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QSAR features for inhibitors of mitochondrial bioenergetics. Anatoly A. Starkov.

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Presentation on theme: "QSAR features for inhibitors of mitochondrial bioenergetics. Anatoly A. Starkov."— Presentation transcript:

1 QSAR features for inhibitors of mitochondrial bioenergetics. Anatoly A. Starkov

2 H+H+H+H+ H+H+H+H+ Oxygen C-III SDH C C-IV C-I FMN IM NADH NAD + Succinate Fumarate Water e e e e e e e CoQ Fuel Supply System Electron transfer in the respiratory chain NADH Oxygen, CoQH 2 e H+H+H+H+ p.m.f. =  +  pH

3 1.What is “uncoupling”? 2.What are “uncouplers”? 3.What are the mechanisms of uncoupling? 4.How much uncoupling is toxic? 5.Is a class-independent QSAR model for uncouplers possible? What descriptors should be selected? 6.What models should be used to test the uncouplers? A. UNCOUPLING.

4 Classical definitions: Uncoupling of oxidative phosphorylation is a process de-coupling oxygen consumption from ATP production. Uncouplers: 1.Stimulate resting respiration. 2.Decrease ATP yield (P:O ratio). 3.Activate latent ATPase.

5 any energy-dissipating process competing for energy with routine mitochondrial functions, thus inducing a metabolically futile wasting of energy. Wallace KB, Starkov AA. Mitochondrial targets of drug toxicity. Annu Rev Pharmacol Toxicol. 2000;40:353-88. UNCOUPLING:.

6 Respiratory chain H+H+ H+H+ AH A-A- A-A- H+H+ H+H+ IM Matrix A-A- AH Respiratory chain H+H+ H+H+ AH A-A- H+H+ H+H+ IM Matrix  pH  HA 2 - AH A-A- Proton shuttling by lipophilic weak acids. substituted phenols trifluoromethylbenzimidazoles salicylanilides carbonylcyanide phenylhydrazones - + - +  pH  1. 2.

7 Blaikie FH, Brown SE, Samuelsson LM, Brand MD, Smith RA, Murphy MP. Targeting dinitrophenol to mitochondria: limitations to the development of a self-limiting mitochondrial protonophore. Biosci Rep. 2006 Jun;26(3):231- 43.

8 Terada H. Uncouplers of oxidative phosphorylation. Environ Health Perspect. 1990 Jul;87:213-8.

9 [uncoupler],  M State 4, nmol O 2 /min/mg State 3 respiration rate [Uncoupler] max

10 Respiratory chain H+H+ H+H+ RN + RN H+H+ H+H+ IM Matrix - +  pH  A-A- Respiratory chain H+H+ H+H+ RNA - H + RN H+H+ H+H+ IM Matrix  pH  A-A- RN - + Proton shuttling by lipophilic weak bases and ion pairs. amine local anesthetics 3. 4.

11 Respiratory chain H+H+ H+H+ AH A-A- A-A- H+H+ H+H+ IM Matrix P Protein –mediated uncoupling by non-permeating anions and protein modifying reagents. P: ATP/ADP translocator, Glutamate transporter Long-chain fatty acids, SDS, 2,4-DNP Respiratory chain H+H+ H+H+ H+H+ H+H+ IM Matrix P P: Uncoupling Protein 1 (UCP1), anion carriers, membrane-active peptides, Permeability transition Pore (mPTP). Long-chain fatty acids, SH-modifying reagents.  pH   pH  - + - + 5. 6.

12 Respiratory chain H+H+ H+H+ 2H + IM Matrix E U  pH  - + Ca 2+ U: Ca2+ uniporter. E: Ca2+ ionophores. 7. Ion cycling. (Variant : U=valinomycin, Ca 2+ =K +, E=nigericine)

13 RC H+H+ H+H+ Ca 2+ 2H + IM Matrix  pH  Ca 2+ 2H + U E precipitate + CypD + + Fuel Supply System PTP Cytosol Ca 2+ signal ER storage + 1. Normal Ca 2+ signaling: Uncoupling due to the permeability transition pore (mPTP). 8.

14 RC H+H+ H+H+ Ca 2+ 2H + IM Matrix  pH  Ca 2+ 2H + U E precipitate + CypD + + Fuel Supply System PTP Cytosol Ca 2+ flooding ER storage + 2. Pathological Ca 2+ flooding opens mPTP:

15 McLaughlin SG, Dilger JP. Transport of protons across membranes by weak acids. Physiol Rev. 1980 Jul;60(3):825-63. Classical efficient uncoupler: 4 { "@context": "http://schema.org", "@type": "ImageObject", "contentUrl": "http://images.slideplayer.com/10/2801376/slides/slide_15.jpg", "name": "McLaughlin SG, Dilger JP. Transport of protons across membranes by weak acids.", "description": "Physiol Rev. 1980 Jul;60(3):825-63. Classical efficient uncoupler: 4

