Pharmacokinetics, Pharmacogenetics, and Drug Transport Flashcards

Course Overview and Administrative Background

• Instructor Profile:

  • Pankaj Desai, PhD (PhD earned from the University of South Carolina, focusing on cancer chemotherapy).

  • 31-year tenure as a faculty member ("Bearcat") at the University of Cincinnati.

  • Primary research specialization: Oncology drug development, specifically focusing on novel therapies for high-grade gliomas and glioblastoma. Operates a research laboratory located in MSB 3212 with an open-door policy.

  • Administrative roles: Former Division Chair of Pharmaceutical Sciences; currently serves as Associate Dean for Graduate Studies and Research.

  • Teaching role: Serving as a substitute/pinch hitter covering lectures previously taught by Dr. Kodagiri, who relocated to the University of Maryland.

• Regional Pharmaceutical & Clinical Research Opportunities:

  • "net pays" (located in Norwood, OH): A multi-billion-dollar asset organization ranked among the top 10 global Clinical Research Organizations (CROs), providing student clinical research, internship, and employment opportunities.

Classroom Logistics and Assessment Structure

• Student Interactions & Attendance:

  • Direct student engagement and roll call acknowledged individuals including Amanda, Morris, Catherine, Akshat, Darshan, and Arya.

• In-Class Quiz Specifications:

  • Platform & Timing: Administered via Dashboard, opening automatically at 11:15 AM and closing at 11:30 AM (15-minute duration).

  • Format & Scope: Comprises 10 broad review questions covering Pharmacokinetics (ADME) and Pharmacodynamics (dose-response relationships). Excludes Lecture 1 (general overview).

  • Grading & Policies: Automatically graded, non-curved assessment serving as a diagnostic self-check one week prior to major exams. Requires charged laptops in class with lids closed upon completion; student disability accommodations are managed individually.

Fundamentals of Pharmacokinetics versus Pharmacodynamics

• Pharmacodynamics (PD):

  • Definition: The quantitative study of what the drug does to the body.

  • Focus: Evaluates drug-receptor interactions, biochemical mechanism of action, potency ($EC_{50}$, $EC_{90}$), and therapeutic versus toxic concentration ranges.

• Pharmacokinetics (PK / ADME):

  • Definition: The quantitative assessment of what the body does to the drug.

  • Focus: Characterizes the rate and extent of drug movement into, through, and out of the body through Absorption, Distribution, Metabolism, and Excretion (ADME), governed by physicochemical properties, formulation factors, and physiological processes.

• Clinical Phases of Drug Development and PK Profiling:

  • Phase 1 Clinical Trials: First-in-human studies conducted in healthy human volunteers housed in a dedicated Clinical Research Unit (CRU) for 4 to 5 days. Permits dense serial blood sampling to construct definitive pharmacokinetic profiles.

  • Phase 2 Clinical Trials: Conducted in patient populations to assess therapeutic efficacy and safety in outpatient/community settings. Employs sparse PK blood sampling during routine outpatient clinic or laboratory visits.

Quantitative Pharmacokinetic Parameters and Concentration-Time Curves

• Key Parameters Derived from Plasma Concentration-Time Profiles:

  • $C_{\max}$ (Peak Plasma Concentration): The maximum drug concentration achieved in systemic circulation; directly correlates with the intensity of therapeutic or adverse drug actions.

  • $T_{\max}$ (Time to Peak Concentration): The time required to reach $C_{\max}$. A shorter $T_{\max}$ is clinically preferred when rapid onset of action is required. At $T_{\max}$, the rate of drug absorption equals the rate of drug elimination.

  • Elimination Rate Constant ($K_{el}$ or $k$): Fractional rate of drug removal per unit time; used to directly calculate elimination half-life.

  • Absorption Rate Constant ($K_a$): Quantitative measure governing the rate of systemic drug absorption and $T_{\max}$.

  • Elimination Half-Life ($t_{1/2}$): The time required for the plasma drug concentration to decrease by 50%. Approximately 5 half-lives ($5 \times t_{1/2}$) are required to eliminate $> 95\%$ of an administered dose from the body. Dictates clinical maintenance dosing intervals ($\tau$) to achieve steady-state concentrations without complete inter-dose clearance.

