Supplemental Information. Therapy-Related Clonal Hematopoiesis in Patients. with Non-hematologic Cancers Is Common

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1 Cell Stem Cell, Volume 21 Supplemental Information Therapy-Related Clonal Hematopoiesis in Patients with Non-hematologic Cancers Is Common and Associated with Adverse Clinical Outcomes Catherine C. Coombs, Ahmet Zehir, Sean M. Devlin, Ashwin Kishtagari, Aijazuddin Syed, Philip Jonsson, David M. Hyman, David B. Solit, Mark E. Robson, José Baselga, Maria E. Arcila, Marc Ladanyi, Martin S. Tallman, Ross L. Levine, and Michael F. Berger

2 Methods S1. Categorization of non-hematologic cancers The clinical cohort included 51 distinct tumor types, which are listed below, followed by number of patients sequenced (A). Given the large number of distinct tumor types, an additional grouping was established to collapse the 51 tumor types into seven broader tumor categories for purposes of analyzing the proportion of patients with CH and CH- PD by tumor categorization (B) as described in Table 1. A. Tumor types 1) Adrenocortical carcinomas: 13 2) Ampullary carcinomas: 14 3) Anal carcinoma: 21 4) Appendiceal carcinoma: 39 5) Biliary cancer (includes intra- and extrahepatic cholangiocarcinomas and gallbladder carcinomas): 137 6) Bladder carcinoma (includes urothelial, squamous, adenocarcinoma, small cell, plasmacytoid, and sarcomatoid histologies, also includes upper tract urothelial carcinomas): 268 7) Breast carcinoma (includes invasive ductal, invasive lobular, mixed invasive ductal/lobular, metaplastic, and adenoid cystic breast carcinomas): 722 8) Breast sarcoma (includes angiosarcoma of the breast and malignant phyllodes tumor of the breast): 5 9) Cancer of unknown primary (includes adenocarcinoma of unknown primary, neuroendocrine of unknown primary, squamous cell of unknown primary): 91 10) Cervical carcinoma (includes cervical adenocarcinoma, cervical adenosquamous carcinoma, cervical squamous carcinoma, etc.): 26 11) Chondrosarcomas: 22 12) Chordomas: 7

3 13) Colorectal carcinomas (includes colon and rectal carcinomas): ) Embryonal tumors (includes neuroblastomas and olfactory neuroblastomas): 50 15) Endometrial carcinomas (includes uterine endometrioid carcinomas, uterine serous carcinomas, uterine carcinosarcoma/uterine malignant mixed mullerian tumors, etc.): ) Ependymoma: 8 17) Esophagogastric carcinoma (includes carcinomas arising in the esophagus, stomach, and gastroesophageal junction): ) Ewing sarcomas: 20 19) Gastrointestinal neuroendocrine tumors: 33 20) Gastrointestinal stromal tumors: 74 21) Germ cell tumors: ) Gestational trophoblastic disease: 8 23) Gliomas (includes astrocytoma, oligodendroglioma, anaplastic astrocytoma, anaplastic oligodendroglioma, glioblastoma multiforme, etc.): ) Head and neck carcinomas (includes oral cavity squamous cell carcinomas, oropharynx squamous cell carcinomas, sinonasal squamous cell carcinomas, nasopharyngeal carcinomas, larynx squamous cell carcinomas, etc.): 70 25) Hepatocellular carcinomas: includes hepatocellular carcinomas and fibrolamellar carcinomas: 61 26) Melanomas (includes acral melanomas, cutaneous melanomas, mucosal melanomas, uveal melanomas, etc.): ) Meningothelial tumors (includes anaplastic and atypical meningiomas): 18 28) Mesotheliomas (includes pleural and peritoneal mesotheliomas): 57

4 29) Miscellaneous cancers, not meeting any of the other categories: 6 30) Nerve sheath tumors: 12 31) Non-small cell lung cancers (primarily adenocarcinoma and squamous cell carcinomas of the lung but also includes large cell neuroendocrine, poorly differentiated NSCLC, sarcomatoid carcinomas of the lung, etc.): ) Osteosarcomas: 16 33) Ovarian carcinomas: ) Pancreatic carcinomas (primarily pancreatic adenocarcinoma but also includes pancreatic neuroendocrine tumors and acinar cell carcinomas): ) Penile carcinomas: 6 36) Pheochromocytoma: 2 37) Pineal tumor: 1 38) Prostate carcinomas (primarily prostate adenocarcinoma but also includes prostate neuroendocrine carcinomas and prostate small cell carcinomas): ) Renal cell carcinomas: ) Retinoblastoma: 1 41) Salivary carcinomas: 64 42) Sellar tumors (includes granular cell tumor and pituicytoma): 2 43) Sex cord stromal tumors (includes granulosa cell tumor and Sertoli-Leydig cell tumor): 13 44) Skin-cancers, non-melanoma (includes basal cell carcinomas, cutaneous squamous cell carcinomas, Merkel cell carcinomas): 73 45) Small bowel carcinoma: 22 46) Small cell lung carcinoma: 37

