rings in (candidate) drugs - case stories

Rings in (Candidate) Drugs – Case Stories

Jonas Boström, CVMD iMED, AstraZeneca R&D, Mölndal, Sweden [email protected]

mailto:[email protected]

A Five Case Story Rollercoaster

key computational approach for designing candidate drugs often the use of shape and electrostatic comparisons between molecules

1) for (purely) geometrical reasons 2) replacing unwanted functional groups (acids and esters) 3) to fine-tune electrostatics for improved properties

2000

CB1 scaffold-hopping –>

only geometry matters 1

LXR scaffold-hopping

and ”luck” 1

P2Y12 ester replacement

heterocycle 2

NOFI acid bioisostere –

> heterocycle 2

now

”Matched-Pair Project X”

heterocyclic electrostatics improved properties 3

Heterocyclic rings…


only geometry matters?

CB1 antagonists – a “me-too” project

2000

now

Rimonabant (Sanofi-Aventis): a selective high-affinity CB1 inverse agonist inhibits food intake in rats, mice, marmosets and man was in clinical studies for treatment of obesity

Cannabis increases food intake Marijuana users are often overweight inverse agonist (or antagonist)?

No CB1 protein X-ray structure ligand-based design

CB1 antagonists – a “me-too” project

Our scaffold-hopping strategy Replace the pyrazole scaffold in Rimonabant with different heterocyclic rings: thiazoles, pyrroles and pyrazines Check chemical feasibility Check shape-complementarity to the putative bioactive conformation of Rimonabant Mini-libraries of 20 cpds/class were synthesized to evaluate the different classes.

Boström et al. “Scaffold hopping, synthesis and structure–activity relationships of 5,6-diaryl-pyrazine-2-amide derivatives: A novel series of CB1 receptor antagonists”. Bioorganic and Medicinal Chemistry, 2007,15, 4077.

Shape-Based Scaffold-Hopping

All classes virtually identical to Rimonabant Shape-overlays (ROCS). Tanimoto> 0.9

All central scaffolds “work”, and are electronically different – essential for geometrical reasons?

(compounds very lipophilic)

AZ-”pyrazines”

AZ-”pyrroles”

AZ-”thiazoles”

19 cpds synthesized 10 < 100nM (4 < 10nM)

33 cpds synthesized 17 < 100nM (4 < 10nM)

17 cpds synthesized 4 < 100nM (2<10nM)

+ from the literature/patents imidazoles, triazoles, ring-closures, etc

Withdrawal of marketing authorization for Rimnabant (FDA/EMEA). AZ Project stopped after candidate drug nomination.

Effect of AZ-”pyrazine” and Rimonabant on body-weight (BW) in cafeteria-diet, obese mice.

Cpd5e

CB1 Summary

AZ-”pyrazine”

AZ-”pyrazine”

any heterocycle

?

“The function of the central scaffold is not to make any direct interactions with the cannabinoid receptor, but rather to place the substituted phenyl rings and the N-carboxyamide fragments in an optimal 3D orientation”

Boström et al. “Scaffold hopping, synthesis and structure–activity relationships of 5,6-diaryl-pyrazine-2-amide derivatives: A novel series of CB1 receptor antagonists”. Bioorganic and Medicinal Chemistry, 2007,15, 4077.


and ”luck” 2015

LXR “Lucky” Scaffold-hopping

The Role of Conformational Analysis in the Design of LXR Agonists

“T0901317” 24 nM

“AZ HTS hit” 70 nM

A novel series of potent nonsteroidal

LXR agonists

LXR “Lucky” Scaffold-hopping Liver X Receptor (LXR) agonists for the treatment of atherosclerotic cardiovascular disease

Conformational ensembles OMEGA, shape-matching ROCS accessed via an interactive web-interface

The design of a novel series LXR agonists from a conformational analysis of two compounds:

Molecular Alignments

LXR Scaffold-hopping Systematic pair-wise shape comparison of low-energy conformation gave a few alignments

Conformational ensembles OMEGA, shape-matching ROCS accessed via an interactive web-interface

hydrophob

hydrophob

acceptor

acceptor

hydrophob

It seemed to make sense…

Tanimoto: 0.86

add one “H-bond acceptor”

Visual inspection of these favored the alignment which the N-substituted side-chains and the phenyl rings (pharmacophores/SAR) are all in approximate coincidence.

