Exosomes and the NKG2D receptor-ligand system in pregnancy and cancer : using stress for survival

Abstract: Although not obvious at first sight, several parallels can be drawn between pregnancy andcancer. Many proliferative, invasive and immune tolerance mechanisms that supportnormal pregnancy are also exploited by malignancies to establish a nutrient supply andevade or edit the immune response of the host. The human placenta, of crucial importancefor pregnancy success, and its main cells, the trophoblast, share several features withmalignant cells such as high cell proliferation rate, lack of cell-contact inhibition andinvasiveness. Both in cancer and in pregnancy, the immune defense mechanisms,potentially threatening the survival of the tumor or the fetus, are progressively blunted oreven turned into tumor- or pregnancy-promoting players. Amongst immune mechanisms that are meant to protect the host from cancer and can be apotential threat to the fetus, the NKG2D receptor-ligand system stands out as the mostpowerful, stress-inducible “danger detector” system that comprises the activating NK cellreceptor NKG2D and its ligands, the MIC (MHC class I Chain-related proteins A and B)and ULBP (UL-16 Binding Proteins) families. It is the major cytotoxic mechanism in thebody promoting surveillance and homeostasis. In the present thesis we investigate theNKG2D receptor-ligand system in human early normal pregnancy and in theleukemia/lymphoma cell lines Jurkat and Raji and ask the questions “How is the NKG2Dreceptor-ligand system functioning in pregnancy and tumor? How is the danger of cytotoxicattack of the fetus avoided? Why is the immunosurveillance function compromised incancer patients?” We developed a method to isolate and culture villous trophoblast from early human normalplacenta and used it to study the NKG2D receptor-ligand system. We discovered that theNKG2D ligand families of molecules MICA/B and ULBP1-5 are constitutively expressedby the syncytiotrophoblast of the chorionic villi. Using immnunoelectron microscopy, westudied the expression of these molecules at the subcellular level and could show for thefirst time that they are preferably expressed on microvesicles in multivesicular bodies(MVB) of the late endosomal compartment and are secreted as exosomes. Exosomes arenanometer sized microvesicles of endosomal origin, produced and secreted by a great7variety of normal and tumor cells. The exosomes are packages of proteins and ribonucleicacids that function as “mail” or “messengers” between cells conveying different biologicalinformation. We isolated and studied exosomes from placental explant cultures. We foundthat they carry NKG2D ligands on their surface and are able to bind and down-regulate thecognate receptor on NK-, CD8+ and T cells. The down-regulation selectively causedimpairment of the cytotoxic response of the cells but did not affect their lytic ability asmeasured by perforin content and gene transcription. Thus, the NKG2D ligand-bearingexosomes suppress the cytotoxic activity of the cells in the vicinity of the placenta, leavingtheir cytolytic machinery intact, ready to function when the cognate receptor isrestored/recycled. These findings highlight the role of placental exosomes in the fetalmaternalimmune escape and support the view of placenta as an unique immunomodulatoryorgan. Next, we studied the expression and exosomal release of NKG2D ligands by tumor cellsusing the leukemia cell lines Jurkat and Raji as a tumor model. We found that NKG2Dligand-bearing exosomes with similar immunosuppressive properties as placental exosomesare constitutively secreted by the tumor cells, as a mechanism to blunt the cytotoxicresponse of the immune cells and thus protect themselves from cytotoxic attack by the host.Interestingly, we found that thermal- and oxidative stress up-regulates the exosomesecretion and the amount of exosome-secreted NKG2D ligands. Our results imply thattumor therapies that cause stress-induced damage, such as thermotherapy and stripping ofoxygen supply to the tumor, might have a previously unrecognized side effect causingenhanced exosome production and secretion, which in turn suppresses the natural antitumorimmune response and thus should be taken into account when designing an optimaltherapy of cancer patients. In conclusion, we describe a novel stress-inducible mechanism shared by placenta andtumors as an immune escape strategy. We found that placenta- and tumor-derived NKG2Dligand-bearing exosomes can suppress immune responses to promote the survival and wellbeing of the fetus or the tumor. Our work comprises an important contribution to theelucidation of the NKG2D ligand-receptor system and its mode of operation in the humanbody and opens new perspectives for designing novel therapies for infertility and cancer.