Biological Evaluation of Compounds

Consult was in GHI479. New data received Feb 9th 2017. Raw data attached. Relevant discussion: GHI484. This post authoured by Mat Todd.

The following compounds were shipped to Syngene yesterday. These may be discussed over at Github Issue 384. 

Strings

MMV693148 OSM-S-346 AEW 284-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(OC(C)C4=CC=C(Cl)C(Cl)=C4)N32 InChI=1S/C20H14Cl2F2N4O2/c1-11(13-4-7-15(21)16(22)8-13)29-18-10-25-9-17-26-27-19(28(17)18)12-2-5-14(6-3-12)30-20(23)24/h2-11,20H,1H3 ILDZQTSEZGQWFR-UHFFFAOYSA-N

MMV693149 OSM-S-347 AEW 285-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(OCC4=CC=C(Cl)C(Cl)=C4)N32 InChI=1S/C19H12Cl2F2N4O2/c20-14-6-1-11(7-15(14)21)10-28-17-9-24-8-16-25-26-18(27(16)17)12-2-4-13(5-3-12)29-19(22)23/h1-9,19H,10H2 CSHOSFVEVPHTHZ-UHFFFAOYSA-N

MMV693150 OSM-S-348 AEW 286-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(OCC4=CC=C(Cl)C=C4)N32 InChI=1S/C19H13ClF2N4O2/c20-14-5-1-12(2-6-14)11-27-17-10-23-9-16-24-25-18(26(16)17)13-3-7-15(8-4-13)28-19(21)22/h1-10,19H,11H2 ROAUWAVERUYYHI-UHFFFAOYSA-N

MMV693151 OSM-S-349 AEW 290-1 FC(C=C1)=C(F)C=C1C2=NN=C3C=NC=C(OCCC4=CC=CC=C4)N32 InChI=1S/C19H14F2N4O/c20-15-7-6-14(10-16(15)21)19-24-23-17-11-22-12-18(25(17)19)26-9-8-13-4-2-1-3-5-13/h1-7,10-12H,8-9H2 DNQWZOZQWWSJTR-UHFFFAOYSA-N

MMV693152 OSM-S-350 AEW 291-1 ClC(C=C1)=C(Cl)C=C1C2=NN=C3C=NC=C(OCCC4=CC=CC=C4)N32 InChI=1S/C19H14Cl2N4O/c20-15-7-6-14(10-16(15)21)19-24-23-17-11-22-12-18(25(17)19)26-9-8-13-4-2-1-3-5-13/h1-7,10-12H,8-9H2 DJLRRTBQGVDDPH-UHFFFAOYSA-N

MMV693153 OSM-S-351 AEW 292-1 ClC(C=C(Cl)C=C1)=C1C2=NN=C3C=NC=C(OCCC4=CC=CC=C4)N32 InChI=1S/C19H14Cl2N4O/c20-14-6-7-15(16(21)10-14)19-24-23-17-11-22-12-18(25(17)19)26-9-8-13-4-2-1-3-5-13/h1-7,10-12H,8-9H2 VJQTVLCLRZKBED-UHFFFAOYSA-N

MMV693154 OSM-S-352 AEW 293-1 FC(C=C1)=CC=C1C2=NN=C3C=NC=C(OCCC4=CC=CC=C4)N32 InChI=1S/C19H15FN4O/c20-16-8-6-15(7-9-16)19-23-22-17-12-21-13-18(24(17)19)25-11-10-14-4-2-1-3-5-14/h1-9,12-13H,10-11H2 QCWQNZCCYWUBQM-UHFFFAOYSA-N

MMV693155 OSM-S-353 AEW 294-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(OCC(C4=CC=CC=C4)CO)N32 InChI=1S/C21H18F2N4O3/c22-21(23)30-17-8-6-15(7-9-17)20-26-25-18-10-24-11-19(27(18)20)29-13-16(12-28)14-4-2-1-3-5-14/h1-11,16,21,28H,12-13H2 MGANJQKOPZQELF-UHFFFAOYSA-N

MMV693161 OSM-S-359 EGT 48-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(SCC4=CC=CC=C4)N32 InChI=1S/C19H14F2N4OS/c20-19(21)26-15-8-6-14(7-9-15)18-24-23-16-10-22-11-17(25(16)18)27-12-13-4-2-1-3-5-13/h1-11,19H,12H2 KEZIWQVAJDNGBM-UHFFFAOYSA-N

