The huKS-IL2 immunocytokine (IC) consists of IL2 fused to a mAb against EpCAM, while the hu14.18-IL2 IC recognizes the GD2 disialoganglioside. They are under evaluation for treatment of EpCAM(+) (ovarian) and GD2(+) (neuroblastoma and melanoma) malignancies because of their proven ability to enhance tumor cell killing by antibody-dependent cell-mediated cytotoxicity (ADCC) and by antitumor cytotoxic T cells. Here, we demonstrate that huKS-IL2 and hu14.18-IL2 bind to tumor cells via their antibody components and increase adhesion and activating immune synapse (AIS) formation with NK cells by engaging the immune cells' IL-2 receptors (IL2R). The NK leukemia cell line, NKL (which expresses high affinity IL2Rs), shows fivefold increase in binding to tumor targets when treated with IC compared to matching controls. This increase in binding is effectively inhibited by blocking antibodies against CD25, the α-chain of the IL2R. NK cells isolated from the peritoneal environment of ovarian cancer patients, known to be impaired in mediating ADCC, bind to huKS-IL2 via CD25. The increased binding between tumor and effector cells via ICs is due to the formation of AIS that are characterized by the simultaneous polarization of LFA-1, CD2 and F-actin at the cellular interface. AIS formation of peritoneal NK and NKL cells is inhibited by anti-CD25 blocking antibody and is 50-200% higher with IC versus the parent antibody. These findings demonstrate that the IL-2 component of the IC allows IL2Rs to function not only as receptors for this cytokine but also as facilitators of peritoneal NK cell binding to IC-coated tumor cells.
"Alternative approaches for IL-2 based immunotherapy e.g. toxin conjugates, antibodies, fusion proteins, gene therapy are therefore currently being explored in various cancers (Eigentler et al., 2011; Telang et al., 2011; Gubbels et al., 2011). However, since IL-2 is essential for the development, survival and function of regulatory T (Treg) cells, which function to inhibit immune responses and prevent autoimmune disease, IL-2 may have a role in promoting T cell tolerance (an important consideration is the dose of IL-2 used as a low dose appears to favour tolerance over autoimmunity). "
[Show abstract][Hide abstract] ABSTRACT: Two candidate preparations of human sequence recombinant Interleukin-2 (IL-2) were formulated and lyophilized at NIBSC prior to evaluation in a collaborative study for their suitability to serve as a replacement international standard. The preparations were tested by eight laboratories using in vitro bioassays and immunoassays. The candidate preparation 86/500 was judged suitable to serve as a replacement international standard based on the data obtained for activity and stability. On the basis of the results reported here, the preparation coded 86/500 was established by the WHO Expert Committee on Biological Standardisation (ECBS) in 2012 as the WHO 2nd IS for human IL-2 with an assigned value for IL-2 activity of 210 IU/ampoule. Calibration of the 2nd IS is primarily based on the bioassay in use in various laboratories and relies exclusively on the estimates calculated relative to the WHO 1st IS for continuity of the IU.
"Some IL-2 centered strategies have already been approved by the FDA for the treatment of metastatic melanoma  and renal cell carcinoma . Intense research (basic, translational, and clinical) is also underway on the use of IL-2 and IL-2-antibody conjugates (immunocytokines) to boost the anti-cancer activities of NK    . Our initial studies on the characterization of the global proteome of naïve and IL-2-stimulated human NK cells reveal a large number of proteins exhibiting changes in expression levels upon IL-2 stimulation. "
[Show abstract][Hide abstract] ABSTRACT: Unlabelled:
Natural killer (NK) cells efficiently cytolyse tumors and virally infected cells. Despite the important role that interleukin (IL)-2 plays in stimulating the proliferation of NK cells and increasing NK cell activity, little is known about the alterations in the global NK cell proteome following IL-2 activation. To compare the proteomes of naïve and IL-2-activated primary NK cells and identify key cellular pathways involved in IL-2 signaling, we isolated proteins from naïve and IL-2-activated NK cells from healthy donors, the proteins were trypsinized and the resulting peptides were analyzed by 2D LC ESI-MS/MS followed by label-free quantification. In total, more than 2000 proteins were identified from naïve and IL-2-activated NK cells where 383 proteins were found to be differentially expressed following IL-2 activation. Functional annotation of IL-2 regulated proteins revealed potential targets for future investigation of IL-2 signaling in human primary NK cells. A pathway analysis was performed and revealed several pathways that were not previously known to be involved in IL-2 response, including ubiquitin proteasome pathway, integrin signaling pathway, platelet derived growth factor (PDGF) signaling pathway, epidermal growth factor receptor (EGFR) signaling pathway and Wnt signaling pathway.
The development and functional activity of natural killer (NK) cells is regulated by interleukin (IL)-2 which stimulates the proliferation of NK cells and increases NK cell activity. With the development of IL-2-based immunotherapeutic strategies that rely on the IL-2-mediated activation of NK cells to target human cancers, it is important to understand the global molecular events triggered by IL-2 in human NK cells. The differentially expressed proteins in human primary NK cells following IL-2 activation identified in this study confirmed the activation of JAK-STAT signaling pathway and cell proliferation by IL-2 as expected, but also led to the discovery and identification of other factors that are potentially important in IL-2 signaling. These new factors warrant further investigation on their potential roles in modulating NK cell biology. The results from this study suggest that the activation of NK cells by IL-2 is a dynamic process through which proteins with various functions are regulated. Such findings will be important for the elucidation of molecular pathways involved in IL-2 signaling in NK cells and provide new targets for future studies in NK cell biology.
Journal of proteomics 06/2013; 91. DOI:10.1016/j.jprot.2013.06.024 · 3.89 Impact Factor
"This bridging results in the formation of an AIS defined by the polarization of effector molecules including adhesion molecules and IL2Rs on the NKL cells to the immune synapse and the subsequent hu14.18-IL2 IC-facilitated NK cell-mediated tumor cell killing (Gubbels et al., 2011). It is possible that the beneficial effect of high affinity CD16 alleles may be less important with IC than with a conventional mAb. "
[Show abstract][Hide abstract] ABSTRACT: Disease recurrence is frequent in high-risk neuroblastoma (NBL) patients even after multi-modality aggressive treatment [a combination of chemotherapy, surgical resection, local radiation therapy, autologous stem cell transplantation, and cis-retinoic acid (CRA)]. Recent clinical studies have explored the use of monoclonal antibodies (mAbs) that bind to disialoganglioside (GD(2)), highly expressed in NBL, as a means to enable immune effector cells to destroy NBL cells via antibody-dependent cell-mediated cytotoxicity (ADCC). Preclinical data indicate that ADCC can be more effective when appropriate effector cells are activated by cytokines. Clinical studies have pursued this by administering anti-GD(2) mAb in combination with ADCC-enhancing cytokines (IL2 and GM-CSF), a regimen that has demonstrated improved cancer-free survival. More recently, early clinical studies have used a fusion protein that consists of the anti-GD(2) mAb directly linked to IL2, and anti-tumor responses were seen in the Phase II setting. Analyses of genes that code for receptors that influence ADCC activity and natural killer (NK) cell function [Fc receptor (FcR), killer immunoglublin-like receptor (KIR), and KIR-ligand (KIR-L)] suggest patients with anti-tumor activity are more likely to have certain genotype profiles. Further analyses will need to be conducted to determine whether these genotypes can be used as predictive markers for favorable therapeutic outcome. In this review, we discuss factors that affect response to mAb-based tumor therapies such as hu14.18-IL2. Many of our observations have been made in the context of NBL; however, we will also include some observations made with mAbs targeting other tumor types that are consistent with results in NBL. Therefore, we hypothesize that the NBL observations discussed here may also be relevant to mAb therapy for other cancers, in which ADCC is known to play a role.
Frontiers in Pharmacology 05/2012; 3:91. DOI:10.3389/fphar.2012.00091 · 3.80 Impact Factor
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.