  • Area Under the Plasma Concentration-Time Curve (AUC): Measures total systemic exposure over time. Represents total presence of active drug in systemic circulation and correlates better with cumulative pharmacological activity than a single snapshot peak concentration ($C_{\max}$).

  • Apparent Volume of Distribution ($V_d$): Proportionality factor relating the total amount of drug in the body to its measured plasma concentration.

  • Clearance ($CL$): The volume of plasma completely cleared of drug per unit time across all elimination pathways.

  • Bioavailability ($F$): The systemic fraction of an administered non-IV dose that reaches systemic circulation unchanged.

• Case Example: Bosotinib and Rifampin Drug-Drug Interaction (DDI):

  • Bosotinib: Small-molecule anticancer tyrosine kinase inhibitor.

  • Rifampin (Rifampicine): Anti-tuberculosis drug and the most potent known inducer of cytochrome P450 3A4 (CYP3A4) enzymes.

  • DDI Mechanism: In Phase 1 clinical DDI trials (typically requiring ~1 year to complete), co-administration of rifampin with bosotinib causes a drastic reduction in bosotinib $C_{\max}$ and total AUC on a log-concentration vs. time plot. Rifampin dramatically induces hepatic and intestinal CYP3A4, accelerating bosotinib elimination and causing significant loss of therapeutic efficacy.

Phases of Oral Drug Action and Gastrointestinal Absorption

• Three Sequential Phases of Oral Drug Action:

  1. Pharmaceutical Phase: Disintegration of the solid dosage form (tablet/capsule) into small particles to increase surface area, followed by dissolution of the active drug in gastrointestinal (GI) fluids.

  2. Pharmacokinetic Phase: Simultaneous absorption, tissue distribution, metabolic transformation, and renal/biliary excretion.

  3. Pharmacodynamic Phase: Interaction of systemic drug molecules with biological target receptors yielding therapeutic or toxic responses.

• Physicochemical Rules for Optimal Oral Drug Absorption:

  • Molecular Weight ($MW$): Optimal absorption occurs when $MW < 500\,g\,mol^{-1}$.

  • Lipophilicity / Partition Coefficient: Ideal octanol-water partition coefficient ($\log(P)$) is approximately 3 (lipophilic solubility is 3-fold higher than aqueous solubility).

  • Solubility vs. Permeability Trade-off: Drugs (mostly weak bases or weak acids) must dissolve in GI fluids (stomach pH 1.2 transitioning to small intestine pH 6.8 to 7.0) in their un-ionized, lipophilic state to permeate lipid membranes via passive diffusion.

• Anatomical Dynamics of GI Absorption:

  • Surface Area Comparison: The stomach provides a surface area of approximately $3,333\,cm^2$, whereas the small intestine expands to nearly $2,000,000\,cm^2$ due to mucosal folds, villi, and microvilli.

  • Site of Maximal Absorption: Although weak acids are un-ionized in acidic stomach fluids according to pH-partition theory, the massive surface area of the small intestine makes it the primary site of absorption for both weak acids and weak bases.

• Mechanisms of Epithelial Transport:

  • Passive Diffusion: Automatic, non-saturable transcellular movement down a concentration gradient governed by Fick's Law, requiring zero energy expenditure. Supported by "sink conditions" where local microvascular blood flow rapidly carries absorbed drug away from the basal membrane to maintain a steep concentration gradient.

  • Paracellular Transport: Passive movement of small, highly water-soluble drugs through tight junctions between adjacent epithelial cells.

  • Carrier-Mediated Active Transport:

    • Facilitated Transport: Transporter-mediated passage down a concentration gradient without energy expenditure.

    • Primary Active Transport: Protein conformational changes move drugs against concentration gradients using direct ATP hydrolysis energy.

    • Secondary Active Transport: Solute movement coupled to electrochemical ion gradients (e.g., $Na^+/K^+$ ATPases).