Then came the LXR X-ray structure… The right geometry (and potency)…but for the wrong reason… Conformation was essentially correct, but the pose wasn’t

PDB: 1PQC

LXR X-ray structures were solved… isothiazol-3(2H)-one 1,1-dioxides containing scaffold superior to HTS hit series

A novel series of potent nonsteroidal

LXR agonists “designed”

17 nM

lucky scaffold hopping

expansion

virtually identical

Project stopped after candidate drug nomination

“AZ HTS hit” 70 nM

Boström et al. “Do structurally similar ligands bind in a similar fashion?”. J.Med. Chem., 2006, 49, 6716.


heterocycle 2

P2Y12 – ethylester replacements

Easy chemistry – library design 160 cpds with variation of substituents on pyridine and phenyl SAR – affinity range for compounds binding/GTPgS

Zetterberg, Bach, Boström ”A novel series of piperazinyl-pyridine ureas as antagonists of the purinergic P2Y12 receptor” Bioorg. Med. Chem. Lett. 21 (2011) 2877–2881

SAR

Thrombosis is the formation of a blood clot inside a blood vessel, obstructing the flow of blood through the circulatory system. May cause stroke and sudden death.

HTS several hits Among the most interesting: the ”piperazinylpyridines” series

P2Y12 – Prevention of Thrombosis

Ethyl ester functionality a potential liability • generally high in vivo clearance due to hydrolysis


Can we replace the ester while retaining activity? …and increase metabolic stability by replacing the ethyl ester with bioisosteres


Non-cyclic ester replacements – inactive in P2Y12 binding and GTPgS

Design Strategy

Replace ethylester functionality by hydrolytically stable 5-membered heterocycle

Around 20 five-membered heterocyclic systems synthetically attainable evaluated.

Shape and electrostatics to classify similarity to ethyl ester

Heterocyclic Rings as Ethylester Replacements

“good”

+1

“mediocre”

+4

“bad”

+ a few

Ten compounds (structures shown) made and tested.

Bach, Boström, Zetterberg “5-Alkyl-1,3-oxazole derivatives of 6-amino-nicotinic acids as alkyl ester bioisosteres are antagonists of the P2Y12 receptor” Future Med. Chem., 2013, 5(17), 2037-2056

P2Y12 Results

Oxazoles a suitable replacement shape and electrostatics (ItsElectric*)

* ItsElectric and EON (OpenEye) are based on the same toolkit (ZAP) and theory (PB electrostatic potentials).

correctly suggests that 5-methyl-oxazole better than the 4-methyl oxazole

Conformations where possible ring-substitutents overlayed with the ethyl moiety selected

weak H-bond

acceptor

strong H-bond

acceptor

Shape & Electrostatic complementarities

vdW

P2Y12 Summary The potential metabolic instability of ethyl esters was addressed by shape and electrostatics rankings to select 5-membered heterocycles for synthesis. Hydrolytically stable 5-methyl derivatives were identified as P2Y12 antagonists. The physical nature of the shape and electrostatic similarity approach ensured that a non-obvious substructure change could be incorporated to mimic the behavior of the ethyl ester functionality. Not likely that other commonly used similarity measures (FPs, MCSS), would have predicted the oxazole fragment as similar to the ethyl ester, let alone correctly suggesting that the 5-methyl-oxazole was a better replacement for the ethyl ester than the 4-methyl oxazole. The levels of clearance and bioavailability made the oxazoles series attractive compounds for further development.

