AIP Conference Proceedings 2344, 040009 (2021); https://doi.org/10.1063/5.0049157 2344, 040009
© 2021 Author(s).
Nidogen-1 (NID1) as specific ligand in natural killer (NK) cells stimulated from advanced ovarian cancer: Immunotherapy review
Cite as: AIP Conference Proceedings 2344, 040009 (2021); https://doi.org/10.1063/5.0049157 Published Online: 23 March 2021
Sanya Khaerunnisa, Lady Feren Pangjaya, Nuzli Fahdia Mazfufah, Retno Lestari Budiman, and Radiana Dhewayani Antarianto
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Nidogen-1 (NID1) as Specific Ligand in Natural Killer (NK) Cells Stimulated from Advanced Ovarian Cancer:
Immunotherapy Review
Sanya Khaerunnisa
1, 2, Lady Feren Pangjaya
1, 2, Nuzli Fahdia Mazfufah
2, Retno Lestari Budiman
3, Radiana Dhewayani Antarianto
2, 4, a)1Biology Undergraduate Program, Faculty of Mathematics and Natural Science, Universitas Indonesia, Kampus UI Depok, West Java, 16424, Indonesia
2Stem Cell and Tissue Engineering Research Cluster, Indonesian Medical Education and Research Institute (IMERI), Universitas Indonesia, Salemba Raya, Jakarta, West Java, 10430, Indonesia
3Department of Biology, Faculty of Mathematics and Natural Science, Universitas Indonesia, Kampus UI Depok, West Java, 16424, Indonesia
4Department of Histology, Faculty of Medicine, Universitas Indonesia, Salemba Raya, Jakarta, West Java, 10430, Indonesia
a)Corresponding author: [email protected]
Abstract. The pathway for the introduction of NK cell into abnormal cells is mediated by the receptor-ligand interaction on the cell surface. Among all the activation receptors, NKP44 has become a unique activation receptor because it has an ITIMs signaling pathway and actively plays an important role in several functions of stimulated NK cells. A potential ligand that can specifically regulate the activity of NKP44 receptors is NID1. NID1 is a novel ligand that can initiate two different regulations on NK cells activity. In addition, NID1 also provides predictions that are compatible with the electrophoretic glycoprotein mobility that has been detected by NKp44 through SDS-PAGE and ELISA tests compared to all proteins in the microenvironment of ovarian cancer. Curently, NID1 has been proven to be able to increase the cytotoxic reaction of non-cancerous NK cells. Further research is needed for the development of ovarian cancer immunotherapy based on the receptor-ligand interactions.
Keywords: ovary cancer, NK cell, immunotherapy, NKP44, NID1
INTRODUCTION
Ovary cancer is the fifth-leading cause of death in 1out of 78 women over the world. The diagnosis of advanced ovarian cancer provides the largest contribution to the number cases of death from ovary cancer. More than 75%
patients are diagnosed with advanced disease (stage III or IV) [1]. Chemotherapy or surgery are types of treatment that are usually given to treat patients with ovarian cancer. However, chemo-resistance is a major challange that can increase the risk of cancer recurrence after treatment [2,3]. Therefore, finding other alternatives to ovarian cancer therapy must be done immediately [2]. Studies show that ovarian cancer is a type of immunogenic disease so immunotherapy is a highly recommended alternative treatmen for patiens with ovarian cancer[2].Immunotherapy is used to strengthen the hosts immune systems. One of the immunotherapy agents that can be used for the treatment of ovarian cancer is natural killer cells (NK). The ligand-receptor binding produces an activation signaling pathway activates NK cells so that cytokines or cytotoxic granules such as perforin and granzyme B are released to trigger apoptosis of target cells [4]. Activation receptor that can recognize many ligands and play an important role in the
cytotoxic reaction of stimulated peripheral blood NK cells is NKP44. Potential ligands that most influence the function of tumor cell recognition in the cancer microenvironment through NKP44 are Nidogen-1 (NID1) [5].
The NKP44 receptor was chosen because NKP44 is a receptor capable of recognizing many ligands and is actively playing the most important role in some of the activated NK cell functions. The NKP44 receptor is expressed on activated peripheral blood NK cells [5]. Micro or extracellular tissue vesicles can produce cytokines or chemokines that induce NKP44 expression. The extracellular vesicles consisting of exosomes, microparticles, immune cells, and fibroblasts were obtained from ultracentrifugation of ovarian tissue. One of the potential ligands from exosomes or microparticles that is known to affect the recognition function of tumor cells through the NKP44 receptor is Nidogen- 1 (NID1) [5].
