Crosstalk Between NK Cell Receptors and Tumor Membrane Hsp70-Derived Peptide: A Combined Computational and Experimental Study

Natural killer (NK) cells are central components of the innate immunity system against cancers. Since tumor cells have evolved a series of mechanisms to escape from NK cells, developing methods for increasing the NK cell antitumor activity is of utmost importance. It is previously shown that an ex vivo stimulation of patient-derived NK cells with interleukin (IL)-2 and Hsp70-derived peptide TKD (TKDNNLLGRFELSG, aa450-461) results in a significant upregulation of activating receptors including CD94 and CD69 which triggers exhausted NK cells to target and kill malignant solid tumors expressing membrane Hsp70 (mHsp70). Considering that TKD binding to an activating receptor is the initial step in the cytolytic signaling cascade of NK cells, herein this interaction is studied by molecular docking and molecular dynamics simulation computational modeling. The in silico results showed a crucial role of the heterodimeric receptor CD94/NKG2A and CD94/NKG2C in the TKD interaction with NK cells. Antibody blocking and CRISPR/Cas9-mediated knockout studies verified the key function of CD94 in the TKD stimulation and activation of NK cells which is characterized by an increased cytotoxic capacity against mHsp70 positive tumor cells via enhanced production and release of lytic granules and pro-inflammatory cytokines. This work investigates the interaction between the functional part of Hsp70 protein (TKD) and NK cell receptors through in silico and in vitro studies. CD94 activating receptor plays a major role in the TKD-mediated NK cell stimulation. The CD94 complex with TKD part of membrane Hsp70 overexpressed on tumor cells is a key step to trigger the cytolytic signaling cascade of NK cells leading to enhanced release of lytic granules. image