Supplementary Information
PD, SD, CM
DISEASE PROGRESSION, DISEASE
Below is the link to the electronic supplementary material.Supplemental Fig. 1. Broad immune characterization of peripheral blood of patients treated with doxorubicin and pembrolizumab at baseline. A) Pre-treatment peripheral blood samples were examined by flow cytometry for lymphocyte composition frequencies, including CD3 + T cells, CD3 − CD56 + natural killer (NK) cells, CD3 + CD56 + natural killer T (NKT) cells, TCRγδ T cells, and CD19 + B cells. B) The composition of NK cells was further assessed for CD56bright (early), CD16bright (mature), and CD56dim CD16dim (terminal) NK cell subsets. C) B cell subsets were assessed as IgD + CD27- (Naïve), IgD- CD27 + (memory), or CD38 + plasmablasts. D) Monocyte subsets were assessed as a fraction of a myeloid cell gate for frequencies of CD14 + CD16- classical monocytes (clMono), CD14 + CD16 + intermediate monocytes (intMono), and CD14- CD16 + non-classical monocytes (ncMono). E) Dendritic cell subsets were similarly assessed as a fraction of a myeloid cell gate for frequencies of CD141 + dendritic cells (cDC1), CD1c + dendritic cells (cDC2), and CD123 + plasmacytoid dendritic cells (pDC). F) Percent change from pre-treatment to C2D1 (purple) and from pre-treatment to post Cycle 3 (green) is shown. Patient with disease progression (PD, n = 1) is depicted in dark blue, stable disease (SD, n = 2) in light blue, confirmed and unconfirmed partial response (PR = 5) in dark red, and complete response (CR, n = 1) in light red. *p < 0.05. Supplemental Fig. 2. Canonical T cell subsets at baseline. CD8 + T cells (A) and CD4 + T cells (B) were further assessed by flow cytometry for frequencies of naïve (CCR7 + , CD45RA +), central memory (CM; CCR7 + , CD45RA-), effector memory (EM; CCR7-, CD45RA-), and effector memory CD45RA + (EMRA, CCR7-, CD45RA +) cell subsets. Frequencies of regulatory T cells (Tregs) were also assessed within CD4 + T cells. Data are shown separately for patients with PD (dark blue), SD (light blue), PR (dark red), and CR (light red). Supplemental Fig. 3. Detailed T cell composition changes over the course of therapy in patients treated with doxorubicin and pembrolizumab. Percentage change of all identified CD8 + T cell subsets (A) and CD4 + T cell subsets (B) from pre-treatment to C2D1 (purple) and pre-treatment to post Cycle 3 (red). Fold change in CD8 + (C) and CD4 + (D) T cell subsets are shown separately for patients with PD (dark blue), SD (light blue), PR (dark red), and CR (light red). Supplemental Fig. 4. Detailed dynamics of proliferating CD8 + T cells in patients treated with doxorubicin plus pembrolizumab. Pre-treatment frequencies (A), fold change from pre-treatment to C2D1 (B), and fold change from pre-treatment to C3D1 (C) are shown for PD-1low Ki-67 + and PD-1high Ki-67 + CD8 + T cells. PD (dark red), SD (light red), PR (light blue), and CR (dark blue). Fold change of all T cell subsets, with the inclusion of identified PD-1low Ki-67 + and PD-1high Ki-67 + populations, from pre-treatment to C2D1 (D) and from pre-treatment to C3D1 (E). (PDF 243 kb)