The NID1 ligand is a ligand that is found in normal tissue and increases in all types of ovarian cancer, so the use of NID1 as an immunotherapy agent is promising [16]. The success of the research conducted by Gaggero et al. in 2018 proved that the NID1 microenvironment of non-cancerous tissues is an extracellular ligand that can regulate NKP44 receptors in non-cancerous NK cells [18]. Single cell immuno-staining is the method of detecting specific NID1 proteins using fluorescence antibodies was carried out to prove the success of the binding of the NID1 ligand and the NKP44 receptor [19].
Here, we summarize the latest advances made in understanding the potential and main challange of using NID1 as a candidate for ovarian cancer-specific immunotherapy agents to enhance the cytotoxic reaction of NK cell in cancer patients.
MATERIALS AND METHODS
This systematic literature search was performed on February-June 2020. A total of 13 articles consisting of review article and original article published in the last 5 years were retrieved. The combinations of terms used for the search included “ovary cancer”, “NK cell”, “immunotherapy”, “NKP44”, and “NID1”.
RESULT AND DISCUSSIONS
NK cells are lymphocytes of innate immunity that are located in the blood and almost all lymphoid organs. NK cell serve as the main defense mecanism that do not required prior sensitization to recognize antigens and mediate their anti-tumor effects. The cytotoxic function of NK cells are regulated by the interaction of activating and inhibitory receptors that bind to the corresponding target cell ligands originating from the cancer microenvironment or cell surface [6]. When major histocompatibility complex (MHC) which is always present on the surface of normal cell dissapear, the NK cell activation receptors will recognize these cells as an abnormal cells that must be killed through cytotoxic reactions [7]. Natural Cytotoxicity Receptors (NCRs), NKP30, NKP44, and NKP46 are activation receptors that play an important role in regulating cytotoxic reactions in NK cell through the signaling pathways of immune cell receptors based on the tyrosine-based activating motifs (ITAMs) to release cytokines or kill target cells directly[8].
NK cell activity was originally identified from human peripheral blood mononuclear cells and NKP44 is an activation receptor that actively plays an important role in several functions of stimulated peripheral blood NK cell [5]. Among the NCRs, only NKP44 can mediate both activating and inhibitory signals to NK cell depends on the ligand it binds because NKP44 is the only NCRs receptor that has tyrosine-based inhibitory motive (ITIMs) besides ITAMs signaling pathways [8]. The existence of this ITIMs signaling pathways makes NKP44 have a high level of effectiveness in tumor cell recognition [5].
The cancer microenvironment becomes an important factor that plays a role in mediating the development and inhibition of tumor cell growth [9]. Cancer microenvironment is a collection of cellular components as well as extracellular matrix components contained in cancer tissues. The constituents of the cancer microenvironment are mixtures of cancer cells, cancer associated fibroblasts (CAFs), immune cells and extracellular matrices which initiate the uptake and release of extracellular vesicles (EVs) filled with ligands, peptides, chamokines/cytokines, or short non coding RNA [9].
Extracellular vesicles (EV) are collections of components released by all types of cells into extracellular space such as the microenvironment of the tissue. Extracellular vesicles play a role in the process of mediating the transfer of proteins, lipids, and nucleic acids through the plasma membrane of tumor tissue and stroma that can be used as components of diagnosis, prognosis, or cancer therapy[10]. Extracellular vesicles consist of exosomes measuring 30- 150 nm and microparticles measuring 100 nm-1μm [11]. Proteins, antigens, lipids, miRNA and mRNA are molecules derived from total lysate of all tissues in the cell compartment such as exosomes. Exosomes in the tissue micro-
environment are produced by normal cells (healthy) and abnormal cells (pathological) [12]. Tissue microenvironment is capable of producing specific cytokines and chemokines which can induce the expression of activation receptors and increase the production of cytokines or NK cell cytotoxic granules. Receptor-ligand interactions in tumor cell recognition play an important role in cytotoxicity mediated by NK cells against tumor cells [5].
