Supplementary Data
Supplementary Figure S1. H-scores for pERK and pS6 in paired baseline and day 15 on-treatment tumor biopsy specimens from BM2 patients treated with D+T+P. H-scores were derived in work by Corcoran et al (1). A paired t test was used to assess the difference between baseline and on-treatment H-scores. Paired biopsy samples from baseline and day 15 were available for n = 15 BM2 patients.
Supplementary Figure S2. Baseline and on-treatment expression of MPAS gene signature in BM1 and BM2 patients. A paired t test was used to assess the difference between baseline and on-treatment expression. Paired biopsy samples from baseline and day 15 were available for n = 34 D+T+P-treated patients.
Supplementary Figure S3. Key baseline gene expression profiles by BM subtype. A, BIM expression at baseline by BM subtype. B, MAPK gene signature expression (MPAS) at baseline by BM subtype. P values were derived using a two-tailed t test (BM1 samples, n = 16; BM2 samples, n = 31 [3 BM2 samples were omitted as not evaluable]).
A
B
Supplementary Figure S4. Elevated expression of cell cycle gene signatures in BM2 subtype samples. Box plots show G2M and E2F signature expression at baseline by BM subtype. P values were derived using a two-tailed t test. (BM1 samples, n = 16; BM2 samples, n = 34).
Supplementary Figure S5. Baseline and on-treatment expression of immune gene signatures associated with response to D+T+P treatment in BM1 and BM2 patients. A paired t test was used to assess the difference between baseline and on- treatment expression. Paired biopsy samples from baseline and day 15 were available for n = 34 D+T+P-treated patients.
Supplementary Table S1. List of specific genes that compose each of the 20 immune gene signatures evaluated. Gene signatures are previously unpublished.
Signature Gene
B-cell CD79B CD19
BLK SPIB
CXCR5 TNFRSF13B
MS4A1 TNFRSF13C
B-cell - memory IGJ IGHD
TNRFSF17 IGLL5
FCRL5 IGHG1
MZB1
Chemokine - myeloid CCL2 CCL4L2
CCL3 CCL8
CCL4 OSM
Chemokine - neutrophil CCL20 CXCL2
CXCL1 CXCL5
CXCL3 IL8
Cytotoxic cell FASLG CCL5
PRF1 GZMA
GZMH NKG7
Fibroblast C3orf80 MEDAG
CTHRC1 COLEC12
EDNRA COL3A1
ADAMTS12 COL10A1
SFRP2 BNC2 COL1A2 FBLN2
COL1A1 VGLL3
RGS4 GXYLT2
COL6A3 MMP2
GPC6 FAP
WISP1 LGI2
GREM1 CLMP
EMILIN1 TNN
LOC100132891 THBS2
ALPK2 MFAP5
FNDC1 ASPN
HAS2 DKK2
POSTN SULF1
CPXM1 COL5A2
KCNE4 ITGA11
TNFAIP6 LRRC15
COL8A1
IFN-γ IFNG CXCL11
CXCL9 GBP1
CXCL10
Langerhans DC CD207 CD1C
FCER1A CLEC10A
CD1A HLA-DQB2
CD1B CCL17 CD1E CCL22
M2 LILRB5 F13A1
CD209 MRC1
FOLR2 CD163L1
IL10 CD163
MS4A4A
Macrophage CSF1R ITGAX
MSR1 ITGAM
FCGR3A FPR1
SIGLEC7 GPR34
TLR8 TLR7
CD84 LST1
CYBB SIGLEC1
LILRB1 VSIG4
LILRB2 FCGR2A
HAVCR2 C1QA