The ester subseries made it to the clinic – but lack of efficacy

…a P2Y12 X-ray structure was published (and PDB deposited) last year

P2Y12 Post-Summary

Where’s the H-bond donor? Ethylester fits tightly, and no room for ”missing” waters. SER-156 could adopt another rotamer (<4.0Å hbond) to carbonyl oxygen No interaction for ether oxygen (making sense…) Electron density quality

P2Y12 Post-Summary

PDB 4ntj

(Resolution 2.6Å)

“NOFI” heterocyclics as carboxylic acid isosteres

Electrostatic Similarities

2000


> heterocycle 2

Protein-protein interfaces provide an important class of drug targets receiving increased attention. The typical design strategy usually involves large molecules (peptides and macrocycles)

A simplistic(!) view of the mechanism

Novel Oral Fibrinolysis Inhibitors (NOFI)

Boström et al “Potent Fibrinolysis Inhibitor Discovered by Shape and Electrostatic Complementarity to the Drug Tranexamic Acid” J.Med.Chem. 2013, 56, 3273

Fibrinolysis is a process that prevents blood clots from growing and becoming problematic

X-ray crystallography – structures known

pdb: 1ceb (and 1cea)

Crystal structures of the recombinant Kringle 1 domain of human plasminogen in complexes with the ligands EACA and TXA (Mathews et al. Biochemistry, 1996, 35, 2567–2576)

Fibrinolysis Inhibition

Project goal: identify novel compounds which can be given at lower doses and less frequently than TXA, by improving some combination of the efficacy, bioavailability and clearance.

TXA and EACA are well-established clinical agents used to reduce blood-loss following surgery and trauma and to treat heavy menstrual bleeding, mild haemophilia and certain forms of von Willebrands disease

Tranexamic acid (TxA) EACA

Modest potency and non-optimal pharmacokinetic properties leading to inconvenient dosing (up to 6 g per day). Associated side-effects are also nausea and vomiting. This clearly limits their use and there is consequently an unmet need for new inhibitors of plasminogen with a more convenient dosing and a more acceptable side-effect profile.

Lysine analoges:

Two Fibrinolysis Inhibitors on the Market

Zooming in on TXA in complex with the recombinant kringle-1 domain of human plasminogen …

prominent electrostatic features zwitterionic...

pdb: 1ceb (TxA) pdb: 1cea (EACA)

Shape-based approaches often successful in virtual screening. In this case not be sufficiently discriminatory due to the simple molecular framework of the zwitterion TXA. This motivated us to compare electrostatic potentials

Low-throughput screening assay for testing putative blood clotting agents; select a small set of compounds

Shape-based screening not sufficiently discriminatory due to simple molecular framework of TXA

At the time, pre-filtering was a necessity (electrostatic calculations not fast enough)

“De-crapping” and visual inspection

Virtual Screen Strategy

Virtual Screening – Results 68 compounds screened – ‘only’ one hit Remaining compounds selected were either weakly active or inactive.

VS

Clot-Lysis plasma assay 0.8 µM Clot-Lysis plasma assay 3.1 µM

The potency of the lead compound 4-PIOL was found to be four times that of TXA, the current drug widely used in the clinic. Surprising that such an active compound from such as a small set of measurements was discovered.

Isoxazolone a bioisostere Visual inspection of the hydrogen-bonding network in the ligand−protein complex indicates that 4-PIOL and TXA bind in their zwitterionic forms (modelled in their neutral state). The shape of the binding site is well-defined, with both ligands filling the binding site.

isoxazolone as a carboxylic acid bioisostere (~5 times potent)

GABA 33nM

GABA 6nM

* Using ligand-based approach ROCS/EON

isoxazolone

ClotLys (Plasma): 0.8µM pKa acid: 4.0




ClotLys (Plasma): >100µM pKa acid: 4.0

Tetrazole the most common carboxylic

acid isostere, but is an imperfect surrogate.

None more potent

Acquired and/or synthesized heterocyclic compounds

Few similar compounds to 4-PIOL available internally and externally at the time*

Acquired and/or synthesized heterocyclic compounds





ClotLys (Plasma): >100µM pKa acid: 4.0

false positive

base acid

tetrazole shorter

Tetrazole vs isoxazolone

8.0Å (vs 9.3Å)

base acid too short

DMPK Profiling

The lipophilicity of 4-PIOL was determined to be low (logD < 0), an atypical liability. Hydrophilicity can be disadvantageous, mainly due to poor cell permeability (Caco-2), resulting in unfavorable DMPK characteristics in general and suggestive of difficulties with creating a successful oral formulation

The pKa of the basic amines in TXA and 4- PIOL are similar (9.7 and 10.5), and that the acid pKa’s are nearly identical (4.1 and 4.0).

