title: TAMs re-polarization using a novel IFN-γ based concept
Creators: name: Berger, Nir
Publisher: name: UNSPECIFIED
Year: 2022

description: Immunotherapies using immune-checkpoint modulators have revolutionized the oncology field in the last decade, far beyond their remarkable clinical efficacy in some patients. Immunotherapy has created radical changes in the evaluation of treatment efficacy and toxicity with a more holistic vision of the patient with cancer. Scientists today consider tumor cells to be an orchestrator of the inflammatory response and not as a lone soldier. T-cell targeted immunomodulators are now used as single agents or in combination with chemotherapies as first or second lines of treatment for about 50 cancer types. Yet, as much as anti CTLA4, anti PD1 and anti PD-L1 are successful, there are always patients who do not respond to treatment in many indications, mainly in “cold” tumors, low in T cells and high in myeloid cells. A key component in checkpoint inhibitors’ efficacy is that they evoke the secretion of IFN-γ by T cells and NK cells in the tumor microenvironment. This IFN-γ can remodel macrophages from M2 like inhibitory state to M1 pro-inflammatory state and help reduce tumor growth and metastasis. In the past 30 years, some attempts were made to explore IFN-γ as a therapeutic concept for several tumor indications, but those attempts met several obstacles: the high sensitivity to the cytokine in humans resulted in high toxicity and inability to inject it I.V. The trapping of IFN-γ in non-target tissues and the endothelium via its high-affinity receptor, IFNγR1, rendered the introduction of tumor specificity antibody arm to be inefficient. Tumor associated macrophages (TAMs) are a major component of the tumor immune population and support low levels of inflammation in the tumor which reduces T cells and NK cells activity against the tumor. TAMs presence in the tumor are generally associated with poor prognosis and are known to support tumor neo-angiogenesis, extravasation and metastatic growth (Chávez-Galán et al., 2015) and are known to highly express PD-L1. In the present study, we try to combine anti PD-L1 “Avelumab” and IFN-γ cytokine in a bi-specific immuno-cytokine construct, that can target both myeloid and tumor cells expressing PD-L1, while remodeling the immune environment to a pro-inflammatory state and polarizing TAMs into M1-like inflammatory macrophages. The design is based on the Merck proprietary SEED IgG1.4 platform, with a bi-specific construct which includes an anti PD-L1 Fab specificity arm, and a single chain IFN-γ cytokine containing alterations and mutations. The mutated IFN-γ was reduces in valency from two to one, resulting in significantly reduced binding to IFNγR1 which should reduce the IFN-γ potency and toxicity while allowing for the anti PD-L1 specificity arm to be dominant and allow accumulation in the tumor. Herein, the bi-specific scIFNγ / anti PD-L1 constructs were able to show in human cell lines, specificity driven potency, which is valency dependent. Constructs were able to polarize M2 macrophages into M1 like pro-inflammatory macrophages. For In-vivo studies, a surrogate mouse single chain IFN-γ was developed, with novel point mutations that resulted in the same valency as the human construct and were able to be tolerated in I.V. injection, show localization and accumulation in RENCA tumors and show efficacy in LLC1 syngeneic tumors. Hence, the presented study demonstrates the potential and risk of using an IFN-γ based immune-cytokine platform with improved tolerability and tumor specificity.