5 47) Soft tissue sarcoma (many types, most common types include undifferentiated pleomorphic sarcomas, dedifferentiated liposarcomas, angiosarcomas, leiomyosarcomas, solitary fibrous tumors, and synovial sarcomas): ) Thymic tumor (includes thymic carcinomas and thymomas): 8 49) Thyroid carcinoma: ) Uterine sarcomas: 66 51) Vaginal/vulvar carcinomas: 3 B. Tumor categories: 1) Lung: includes Non-small cell lung cancers, small cell lung cancers, and mesotheliomas 2) Breast: Breast cancers only 3) Gastrointestinal (GI): includes colon cancer, rectal cancer, pancreatic adenocarcinoma, neuroendocrine cancers arising in GI tract, anal cancers, ampullary cancers, gallbladder cancers, cholangiocarcinomas, hepatocellular cancers, fibrolamellar hepatocellular cancers, esophageal cancers, gastric cancers, gastroesophageal junction cancers, appendical cancers, small bowel cancers 4) Sarcomas: includes soft tissue sarcomas, osteosarcomas, gastrointestinal stromal tumors, breast sarcomas, chondrosarcomas, Ewing sarcomas, 5) Genitourinary (GU): includes germ cell tumors, renal cell carcinomas, prostate cancers, penile cancers, urothelial cancers (arising in bladder and upper tract) 6) Gynecological (Gyn): includes cervical cancers, endometrial cancers, ovarian cancers, vaginal and vulvar cancers, gestational trophoblastic disease

6 7) Other: includes all other cancers types with most prominent subtypes including cancers of unknown primary site, cancers arising from the skin (melanomas, basal cell carcinomas, cutaneous squamous cell carcinomas, Merkel cell carcinomas), thyroid cancers, central nervous system cancers, head and neck cancers

7 Supplemental data Table S1. Related to Table 1. Patients who were excluded from analyses due to presence of an active hematologic malignancy (or known precursor condition) at time of matched normal blood sampling. VAF= variant allele fraction. H&N= Head and Neck squamous cell carcinoma. NSCLC= non-small cell lung carcinoma. SCC= squamous cell carcinoma. CLL= chronic lymphocytic leukemia. CML= chronic myeloid leukemia. MGUS= monoclonal gammopathy of undetermined significance. WM= Waldenstrom s macroglobulinemia. MDS= myelodysplastic syndrome. ET= essential thrombocythemia. MBL= monoclonal B-cell lymphocytosis. MZL= marginal zone lymphoma. ENMZL= extranodal MZL. DLBCL= diffuse large B-cell lymphoma. PV= polycythemia vera. AML= acute myeloid leukemia. PTLD= post-transplant lymphoproliferative disorder. CMML= Chronic myelomonocytic leukemia. FL= follicular lymphoma. ^This patient carried a diagnosis of low grade lymphoma in the setting of an IgM monoclonal gammopathy so suspect the diagnosis was WM. Median VAF in the blood for these patients was 0.10 and median VAF in the tumor was Pt no. Age Sex Primary cancer diagnosis 1 68 M Prostate CLL 2 67 M Prostate CLL Hematologic diagnosis Mutation on VAF VAF Died? NGS blood tumor TP53 p.c135y N TP53 p.s241c TP53 p.v173l N TP53 p.v272m M SCC of the skin CML N N SCC, unknown 4 73 M primary CLL N Y DNMT3A 5 79 F NSCLC MGUS p.v386fs N CBL p.c404s F NSCLC Aplastic anemia N N Composite 7 75 M NSCLC lymphoma (FL and MZL) PGR p.t498p N NOTCH2 p.p2280fs Low grade 8 81 F Urothelial lymphoma N N 9 75 M Colon CLL TP53 p.w53* N M Glioblastoma TET2 Multiple myeloma p.h1904r N M Melanoma TET2 CLL p.q744* N DNMT3A p.r379h M Esophageal CLL N Y M Prostate Multiple myeloma N N M Glioblastoma MDS N Y JAK F Oligodendroglioma ET p.v617f N M H&N CLL EED p.h235y N M NSCLC FL N Y F Thyroid CLL N N M Urothelial MBL N N F Breast FL N Y F NSCLC MGUS N N

