Date: Tue August 12th 2025
Time: 3pm UK time (other times)
Place: Zoom link
Previous Meeting: issue #27
Who can come?: Anyone. No need to say anything unless you'd like to. If you'd like to contribute, please join Github.
Attendees: Dr Eve Carter (UCL), Dr Madison Edwards (Uni Toronto), Prof Matt Todd (UCL), Prof Karl Swann (Uni Cardiff), Prof Matilda Katan (UCL), Dr Claudia Tredup (Goethe-University Frankfurt), Dr Jeffrey Mowat (Janssen Pharmaceutica NV), Anna Girtle (Uni Toronto).
Decks: Eve's presentation
Agenda:
In meeting:
Eve reviewed the progress made thus far, specifically the identification of 10 compounds that appear to bind PLCz1 with specificity from an ASMS screen performed by SGC Toronto. Eve further investigated these compounds by screening commercially available analogues in the Aldol assay, while Madison performed SPR and Anna from Nuvisan screened the same compounds in the XY-69 and IP-one assays. Across all of these assays, 4 analogues from the “11 series” of compounds appear most promising. However, the results across assays are variable and no single compound satisfies all of the assays. All 4 compounds consist of an identical acid bound to a variable but similar amine, leading Eve to explore the possibility of generating new analogues following this pattern. She purchased the common acid and a range of similar looking amines to generate new amides. She synthesized 15 amides using this approach and screened them all against chicken PLCz1 in the Aldol assay. Based on these results she took the top 8 compounds and performed dose response measurements to try to get IC50s (range from 27-128 µM). Interestingly, the 2 compounds with the lowest IC50s are enantiomers of each other, but the racemic compound was screened previously and was not identified as a top hit. She suggested this could be due to an impurity or decomposition in the commercial compound, but will go back and explore this phenomenon in the next month.
Mat agreed that the compounds Eve synthesized are likely of higher quality and suggested she could make her own sample of the racemic compound to screen in the Aldol assay to resolve this. Jeffrey asked if there were plans to substitute the left hand side of the molecules, which is something that would be nice to do if there is more time. Madison commented that they previously had slightly adjusted the left hand side of some of the 11 series compounds and found that they had significantly reduced binding by SPR, which is why Eve started with alterations to the right hand side of the molecules. Mat commented that running SPR on the newly synthesized 11 series compounds will be useful to see if binding affinity is improved. Overall, the results Eve got from this approach are slightly better than the purchased compounds. Next steps include shipping the top synthesized compounds to Madison to screen in SPR and potentially ship to Nuvisan to have them screen the compounds in their biochemical assays.
Madison plans to start with SPR for these new compounds, and may also try ITC as an alternative binding assay in the future. Matilda commented that she has gotten the best insight into binding with NMR, as it is very clear when there is or isn’t binding. She suggested this may be a useful approach since she has had success screening low binders this way, although it only gives yes/no binding rather than being quantifiable.
Claudia asked whether the ASMS data from other PLCs had been analyzed to see if unique binders emerged after excluding those from PLCd and human PLCz, or if the same compounds were ultimately identified. Matt commented that a master’s student had reviewed the PLC-related screens and found two compounds selective for chPLCz1 that had initially been missed. One of these Eve has ordered for testing and the other was not available commercially. However, there was no evidence of pan-PLC inhibitors or any compounds showing strong enrichment across PLCs.
Matt then revisited last month's conversation about injecting molecules into cells. Karl explained that this approach is generally ineffective as small molecules diffuse out or are actively pumped out of cells. Jeffrey added that direct injection also seemed unnecessary, since the compounds were lipophilic enough to enter cells, but their binding was simply too weak to yield useful results. He noted that even accounting for medium binding and cell shift effects, the compounds were well above the potency needed for cellular activity. Karl confirmed they had attempted injection with the Nuvisan compound alongside fluorescent calcium dyes and PLCz, but saw no effect. He suggests this is likely because the drug was expelled too quickly.
Matilda shared that a previous suggestion to test competition at the PLC gamma active site had generated useful NMR data. Using IP3 as a reference, they showed the test molecules did not compete at the active site, which might be relevant to this project.
To do:
L'esprit de l'escalier
If you'd like to follow up after the meeting, please comment below. You can also email, but please be clear if anything in the email should not be public domain - the default is open.