Summary • 4-PIOL was identified as a four-fold more potent fibrinolysis inhibitor than TXA using a low

throughput screen where the compound selection was made using computational

techniques.

• The key computational approach to our contribution for finding the right lead compound

was shape and electrostatic comparisons between molecules.

• It is clear that the computational approach described in the present work identified a non-

trivial bioisostere of TXA as a high quality lead for a subsequent lead optimization program.

• It is difficulty to accurately assigning correct ionization states to a large collection of

molecules, supporting our approach of modeling all compounds in their neutral state.

• 4-PIOL served as an excellent starting point for subsequent lead optimization.

Analogs to 4-PIOL selective against GABA-A, with good cell permeability and retained potency with respect to plasminogen inhibition were designed, by means of two approaches, keeping the graph/framework.

• A “methyl-scanning” exercise on 4-PIOL and • molecular overlays of two structurally different series Leading to the same observation: substitution of the 2’-position increases GABA-A selectivity and improved cell permeability

Lead Optimization

Challenging synthetic chemistry issues were solved Project stopped after candidate drug nomination.

“4-PIOL served as an excellent starting point for subsequent lead optimization”

* Concerns over commercial value especially after Lysteda (TxA in a sustained slower-release formulation) launch in US.

Molecular Matched-Pair Project X

”Matched-Pair Project X” fine-tune heterocyclic electrostatics

improved properties 3

https://twitter.com/jmedchem background from MMP oxadiazole paper: Boström et al J. Med. Chem., 2012, 55, 1817

https://twitter.com/jmedchem

• “Molecules that differ only by a small structural change” (remainder of the molecule is exactly the same).

For example • The assumption underlying the matched-pair approach is that it is easier to predict

differences in the values of a property than it is to predict the value of the property itself (one reason is cancellation of errors).

• The observation of an effect across several chemical series increases our confidence that the effect is real -> general “rules of thumbs” -> future design

• One limitation is that it can only make predictions about structural features that have precedent in the given assay.

What Are Molecular Matched-Pairs?

Prazosin Terazosin Solubility: 1.1 mg/ml Solubility: 28.1 mg/ml

Oxadiazoles in an in-house AZ project

O

O N

NR1R2

O

N N

OR1R2

Binding Affinity: 80 nM Log D: 3 Solubility: NA HLM Clint: 20 µl/min/mg hERG: 2 µM

Binding Affinity: 40 nM Log D: 2 Solubility: 100 mM HLM CLint: 10 µl/min/mg hERG: 10 µM

1,3,4-oxadiazoles showed better properties than 1,2,4-oxadiazoles

in terms of lipophilicity, solubility, HLM Clint and hERG

Goal: to develop a better understanding for the generality of the observed effects, as well as if possible rationalize the effect.

1,3,4-oxadiazoles 1,2,4-oxadiazoles

Oxadiazoles are five-membered heterocycles containing two carbons, two nitrogens and one oxygen atom and they exist in several different regioisomeric forms

R' O

N N

R''

R' N

O N

R''

R'

N NO

R''

R' N

N O

R''

1,2,4-oxadiazoles

1,3,4-oxadiazole1,2,5-oxadiazole

CSD: NAXDIZ

CSD: ZZZTQC01

Side chains (R-groups) will have the same exit vector for 1,2,4 and 1,3,4 regioisomers: same overall shape

(but the 1,2,5 regioisomers differs)

Year

Freq

uen

cy o

f o

ccu

rren

ce o

f o

xad

iazo

les

in P

har

mac

euti

cal P

rep

arat

ion

s

Oxadiazoles in drug projects

Oxadiazole rings are used in drug discovery programs for different purposes

• contributing to the binding interactions with the target.

• modulates molecular properties through its position on the periphery of the molecule

• oxadiazoles has been used as replacements for carbonyl containing compounds (esters, carbamates, hydroxamic esters)

• as linker to orient its substituents appropriatly.

N

SNH

N

N

OO

O

ON

N

OH

ONO

N

F

1 (zibotentan) 2 (ataluren)

Structures of oxadiazole compounds in late stage clinical development or launched

F

NH

O

N

N

O

OH

NH

O

NN

O

3 (raltegravir)

Oxadiazole data set Using query-specific matched-pair tool 150 pairs.

