OpenSourceMalaria:Triazolopyrazine (TP) Series
Open Source Malaria Series 4: The Triazolopyrazine (TP) Series
The Triazolopyrazine Series is the newest of the OSM series. It was announced on September 10th 2013 and is sometimes referred to as the TP Series, or OSM Series 4.
The series arises from industrial work that cannot be fully disclosed which was followed by some hit-to-lead work funded directly by MMV and performed by a CRO which can.
A great deal of exploration of the series has been done, with significant diversity in the core and pendant groups. The series includes many potent compounds.
There is evidence from parasite ion regulation assays (below) that these compounds may be PfATP4 inhibitors. Such evidence distinguishes Series 4 from Series 1-3 where there was no experimental evidence for a mechanism of action.
As with everything involved in OSM, suggestions can be given in multiple ways.
Start of the Campaign
A briefing document written for MMV was the initial knowledgebase. This was accompanied by a PDF summary of pharmacokinetics and efficacy. This is being folded into the sections below, then supplemented.
The aim of the campaign at the outset was to improve the metabolic stability and the pharmacokinetic properties of this series in rat so as to meet the once-dosing criteria (TCP1) set by MMV. New chemistry directed towards blocking the putative metabolic sites was a major part of the research prior to the data being contributed to OSM.
Sources of Data on the TP Series
Initial briefing document (A minor error in the briefing document referring to the amides has been clarified.)
PDF summary of pharmacokinetics and efficacy
The current synthetic lab notebook
Raw Data Spreadsheet on Compounds Prior to Donation to OSM
Summary of Data on Amides in the Series
Notable Points about the TP Series
- Compounds in this series have been identified down to 16 nM potency.
- Seems to have good in vitro HLM and hHEP stability Clint < 8.1 is compatible with 10 nM potency.
- RLM remains stubbornly high, particularly for the more potent analogues translating to short half-lives in rat PK.
- Series appears to have little polypharmacology or cytotoxicity.
- Not so far challenged the hypothesis that rat metabolism may not be a great model for human metabolism for this series.
- The series shows activity in Kieran Kirk’s PfATP4 assay which goes away for Pfal inactives in the series.
Concerns about the TP Series
- Although dofetilide binding looks weak or nil, the series has shown activity in a patch clamp assay at Essen (1-10 uM) which is quite potent though with a window of >100 fold over Pfal potency.
- In Kip Guy’s resistant mutants the picture is mixed, but there is still support for the idea that some members of the series are weaker in the resistant strains. The series has no or weak >>1uM activity against gametocytes, no activity against Winzeler’s Pb liver stage and may have weak activity against ookinetes but the dose-response data has not been completed.
Project Strands of Current Interest
The biggest issue is metabolic stability, as measured in rat in particular. There are few toxicity concerns. Thus possible future directions:
- Small scale changes around the side chains, particularly phenethyl to attempt to balance potency and metabolism. Other possibilities: a) N is tolerated in the ring, hasn’t been explored much recently. b) Is 3,4-diF the best substitution pattern? c) Some evidence (eg MMV669848) that the phenethyl side chain can be rigidified, perhaps the iso-indoline of that compound could be improved on with other ring systems and by more optimal substitution of the aromatic benzene ring of the isoindoline. d) The amide MMV670944 is interesting and shows good RLM stability, but many other amides failed to match its potency.
- Incorporation of a basic centre to increase volume as a potential fix for half-life. However, this might come at the expense of plasma concentration so would require high potency. Of the 29 compounds with a basic centre only one (MMV670437, below) has a measured potency < 100 nM (actually 44 nM).
- More significant structural changes. Of the changes made to the basic skeleton, the most successful might be the recent evaluation of the substitution position changes (e.g., MMV670945), possibly in combination with modifying the disposition of the N atoms in the core. Related compounds have been made by others and it would be wise to incorporate the learnings from these series into any plans to explore this substitution pattern further. The first few compounds look similar in terms of metabolic stability.
Cycloaliphatic Triazole Substituent
Attempts at lowering the lipophilicity of the TP compounds by replacing the triazole aryl substituent with a cyclo(hetero)aliphatic group, linked either by the heteroatom or otherwise (e.g. piperidine, tetrahydropyran, indoline or isoindoline) lowered the potency against PfNF54, as did an aniline substituent.
