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TLR9 expression in Chronic Lymphocytic Leukemia identifies a pro-migratory subpopulation and novel therapeutic target
journal contribution
posted on 2023-06-09, 22:43 authored by Emma KennedyEmma Kennedy, Eve Marie Coulter, Emma Halliwell, Nuria Profitos-Peleja, Elisabeth Walsby, Barnaby Clark, Elizabeth H Phillips, Thomas Burley, Simon MitchellSimon Mitchell, Stephen Devereux, Chris JonesChris Jones, Rosalynd Johnston, Timothy ChevassutTimothy Chevassut, Christopher PepperChristopher Pepper, Andrea PepperAndrea Pepper, othersCLL remains incurable despite BCR-targeted inhibitors revolutionizing treatment. This suggests that other signaling molecules are involved in disease escape mechanisms and resistance. Toll-like receptor 9 (TLR9) is a promising candidate, which is activated by unmethylated CpG-DNA. Here, we show that plasma from CLL patients contains significantly more unmethylated DNA than plasma from healthy controls (p<0.0001) and that cell-free DNA levels correlate with the prognostic markers CD38, b2-microglobulin and lymphocyte doubling time. Furthermore, elevated cell-free DNA was associated with shorter time to first treatment (TTFT: p=0.003, HR=4.0). We went on to show that TLR9 expression was associated with in-vitro CLL cell migration (p<0.001) and intracellular endosomal TLR9 strongly correlated with aberrant surface expression ((sTLR9); r=0.9). In addition, lymph node-derived CLL cells showed increased sTLR9 (p=0.016) and RNA sequencing of paired sTLR9hi and sTLR9lo CLL cells revealed differential transcription of genes involved in TLR signaling, adhesion, motility and inflammation in sTLR9hi cells. Mechanistically, the TLR9 agonist, ODN2006, promoted CLL cell migration (p<0.001) that was mediated, by p65 NF-kB and STAT3 transcription factor activation. Importantly, autologous plasma induced the same effects, which were reversed by a TLR9 antagonist. Furthermore, high TLR9 expression promoted engraftment and rapid disease progression in a NSG mouse xenograft model. Finally, we showed that dual targeting of TLR9 and BTK was strongly synergistic (median CI=0.2 at ED50), which highlights the distinct role for TLR9 signaling in CLL and the potential for combined targeting of TLR9 and BTK as a more effective treatment strategy in this incurable disease.
Funding
Equipment Grant - Flow cytometer for cancer research; G2599; SUSSEX CANCER FUND FOR TREATMENT AND RESEARCH
Mining the Wnt signalling-responsive surfaceome for drug targets in acute myeloid leukaemia; G3090; WELLCOME TRUST
Overcoming Ibrutinib and Venetoclax resistance in Chronic Lymphocytic Leukaemia; G3247; MRC-MEDICAL RESEARCH COUNCIL
How does SARS CoV-2 infect blood vessels?; G3146; UK RESEARCH AND INNOVATION; MR/V036750/1
Modelling and targeting Acute Myeloid Leukaemia cells in the Bone Marrow protective niche; G3106; SUSSEX CANCER FUND FOR TREATMENT AND RESEARCH
Drug-induced selective lethality in populations of DNMT3A knockdown cells; G2782; WELLCOME TRUST; 218435/Z/19/Z
In vitro modelling and therapeutic targeting of tumour cell migration in chronic lymphocytic leukaemia.; G2544; BLOODWISE; 18005
History
Publication status
- Published
File Version
- Accepted version
Journal
BloodISSN
0006-4971Publisher
American Society of HematologyExternal DOI
Page range
1-32Department affiliated with
- Clinical and Experimental Medicine Publications
Full text available
- No
Peer reviewed?
- Yes
Legacy Posted Date
2021-01-12First Compliant Deposit (FCD) Date
2021-01-11Usage metrics
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