The compounds essentially show the same characteristics as compounds with typical drug-like properties.

The data set covers molecules with different ionization states at physological pH;

and several structural series; ~25 clusters. ClogP

H-bond don+acc

MW

Check differences in lipophilicity ordered exp. logD for the 148 matched-pairs

1,3,4-oxadiazole are systematically less lipophilic than their 1,2,4-oxadiazole partner by one log unit

R' O

N N

R''R' N

N O

R''

1,2,4-oxadiazoles 1,3,4-oxadiazole

The median logD value for the 1,2,4-isomers

is 4.4, whereas it is 3.2 for the 1,3,4-isomers.

y = 0.99 + 1.06*x r2 = 0.97

Boström et al ”Oxadiazoles in Medicinal Chemistry” J. Med. Chem., 2012, 55, 1817

Matched Pairs Consequences

Lipophilicity is a cardinal property in drug discovery. It generally affects many other properties relevant in lead optimization. How would the difference in lipophilicity between the oxadiazole regioisomers impact other molecular properties? We investigated for example:

Solubility

hERG (and pKa)

CYP inhibition

HLM CLint

De

sire

d p

rop

ert

y

Lipophilicity/undesired property

1

2

3

?

Impact on solubility (subset of 55 pairs)

Lipophilicity can influence many other molecular properties, and perhaps the most obvious connection exists between lipophilicity and solubility.

R2=0.4

N=116

In general the 1,3,4-oxadiazole is more soluble than the 1,2,4-oxadiazole partner


In addition, the 1,3,4-oxadiazoles generally show better metabolic stability, in terms of lower HLM CLint values (34 matched-pairs).

Impact on CYP inhibition

For various CYP enzymes the inhibitory potency of the less polar 1,2,4-oxadiazole derivatives is typically more pronounced than that of the more polar 1,3,4-conterparts; this is particularly true for CYP3A4 (recognizing lipophilic substrates) and CYP1A2 (recognizing planar heterocycles).


Inhibition of the hERG channel can trigger Torsades de Pointes and arrhythmia One design strategy to reduce hERG inhibition is to decrease lipophilicity. In the majority of cases (N=11), the less lipophilic isomer (1,3,4) is also less potent at hERG.

All matched pairs in this subset have a basic functionality near the oxadiazole unit. There’s little or no change in pKa upon switching between the regioisomers. That is, the regioisomers do not have a significant effect on the basicity of the nearby groups.

Impact on hERG


Using electrostatics to rationalize Why are 1,3,4-oxadiazole less lipophilic? Ask your MedChemists (none knew) and calculate…

Dipole calculations at B3LYP/6-31G**. Experimental data confirms.

Kenny P. et al. J. Med. Chem. 2008, 51, 3720–3730, and Kenny P J. Chem. Soc. Perkin Trans 2 1994, 199-202

1,3,4-oxadiazole show significantly larger dipoles than their 1,2,4-oxadiazole partner The greater the dipole the greater polarity.

N

ON

O

N N

Minimized molecular electrostatic potential (Vmin) has been shown to be an effective predictor of the of hydrogen bond acceptors

- hydrogen bond acceptor strength changes and moves around

Other (unexpectedly) large differences

thiadiazole regioisomers

THF vs furan

We have for example investigated:

Thiadiazoles regioisomeric pairs

Calculated electrostatics show analogous differences to oxadiazole regioisomers

(3.7 vs 2.5D) although the difference is slightly smaller (3.3 vs 2.6D).

In thiadiazoles the oxadiazole oxygen atom is ‘replaced’ by a sulfur atom

Prediction: a 1:1 relationship between regioisomeric pairs, and that the 1,3,4-

thiadiazoles is less lipophilic than 1,2,4 regioisomers.