Drug Transporters in Absorption and Disposition

• Solute Carrier (SLC) Superfamily:

  • Organic Anion Transporting Polypeptides (OATP): Influx transporters facilitating uptake of weakly acidic or anionic drugs across GI, hepatic, and renal membranes.

  • Organic Cation Transporters (OCT): Influx transporters handling weakly basic or cationic drugs.

  • Clinical Substrate Example: Metformin (anti-diabetic agent) relies heavily on OCTs for renal clearance, requiring precise clinical dose adjustments based on renal function.

• ATP-Binding Cassette (ABC) Superfamily (Efflux Transporters):

  • P-glycoprotein (P-gp / ABCB1 / MDR1): Discovered in 1987 in drug-resistant cancer cells; functions as a cellular "bouncer". Utilizes energy from ATP hydrolysis ($ATP \rightarrow ADP$) to pump hydrophobic and amphipathic drugs out of cells back into the GI lumen, bile, urine, or capillary blood.

  • Breast Cancer Resistance Protein (BCRP / ABCG2): Efflux transporter present in intestinal mucosa, blood-brain barrier, liver, and breast tissue.

  • Representative Substrates: Cyclosporine, loperamide, digoxin, HIV protease inhibitors, and various immunosuppressants.

• First-Pass Metabolism and Oral Bioavailability ($F$):

  • Bioavailability Equation:         $F = \frac{\text{AUC}_{oral}}{\text{AUC}_{IV}}$         (calculated by comparing equal oral and intravenous doses).

  • First-Pass Hepatic/Intestinal Effect: Splanchnic blood circulation draining the GI tract flows directly into the portal vein and through the liver prior to entering systemic circulation. Pre-systemic elimination by gut wall enzymes (CYP3A4) and hepatic metabolism significantly reduces $F$

  • Bioavailability Spectrum Examples:

    • Letrozole: 100% bioavailability ($F = 1.0$); $\log(P) \approx 3$, not a P-gp substrate, half-life $\approx 60\,hours$.

    • Propranolol: Low oral bioavailability ($F = 0.25$) due to extensive first-pass hepatic metabolism.

    • Actonel (risedronate): Osteoporosis therapy developed by Procter & Gamble with an extremely low oral bioavailability of $\approx 0.5\%$ ($F = 0.005$).

Principles of Drug Distribution and Body Fluid Compartments

• Determinants of Tissue Distribution:

  • Organ Perfusion Rate: High-cardiac-output organs (liver receiving ~25% of cardiac output, kidneys ~20%, heart, lungs) rapidly reach distribution equilibrium. Low-perfusion tissues (skin, bone, adipose tissue) equilibrate slowly.

  • Plasma Protein Binding: Reversible binding to circulating plasma proteins such as albumin ($MW \approx 69,000\,Da$) or $\alpha_1$-acid glycoprotein. Only free (unbound) drug can permeate capillary walls to reach target tissues. Bound drug serves as a circulating reservoir that continuously dissociates to replenish free drug as it is cleared.

• Specialized Biological Barriers and Non-Invasive Matrix Sampling:

  • Blood-Brain Barrier (BBB): Endothelial cells featuring continuous tight junctions and dense luminal P-gp expression, restricting access to small, lipophilic, non-P-gp substrate molecules.

  • Placental Barrier: Evaluated in pre-clinical animal models (e.g., sheep) during drug development to prevent fetal teratogenicity.

  • Salivary Excretion: Free, un-ionized weakly basic drugs partition into saliva. Non-invasive sampling via "Salivette" cotton swab devices under the tongue provides a practical PK tool for pediatric populations, requiring high-sensitivity LC-MS/MS bioanalysis at nanogram concentrations.

• Apparent Volume of Distribution ($V_d$):

  • Anatomical Fluid Volumes (70 kg adult; Total Body Water $\approx 60\% = 42\,dm^3$):

    • Plasma Volume: $\approx 3.5\,dm^3$ (Total Blood Volume $\approx 5.0\,dm^3$).

    • Extracellular Fluid (ECF): $\approx 15.0\,dm^3$ (plasma + interstitial fluid).

    • Intracellular Fluid (ICF): $\approx 27.0\,dm^3$.

