Abstract 3060: Temporal analysis of type 1 interferon activation in tumor cells following external beam radiotherapy or targeted radionuclide therapy
Authors: Jagodinsky Justin C. , Bates Amber M. , Hernandez Reinier , Grudzinski Joseph J. , Marsh Ian R. , Chakravarty Ishan , Arthur Ian S. , Zangl Luke M. , Brown Ryan J. , Nystuen Erin J. , Emma Sarah E. , Kerr Caroline , Jin Won Jong , Carlson Peter M. , Engle Jonathan W. , Aluicio-Sarduy Eduardo , Barnhart Todd E. , Le Trang , Kim KyungMann , Bednarz Bryan P. , Weichert Jamey P. , Patel Ravi B. , Morris Zachary S. 
Published: July 2021

Abstract
Background: Radiation (RT) activates a type 1 interferon (IFN1) response and this is critical to the effect of RT in priming a response to immune checkpoint blockade (ie anti-CTLA4). However, little is known about the time course of this effect. Clinical interest in utilizing systemically administered targeted radionuclide therapy agents (TRT) is growing as these agents can be used to target multiple sites of disease including micro-metastases. It is unclear how IFN1 activation induced by continuous delivery of RT during exponential decay of a TRT source will compare to that induced following instantaneous external beam RT (EBRT). Here we report the time course of IFN1 response following RT in vitro and in vivo.
Methods: For in vitro studies, we utilized murine models of melanoma (B16, B16 STING knockout, B78), and head and neck squamous cell carcinoma (MOC2). EBRT was prescribed to 2.5 Gy, 12 Gy, or 3 fractions of 8 Gy. For in vivo studies, syngeneic C57BL/6 mice were engrafted with either B78 or MOC2 cells on the flank and RT was delivered when mean tumor size was 100-150 mm3. EBRT was prescribed to 2.5 Gy or 12 Gy. For TRT, we used 90Y conjugated to NM600, an alkylphosphocholine analog that exhibits selective uptake and retention in tumor cells of nearly any type, including B78 and MOC2. Tumor-specific dosimetry for 90Y-NM600 was determined using sequential 86Y-NM600 PET/CT imaging (3h, 24h, 48h, 72h) and a Monte Carlo based dose calculation platform. TRT was prescribed to a cumulative absorbed dose of 2.5 Gy or 12 Gy. Following delivery of RT, cells or tumors were harvested at 1d, 7d, and 14d post RT and RNA was isolated. Gene expression of Ifn-β and IFN response elements was quantified by qPCR and normalized to untreated controls.
Results: We observed significant IFN1 activation in all cell lines, with peak activation in B78, B16, and MOC2 cell lines occurring 7, 7, and 1 days, respectively, following RT for all doses. This effect was STING-dependent. Select IFN response genes remained upregulated at 14 days following RT. IFN1 activation following STING agonist treatment in vitro was identical to RT suggesting time course differences between cell lines were mediated by STING pathway kinetics and not DNA damage susceptibility. In vivo delivery of EBRT and TRT to B78 and MOC2 tumors resulted in a comparable time course and magnitude of IFN1 activation. In the MOC2 model, the combination of 90Y-NM600 and anti-CTLA-4 therapy reduced tumor growth and prolonged survival compared to single agent therapy.
Conclusions: We report the time course of the STING-dependent IFN1 response following RT in multiple murine tumor models. We show the potential of TRT to stimulate IFN1 activation that is comparable to that observed with EBRT of equivalent cumulative dose. Further evaluation of the timing and magnitude of IFN1 response following EBRT and TRT may be critical to the optimal integration with immunotherapies.
Citation Format: Justin C. Jagodinsky, Amber M. Bates, Reinier Hernandez, Joseph J. Grudzinski, Ian R. Marsh, Ishan Chakravarty, Ian S. Arthur, Luke M. Zangl, Ryan J. Brown, Erin J. Nystuen, Sarah E. Emma, Caroline Kerr, Won Jong Jin, Peter M. Carlson, Jonathan W. Engle, Eduardo Aluicio-Sarduy, Todd E. Barnhart, Trang Le, KyungMann Kim, Bryan P. Bednarz, Jamey P. Weichert, Ravi B. Patel, Zachary S. Morris. Temporal analysis of type 1 interferon activation in tumor cells following external beam radiotherapy or targeted radionuclide therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 3060.