DOI: 10.5061/dryad.n5tb2rbwc
title: Innate immune activation by checkpoint inhibition in patient-derived lung cancer tissues

description: Although Pembrolizumab-based immunotherapy has significantly improved lung
 cancer patient survival, many patients show variable efficacy and
 resistance development. A better understanding of the drug’s action is
 needed to improve patient outcomes. Functional heterogeneity of the tumor
 microenvironment (TME) is crucial to modulating drug resistance;
 understanding of individual patients’ TME that impacts drug response is
 hampered by lack of appropriate models. Lung organotypic tissue slice
 cultures (OTC) with patients’ native TME procured from primary and
 brain-metastasized (BM) non-small cell lung cancer (NSCLC) patients were
 treated with Pembrolizumab and/or beta-glucan (WGP, an innate immune
 activator). Metabolic tracing with 13C6-Glc/13C5,15N2-Gln, multiplex
 immunofluorescence (mIF), and digital spatial profiling (DSP) were
 employed to interrogate metabolic and functional responses to
 Pembrolizumab and/or WGP. Primary and BM PD-1+ lung cancer OTC responded
 to Pembrolizumab and Pembrolizumab + WGP treatments, respectively.
 Pembrolizumab activated innate immune metabolism and functions in primary
 OTC, which were accompanied by tissue damage. DSP analysis indicated an
 overall decrease in immunosuppressive macrophages and T cells but revealed
 microheterogeneity in immune responses and tissue damage. Two TMEs with
 altered cancer cell properties showed resistance. Pembrolizumab or WGP
 alone had negligible effects on BM-lung cancer OTC but Pembrolizumab + WGP
 blocked central metabolism with increased pro-inflammatory effector
 release and tissue damage. In depth metabolic analysis and multiplex TME
 imaging of lung cancer OTC demonstrated overall innate immune activation
 by Pembrolizumab but heterogeneous responses in the native TME of a
 patient with primary NSCLC. Metabolic and functional analysis also
 revealed synergistic action of Pembrolizumab and WGP in OTC of metastatic
 NSCLC.
description: The polar extracts were split for NMR and/or IC-UHR-FTMS analysis. 1D <sup>1</sup>H and <sup>1</sup>H(Albers et al.) heteronuclear single quantum coherence (HSQC) NMR analyses were performed on a Bruker Avance III 16.45 T spectrometer or an Agilent 14.1 T DD2 spectrometer outfitted respectively with a 1.7 mm cryoprobe or a 3 mm coldprobe as described previously (K. Sellers et al., 2015). IC-UHR-FTMS analysis was done as described previously (T. W. Fan et al., 2016) on a Dionex ICS-5000 ion chromatography system coupled to a Thermo Orbitrap Fusion Tribrid Fourier transform mass spectrometer with a mass resolving power of 370,000 at 400 m/z. Experimental details of sample preparation are provided in the manuscript (accepted).
description: Data uploads of curated FT-MS data files as .xlsx pivot tables.
 One file per tissue type.
 UK2035_OTC_13C6Glc_WGP_Pembro_MS.xlsx : Patient number 2035
 (metaststic lung cancer) tissue slices treated with 13C glucose
 ±WGP, pembrolizumab. Mass spectrometry.
 UK096_Mf_WGP_6well_2d_pol_13C6Glc_MS.xlsx: Patient number 96
 macrophages ±WGP 2D culture.
 UK096_Mf_3d_pol_13C15NQ_MS.xlsx: : Patient number 96 macrophages
 ±WGP 3D culture. UK131_OTC_13C6Glc_MS_data.xlsx:
 Patient number 131 tissue slices 13C Glucose tracer.
 UK131_OTC_13C15N_MS_data.xlsx : Patient number 131 tissue
 slices 13C, 15N Glutamine trace.