  • Mathematical Calculation:         $V_d = \frac{\text{Dose}_{IV}}{C_{p0}}$         Where $C_{p0}$ is the extrapolated initial plasma concentration at time zero post-IV bolus.

  • Interpretation of Apparent $V_d$: $V_d$ is an apparent parameter rather than a real physiological volume. Drugs that bind extensively to tissue proteins or intracellular structures leave very little free drug in circulation ($C_{p0}$ is extremely small), yielding calculated $V_d$ values that far exceed total body water (e.g., $400\,dm^3$; analogous to adding active charcoal to a beaker to adsorb free drug out of solution).

Mechanisms of Drug Clearance and Excretion

• Primary Pathways for Top 200 Drugs:

  • Hepatic Metabolism (CYP450 enzymes): Eliminates $\approx 75\%$ of top clinical drugs by converting lipophilic parent compounds into hydrophilic metabolites.

  • Renal Excretion: Eliminates $\approx 20\%$ unchanged.

  • Biliary/Fecal Excretion: Eliminates $\approx 5\%$.

• Renal Excretion Mechanics:

  • Net Excretion Equation:         $\text{Amount Excreted} = \text{Amount Filtered} + \text{Amount Secreted} - \text{Amount Reabsorbed}$

  • Functional Steps: Passive glomerular filtration of unbound plasma drug, active tubular secretion driven by renal SLC transporters (OATP, OCT), and passive or active tubular reabsorption back into peritubular capillaries.

Pharmacogenetics: Tripartite Classification Framework

• Genomic Foundations:

  • The human genome contains $3 \times 10^9$ base pairs; individuals are 99.7% identical, with 0.3% inter-individual single nucleotide polymorphisms (SNPs) and structural variants driving differential drug responses.

• Tripartite Classification Scheme:

  1. Drug Target Pharmacogenetics (PD): Genetic variation in drug receptors, enzymes, or signaling proteins. Inactivating null mutations are extremely rare because drug targets are essential for normal physiological function. Minor amino acid substitutions or promoter SNPs produce continuous bell-shaped Gaussian population distributions.

  2. Drug Transport Pharmacogenetics (PK): Variations in influx/efflux transporter proteins. Inactivating mutations are generally incompatible with normal cellular transport; variations produce continuous bell-curve population shifts.

  3. Drug Metabolism Pharmacogenetics (PK): Variations in drug-metabolizing enzymes (CYP450s). Inactivating/null mutations ARE common due to functional gene redundancy (gene deletions, premature stop codons), creating discrete, bimodal or multimodal population sub-groups (represented as separate loss-of-function bumps outside the main bell curve).

Drug Target Pharmacogenetics Examples and Clinical Nomenclature

• Drug Class Nomenclature Suffixes:

  • -olol: Beta-adrenergic receptor antagonists / Beta-blockers (e.g., metoprolol).

  • -mab: Monoclonal antibodies (e.g., trastuzumab).

  • -prazole: Proton pump inhibitors for gastric acid suppression.

  • -navir: HIV protease inhibitors (e.g., ritonavir, saquinavir).

• $\beta_1$-Adrenergic Receptor ($\beta_1$-AR) Polymorphisms:

  • Structure: 7-transmembrane GPCR regulating heart rate and cardiac contractility.

  • Polymorphic Site 49 (Extracellular N-terminus): Alters ligand binding affinity.

  • Polymorphic Site 389 (Intracellular C-terminus): Arginine (Arg389) vs. Glycine (Gly389).

  • Functional Effect: Arg389 triggers strong G-protein coupling and robust adenylyl cyclase activation, generating elevated cyclic AMP (cAMP) and high blood pressure. Gly389 produces a weak intracellular signal.

  • Clinical Significance: Patients harboring the Arg389 allele show a significantly greater antihypertensive response to metoprolol than Gly389 patients because the Arg389 receptor is a major pathophysiological driver of their hypertension.

• HIV Pharmacotherapy and Chemokine Co-Receptor Genetics:

  • Central Dogma Paradigm Shift: Discovery of reverse transcriptase by David Baltimore proved genetic information can flow backward from RNA to DNA in retroviruses.

