Combining Immunocytokine and Ex Vivo Activated NK Cells as a Platform for Enhancing Graft-Versus-Tumor Effects Against GD2+ Murine Neuroblastoma

Abstract

Management for high-risk neuroblastoma (NBL) has included autologous hematopoietic stem cell transplant (HSCT) and anti-GD2 immunotherapy, but survival remains around 50%. The aim of this study was to determine if allogeneic HSCT could serve as a platform for inducing a graft-versus-tumor (GVT) effect against NBL with combination immunocytokine and NK cells in a murine model. Lethally irradiated C57BL/6 (B6) x A/J recipients were transplanted with B6 bone marrow on Day +0. On day +10, allogeneic HSCT recipients were challenged with NXS2, a GD2+ NBL. On days +14-16, mice were treated with the anti-GD2 immunocytokine hu14.18-IL2. In select groups, hu14.18-IL2 was combined with infusions of B6 NK cells activated with IL-15/IL-15Rα and CD137L ex vivo. Allogeneic HSCT alone was insufficient to control NXS2 tumor growth, but the addition of hu14.18-IL2 controlled tumor growth and improved survival. Adoptive transfer of ex vivo CD137L/IL-15/IL-15Rα activated NK cells with or without hu14.18-IL2 exacerbated lethality. CD137L/IL-15/IL-15Rα activated NK cells showed enhanced cytotoxicity and produced high levels of TNF-α in vitro, but induced cytokine release syndrome (CRS) in vivo. Infusing Perforin-/- CD137L/IL-15/IL-15Rα activated NK cells had no impact on GVT, whereas TNF-α-/- CD137L/IL-15/IL-15Rα activated NK cells improved GVT by decreasing peripheral effector cell subsets while preserving tumor-infiltrating lymphocytes. Depletion of Ly49H+ NK cells also improved GVT. Using allogeneic HSCT for NBL is a viable platform for immunocytokines and ex vivo activated NK cell infusions, but must be balanced with induction of CRS. Regulation of TNFα or activating NK subsets may be needed to improve GVT effects.

The immune landscape of solid pediatric tumors

“…However, immune-mediated cancer cell recognition and killing might occur independently of the availability of mutated antigen, and other forms of immunotherapy might be exploited in tumors with low mutational load [ 16 ], including pediatric cancers. These include for instance T-cell or NK-cell based adoptive therapy [ 17 ], vaccines directed at non-mutated antigens, and oncolytic therapy, alone or in combination with checkpoint inhibitors [ 18 ] [ 19 ]. These approaches have demonstrated encouraging results in pre-clinical models [ 17 – 19 ], but clinical successes using immunotherapy have been only obtained in neuroblastoma, a tumor type for which observed spontaneous remissions are likely mediated by cellular immunity [ 20 , 21 ].…”

“…These include for instance T-cell or NK-cell based adoptive therapy [ 17 ], vaccines directed at non-mutated antigens, and oncolytic therapy, alone or in combination with checkpoint inhibitors [ 18 ] [ 19 ]. These approaches have demonstrated encouraging results in pre-clinical models [ 17 – 19 ], but clinical successes using immunotherapy have been only obtained in neuroblastoma, a tumor type for which observed spontaneous remissions are likely mediated by cellular immunity [ 20 , 21 ]. In high-risk patients, complementing standard therapy with dinutuximab, which targets the NBL-associated antigen GD2, interleukin-2 (IL-2) and granulocyte-monocyte colony-stimulating factor (GM-CSF) increased event-free and overall survival [ 22 ] but is not curative for the majority of patients who will ultimately relapse and die [ 17 ].…”

Strategies for Potentiating NK-Mediated Neuroblastoma Surveillance in Autologous or HLA-Haploidentical Hematopoietic Stem Cell Transplants

“…Benefits include the possibility to use haplo healthy donors as a source of NK cells without significant GVH, and the constitutive surface expression of activating NK receptors (NCR, NKG2D, DNAM-1) recognizing tumor antigens and co-operating with CARs [ 19 , 182 ]. All the available CAR-based platforms, some of which are designed to favor CAR-cell persistence and tumor invasiveness [ 183 , 184 , 185 , 186 ], have the risk of exacerbating the GVHD [ 187 ]. This event, however, could be controlled with approaches including the use of CAR construct holding suicide genes [ 188 ] or the infusion of ex vivo expanded donor-derived regulatory lymphocyte subpopulations [ 72 ].…”

A Survival Analysis on the Immune Landscape of Paediatric Solid Tumours.

“…T-cell or NK-cell-based adoptive therapy [19], vaccines targeting non-mutated antigens, and oncolytic therapy, either alone or in conjunction with other treatments, are a few examples of these. Inhibitors of checkpoints [20] [21].…”

“…Inhibitors of checkpoints [20] [21]. These methods have revealed intriguing outcomes in pre-clinical models [19][20][21], although immunotherapy clinical successes have only been attained in tumor-type neuroblastoma with the likeli-hood of the reported spontaneous remissions cellmediated immunity [22,23]. In high-danger patients receiving dinutuximab, which inhibits the NBL-associated antigen GD2, in addition to conventional therapy Granulocyte-monocyte colonystimulating factor (GM-CSF) and interleukin-2 (IL-2), increased event-free and overall survival [24], but is not generally curative who will eventually relapse and pass away [19].…”