To obtain cancer microenvironment EVs, isolation or extraction of EVs by ultracentrifugation methods deemed necessary. Homogenization of ovary cancer tissue must first be carried out in specific buffer called radioimmunoprecipitation assay buffer (RIPA buffer). RIPA buffer will stabilize the protease-cofactor and maintain the pH of the tissue. It allows extraction of proteins from membranes, cell nuclei, and cytoplasm [13]. In addition, the ultracentrifugation process is also carried out at low temperatures (generally 4 °C) for 30 minutes at a speed of 104000xg (30,000 rpm) to obtain lysates or pellets consisting of cellular structures, microvesicles, exosomes, and proteins [14]. Ultrasentrifugation lysates containing exosomes, microparticles, and nucleic acids can then be used for further analysis such as SDS-PAGE, western-blot, electron microscopy, flow cytometry, and qRT-PCT [15].
NKP44 is able to recognize many ligands, especially those found in the tumor microenvironment. It is on this basis that the NKP44 receptor is a strong choice for fulfilling various functions and adjusting changes in tumor cells. The interaction of NKP44 receptors and their ligands in activating or regulating immunological processes can be utilized in the design of NK-based immunotherapy with various pathological conditions [5].
One potential ligand that is known to influence tumor cell recognition function through NKP44 towards cancer microenvironment receptors is Nidogen-1 (NID1) [5]. NID1 is an important component of the basement membrane that act as a linker between collagen IV, laminins, and perlecan to maintain the tissue architecture and structure.
Basement membrane is a thin layer that controls cellular activity. Research carried out by Zhou et al. in 2017 about the role of NID1 in cancer cell growth reported that the presence of NID1 in ovarian cancer tissue is closely related to the increased metastasis and therapy resistance that occur when cancer cells has successfully cross the basement membrane and spread to other organs [16]. Other studies conducted by Li et al. in 2015 showed that NID1 levels of advanced disease (stage III or IV) were significantly increased. These results indicate that NID1 can be a useful biomarker in ovarian cancer because the high expression of NID1 is highly correlated with poor survival in patients [17].
Currently, Nidogen-1 (NID1) is known as novel ligand that bind specifically to NKP44. Recent studies have expanded our knowledge about NID1 function for e immunotherapy mediated by NK cells. Research carried out by Gaggero et al. in 2018 reported that NID1 is an extracellular ligand that provides two different regulations on non- cancerous NK cell activity both activating or inhibiting through the production of IFN- γ cytokines [18]. Research by Gaggero et al. in 2018 conducted using normal NK cells showed that there are two forms of NID1 that can have different effects on the cytotoxic function of NK cells, namely, dissolved NID1 in the extracellular environment and NID1 attached to the surface of tumor cells. Dissolved NID1 can reduce NK cell cytokine production [18]. In contrast to NID1 in dissolved form, NID1 attached to the surface of tumor cells is able to increase the production of IFN-γ cytokines by NK cells [18]. This is different from the research conducted by Zhou et al. in 2015 with the results of studies showing that NID1 present on the surface of cancer cells played a role in increasing invasion, corrosion, and migration of ovarian cancer cells [16]. Dissolved NID1 can also be found in extracellular fluid according to the research of Barrow et al. 2019 it played a role in regulating the activity of activated blood NK cells [8].
FIGURE 1Ǥ Interaction between NKP44 and NID1ǤTw types f NID1, sNID1 as the dissolved form f NID1 and membrane bound NID1
NID1 NID1
NKP44 NKP44
NK cell
Cancer cell
NK cell
sNID1
These results provide an opportunity that the use of NID1 as an inducing agent can enhance the cytotoxic function of exhausted or relatively suppressed ovarian cancer NK cell. Furthermore, immunostaining experiments to prove the success of specific binding of NKP44 activation receptors and NID1 ligands in total ovary lysates need to be done first to support further research [18,19].
CONCLUSSION
In conclussion, b ǡNID1 may represent as a candidate prognostic indicator and a potential therapeutic target of ovarian cancer that can improve the efficiency of adaptive immunotherapy mediated NK cells. Further research is needed to better understand the ability of NID1 which is able to provide two regulation of NK cell activity for the purpose of developing ovarian cancer immunotherapy based on the receptor-ligand interactions.
ACKNOWLEDGMENTS
Funding for this research is obtained from Dana Masyarakat (DAMAS) UI through PUTI Universitas Indonesia Research Grant 2020 contract number ND-47/UN2.F1.DEPT.1/SDM.01.01/2020 and with principal investigator Radiana Dhewayani Antarianto.
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