  • HAART Therapy: Highly Active Antiretroviral Therapy combines 2–3 nucleoside reverse transcriptase inhibitors (NRTIs), 1–2 protease inhibitors (-navir), and 1 non-nucleoside reverse transcriptase inhibitor (NNRTI).

  • Chemokines: Small (8–10 kDa) chemotactic cytokines categorized by cysteine structural motifs (CC, CXC, CXXXC, C) that signal through GPCRs to drive directional cell migration (chemotaxis).

  • CCR5 Co-Receptor Discovery: HIV cell entry requires binding to CD4 and a chemokine GPCR co-receptor (CCR5 or CXCR4). Genomic studies of high-risk African sex worker cohorts resistant to HIV identified a protective homozygous deletion mutation in CCR5 ($\Delta 32$), leading to the development of CCR5 antagonist drugs (e.g., maraviroc).

Theranostics, Companion Diagnostics, and Geno-Stratification

• Theranostics Paradigm: The integration of targeted therapeutics with companion diagnostic testing to identify candidate patient populations prior to treatment.

• HER2-Positive Breast Cancer Model:

  • $\approx 75\%$ of breast cancers are estrogen-dependent; $\approx 25\%$ overexpress Human Epidermal Growth Factor Receptor 2 (HER2, an EGF receptor family member).

  • $\approx 5\%$ of breast cancers are "triple-negative" (lacking estrogen, progesterone, and HER2 receptors; hardest to treat).

  • Herceptin (trastuzumab, anti-HER2 mAb) was co-developed alongside the companion diagnostic Hercept-test (immunohistochemical tissue staining) to identify HER2-overexpressing patients.

• Geno-Stratification in Clinical Trials:

  • Step 1: Perform genomic stratification screening to exclude non-responders lacking the molecular target.

  • Step 2: Screen responder sub-groups for genetic markers associated with adverse drug reactions to exclude individuals at high risk for toxicity.

Pharmacogenetics of Drug Transporters: ABCB1 / PGP / MDR1

• Nomenclature and Tissue Expression of ABCB1:

  • Synonymous Names: P-glycoprotein (PGP), Multidrug Resistance 1 (MDR1), and ATP-Binding Cassette Subfamily B Member 1 (ABCB1).

  • Localization: ATP-dependent efflux pump located in intestinal epithelium, liver canalicular membranes, renal proximal tubules, blood-brain barrier, blood-testis barrier, placenta, pancreas, and adrenal glands.

• Substrate Promiscuity:

  • Transports a broad array of hydrophobic and amphipathic molecules, including anticancer agents, cardiac glycosides, HIV protease inhibitors (-navir), immunosuppressants (cyclosporine, FK506, corticosteroids), antibiotics, antihistamines, and endogenous steroids/lipids.

• Genomic Variability and Clinical Impact:

  • Contains 16 to 19 polymorphic sites (e.g., C3435T) that alter P-gp expression levels and intestinal efflux activity.

  • Drives extensive inter-individual variability in the oral bioavailability and systemic blood levels of narrow therapeutic index drugs (e.g., organ transplant immunosuppressants), creating severe clinical dosing challenges.

• Digoxin Drug-Drug Interactions and Historical Case Study:

  • Digoxin is a cardiac glycoside substrate for P-gp.

  • Quinidine Interaction: Inhibits P-gp, competing for transport and increasing plasma digoxin concentrations 2- to 3-fold.

  • Rifampin Interaction: Induces P-gp expression, increasing intestinal/hepatic efflux and decreasing plasma digoxin concentrations to sub-therapeutic levels.

  • Historical Application: Serbian leader Slobodan Milošević, on trial for war crimes (including the Srebrenica massacre) at the International Criminal Court in The Hague, was treated with digoxin for severe hypertension. Forensic blood testing revealed unprescribed rifampin, which blunted his digoxin therapy. The drug interaction was orchestrated by associates to manufacture an untreatable medical crisis in an attempt to secure transfer to Moscow, Russia. Milošević died in his cell prior to trial completion; co-conspirators Radovan Karadžić and Ratko Mladić ("butcher of Srebrenica") were later convicted.