Protein binding unit 4 bppk pe 520 (final) [autosaved] (1)

bp/pk 520

1. Protein binding Unit- 4 biopharmaceutics and pharmacokinetics (pe- 520) Priyansha singh (m.s. pharm- pharmacology & toxicology, niper Guwahati )

2. Basic diagram of protein binding

3. What is protein binding A drug in the body can interact with several tissue components of which the two major categories are blood and extravascular tissues. The interacting molecules are generally the macromolecules such as proteins, DNA or adipose. The proteins are particularly responsible for such an interaction . The phenomena of complex formation with proteins is called as protein binding of drugs . The importance of such a binding derives from the fact that the bound drug is both pharmacokinetically as well as pharmacodynamically inert i.e. a protein bound drug is neither metabolized nor excreted nor is it pharmacologically active. A bound drug is also restricted since it remains confined to a particular tissue for which it has greater affinity. Moreover, such a bound drug, because of its enormous size, cannot undergo membrane transport and thus its half-life is increased .

4. mechanism behind protein binding Binding of drugs generally involves weak chemical bonds such as hydrogen bonds, hydrophobic bonds, ionic bonds or van der Waal 's forces and , therefore, is a reversible process . 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 ; for example, covalent binding of chloroform and paracetamol metabolites to liver results in hepatotoxicity . Binding of drugs falls into 2 classes: 1 . Binding of drugs to blood components like-a) Plasma proteins & b) Blood cells 2. Binding of drugs to extravascular tissue proteins, fats, bones, etc.

5. BINDING OF DRUGS TO BLOOD COMPONENTS Pharmacological class- I mipramine- TCA Antidepressant Lidocaine- Local Anesthetic (surface linked/soluble and amide linked), Antiarrhythmic (class 1B) Quinidine- Class 1A antiarrhythmic Chlorpromazine- phenothiazine antipsychotic Cyanocobalamine- Vitamin B12 Cupric ion = Cu+ ions

6. BINDING OF DRUGS TO BLOOD COMPONENTS The main interaction of drug, in the blood compartment is with the plasma proteins which are present in abundant amounts and in large variety. The binding of drugs to plasma proteins is reversible. The extent or order of binding of drugs to various plasma proteins is: albumin > alpha 1-acid glycoprotein > lipoproteins > globulins. A short concept- all proteins have some pockets/ specific sites in which only a specific drug binds depending upon the molecular weight, lipophilicity and concentration of the protein present. Now we will have a look at binding of drugs with Human Serum Albumin (HSA) Alpha 1- acid glycoprotein Lipoproteins Globulins RBC components

7. a. Binding of drugs to human serum albumin It is one of the most abundant plasma protein (upto 60%), having M.W. of 65k with large binding capacity. Both endogenous compounds such as fatty acids, bilirubin and tryptophan as well as drugs bind to HSA. A large variety of drugs ranging from weak acids, neutral compounds to weak bases bind to HSA. A drug can bind to more than one site in which case the main binding site is called as the primary site and the other as the secondary site ; for example , site I is the primary site for dicoumarol and site II is then the secondary site Groups of drugs that bind to the same site, compete with each other for binding, but drugs that bind to one site do not competitively inhibit binding of drugs to other sites. However, they may either promote or retard binding of a drug to another site by energetic coupling mechanisms . Pharmacological class Warfarin - oral anticoagulant (coumarin derivatives) Azapropazone- NSAID Diazepam- Benzodiazepines (intermediate acting) Digitoxin- Cardiac glycoside Tamoxifen- SERM used against breast cancer

8. b. Binding of drugs with alpha 1 glycoprotein (Beta blocker)

9. c. Binding to lipoproteins Why is HDL good Pharmacological class Diclofenac - NSAID (phenylacetic acids- non selective COX inhibitor) Cyclosporin A- immunosuppressant

10. d. Binding to globulin

11. e. Binding o f drugs with rbc and its components Pharmacological class- Phenytoin- antiepileptic (hydantoin) Pentobarbital- Barbiturate (short acting) Acetazolamide - Diuretics (carbonic anhydrase inhibitor) Chlorthalidone- thiazide diuretic

