Figure 7. Expansion of CD71⁺ neutrophils in cancer patients (see also Fig. S7).

A) Representative contour plot of CyTOF analysis of 14 melanoma samples and 5 healthy controls to identify CD71 positive cells among CD66b⁺CD15⁺CD16⁻CD10⁻neutrophils in blood of healthy donors and melanoma patients suggested expansion of neutrophil progenitor subset N1 in blood of cancer patients. Gating also included exclusion of dead cells and cells expressing CD3, CD19, CD56 and CD14 (not shown).

B) Frequency of neutrophil progenitor subset N1 within all leukocytes or ratio of subset N-to-CD66b⁺neutrophils in blood of healthy donors and melanoma patients. Data analyzed according to results shown in panel A. n = 5 healthy donors, n = 14 melanoma patients. Welch’s t-test was performed and p-value shown.

C) Differentially expression test of protein marker expression between CD71− and CD71+ neutrophils (Wilcox rank sum test, Fig. S7C) revealed a number of important markers including progenitor markers CD38, CD48, CD49d, maturation markers CD16, CD10, CD35, CD101, antigen-presenting markers CD86, HLA-DR, CD64 and angiogenesis-associated marker CD304 (star indicating FDR-corrected p values < 0.01, 2-fold change difference in the expression median, Fig. S7C). Expression value was normalized from 0–1 for each marker.

D) CD71⁺ eNeP are found in the blood of NSCLC patients and in lung tumor samples (from our reanalysis of single cell RNA sequencing data sets (Zilionis et al. 2019)). Each dot is one cell, red dots representing positive CD71 expression.

E) Violin plot from scRNA-Seq of CD71⁺ neutrophils and other neutrophils (CD71⁻, data set also depicted in D).