11

Phosphodiesterase inhibitors:Phosphodiesterase inhibitors:selection of characteristics selection of characteristics

determining biological activitydetermining biological activity

A. KnyshA. KnyshV.A. PotemkinV.A. Potemkin20112011

22

PDE4 inhibitors are used in treatment of:PDE4 inhibitors are used in treatment of:

• Bronchial asthma Bronchial asthma • Chronic obstructive pulmonary disease (emphysema + chronic bronchitis)Chronic obstructive pulmonary disease (emphysema + chronic bronchitis)

AsthmaAsthma is characterized by is characterized by reversible bronchoconstrictionreversible bronchoconstriction caused by airway caused by airway hyper-responsiveness to a variety of stimuli. They include immunologic reactions hyper-responsiveness to a variety of stimuli. They include immunologic reactions to environmental allergens, viral infections and inhaled air pollutants.to environmental allergens, viral infections and inhaled air pollutants.

Chronic bronchitisChronic bronchitis is characterized by is characterized by hypersecretionhypersecretion ofof mucusmucus,, inflammation and inflammation and decrease of airway lumendecrease of airway lumen. Although the most important cause is cigarette . Although the most important cause is cigarette smoking, other air pollutants, such as sulfur dioxide and nitrogen dioxide, may smoking, other air pollutants, such as sulfur dioxide and nitrogen dioxide, may contribute. contribute.

33

N

N N

N

O

O

CH3

H3C

CH3

N

N N

HN

O

O

CH3

H3C

Methylxanthines as first PDE inhibiting agents Methylxanthines as first PDE inhibiting agents

theophyllinetheophylline caffeinecaffeine

• non-selective (inhibits all 5 types of PDE)non-selective (inhibits all 5 types of PDE)• inhibits high-polymeric fraction of PDE inhibits high-polymeric fraction of PDE whose activity is increased only during the whose activity is increased only during the asthmatic attackasthmatic attack• theophylline derivatives with higher PDE theophylline derivatives with higher PDE inhibitory activity are often proved to be inhibitory activity are often proved to be inefficient bronchial spasmolytics inefficient bronchial spasmolytics • theophylline spasmolytic effect is theophylline spasmolytic effect is determined not only by PDE inhibitory activitydetermined not only by PDE inhibitory activity• antagonist of A1, A2 adenosine receptorsantagonist of A1, A2 adenosine receptors• inhibits slow calcium channelsinhibits slow calcium channels• inhibits free oxygen radical formation and inhibits free oxygen radical formation and releasing of inflammatory agentsreleasing of inflammatory agents• may cause side effectsmay cause side effects

• is used as a stimulating agent for is used as a stimulating agent for nervous systemnervous system• also inhibits PDE and adenosine also inhibits PDE and adenosine receptors which is considered to be receptors which is considered to be a mechanism of its stimulating effecta mechanism of its stimulating effect• is not used for treatment of is not used for treatment of bronchial asthma because of its low bronchial asthma because of its low selectivity and efficiency in selectivity and efficiency in comparison with theophyllinecomparison with theophylline• PDE inhibitory effect is reported to PDE inhibitory effect is reported to show in concentrations which are show in concentrations which are behind the therapeutic corridorbehind the therapeutic corridor• stimulating effect is much more stimulating effect is much more evident than that of theophyllineevident than that of theophylline

44

Mechanism of actionMechanism of action

cAMPcAMPadenylate cyclaseadenylate cyclase phosphodiesterasephosphodiesterase

ATPATP AMPAMP

receptorreceptor+ agonist+ agonist PDE4 inhibitorsPDE4 inhibitors

CaCabasophilebasophile

smooth smooth musclemuscle

cellcell

55

Enzyme phosphodiesterase 4 as molecular targetEnzyme phosphodiesterase 4 as molecular target

• Hydrolase 3.1.4.17Hydrolase 3.1.4.17• 4 chains A, B, C, D4 chains A, B, C, D• Zn2+, Mg2+Zn2+, Mg2+• Catalytic domainCatalytic domain

Investigation of theInvestigation of the crystal structures of the PDE4D2 catalytic domain in crystal structures of the PDE4D2 catalytic domain in complex with (R)- or (R,S)-rolipram complex with (R)- or (R,S)-rolipram suggests suggests that inhibitor selectivity is that inhibitor selectivity is determined by the chemical nature of determined by the chemical nature of aminoacidsaminoacids and subtle and subtle conformational changes of the binding pockets. The conformational conformational changes of the binding pockets. The conformational states of states of Gln369Gln369 in PDE4D2 may play a key role in inhibitor recognition. in PDE4D2 may play a key role in inhibitor recognition.