12. f. Binding of drugs to tissue proteins The body tissues, apart from HSA, comprise 40% of the body weight which is l00 times that of HSA. Hence, tissue-drug binding is much more significant than thought to be. A drug can bind to one or more of the several tissue components. Tissue-drug binding is important in distribution from two viewpoints: firstly , it increases the apparent volume of distribution of drugs in contrast to plasma protein binding which decreases it; this is because the parameter is related to the ratio of amount of drug in the body to the plasma concentration of free drug and the latter is decreased under conditions of extensive tissue binding of drugs. S econdly , tissue-drug binding results in localization of a drug at a specific ·site in the body (with a subsequent increase in biological half-life). This is more so because a number of drugs bind irreversibly with the tissues (contrast to plasma protein-drug binding); for example, oxidation products of paracetamol, phenacetin , chloroform, carbon tetrachloride and bromobenzene bind covalently to peptic tissues .

13. f. Binding of drugs to tissue proteins Factors influencing localization of drugs in tissues include lipophilicity and structural features of the drug, perfusion rate, pH differences, etc. extensive tissue-drug binding suggests that a tissue can act as the storage site for drugs. Drug that binds to both tissue and plasma components result in competition between drug binding sites . For majority of drugs that bind to extra vascular tissues, the order of binding is liver > kidney > lung > muscle . Several examples of extravascular tissue-drug binding are : Paracetamol- acetaminophen NSAID

14. f. Binding of drugs to tissue proteins Pharmacological classification M etallothionin- cysteine-rich, low molecular weight proteins. Chloroquine- antimalarial and amebicides Phenothiazine- antipsychotic drugs Tetracycline side effect- fanconi syndrome, discoloration of bone and teeth Aminoglycosides side effects- ototoxicity, nephrotoxicity

15. Comparision between plasma protein and tissue protein drug binding

16. Factors affecting protein binding Factors relating to the drug:- a. Physicochemical characteristics of the drug b. Concentration of drug in the body c. Affinity of a drug for a particular binding component Factors relating to the protein and other binding components:- a. Physicochemical characteristics of the protein or binding agent b. Concentration of protein or binding component c. Number of binding sites on the binding agent Drug interactions a. Competition between drugs for the binding site (displacement interactions ) b. Competition between drugs and normal body constituents c. Allosteric changes in protein molecule Patient related factors a. Age b. Inter-subject variations c. Disease states

17. Factors relating to the drug:-

18. Influence of physicochemical properties of drugs on protein binding

19. Factors relating to the protein and other binding components:-

20. Drug interactions

21. Age differences from neonates to baby

22. Patient related factors

23. Kinetics of protein drug binding Are drugs bound to the protein in a reversible or irreversible or both manners- majorly drugs are reversibly binding with drugs which is why it shows reversible kinetics The drugs which show irreversible binding are totally out of the question of being in equilibrium with free drug. It is an irreversible equation then.

24. Kinetics of protein drug binding

25. Kinetics of protein drug binding

26. Kinetics of protein drug binding How do we come to know by looking at the equation 4.17 that r has to be taken on x axis and D on y axis?  R is a number basically determining the ratio of no. of moles of bound drug to the total protein present. Therefore R is supposed to be on Y axis

27. Kinetics of protein drug binding

28. Kinetics of protein drug binding

29. Implications of protein binding on pharmacokinetic properties Plasma and tissue protein binding of drugs is a major factor that affects both pharmacokinetics and pharmacodynamics of the drug. VOLUME OF DISTRIBUTION ( Vd) - more the protein binding, lesser the volume of distribution and vice versa CLEARANCE - clearance again depends upon volume of distribution therefor more the protein binding, lesser the clearance of the drug and vice versa HALF LIFE- more the drug is bound to protein and especially to plasma protein , more will be the residence time and hence the half life of the drug increases and vice versa. BIOAVAILABILITY - more the drug is bound to plasma protein lesser it is pharmacologically active and hence the bioavailability. CONCENTRATION - alteration in the concentration of drug which is administered will directly affect the protein binding of that drug. More protein binding with drug, lesser the amount of pharmacologically active concentration of the drug. DURATION OF DRUG- greater the amount of protein binding of a particular drug, longer is the half life and hence longer is the duration of action (as well as its toxicity)

30. Significance of protein binding

31. Significance of protein binding