Core Modification of Pyrazine Ring
Based on an assumption that the pyrazine moiety of the TP could undergo AO metabolism at positions alpha- to the nitrogen a few compounds were made with different R groups (Cl, Me, NH2, NEt2). However, all these compounds lost potency against PfNF54.
Core Modification of Triazole Ring
Two compounds based on imadazopyrazines were made (MMV669846 and MMV670250, below). Both showed reduced potency against PfNF54 vs. the corresponding TP compound. The RLM stability of one was found to be poor. Approx 20 structures were made with variations to the 6,5 core system. MMV669846 was the most potent. As most of the analogues were >1 μM potency, fewer were tested in RLM (quite a few in HLM). Of the 4 tested in RLM, the greatest stability had a Clint of 109, (HLM 9.5), several had HLM Clint 8 or less, particularly after moving or removing the N from the pyrazine ring.
(Note - original briefing document contained two entries for MMV669846 at this point with different potencies - need check of original spreadsheet, to verify the above is correct)
What Can the Community Do Now?
Those wishing to contribute to OSM Series 4 should in the first instance read the briefing document. The first question is: Which 10 compounds ought to be made in Sept/Oct 2013 for evaluation?
Resources Needed: Chemists to either make new molecules, or help obtain existing compounds that might be relevant to this series.
1. Hit-to-lead synthesis and biological evaluation, to improve potency and drug likeness.
2. Validation of PfATP4 activity.
Current Questions for the Community
Starting-point questions are listed at the end of the briefing document. These translate into the initial question of Which 10 compounds ought to be made first? (also asked here and Does anyone possess compounds that could be relevant to this series already? (asked here and there is a separate section for results below).
Prior Knowledge of the Series
Potency of Hits and Analogs
(Detail will be placed here, but for the moment please browse the briefing document).
Possible PfATP4 Activity Deduced from Parasite Ion Regulation Assays
The following five compounds were evaluated in parasite ion regulation assays in the Kirk Laboratory; the hypothesis is that PfATP4 is a Na+ ATPase that exports Na+ and imports H+ (or equivalent) and that the effects of the compounds on Na+ concentration and pH are attributable to inhibition of this activity. Structures, potency, metabolism/solubility and raw PfATP4 assay data are here.
MMV669000: no (potency: inactive)
MMV669304: yes (potency: 280 nM)
MMV669360: yes (potency: 356 nM)
MMV669542: yes (potency: 185 nM)
MMV669848: yes (potency: 114 nM)
MMV669000 did not dissipate the plasma membrane Na+ gradient or increase the plasma membrane pH gradient consistent with it not inhibiting PfATP4 at the concentration tested.
The other compounds dissipated the plasma membrane Na+ gradient and increased the plasma membrane pH gradient at a concentration of 2 μM, consistent with them being PfATP4 inhibitors.
(i.e. note the correlation: compound inactive in these assays is the inactive analog in the plasmodium screen)
Other Sources of Compounds in this Series
Strings for Google
Use this section to paste strings to make the page more discoverable.
