Összes szerző

Vörös Orsolya

az alábbi absztraktok szerzői között szerepel:

Ghofrane Medyouni
Role of ion channels in CAR T-cell

Aug 30 - szerda

15:30 – 17:00

II. Poszterszekció

P42

Role of ion channels in CAR T-cell

Ghofrane Medyouni¹, Vivien Jusztus¹, Orsolya Vörös¹, Maria Eduarda Lima^1,2, György Panyi¹, Péter B. Hajdu^{1, 2}

¹ University of Debrecen, Faculty of Medicine, Department of Biophysics and Cell Biology

² University of Debrecen, Faculty of Dentistry, Division of Dental Biochemistry

Cancer immunotherapy partly relies on the reprogramming of host immune cells to eliminate cancer cells. Genetic modification of T cells to express chimeric antigen receptors (CARs) is utilized in the treatment of hematological malignancies. Despite its success, many challenges remain to improve the efficacy and safety of this therapy. Ion channels in T-cells participate in the regulation of Ca²⁺-dependent activation pathway and play a role in various effector functions inevitable for target cell abolition. Hence, modification of ion channels’ function can contribute to successful immune therapy. However, no study has been reported about functional role of CAR T-cell ion channels yet.

We established a 3^rd-generation CAR expressing cell line (CD19-CAR cells) from Jurkat cells. We used the whole-cell patch-clamp technique and FURA-2-based Ca²⁺-imaging to determine the biophysical properties of Kv1.3 and Ca²⁺-response of CD19-CAR cells, respectively. We adapted a Calcein Red based killing assay to test CD19-CAR cells’ target cell killing capacity. We assessed the localization of Kv1.3 in standalone and in the CAR-synapse engaged CD19-CAR cells.

We showed that Kv1.3 activation and inactivation kinetics are the same in non-transfected and CD19-CAR cells, while voltage-dependence of activation was different. Thapsigargin-induced Ca²⁺-response of CD19-CAR cells was lower as compared to the control. We showed that Kv1.3 channels are co-localized with CARs in standalone CD19-CAR cells, and they redistribute to the contact region between a CD19-CAR cell and a target cell (Raji B cell). Upon Vm24 addition (specific Kv1.3 inhibitor, 1 nM) the target cell ability of CD19-CAR cells was impaired. Based on these results, we suppose that ion channels can affect the outcome of immunotherapy, and further experiments are needed to clarify their functional role.

Acknowledgment

This work was supported by Stipendium Hungaricum Scholarship to M.G.t and NKFIH (K128525, P.H.).

Jusztus Vivien
Ion channel expression of CD8+ T cells in ovarian cancer

Aug 30 - szerda

15:30 – 17:00

II. Poszterszekció

P43

Ion channel expression of CD8⁺ T cells in ovarian cancer

Vivien Jusztus¹, Ghofrane Medyouni¹, Orsolya Vörös¹, Zsolt Szabó¹, Rudolf Lampé³, György Panyi¹, Orsolya Matolay³, Eszter Maka³, Zoárd Krasznai³, Péter Hajdu^1,2

¹ University of Debrecen, Faculty of Medicine, Department of Biophysics and Cell Biology

² University of Debrecen, Faculty of Dentistry, Division of Dental Biochemistry

³ University of Debrecen, Faculty of Medicine, Department of Gynecology and Obstetrics

The ion channels of T lymphocytes have an important role in effector functions such as activation, cytokine production and tumor cell elimination. T cells recognise and kill cancer cells during continuous monitoring. Although tumor infiltrating lymphocytes (TILs) are able to penetrate the tumor, they cannot fulfil their effector function due to the suppressive nature of the tumour microenvironment. K⁺ channels, such as Kv1.3 and KCa3.1, stabilize the negative membrane potential of T cells to control Ca²⁺-influx through CRAC channels and Ca²⁺-dependent signaling. In the present study, we determined the expression of T cell ion channels from peripheral blood of untreated ovarian cancer patients and healthy donors.

PBMCs were isolated from blood of ovarian patients and healthy donors using Ficoll-Paque density gradient method. Cells were activated with CD3/CD28 antibodies. Whole-cell current was measured in activated CD8⁺ T cells using patch-clamp technique. Ca²⁺ response of CD8⁺ cells was evaluated with FURA-2 Ca²⁺-imaging method.

KCa3.1 expression level in blood CD8⁺ cells from malignant tumor patients were lower than in healthy and benign tumor groups. Contrary, the Kv1.3 conductance of CD8⁺ T cells were significantly higher in malignant tumor patients as compared to other two groups. To asses Ca²⁺ response of cells, we determined the quotient of FURA-2 ratios measured in 2 mM Ca²⁺ and 0 mM Ca²⁺ after thapsigargin addition: there was no differences between the groups.

In summary, we suppose that down-regulation of KCa3.1 expression and Kv1.3 level upregulation in blood CD8⁺ cells could be a reporter on the presence of malignancy, as we reported before for head and neck cancer patients [1].

Acknowledgment

This work was supported by Stipendium Hungaricum Scholarship to M.G. and NKFIH (K128525, P.H.).

References

[1] Chimote AA, Balajthy A, Arnold MJ, Newton HS, Hajdu P, Qualtieri J, et al. (2018) Sci Signal. 11(527): 1–12