Congratulations Cassie and team! Great to see it out, well done.

And we did make it! Amazing day! We’re so grateful to all the friends, family and colleagues who helped us reach our fundraising goal!

Extremely hot #BigHike

@cancerresearchuk.org

Thanks! ☺️

Thanks if you made it this far! This has been a really fun project with lots of twists and turns. Do check it out if it takes your interest! A lot of hard work from Alastair and super collaboration with @bendinglab.bsky.social @sarahdimeloe.bsky.social and thanks to the input from all authors 12/12

And this mechanism also fits for E.coli. EV5 shows E. coli Nissle, another popular probiotic and BCT vector also causes aborted T cell activation - so caution I guess if you wanna couple this with checkpoint blockade (as there are no checkpoints to block if T cells are disabled). 11/12

Before you go - a couple of fun side notes from the sup figures. FIRST, we could have missed this phenotype without the Timer system, EV1 shows comparison with the Nr4a1-GFP and Nr4a1-Timer. GFP gives the impression all is fine bc it sticks around-don't get the info of seeing it turned off. 10/12

What about back in vivo, can it still work? Well the Asnase-deficient STm still controls tumour growth, despite losing some effect on tumour stemness. SO, bringing T cells back on board is a good thing. That's where this story ends, for now! Much to unpick still! 9/12

...not quite. Would have been a neat bundle of joy as is, but then we wondered about the effect on the tumour cell...so we looked and yup, Asn depletion is also reducing tumour c-myc...but that's a good thing! Use of the Asnase-KO STm restores tumour c-Myc and 'stemness' 8/12

After a few rabbit holes we finally pin it down to ASPARAGINE! STm express an asparaginase (as do E. coli), and gobble up asparagine in the TME. T cells REALLY need it (c-myc really needs it). So we remove Asnase and T cells now activate beautifully - JOB DONE?! 7/12

What's going on? TCR signal transduction is fine...mTOR/AKT are fine. Finally (took us a while!) we found c-myc was the culprit - T cells in presence of STm TCM could not upreg protein levels required for metabolic switch, so activation aborts....cool....but why? 6/12

Turn to RNAseq and by 16hrs stim CD4's have failed metabolic reprogramming -> check functionally by seahorse - yep T cells can't upreg glycolysis, oxphos also impaired. This occurs even with just the conditioned medium of STm-infected org's, so it's soluble 5/12

T cells are dysfunctional. IFNy doesn't correlate with TCR activity, ↓PD-1 & ↓ polyfunctionality. Tumour organoid--T cell co-culture shows arrested TCR signalling after 24-48 hrs aCD3/CD28 stim. What is going on? 4/12