Next meeting:
Date: Tue August 12th 2025
Time: 3pm UK time (other times)
Place: Zoom link
Previous Meeting: issue #27
Who can come?: Anyone. No need to say anything unless you'd like to. If you'd like to contribute, please join Github.
Attendees: Dr Eve Carter (UCL), Dr Madison Edwards (Uni Toronto), Prof Matt Todd (UCL), Prof Karl Swann (Uni Cardiff), Prof Matilda Katan (UCL), Dr Claudia Tredup (Goethe-University Frankfurt), Dr Jeffrey Mowat (Janssen Pharmaceutica NV), Anna Girtle (Uni Toronto).
Decks: Eve's presentation
Agenda:
In meeting:
Eve reviewed the progress made thus far, specifically the identification of 10 compounds that appear to bind PLCz1 with specificity from an ASMS screen performed by SGC Toronto. Eve further investigated these compounds by screening commercially available analogues in the Aldol assay, while Madison performed SPR and Anna from Nuvisan screened the same compounds in the XY-69 and IP-one assays. Across all of these assays, 4 analogues from the “11 series” of compounds appear most promising. However, the results across assays are variable and no single compound satisfies all of the assays. All 4 compounds consist of an identical acid bound to a variable but similar amine, leading Eve to explore the possibility of generating new analogues following this pattern. She purchased the common acid and a range of similar looking amines to generate new amides. She synthesized 15 amides using this approach and screened them all against chicken PLCz1 in the Aldol assay. Based on these results she took the top 8 compounds and performed dose response measurements to try to get IC50s (range from 27-128 µM). Interestingly, the 2 compounds with the lowest IC50s are enantiomers of each other, but the racemic compound was screened previously and was not identified as a top hit. She suggested this could be due to an impurity or decomposition in the commercial compound, but will go back and explore this phenomenon in the next month.
Mat agreed that the compounds Eve synthesized are likely of higher quality and suggested she could make her own sample of the racemic compound to screen in the Aldol assay to resolve this. Jeffrey asked if there were plans to substitute the left hand side of the molecules, which is something that would be nice to do if there is more time. Madison commented that they previously had slightly adjusted the left hand side of some of the 11 series compounds and found that they had significantly reduced binding by SPR, which is why Eve started with alterations to the right hand side of the molecules. Mat commented that running SPR on the newly synthesized 11 series compounds will be useful to see if binding affinity is improved. Overall, the results Eve got from this approach are slightly better than the purchased compounds. Next steps include shipping the top synthesized compounds to Madison to screen in SPR and potentially ship to Nuvisan to have them screen the compounds in their biochemical assays.
Madison plans to start with SPR for these new compounds, and may also try ITC as an alternative binding assay in the future. Matilda commented that she has gotten the best insight into binding with NMR, as it is very clear when there is or isn’t binding. She suggested this may be a useful approach since she has had success screening low binders this way, although it only gives yes/no binding rather than being quantifiable.
Claudia asked whether the ASMS data from other PLCs had been analyzed to see if unique binders emerged after excluding those from PLCd and human PLCz, or if the same compounds were ultimately identified. Matt commented that a master’s student had reviewed the PLC-related screens and found two compounds selective for chPLCz1 that had initially been missed. One of these Eve has ordered for testing and the other was not available commercially. However, there was no evidence of pan-PLC inhibitors or any compounds showing strong enrichment across PLCs.
Matt then revisited last month's conversation about injecting molecules into cells. Karl explained that this approach is generally ineffective as small molecules diffuse out or are actively pumped out of cells. Jeffrey added that direct injection also seemed unnecessary, since the compounds were lipophilic enough to enter cells, but their binding was simply too weak to yield useful results. He noted that even accounting for medium binding and cell shift effects, the compounds were well above the potency needed for cellular activity. Karl confirmed they had attempted injection with the Nuvisan compound alongside fluorescent calcium dyes and PLCz, but saw no effect. He suggests this is likely because the drug was expelled too quickly.
Matilda shared that a previous suggestion to test competition at the PLC gamma active site had generated useful NMR data. Using IP3 as a reference, they showed the test molecules did not compete at the active site, which might be relevant to this project.
To do:
L'esprit de l'escalier
If you'd like to follow up after the meeting, please comment below. You can also email, but please be clear if anything in the email should not be public domain - the default is open.
Next meeting: