Article Text
PDF
Abstract
Background Immune checkpoint blockade targeting the PD-1/PD-L1 axis has revolutionized the treatment of many solid tumors. PD-L1 expression is known to be modulated by both infiltrating T cells and by tumor intrinsic mechanisms. Therapeutics targeting the macrophage checkpoints CD47 and SIRPα are now being evaluated clinically, but little is known about how CD47 expression is influenced by tumor molecular subtypes or the presence of immune cells, particularly macrophages.
Methods Formalin fixed and paraffin embedded samples of primary, resectable tumors from patients with HNSCC (N=44), TNBC (N=48) and CRC (N=48) were commercially procured and evaluated for CD47 (Abcam clone EPR21794) and SIRPα (LSBio LS-B551) expression by IHC. DNA/RNA were extracted from all samples and subjected to RNA-Seq and whole exome sequencing (WES). Multiplex immunofluorescence (mIF) was performed on 26 samples from each indication utilizing two custom 4-plex immunofluorescence assays. A macrophage panel evaluated CD68, CD163, PD-L1 and cytokeratin and a tissue resident memory T cell panel evaluated CD3, CD8, CD103 and cytokeratin.
Results CD47 expression was upregulated in the tumor versus the surrounding stroma in all three indications and was highest in HNSCC, followed by TNBC and CRC (figure 1). CD47 expression as measured by IHC was used to subdivide samples into high and low CD47 expression categories using the median within each indication. A phagocytic tumor associated macrophage (TAM) gene signature1 was significantly higher in the high CD47 samples from HNSCC and CRC (figure 1). Characterization of macrophages (mIF panel) indicated that CD163+/CD68- macrophages were the most abundant macrophages in all three indications, demonstrated increased prevalence within the tumor stroma, and correlated strongly with the phagocytic TAM gene signature. PD-L1 expression was predominantly found on CD68+/CD163- macrophages in all tumor indications. We classified samples using molecular subtyping for CRC,2 and unsupervised clustering for HNSCC and TNBC. CD47 expression was generally not differentiated across the CRC molecular subtypes.
Conclusions CD47 targeting therapies are currently being evaluated in clinical trials in multiple solid tumor indications. Here were demonstrate that CD47 expression is higher in tumor compared to stroma in HNSCC, TNBC and CRC and correlates with an increased phagocytic TAM signature. These data suggest that some tumors may avoid immune surveillance by upregulating CD47 in the presence of phagocytic macrophages, and that tumors with these characteristics could be rationale candidates for evaluation of CD47 based therapies.
References
Zhang L, Li Z, Skrzypczynska KM, Fang Q, Zhang W, O’Brien SA, He Y, Wang L, Zhang Q, Kim A et al. Single-Cell Analyses Inform Mechanisms of Myeloid-Targeted Therapies in Colon Cancer. Cell 2020, 181(2):442–459 e429.
Dienstmann R, Vermeulen L, Guinney J, Kopetz S, Tejpar S, Tabernero J: Consensus molecular subtypes and the evolution of precision medicine in colorectal cancer. Nat Rev Cancer 2017, 17(2):79–92.
This is an open access article distributed in accordance with the Creative Commons Attribution Non Commercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited, appropriate credit is given, any changes made indicated, and the use is non-commercial. See http://creativecommons.org/licenses/by-nc/4.0/.