MMV693162 OSM-S-360 EGT 45-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(S(CC4=CC=CC=C4)=O)N32 InChI=1S/C19H14F2N4O2S/c20-19(21)27-15-8-6-14(7-9-15)18-24-23-16-10-22-11-17(25(16)18)28(26)12-13-4-2-1-3-5-13/h1-11,19H,12H2 HSYPMILIAZBMAQ-UHFFFAOYSA-N

MMV693163 OSM-S-361 EGT 39-1 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(S(CC4=CC=CC=C4)(=O)=O)N32 InChI=1S/C19H14F2N4O3S/c20-19(21)28-15-8-6-14(7-9-15)18-24-23-16-10-22-11-17(25(16)18)29(26,27)12-13-4-2-1-3-5-13/h1-11,19H,12H2 OCXPXTXAACVDON-UHFFFAOYSA-N

MMV693164 OSM-S-362 EGT 51-4 FC(F)OC(C=C1)=CC=C1C2=NN=C3C=NC=C(N4N=NC(C5=CC=CC=C5)=C4)N32 InChI=1S/C20H13F2N7O/c21-20(22)30-15-8-6-14(7-9-15)19-26-25-17-10-23-11-18(29(17)19)28-12-16(24-27-28)13-4-2-1-3-5-13/h1-12,20H WUWMNWSXMOOYGG-UHFFFAOYSA-N

Four OSM Series 1 compounds were evaluated in a P. falciparum dual gamete formation assay, with data in the attached report. Literature on this assay is in this paper. The assay was performed by the original Imperial team. 

All compounds were found to be inactive (low activity) against both male and female gametocytes, though the levels of activity for some compounds were higher vs. female than any antimalarials originally tested in the above-linked paper (where the maximum % inhibition at this concentration was 27%).

These data are a little surprising given that several of these compounds had previously displayed very high levels of activity in a late stage gametocyte assay:

Late stage gametocyte testing of a sample set of GSK-Arylpyrrole Series

Late Stage Gametocyte Assay (OSM-S-111)

it might have been expected that there would have been more activity vs (the more susceptible) males.

Compounds tested:

OSM-S-5

CC(N1C2=CC=C(F)C=C2)=C(C(OCC(N)=O)=O)C=C1C InChI=1S/C15H15FN2O3/c1-9-7-13(15(20)21-8-14(17)19)10(2)18(9)12-5-3-11(16)4-6-12/h3-7H,8H2,1-2H3,(H2,17,19) YSUCFIZUNLQZDX-UHFFFAOYSA-N

OSM-S-38

CC1=CC(/C=C(C(N/2)=O)\SC2=N/C3=CC=CC=C3)=C(C)N1C(C=C4)=CC=C4C(F)(F)F InChI=1S/C23H18F3N3OS/c1-14-12-16(15(2)29(14)19-10-8-17(9-11-19)23(24,25)26)13-20-21(30)28-22(31-20)27-18-6-4-3-5-7-18/h3-13H,1-2H3,(H,27,28,30)/b20-13- YBBWTVGRVHTTDD-MOSHPQCFSA-N

OSM-S-39

CC1=CC(/C=C(C(N/2)=O)\SC2=N/C3=CC=CC=C3)=C(C)N1C4=CC(C(F)(F)F)=CC(C(F)(F)F)=C4 InChI=1S/C24H17F6N3OS/c1-13-8-15(9-20-21(34)32-22(35-20)31-18-6-4-3-5-7-18)14(2)33(13)19-11-16(23(25,26)27)10-17(12-19)24(28,29)30/h3-12H,1-2H3,(H,31,32,34)/b20-9- GVGNOLWIUGQIHW-UKWGHVSLSA-N

OSM-S-111

CC1=CC(/C=C(C(N/2)=O)\SC2=N/C3=CC=CC=C3)=C(C)N1C4=CC=C(OC)C=C4 InChI=1S/C23H21N3O2S/c1-15-13-17(16(2)26(15)19-9-11-20(28-3)12-10-19)14-21-22(27)25-23(29-21)24-18-7-5-4-6-8-18/h4-14H,1-3H3,(H,24,25,27)/b21-14- KXIVXNPEYYNDHE-STZFKDTASA-N

This post originally authored by Mat Todd

Recent analysis was undertaken by Corey Nislow (see GHI20) on predicting the mechanism of action of Series 1 compounds. In discussions that took place by email (sorry) which are being incorporated into the final draft of the paper about to be submitted on Series 1, a suggestion was made by Corey that the different compounds in Series 1 may have different MoAs. To verify whether this might be true, one arylpyrrole and several Near Neighbours were sent for evaluation in a Parasite Reduction Ratio (PRR) (colloquially, Rate of Killing) assay at GSK Tres Cantos. Thank you to Benigno Crespo and Laura Sanz for doing this, and to Javier Gamo for signing off on the report, which is attached.

The PRR assay can reveal possible MoA based on a comparison of the killing profile of the compounds. If the pattern of parasite ratio, followed over several days, matches the profile found using an existing compound, it's possible they may share a MoA.

Here are the compounds analysed:

Here are the combined results.

Conclusions

Within the noise of this experiment it would seem as though the compounds share a common MoA. A suggestion was made by Corey that, based on the HipHop results, one compound (OSM-S-39) may share commonalities with artemisinin. Though artemisinin was not evaluated in the present assay run, it is known to possess a very rapid rate of killing that would not match the profiles obtained  for the present compounds.

Compounds examined:

Aryl Pyrrole:

MMV019247 OSM-S-5

Near Neighbours:
MMV689017 OSM-S-10
MMV689018 OSM-S-35
MMV689019 OSM-S-37
MMV689020 OSM-S-39
MMV689021 OSM-S-51

Compound Strings:

MMV019247 OSM-S-5 FC1=CC=C(N2C(C)=CC(C(OCC(N)=O)=O)=C2C)C=C1 InChI=1S/C15H15FN2O3/c1-9-7-13(15(20)21-8-14(17)19)10(2)18(9)12-5-3-11(16)4-6-12/h3-7H,8H2,1-2H3,(H2,17,19) YSUCFIZUNLQZDX-UHFFFAOYSA-N

MMV689017 OSM-S-10 FC1=CC=C(N2C(C)=CC(/C=C3S/C(NC\3=O)=N\C4=CC=CC=C4)=C2C)C=C1 InChI=1S/C22H18FN3OS/c1-14-12-16(15(2)26(14)19-10-8-17(23)9-11-19)13-20-21(27)25-22(28-20)24-18-6-4-3-5-7-18/h3-13H,1-2H3,(H,24,25,27)/b20-13- IMCPBTMVWRMBHR-MOSHPQCFSA-N

MMV689018 OSM-S-35 CC1=C(/C=C2S/C(NC\2=O)=N\C3=CC=CC=C3)C=C(C)N1C4=CC=CC=C4 InChI=1S/C22H19N3OS/c1-15-13-17(16(2)25(15)19-11-7-4-8-12-19)14-20-21(26)24-22(27-20)23-18-9-5-3-6-10-18/h3-14H,1-2H3,(H,23,24,26)/b20-14- QTDBAHFMTAMGMZ-ZHZULCJRSA-N

MMV689019 OSM-S-37 CC1=C(/C=C2S/C(NC\2=O)=N\C3=CC=CC=C3)C=C(C)N1C4=CC=C(C)C=C4 InChI=1S/C23H21N3OS/c1-15-9-11-20(12-10-15)26-16(2)13-18(17(26)3)14-21-22(27)25-23(28-21)24-19-7-5-4-6-8-19/h4-14H,1-3H3,(H,24,25,27)/b21-14- XCTPMKNSBDOHRB-STZFKDTASA-N

MMV689020 OSM-S-39 CC1=C(/C=C2S/C(NC\2=O)=N\C3=CC=CC=C3)C=C(C)N1C4=CC(C(F)(F)F)=CC(C(F)(F)F)=C4 InChI=1S/C24H17F6N3OS/c1-13-8-15(9-20-21(34)32-22(35-20)31-18-6-4-3-5-7-18)14(2)33(13)19-11-16(23(25,26)27)10-17(12-19)24(28,29)30/h3-12H,1-2H3,(H,31,32,34)/b20-9- GVGNOLWIUGQIHW-UKWGHVSLSA-N

MMV689021 OSM-S-51 CC1=C(/C=C2S/C(NC\2=O)=N\C3=CC=CC=C3)C=C(C)N1C4=NC=CC=C4 InChI=1S/C21H18N4OS/c1-14-12-16(15(2)25(14)19-10-6-7-11-22-19)13-18-20(26)24-21(27-18)23-17-8-4-3-5-9-17/h3-13H,1-2H3,(H,23,24,26)/b18-13- WFGBKPBKZGJAHY-AQTBWJFISA-N

What are the best strategies for minimizing metabolic clearance in OSM Series 4? This has been, and remains, a key scientific issue for us, along with maintaining potency and improving solubility.

