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Chakraverty Group: Identifying novel CD4+ T cell platforms for immunotherapy

Supervisor: Ronjon Chakraverty

This project brings together laboratories with established international reputations in T cell immunotherapy (Prof. Chakraverty) and the biology and treatment of acute myeloid leukaemia (AML, Prof. Vyas). The overall aim of the project is to determine how a sub-population of CD4+ T cells develops cytotoxic functions capable of eliminating AML.

About the Research 

Although CD4+ T cells have been classically viewed as providing helper-like function, CD4+ T cells with cytotoxic potential are increasingly recognised as important in mediating anti-tumour immunity.  Indeed, the Vyas lab have identified T cell receptors (TCR) that recognize AML cells in patients undergoing transplantation and found that the bulk elicit CD4+ cytotoxic T cell responses.  However, few cell surface markers have been identified that unambiguously define this population and this limits the potential to select and expand these cells for therapeutic use.  Our laboratory is now using a combination of transcriptomic, proteomic and functional approaches to characterise cytotoxic CD4+ T cell populations at a higher level of resolution with the aim of developing new strategies for their differentiation, expansion and eventual therapeutic use.   

1. Unbiased proteomics to identify novel cytotoxic T cell markers:  The candidate will use high-throughput, unbiased proteomics in combination with a recently established computational pipeline to identify candidate cell surface markers that correlate with human cytotoxic CD4+ T cell differentiation.  A combination of in vitro flow cytometric and functional killing assays will be used to validate and prioritise candidate markers for further study, including additional testing in clinical samples from patients receiving T cell immunotherapies.

2. CRISPR screens to delineate regulators of cytotoxic CD4+ T cell function:  To identify positive or negative regulators of cytotoxic CD4+ T cell differentiation, the applicant will perform lentivirus-based, targeted CRISPR activation and interference in primary T cells using gRNAs directed at T cell-expressed transcription factors (TF).  Candidate TFs will be further validated through functional assays before further interrogation of the molecular pathways involved using an experimental interrogation of TF-binding, chromatin interactions and transcription.  

3. Test the therapeutic potential of cytotoxic CD4+ T cells in AML cells:  The candidate will systematically evaluate methods for the selection and expansion of cytotoxic CD4+ T cells from healthy volunteers and from patients. Using TCR, we have already identified with reactivity versus AML, they will then test TCR-T cells for cytotoxicity in vitro and in vivo using cell lines and primary AML cells.

Translational Potential. This project aims to identify the optimal T cell platforms that could be employed for TCR-based therapeutics of AML and myeloid cancers.

This project is not suitable for part-time research 

Training Opportunities 

The DPhil student will be trained in: (i) fundamental aspects of immunology, specifically flow cytometry, cell culture and functional assays in vitro (2) computational biology (3) cellular immunology, including testing T cell-based immune therapeutics using in vitro and in vivo models.

The training will be focused on specific skill sets that are critical for developing immune therapies.

Key methods include:

  1. Complex multi-colour flow cytometric analysis and FACS-sorting. All students will independently use flow analysers and sorters.
  2. Lentiviral viral transduction of T cells.
  3. In vitro and in vivo assays of T cell function, including biochemical analysis of TCR signalling.
  4. CRISPR gene editing of T cells.
  5. Computational analysis, including exposure to coding using Python and R.
  6. NGS libraries for RNA-seq, TCR sequencing, ATAC-seq in single cells and highly purified cell populations.

Students will be enrolled on the MRC Weatherall Institute of Molecular Medicine DPhil Course, which takes place in the autumn of their first year. Running over several days, this course helps students to develop basic research and presentation skills, as well as introducing them to a wide range of scientific techniques and principles, ensuring that students have the opportunity to build a broad-based understanding of differing research methodologies.

Generic skills training is offered through the Medical Sciences Division's Skills Training Programme. This programme offers a comprehensive range of courses covering many important areas of researcher development: knowledge and intellectual abilities, personal effectiveness, research governance and organisation, and engagement, influence, and impact. Students are actively encouraged to take advantage of the training opportunities available to them.

As well as the specific training detailed above, students will have access to a wide range of seminars and training opportunities through the many research institutes and centres based in Oxford.

The Department has a successful mentoring scheme, open to graduate students, which provides an additional possible channel for personal and professional development outside the regular supervisory framework. We hold an Athena SWAN Silver Award in recognition of our efforts to build a happy and rewarding environment where all staff and students are supported to achieve their full potential.

