Patrick: An Introduction to Medicinal Chemistry 4e Case Study 1 STATINS ANTI-CHOLESTEROL AGENTS

1. Cholesterol Notes Important in biosynthesis and cell membrane structure Excess cholesterol leads to cardiovascular disease Fatty molecule transported round blood supply by low-density and high-density lipoproteins (LDLs and HDLs) LDLs carry cholesterol to cells HDLs carry cholesterol from cells to liver Mortality is associated with high levels of LDLs or low levels of HDLs Cholesterol can cause fatty plaques in arteries leading to a risk of artherosclerosis, clot formation, stroke and heart attack

2. Target for statins Target enzyme Notes Inhibit biosynthetic pathway to cholesterol Prevent synthesis of cholesterol within cells but not from diet Target the enzyme catalysing the rate limiting step in the biosynthetic pathway

3. Catalytic mechanism Notes Involves two hydride transfers Two molecules of cofactor required (NADPH)

3. Catalytic mechanism Substrate binding

3. Catalytic mechanism Notes Lys, His, Glu and Asp are involved in reaction mechanism Histidine acts as acid catalyst Lysine stabilises negatively charged oxygen of mevaldyl-CoA and transition state leading to it Lowers activation energy for first step

3. Catalytic mechanism Notes Glutamic acid acts as an acid catalyst Aspartate residue stabilises uncharged Glu-559 and charged Lys-691

4. Identification of a Lead Compound Compactin (Mevastatin) IC50 = 23 nM Notes Screening of compounds produced by microorganisms Microbes lacking HMGR might produce HMGR inhibitors to inhibit microbes having HMGR - chemical warfare Compactin (Mevastatin) isolated from Penicillium citrinum 10,000 higher affinity for enzyme than substrate Never reached the market

5. Type I Statins Notes Lovastatin isolated from Aspergillus terreus IC50 = 24 nM Simvastatin Pravastatin IC50 = 1900 nM Notes Lovastatin isolated from Aspergillus terreus First statin to be marketed (Merck; 1987) Revolutionised treatment of hypercholesterolaemia Simvastatin introduced in 1988 as semi-synthetic analogue of lovastatin Pravastatin derived from compactin by biological transformation (1991)

5. Type I Statins Notes General structure of type I statins contains a polar head and a hydrophobic moiety including a decalin ring Lovastatin and simvastatin are prodrugs where lactone ring is hydrolysed to give the polar head

5. Type I Statins * Disadvantages of Type I statins = asymmetric centres Disadvantages of Type I statins Various side effects Difficult to synthesise Large number of asymmetric centres

6. Type II Statins Notes Synthetic agents Fluvastatin IC50 = 28 nM Atorvastatin IC50 = 8 nM Cerivastatin IC50 = 10 nM Notes Synthetic agents Contain larger hydrophobic moiety with no asymmetric centres Easier to synthesise Fluvostatin (1994), atorvastatin (1997), cerivastatin (1998), rosuvastatin (2003)

6. Type II Statins Rosuvastatin IC50 = 5 nM Pitavastatin IC50 = 6.8 nM Notes Structures share a number of similar features (‘me too drugs’) Rosuvastatin is the most potent - related to sulfonamide group Cerivastatin is the most hydrophobic Pravastatin and rosuvastatin are the least hydrophobic

6. Type II Statins Notes Statins with lower hydrophobic character target liver cells and have lower side effects Less hydrophobic statins do not cross cell membranes easily Liver cells have transport proteins for statins whereas other cells do not Majority of cholesterol synthesis takes place in liver cells Side effects thought to be due to inhibition of HMGR in other cells such as muscle cells Common side effect is myalgia (muscle pain) Rhabdomyolysis = severe muscle toxicity which can be fatal Cerivastatin withdrawn in 2001 due to rhabdomyolysis and 50 fatalaties

7. Statins - Mechanism of action = Notes Competitive inhibitors of HMGR Polar head group mimics the natural substrate (HMG-SCoA) Same binding interactions for polar head group as natural substrate Hydrophobic moiety forms additional binding interactions Binds more strongly than natural substrate, but does not undergo reaction - no leaving group

7. Statins - Mechanism of action Notes Statins are closer mimics of the first reaction intermediate mevaldyl CoA than the substrate Statins likely to bear a resemblance to the transition state for the first stage of the reaction mechanism Can be viewed as transition-state analogues

8. Statins - Binding interactions Notes Polar head group binds in similar manner to substrate Hydrophobic moiety does not bind to the pocket for SCoA Enzyme is flexible and alters shape to accommodate statins Hydrophobic pocket is created to bind the hydrophobic moiety

8. Statins - Binding interactions methylethyl substituent Methylethyl substituent of Type II statins binds to same region as decalin ring of type I statins

8. Statins - Binding interactions fluorophenyl substituent Arg-590 forms important polar interaction with fluorophenyl substituent Planar guanidium group is also stacked over the phenyl ring

8. Statins - Binding interactions amide group Amide group forms an additional hydrogen bonding interaction with Ser-565 Additional interaction not formed with other statins other than rosuvastatin

8. Statins - Binding interactions sulfone group Notes Rosuvastatin forms additional H-bonding interactions Sulfone oxygen forms a hydrogen bonding interaction with Ser 565 Sulfone group also interacts uniquely with Arg-568 Explains why rosuvastatin is most potent statin Sulfone group important for binding as well as selectivity

9. Other mechanisms of action for statins Notes Statins inhibit HMGR in liver cells Lowers the levels of cholesterol in liver cells Causes an increase in the synthesis of hepatic LDL receptors Increases the number of LDL receptors in the cell membrane of liver cells Increases the amount of LDL-cholesterol cleared from the plasma Crucial to the effectiveness of statins