We're devising the next set of compounds to make (also GHI301) and we're meeting next week (GHI317), so this is a good time to re-engage with metabolic clearance.

Separate Issues will be created for each relevant To Do item on Github, but here are all the background data:

1) All the current clearance data are in the wiki. Some data inherited, others obtained more recently. For the moment we are deprioritising the amide series (top row of data in the main wiki scheme) because of a probable hERG liability. So let's focus mostly on the non-amides.

2) Chris Swain predicted metabolic hotspots (in red, below) for three Series 4 compounds, MMV669844, MMV669848 and MMV670936.

When Chris ran the same compounds through a Cyp predictor the same sites came up minus those on the ring, suggesting the ring could be a substrate for another enzyme such as aldehyde oxidase. However, measured AO clearance values suggest MMV669844, for example, is not an AO substrate (see below).

3) Met and MetID work on these same three compounds above from Sue Charman's lab suggested that oxidation on the triazolopyrazine-aromatic system was occurring in most cases, with other liabilities shown above. The analog of MMV669844 with a CHF₂ group in place of the CH₃ (MMV670652, 17 nM) was tested in microsomal studies, giving CLint HLM<8, RLM 30.

4) Aldehyde Oxidase Screen. Performed, with summary data. Amides appeared to be more susceptible to AO oxidation, but in general several potent members of the ether class were found not to be substrates for AO. We should both not worry about AO for this series and yet periodically re-assess Series 4 compounds in the assay.

5) The nature of the core heterocyclic ring has a major effect on clearance - compare the parent triazolopyrazine MMV639565 with the imidazopyrazines MMV669846 and MMV670250.

The potency and reasonable clearance of MMV669846 make it interesting to look at again. The very high clearance of MMV670250 suggests...what?

6) We know that blocking the pyrazine ring in the 8-position reduces potency and that the N-oxide in the pyrazine ring is inactive (below), and we also now know that replacement of the pyrazine southwest N with CH lowers potency. Is there any reason to investigate these further, e.g. for metabolic clearance rate on these compounds?

6) To Do/Questions

i) Benzylic Blocking: Are there blocking groups in the northwest benzylic position we ought to include that might improve metabolic stability? Is it possible to run some of these through predictive software to see if such structures might help?

ii) Acquiring New Metabolic Data on Existing Compounds: MMV670652 (above) has not yet been studied in hepatocytes, and has not had MetID performed. Is there justification for this, to confirm that we see improvements over MMV669844, or can this be assumed?

iii) Acquiring New Metabolic Data on New Compounds: Of the newer compounds synthesised (1, 2, 3), are there any that ought to be evaluated at the Charman lab for microsomal stability (GHI213) (and solubility: GHI260)? Should we re-run Chris Swain's analysis on any new target compounds?

iv) Trends in the Data: In general the newer Monash data (in green in the wiki) shows that a number of the analogs with blocking groups in the northwest benzylic position and OCHF₂ in the northeast have higher than expected clearance rates vs. what might be expected given the data for the analogs with similar blocking groups in the benzylic position and a cyano group in the northeast. Is it possible that the northeast Ar substituent is playing a role in metabolic liability of the aromatic system?

v) Triazole Mods: We should resynthesise the imidazopyrazine MMV669846 shown above to verify potency and microsomal stability. Underway. Are there other groups that could be entertained on the triazole, or in place of it? What is known of good mimics of such a ring system?

vi) Pinning Down Site of Oxidation: Is it worth establishing where on the triazolopyrazine-Ar structure the oxidation is occurring, given the prediction that Cyp will not oxidise on the TP ring and given the low AO clearance experimentally determined? (Related to GHI124).

Post originally authored by Mat Todd