Additional Supervisors 

1. Paresh Vyas

Publications

  1. Cheminant M, Fox TA, Alligon M, Bouaziz O, Neven B, Moshous D, Blanche S, Guffroy A, Fieschi C, Malphettes M, Schleinitz N, Perlat A, Viallard JF, Dhedin N, Sarrot-Reynauld F, Durieu I, Humbert S, Fouyssac F, Barlogis V, Carpenter B, Hough R, Laurence A, Marçais A, Chakraverty R, Hermine O, Fischer A, Burns SO, Mahlaoui N, Morris EC, Suarez F.   2023 Allogeneic stem cell transplantation compared to conservative management in adults with inborn errors of immunity. Blood 141:60-71.
  2. Zeiser R, Polverelli N, Ram R, Hashmi SK, Chakraverty R, Middeke JM, Musso M, Giebel S, Uzay A, Langmuir P, Hollaender N, Gowda M, Stefanelli T, Lee SJ, Teshima T, Locatelli F; REACH3 Investigators. 2021. Ruxolitinib for Glucocorticoid-Refractory Chronic Graft-versus-Host Disease. New England Journal of Medicine, 385;228-238.
  3. Cuadrado MM, Szydlo RM, Watts M, Patel N, Renshaw H, Dorman J, Lowdell M, Ings S, Anthias C, Madrigal A, Mackinnon S, Kottaridis P, Carpenter B, Hough R, Morris E, Thomson K, Peggs KS, Chakraverty R. 2020. Predictors of recovery following allogeneic CD34+-slected infusion without conditioning to correct poor graft function. Haematologica, 105:2639-2646.
  4. Dertschnig S., Evans P., Santos e Sousa P., Manzo T., Ferrer I.R., Stauss H.J., Clare Bennett C. and Chakraverty R.  2020. Graft-versus-host disease reduces lymph node expression of tissue-restricted antigens and promotes autoimmunity. Journal of Clinical Investigation 130:1896-1911
  5. Ferrer IR, West HC, Henderson S, Ushakov DS, Santos E Sousa P, Strid J, Chakraverty R, Yates AJ, Bennett CL. 2019. A wave of monocytes is recruited to replenish the long-term Langerhans network after immune injury. Science Immunology 2019 4(38); pii: eaax8704
  6. Khan AB, Carpenter B, Sousa PSE, Pospori C, Khorshed R, Griffin J, Velica P, Zech M, Ghorashian S, Forrest C, Thomas S., Gonzalez Anton S., Ahmadi M., Holler A’, Flutter B., Ramirez-Ortiz Z., Means T.K., Bennett C.L., Stauss H., Morris E., Lo Celso C., Chakraverty R.  2018. Redirection to the bone marrow improves T cell persistence and antitumor functions. Journal of Clinical Investigation 128:2010-2024
  7. Santos ESP, Cire S., Conlan T., Jardine L., Tkacz C., Ferrer I.R., Lomas C., Ward S., West H., Dertschnig S., Blobner S., Means T.K., Henderson S., Kaplan D.H., Collin M., Plagnol V., Bennett C.L. and Chakraverty R.  2018. Peripheral tissues reprogram CD8+ T cells for pathogenicity during graft-versus-host disease. Journal of Clinical Investigation Insight 3(5) pii 90711
  8. Mathew, N. R., F. Baumgartner… R. Chakraverty….E. Pearce, B. R. Blazar, and R. Zeiser. 2018. 'Sorafenib promotes graft-versus-leukemia activity in mice and humans through IL-15 production in FLT3-ITD-mutant leukemia cells', Nat Med, 24: 282-91.
  9. Galleu, A., Y. Riffo-Vasquez, C. Trento, C. Lomas, L. Dolcetti, T. S. Cheung, M. von Bonin, L. Barbieri, K. Halai, S. Ward, L. Weng, R. Chakraverty, G. Lombardi, F. M. Watt, K. Orchard, D. I. Marks, J. Apperley, M. Bornhauser, H. Walczak, C. Bennett, and F. Dazzi. 2017. Apoptosis in mesenchymal stromal cells induces in vivo recipient-mediated immunomodulation, Science Translational Medicine, 9 (416)
  10. D’Aveni M, Rossignol J., Coman S., Sivakumaran S., Henderson S., Manzo T., Santos e Sousa P., Bruneau J., Fouquet G., Zavala F., Alegria-Prévot O., Garfa-Traoré M., Suarez F., Trebeden-Nègre H., Mohty M., Bennett C. L., Chakraverty R., Hermine O. and Rubio M.T.  2015. G-CSF mobilizes CD34+ regulatory monocytes that inhibit Graft-versus-Host Disease. Science Translational Medicine, 7: 281ra42
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