LILRB4 C1QB
CD86 C1QC
FCER1G
CD68
Mast cell TPSAB1 HDC
CDK15 C1orf186
TPSD1 CPA3
TPSB2 SIGLEC17P HPGDS RGS13
MS4A2 SIGLEC6
Osteoclast-like ATP6V0D2 DCSTAMP
DNASE2B HS3ST2
CCL18 CHIT1 pDC-like PTCRA LSP1
LGALS9 DOK2
LST1
Phagocytic CLEC9A PLD4 CLEC9A
CX3CR1 SIGLEC8
GAPT
PMN 1 PROK2 CXCR2
FCGR3B CXCR1
FFAR2
PMN 2 FCAR LILRA5
FPR1 MEFV
FPR2
CD300E
PMN 3 PLAUR TREM1
IL1RN AQP9
CCL7
SLC11A1
T-cell CD8A CD3G
CD8B CD247 PYHIN1 TCRVB
TRAT1 CD3D
GZMK CD3E
SH2D1A CD2
CXCR6 SIRPG
Treg CRTAM TIGIT
FOXP3 CXCL13
LAIR2 CTLA4
ICOS TNIP3
LTA ZBED2
Type I IFN SP100 IFIT2
BST2 IFI6
MX1 ISG15
MX2 OASL
USP18 RSAD2
IFIH1 IFI44L
IFIT1 OAS2
Supplementary Table S2. Demographics and disease characteristics of BM1 and BM2 patients treated with D+T+P
BM1 BM2 (n = 16) (n = 34) 62.5 63.5 Age, median (range), y (35 - 77) (42 - 74) Gender, n(%)a Female 11 (69) 17 (50) Male 5 (31) 17 (50) ECOG performance status, n (%) 0 7 (44) 15 (44) 1 9 (56) 19 (56) Prior lines of therapy, n (%) 0 2 (13) 4 (12) 1 8 (50) 12 (35) 2 5 (31) 14 (41) 3 1 (6) 3 (9) 4 0 1 (3) Prior anti-EGFR therapy, n (%) Yes 3 (19) 6 (18) No 13 (81) 28 (82) Primary tumor location, n (%) Colon 13 (81) 26 (76) Left Side 3 (23) 10 (38) Right Side 10 (77) 16 (62) Rectum 3 (19) 8 (24) ECOG, Eastern Cooperative Oncology Group. a P = 0.24 by Fisher exact test, indicating that BM subtype is not related to gender.
Supplementary Table S3. On-treatment modulation of 20 immune metagenes in BM1 and BM2 patients treated with D+T+P. A paired t test was used to assess the difference between baseline and on-treatment expression, and P < 0.05 was considered significant.
Mean Mean Fold Expression Expression P Value Change Signature (Day 1) (Day 15)
B-cell 0.0466 0.4143 0.3677 0.287
B-cell - memory 0.1511 0.6790 0.5279 0.057
Chemokine - -1.0155 -0.8076 0.2078 0.429 myeloid
Chemokine - 0.9898 0.6128 -0.3770 0.276 neutrophil
Cytotoxic cell -0.1334 0.9518 1.0851 0.003
DC_Langerhans -2.4935 -2.4015 0.0919 0.739
Fibroblast 3.4008 3.5765 0.1757 0.486
IFN-γ 0.8993 0.6137 -0.2856 0.412
M2 2.0088 2.5293 0.5205 0.070
Macrophage 2.8103 3.3017 0.4914 0.024
Mast cell -1.2541 -0.5097 0.7444 0.011
Osteoclast-like 0.4456 1.1551 0.7095 0.084 pDC-like 2.0548 2.5640 0.5092 0.003
Phagocytic CLEC9A -0.9702 -0.0405 0.9297 0.001
PMN 1 -1.9547 -1.5361 0.4186 0.123
PMN 2 -0.7200 -0.3391 0.3809 0.193
PMN 3 1.8373 1.5294 -0.3079 0.109
T-cell 1.2204 1.8931 0.6727 0.018 Treg -0.7021 -0.5706 0.1315 0.594
Type I IFN 4.2423 4.2090 -0.0333 0.835
Supplementary Reference
1. Corcoran RB, Andre T, Atreya CE, Schellens JHM, Yoshino T, Bendell JC, et al. Combined BRAF, EGFR, and MEK Inhibition in Patients with BRAF(V600E)- Mutant Colorectal Cancer. Cancer Discov 2018;8(4):428-43.