66

AimsAims

using data on biological activity (% of in vitro inhibited enzyme), using data on biological activity (% of in vitro inhibited enzyme), and calculating molecular characteristics with special programs, and calculating molecular characteristics with special programs, to find, what this activity depends onto find, what this activity depends on

Structure (x)Structure (x)knownknown

BiologicalBiologicalactivity (y)activity (y)

knownknown??functionfunction

unknownunknown

77

Training setTraining set

73 compounds, taken from Online Chemical Modeling Database 73 compounds, taken from Online Chemical Modeling Database (www.ochem.eu Helmholtz Zentrum, Munich, Germany)(www.ochem.eu Helmholtz Zentrum, Munich, Germany)

Basic Basic structure structure (group)(group)

Benzothiadiazine Benzothiadiazine derivatives derivatives

Phtalasine Phtalasine derivativesderivatives

Theophylline Theophylline derivativesderivatives Oxime derivativesOxime derivatives

FormulaFormula

NumberNumber 2929 1212 88 1414

Mean Mean activityactivity 70%70% 23%23% 41%41% 50%50%

N

NHN

N

O

O

R

R = NHx(SO2C6H5)2-x,NHSO2C6H4CF3/OCH3,CH2C6H2(OCH3)3

N

SN

O

O

R

R

RO

R = NO2, Cl, OCH3, OH

N

NHO

N

Cl

Cl

R

R = N(CH3)2, CO, CON(CH3)2

O

O

NO

OR

R'

R = H, COCH3, COC6H5R' = N(CH2CH2OH)2,C(CH3)2CH2CH2OH,morpholine, piperidine,pyrrolidine, piperazine

88

methods/programsmethods/programs

• HyperChem – visualization of molecules: before HyperChem – visualization of molecules: before calculation (using just one method of optimization, offered calculation (using just one method of optimization, offered by HyperChem); after calculations, including complexes by HyperChem); after calculations, including complexes with virtual receptor, presented by set of pseudoatoms) with virtual receptor, presented by set of pseudoatoms)

• MERA, MERA_MAGIC, Prok1: calculation of different MERA, MERA_MAGIC, Prok1: calculation of different descriptors descriptors

• BiS: calculations of virtual receptor, using molecule set BiS: calculations of virtual receptor, using molecule set in consecutive order to reconstruct the generalized self-in consecutive order to reconstruct the generalized self-consistent complementary receptor for the full datasetconsistent complementary receptor for the full dataset

• STATISTICA, FieSta: mathematical processing: STATISTICA, FieSta: mathematical processing: correlation coefficients (>0,7; >0,5), 1,2,3 factor analysis; correlation coefficients (>0,7; >0,5), 1,2,3 factor analysis; regression models: graphs and equationsregression models: graphs and equations

99

original dataoriginal data

visualization visualization before before

calculationcalculation

CCAALLCCLLAATTIIOONN

visualization visualization after calculationafter calculation

virtual receptorvirtual receptor

statisticsstatistics

1010

Interpretation of resultsInterpretation of results

• bond stretch energy (‘bond’); bond stretch energy (‘bond’); • geometric sectiongeometric section (‘Ge_Sec3’); (‘Ge_Sec3’);• number of types of molecular associates (‘NC’);number of types of molecular associates (‘NC’);• molecule surface areamolecule surface area (‘S’); (‘S’);• sum of atomic volumessum of atomic volumes (‘VOINO1R’); (‘VOINO1R’);• sum of free (non-overlapping) carbon atomic volumessum of free (non-overlapping) carbon atomic volumes (‘vf0ug’); (‘vf0ug’);• sum of free carbon atomic surfacessum of free carbon atomic surfaces (‘snefug’) (‘snefug’)

Inhibitory activity mainly (correlation coefficients>0,7) depends on:Inhibitory activity mainly (correlation coefficients>0,7) depends on:

R =R = 0.705; inh = 0.117128*0.705; inh = 0.117128*bondbond + + 0.118559 (1 factor analysis)0.118559 (1 factor analysis)R = 0.776; inh = R = 0.776; inh = −−0.0083120.008312**Ge_Sec3Ge_Sec3 + 0.019009* + 0.019009*NCNC + + 0.675160 (2)0.675160 (2)R = 0.812; inh = 0.069814*R = 0.812; inh = 0.069814*bondbond −−0.0189680.018968**SS + 0.009161* + 0.009161*NCNC + + 1.644785 (3)1.644785 (3)R = 0.707; inh = R = 0.707; inh = −−14.372214.3722**VOIN01RVOIN01R + + 17.6065 (1)17.6065 (1)R = 0.818; inh = R = 0.818; inh = −−0.0783800.078380**vf0ugvf0ug + 0.588445* + 0.588445*snefugsnefug + + 0.858496 (2)0.858496 (2)

1111

Inhibitory activity of the molecule Inhibitory activity of the molecule depends on its depends on its geometrygeometry to a to a marked degreemarked degree

Which is not of great wonder becauseWhich is not of great wonder because

Most effective molecule - target interactions (which usually means the highest activity) occur with molecules whose size let them complementary fill the binding site and have the strongest interactions with the atoms at the inlet part

1212

Inhibition/bond stretch energyInhibition/bond stretch energy Inhibition/electron-electron interactionsInhibition/electron-electron interactions

Inhibition/resonanceInhibition/resonanceInhibition/number of molecular associatesInhibition/number of molecular associates

NNOONNGGOOMMEETTRRIICC

FFAACCTTOORRSS

1313

Inhibition/sum of atomic volumesInhibition/sum of atomic volumes Inhibition/geometric sectionInhibition/geometric section

Inhibition/molecule surface areaInhibition/molecule surface areaInhibition/sum of free carbon Inhibition/sum of free carbon

atomic surfacesatomic surfaces

GGEEOOMMEETTRRIICC

FFAACCTTOORRSS

1414

Pseudoatoms of virtual receptorPseudoatoms of virtual receptor

The most active benzothiadiazineThe most active benzothiadiazine structure 95%structure 95%

Pde417

Theophylline 26%Theophylline 26%

Pde448