1. TNBC Supplementary Figure 1 d e f HD PEST Notch pathway activation (TNBC cases only) c a b NOTCH1 NOTCH2 NOTCH3

2. Supplementary Figure 2 NOTCH4 a b

3. Supplementary Figure 3 TCGA-BH-A1FC, ∆exon21-27 Canonical NOTCH1 transcript 34 33 32 31292725232119 302826 242220 18 TCGA-A2-A0T0, chr9:139,390,528-139,390,695 deletion TCGA-A8-A08X, chr9:139,391,422 inter-chromosomal translocation Canonical NOTCH2 transcript TCGA-AO-A0J6, chr1:120,458,725-120,458,786 deletion a b 34 33 32 31292725232119 302826 242220 18 To: chr14:30,447,409 c TCGA-AN-A0AR, BRD4(exon1)-NOTCH3(exon 26) fusion & chr19:15,270,914-15,272,239 deletion 33 32 31 29 27 30 28 26 25 NOTCH3BRD4EPHX3

4. TCGA-BH-A1FC Tumor TCGA-BH-A1FC Matched Normal a bc d 9q34.3 27 234 28 1 NOTCH1 2120 234 28 1 20 Exon 21-27 deletion S1S2S3 Supplementary Figure 4 ef

5. 19p13.13 332625 21 3326 12 1 50Kb NOTCH3 BRD4 1226bp deletion ab c Supplementary Figure 5 S1S2S3 Notch3 wild-type Mutant Notch3 in TCGA-AN-A0AR Notch3 p.M1583Notch3 p.P2067Rfs*84 d

6. Pfizer Confidential │ 6 TCGA-A2-A0T0TCGA-A8-A08XTCGA-A8-A0J6 RNA-Seq in tumor WXS in tumor WXS in matched normal abc Supplementary Figure 6

7. a b Supplementary Figure 7

8. Chromosome Position (MB) log2 CN ratio c a b d Supplementary Figure 8 e f g TGTGAGCAGA CCCGCTGGTT h i Exon 34 Intron 30 Junction CCACTGGGCCCC CTGGGGTGATACCGCAGG j k l 1p121q21.1 chr1: 144548815 chr1: 120458385 NOTCH2 34 272 1 Intronic 34* 272 1 chr1: 144548815 127293133 1p121q21.1 2 28 30 3234 1272931332 28 30 32 34’ * 5’ - - 3’ NOTCH2Intronic 9q34.3 27 2 2 34 30 31 34 34*31 3031 1 1 NOTCH1 chr9:139390847 chr9:139396004 272931332 28 30 3234 9q34.3 NOTCH1 1 5’ - - 3’ 1272931332 28 30 32 30 31 34’ * chr9:139396004

9. Supplementary Figure 9 TCTCGAGATGGAGATTGGAGATTG wt deletion GCATTCCAACTTTAGCACAAGG CCAGCTACAGCTATGCTGA AGCGACGTGGGCTCTCCTGGGGC GGCAGCGACGTGGGCAGGGCGGGGCTCTCCTGGGGC c a b d e f g 9q34.3 27 234 28 1 NOTCH1 34* 28 1 27* 4q32.3 chr4:166105738 Intergenic chr9:139390607chr9:139397768 chr9:139439295 9q34.3 4q32.3 Intergenic 1272931332 28 30 3234 NOTCH1 1293133 28 30 32 5’ - - 3’ 34’ * chr4:166105738 Alt. splicing chr9:139390682 – chr4:166107169 27’ unsplicedspliced

10. Supplementary Figure 10 a b HCC1937 HBCx5 HBCx17 HBCx12B AA0869 MDA-MB-231 BT-474 HCC1806

11. Supplementary Figure 11 a b 80 KDa 110 KDa 160 KDa NICD1 MDAMB231 - -++ HBCx14 - -++ PF4014 HCC1599 - -++ GAPDH 40 KDa c HBCx14 HCC1599HBCx12B - -++ - -++ - + - + PF4014 110 KDa 160 KDa NICD1 GAPDH 40 KDa 110 KDa 160 KDa NICD1 HCC1937 HBCx14 HCC1599 - ++ -- ++ PF4014 GAPDH 40 KDa -

12. Notch wt Notch mutant Notch wt Notch mutant Notch wt Notch mutant Notch wt Notch mutant Supplementary Figure 12 PF-4014 * * * * * ** * *

13. Notch wt Notch mutant Notch wt Notch mutant Notch wt Notch mutant Notch wt Notch mutant Supplementary Figure 13 PF-4014 * * * * ** * * * * * * * *