16 McLaughlin SG, Dilger JP. Transport of protons across membranes by weak acids. Physiol Rev. 1980 Jul;60(3):825-63.

17 Steps:Mechanism typedescriptors Acquire H+1-5, 7pKa Adsorb to the membrane1-5, 7, (6)D(water-membrane) (partition coefficient) Partition into the membrane1-7, (6)K,K’(surface-core) (species partition coefficient) Cross the membrane1-5, 7k,k’(species) (translocation rate constant) Release H+ inside matrix1-5, 7pKa Cross the membrane1-5, 7k’(species) (translocation rate constant) Acquire H+1-5, 7pKa’ Information on the surrounding: pH out and in, lipid phase volume, lipid phase(s) dielectric constants and viscosity, gradient of the electrical membrane potential across the membrane, total amount of a compound. Minimum reasonable set of parameters to consider: Classical: 4 { "@context": "http://schema.org", "@type": "ImageObject", "contentUrl": "http://images.slideplayer.com/10/2801376/slides/slide_17.jpg", "name": "Steps:Mechanism typedescriptors Acquire H+1-5, 7pKa Adsorb to the membrane1-5, 7, (6)D(water-membrane) (partition coefficient) Partition into the membrane1-7, (6)K,K’(surface-core) (species partition coefficient) Cross the membrane1-5, 7k,k’(species) (translocation rate constant) Release H+ inside matrix1-5, 7pKa Cross the membrane1-5, 7k’(species) (translocation rate constant) Acquire H+1-5, 7pKa’ Information on the surrounding: pH out and in, lipid phase volume, lipid phase(s) dielectric constants and viscosity, gradient of the electrical membrane potential across the membrane, total amount of a compound.", "description": "Minimum reasonable set of parameters to consider: Classical: 4

18 Spycher S, Smejtek P, Netzeva TI, Escher BI. Toward a class-independent quantitative structure--activity relationship model for uncouplers of oxidative phosphorylation. Chem Res Toxicol. 2008 Apr;21(4):911-27.

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20 McLaughlin SG, Dilger JP. Transport of protons across membranes by weak acids. Physiol Rev. 1980 Jul;60(3):825-63. Black lipid membranes as a model to test the intrinsic efficiency of uncouplers:

21 Ilivicky J, Casida JE. Uncoupling action of 2,4-dinitrophenols, 2- trifluoromethylbenzimidazoles and certain other pesticide chemicals upon mitochondria from different sources and its relation to toxicity. Biochem Pharmacol. 1969 Jun;18(6):1389-401. Isolated mammalian mitochondria as a model to test the toxicity of uncouplers

22 1.What do they do? – inhibit electron transport thereby suppressing  H + generation and stimulating ROS production. 2.How many are known? – a few hundreds of natural compounds and a gazillion of synthetic chemicals. 3.Are there some common chemical features in these compounds? – yes and no. 4.Is their MOA similar? – yes and no. 5.Is a class-independent QSAR model for the RC inhibitors possible? – Perhaps, but not there yet. 6.Why it is so? – insufficient knowledge of RC complexes and their structural diversity. 7.What models should be used to test the RC inhibitors? – isolated mammalian mitochondria. B. Inhibitors of the respiratory chain complexes.

23 Oxygen C-III SDH C-I FMN C C-IV IM NADH NAD + Water e e e e e CoQ ROSROS ROS ROSROS Succinate Fumarate e e Fuel Supply System

24 Q i site Q o site b low b high e e e e e e ISP Cyt.c1 Cyt.c Myxothiazol Antimycin Stigmatellin QiQi QoQo QH 2 Q Matrix side -  +  IM A QH 2 CoQ:Cytochrome c reductase (RC Complex III)

25 Antimycin A Myxothiazol Stigmatellin Classical inhibitors of CoQ:Cytochrome c reductase (RC Complex III)

26 N3 FMN N1b N4 N5 N7 N2 N6a N6b N1a Complexity of mammalian NADH:CoQ reductase (RC Complex I)

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28 Schuler F, Casida JE. The insecticide target in the PSST subunit of complex I. Pest Manag Sci. 2001 Oct;57(10):932-40 PSST subunit of Complex I is a common target for many and various inhibitors. Inhibitor binding site

29 Different classes of the Q site Complex I inhibitors. Degli Esposti M. Inhibitors of NADH-ubiquinone reductase: an overview. Biochim Biophys Acta. 1998 May 6;1364(2):222-35.

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31 Future developments toward QSAR model of mitochondrial poisons: 1. Create a realistic biophysical model of the inner mitochondrial membrane; 2. Obtain more detailed information on the molecular structure of mitochondrial proteins targeted by toxins; 3. Create a unified database of mitochondrial toxins and analyze it toward both their molecular properties and the mechanisms of intrinsic activity; 4. Create a good team of researchers with proper expertise (and funding) to develop and validate QSAR models in a relevant biological model (isolated mitochondria) under physiologically meaningful conditions.

32 [O 2 ]=0 Coupled respiration ADP Mito O 2 consumed 50 nmol O 2 1 min State 4 State 4’ State 3  ADP [O 2 ]=0 100 nmol ATP 1 min ADP [ATP]  (~20 mV) V state 3 V state 4,4’ ADP:O

33 50 nmol O 2 1 min ADP [O 2 ]=0 2,4-DNP Mito O 2 consumed Uncoupled respiration  ADP [O 2 ]=0 100 nmol ATP 1 min ADP [ATP]  (~20 mV) 2,4-DNP V(u) state 3 V(u) state 4,4’ ADP:O(u)

34 < > = UncoupledCoupled Uncoupling: less ATP for the same O 2 and substrates V state 3 V state 4,4’ ADP:O V(u) state 3 V(u) state 4,4’ ADP:O(u)

35 Ilivicky J, Casida JE. Uncoupling action of 2,4-dinitrophenols, 2- trifluoromethylbenzimidazoles and certain other pesticide chemicals upon mitochondria from different sources and its relation to toxicity. Biochem Pharmacol. 1969 Jun;18(6):1389-401.

36 ROS production is regulated by  H 2 O 2 emission, % of max H 2 O 2 emission, pmol/min/mg

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