N

SN

S

N N

3.3 D

2.6 D

level of theory: HF/3-21G*

exit vectors differ somewhat

0

1

2

3

4

5

6

7

8

0 1 2 3 4 5 6 7 8

exp

lo

gD

1,2

,4-t

hia

dia

zole

s

exp logD

1,3,4-thiadiazoles

Differences in lipophilicity

1,3,4-thiadiazole are in general less lipophilic than their 1,2,4-thiadiazole partner

R' S

N N

R''R' N

N S

R''

1,2,4-oxadiazoles 1,3,4-oxadiazole

R2=0.92

log

D

The median logD value for the 1,2,4-isomers

is 3.2, whereas it is 2.3 for the 1,3,4-isomers.

N = 33

ordered exp. logD for the 80 compounds

1,2,4-thiadiazoles 1,3,4-thiadiazoles

Alpha-blocker

Solubility: 1.1 mg/ml

Log D (shake-flask): 1.8

ACDlogD: 0.0

clogP: 2.0

Alpha-blocker

Solubility: 28.1 mg/ml

Log D (shake-flask): 0.9

ACDlogD: -1.0

clogP: 2.2

A “me-too” observation: furan vs THF Modification of the hybridization state of carbon atoms can be useful to control molecular properties.

Prazosin (Minipress™) – Terazosin (Hytrin™) serves as a matched-pair “me-too” example.*

* Water solubility for Terazosin increases significantly when going from furan to THF. As a result, improved

bioavailability (90% vs 57%) and half-life (2–3 times) afford longer duration of action, and allow once-daily

administration. In addition, there is a safety concern associated with furans, since they are often anticipated to be

carcinogentic in humans.

Giordanetto, Boström and Tyrchan “Follow-on drugs: how far should chemists look?” Drug Discovery Today, 2011, 16 (15-16),722-732. (doi:10.1016/j.drudis.2011.05.011)

Prazosin US Reg Date: 1976-06-23

Terazosin US Reg Date: 1987-08-07

http://www.sciencedirect.com/science/article/pii/S1359644611001711














Furan vs tetrahydrofurans: Dsolubility

average pSolubility: 4.0 (100µM) average pSolubility: 4.7 (20µM)

D pSol: 0.7

A data set of 50 matched-pairs – measure aqueous solubility for the 100 compounds

The THF containing compound is more soluble in 85% of the matched-pairs.

TH

F

pS

olu

bili

ty

pSolubility (furan)

The observered difference in solublity for furan/THF pairs seems to be general.

R R

Possible rationale for differences

Larger dipole for THF than furan – the greater the dipole the greater polarity. The aromaticity in furan obviously makes it different from ethers, especially compared to more strained systems like oxatanes and

tetrahydrofurans. Consequently tetrahydrofurans seem to be less lipophilic than their open-chain ether partners.

Furan:

0.6 Debye

THF:

1.7 Debye

FURAOX LIBVIC

Different geometries – “escaping flatland”

two matched-pairs found in CSD

A forgotten furan-bioisoster? Furan-to-Thiophene/Benzene is way more frequent than furan-to-THF.

Matched-Pairs Summary It can be very powerful to use molecular matched-pairs in drug design.

• Intuitive, interpretable and can provide data-driven, interpretable guidelines for design/analysis

• Can extract tacit knowledge from accumulated data beyond series, projects and departments

Analyzing 1,2,4- and 1,3,4-oxadiazole regioisomers revealed systematic trends

the 1,3,4-oxadiazole isomer show an order of magnitude of lower logD, compared to the 1,2,4

the favorable effect is also observed in increased aqueous solubility, lower HLM Clint and hERG.

The distinct profile difference is likely due to their different charge distributions.

Methodology can be used to generalize other important properties used for decision-making.

thiadiazoles regioisomers, THF vs furan.

direct impact in in-house AZ projects.

Thanks to all my colleagues!

key computational approach for designing candidate drugs often the use of shape and electrostatic comparisons between molecules

1) for (purely) geometrical reasons 2) replacing unwanted functional groups (acids and esters) 3) to fine-tune electrostatics for improved properties

2000


only geometry matters 1


and ”luck” 1


heterocycle 2


> heterocycle 2

now

”Matched-Pair Project X”

heterocyclic electrostatics improved properties 3

Heterocyclic systems needs special attention – a large number of combinations

rings in (candidate) drugs - case stories

Science

scaffold hopping

lxr scaffold

pyrazole scaffold

shapebased scaffold

az project

nm az hts

novel series lxr agonist

effect of az