based on unbound drug concentrations. Therapeutic Monitoring : Understanding the extent of protein binding helps in accurate therapeutic drug monitoring and dose adjustments. This is crucial in conditions where albumin levels are low, such as in patients with cirrhosis or renal disease, affecting the binding dynamics. 34

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