MMV668955 FC(F)OC(C=C1)=CC=C1C2=NN=C3N2C(CCO)CNC3; InChI=1S/C14H16F2N4O2/c15-14(16)22-11-3-1-9(2-4-11)13-19-18-12-8-17-7-10(5-6-21)20(12)13/h1-4,10,14,17,21H,5-8H2
MMV668960 FC(F)OC(C=C1)=CC=C1C2=NN=C3N2CCNC3; InChI=1S/C12H12F2N4O/c13-12(14)19-9-3-1-8(2-4-9)11-17-16-10-7-15-5-6-18(10)11/h1-4,12,15H,5-7H2
MMV668962 FC(F)OC(C=C1)=CC=C1C2=NN=C3N2CCN(C(CN)=O)C3; InChI=1S/C14H15F2N5O2/c15-14(16)23-10-3-1-9(2-4-10)13-19-18-11-8-20(12(22)7-17)5-6-21(11)13/h1-4,14H,5-8,17H2
MMV669000 O=C(N1CC(C=CC=C2)=C2C1)C3=CN=CC4=NN=C(C5=CC=C(OC(F)F)C=C5)N43; InChI=1S/C21H15F2N5O2/c22-21(23)30-16-7-5-13(6-8-16)19-26-25-18-10-24-9-17(28(18)19)20(29)27-11-14-3-1-2-4-15(14)12-27/h1-10,21H,11-12H2
MMV669025 FC1=C(F)C=CC(CCOC2=CNC(C3=NN=C(C4=CC=C(OC(F)F)C=C4)N32)=O)=C1; InChI=1S/C20H14F4N4O3/c21-14-6-1-11(9-15(14)22)7-8-30-16-10-25-19(29)18-27-26-17(28(16)18)12-2-4-13(5-3-12)31-20(23)24/h1-6,9-10,20H,7-8H2,(H,25,29)
MMV669304 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(CCCC4=CC=CC=C4)N32; InChI=1S/C21H18F2N4O/c22-21(23)28-18-11-9-16(10-12-18)20-26-25-19-14-24-13-17(27(19)20)8-4-7-15-5-2-1-3-6-15/h1-3,5-6,9-14,21H,4,7-8H2
MMV669310 FC(F)OC(C=C1)=CC=C1C2=NN=C3N2C(CCCC4=CC=CC=C4)CNC3; InChI=1S/C21H22F2N4O/c22-21(23)28-18-11-9-16(10-12-18)20-26-25-19-14-24-13-17(27(19)20)8-4-7-15-5-2-1-3-6-15/h1-3,5-6,9-12,17,21,24H,4,7-8,13-14H2
MMV669360 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(COCC4=CC=C(F)C(F)=C4)N32; InChI=1S/C20H14F4N4O2/c21-16-6-1-12(7-17(16)22)10-29-11-14-8-25-9-18-26-27-19(28(14)18)13-2-4-15(5-3-13)30-20(23)24/h1-9,20H,10-11H2
MMV669542 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(C(NC4=CC=CC(Cl)=C4)=O)N32; InChI=1S/C19H12ClF2N5O2/c20-12-2-1-3-13(8-12)24-18(28)15-9-23-10-16-25-26-17(27(15)16)11-4-6-14(7-5-11)29-19(21)22/h1-10,19H,(H,24,28)
MMV669846 FC1=C(F)C=CC(CCOC2=CN=CC3=NC=C(C4=CC=C(Cl)C=C4)N32)=C1; InChI=1S/C20H14ClF2N3O/c21-15-4-2-14(3-5-15)18-10-25-19-11-24-12-20(26(18)19)27-8-7-13-1-6-16(22)17(23)9-13/h1-6,9-12H,7-8H2
MMV669848 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(CN4CC(C=CC=C5)=C5C4)N32; InChI=1S/C21H17F2N5O/c22-21(23)29-18-7-5-14(6-8-18)20-26-25-19-10-24-9-17(28(19)20)13-27-11-15-3-1-2-4-16(15)12-27/h1-10,21H,11-13H2
MMV670250 FC1=C(F)C=CC(CCOC2=CN=CC3=CN=C(C4=CC=C(Cl)C=C4)N32)=C1; InChI=1S/C20H14ClF2N3O/c21-15-4-2-14(3-5-15)20-25-11-16-10-24-12-19(26(16)20)27-8-7-13-1-6-17(22)18(23)9-13/h1-6,9-12H,7-8H2
MMV670437 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(OC[C@H](N(C)C)C4=CC(F)=C(F)C=C4)N32; InChI=1S/C22H19F4N5O2/c1-30(2)18(14-5-8-16(23)17(24)9-14)12-32-20-11-27-10-19-28-29-21(31(19)20)13-3-6-15(7-4-13)33-22(25)26/h3-11,18,22H,12H2,1-2H3/t18-/m0/s1
MMV672939 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=CC(OCCC4=CC(F)=C(F)C=C4)=NN32; InChI=1S/C20H14F4N4O2/c21-15-6-1-12(11-16(15)22)9-10-29-18-8-7-17-25-26-19(28(17)27-18)13-2-4-14(5-3-13)30-20(23)24/h1-8,11,20H,9-10H2
MMV672942 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=CC(NCCC4=CC=C(F)C=C4)=NN32; InChI=1S/C20H16F3N5O/c21-15-5-1-13(2-6-15)11-12-24-17-9-10-18-25-26-19(28(18)27-17)14-3-7-16(8-4-14)29-20(22)23/h1-10,20H,11-12H2,(H,24,27)
MMV672992 N#CC(C=C1)=CC=C1C2=NN=C3C=CC(NCCC4=CN=CC=C4)=NN32; InChI=1S/C19H15N7/c20-12-14-3-5-16(6-4-14)19-24-23-18-8-7-17(25-26(18)19)22-11-9-15-2-1-10-21-13-15/h1-8,10,13H,9,11H2,(H,22,25)