8 22 75 M Pancreas TET2 CLL p.t1280fs N F Endometrial DNMT3A splicing Marginal zone (c lymphoma 2A>T) N F Glioblastoma SF3B1 MDS p.k700e Y M Colon "Low grade lymphoma ^ N N M NSCLC SF3B1 MDS p.k700e Y TET2 p.h1382r F Sarcoma ET N Y F Breast FL N N F SCC of the skin Gastric ENMZL N N M Urothelial MGUS N Y M Prostate ASXL1 DLBCL p.a861fs N SF3B1 p.k666n ASXL1 p.p763fs JAK2 p.v617f M H&N ET N N M Glioblastoma JAK2 PV p.v617f Y TET2 p.g1288s TET2 p.r1241s CBL p.k382n F Colon CLL N Y Merkel cell M carcinoma CLL N N M Thyroid CLL N N Adrenal cortical DNMT3A M carcinoma CLL p.y735c Y ARID1B p.q127l MCL1 p.l97p MCL1 p.p107a MCL1 p.p142a M Melanoma FL N N M SCC of the skin CLL N Y JAK F Breast PV p.v617f Y Merkel cell M carcinoma CLL N Y JAK M Prostate ET p.v617f N M Melanoma MDS N N Adenocarcinoma M unknown primary MGUS N N M Pancreas FL N N M NSCLC ET N N M Prostate MGUS N N

9 48 70 F Thyroid ET Merkel cell F carcinoma ET U2AF1 p.q157p Y JAK2 p.v617f Y ASXL1 p.p874fs M Pancreas MGUS N Y F NSCLC FL N N JAK F Urothelial PV p.v617f N FOXO1 p.l434w PTPN M H&N CLL p.e69k N F SCC of the skin AML N Y F NSCLC PTLD N N M Germ cell tumor CMML N Y Low grade B-cell M Pancreas lymphoma (FL vs CLL) N N F JAK2 Cholangiocarcinoma ET p.v617f N F H&N MDS N Y F Pancreas MGUS N N M Prostate TET2 MGUS p.q1414fs N TET2 p.p929fs ZRSR2 splicing (c.312+1g>a) SETD2 p.l1475i M Sarcoma JAK2 ET p.v617f N MST1R splicing (c a>g) F Melanoma CLL N N DNMT3A M Melanoma CLL p.w860r N RASA1 p.r391q ATM p.v2424e

10 Table S2. Related to Table 1. Characteristics of CH variants identified across 5,649 patients for recurrently mutated genes. P-values for comparisons are for mutants in any given gene for non-mutants for same gene. Overall ^ DNMT3A P-value TET2 P-value PPM1D ^ P-value ASXL1 P-value TP53 P-value Number of 5, patients Age < < < < <0.001 Gender 49% 45% % % % % 0.12 (%Male) WBC ANC ALC AMC NLR >4 38% 36% % % % % Hemoglobin MCV < <0.001 RDW 14.7% 14.7% % % < % % 0.38 Platelet count Prior chemotherapy 64% 61% % % < % % Prior RT 37% 41% % % < % % <0.001 Current/ 46% 52% % % % < % former tobacco use RBC 22% 21% % % % % 0.29 transfusion Platelet 6% 4% % % % % 0.99 transfusion Growth factor 28% 24% % % < % % 0.45 ^Comparisons for PPM1D were performed only in patients who underwent sequencing of this gene (N=3,737). Note that the Overall column is synonymous with respect to the entire 5,649 patient cohort with the following exceptions pertaining to the 3,737 cohort that were used for PPM1D comparisons: Gender is 51% Male, WBC is 7.2, ALC is 1.39, AMC is 0.47, NLR>4 is 37%, Hemoglobin is 12.5, RDW is 14.5%, Platelet count is 252, Prior chemotherapy is 61%, Prior RT is 34%, RBC transfusion is 19%, Platelet transfusion is 5%, and Growth factor is 26%. No adjustments were made for multiple comparisons.

11 Table S3. Related to Figure 3. Details on patients who developed a new hematologic cancer following collection of matched normal blood sample. a Patient did not undergo molecular profiling for newly diagnosed hematologic cancer. b Patient underwent testing with a limited molecular panel which was negative though panel did not include testing for genes previously identified as CH c Patient underwent repeat testing using identical panel which confirmed absence of TET2 mutation in newly diagnosed hematologic cancer. Pt no. Age Sex Initial cancer diagnosis Prior chemo Prior RT Hematologic cancer diagnosis Latency (days) Mutation(s) on NGS 1 54 M Urothelial Y Y ET 493 U2AF1 p.s34y JAK2 p.v617f PTPRS p.t1087m 2 56 F Melanoma N N MDS-RCMD 286 DNMT3A p.f640fs 3 78 M Lung-NSCLC Y Y DLBCL 2 TET2 p.q1545* 4 66 M Lung-NSCLC Y Y MDS 123 SPOP p.k28* PPM1D p.l450v TP53 p.h193r U2AF1 p.q157p TP53 p.g266r PPM1D p.l450* PPM1D E527fs 5 69 M Mesothelioma Y Y MDS 259 TP53 p.v216m DNMT3A p.l347fs 6 81 M Prostate N N MDS-RCMD 283 U2AF1 p.s34f ARID1A p.p223a ARID1A p.y222_p22 3insV U2AF1 p.q157r KDM6A p.223a 7 67 F Lung-NSCLC Y N PV 98 JAK2 p.v617f TET2 p.w1847* 8 20 M Nonseminoma Y N MDS/MPN, histiocytic sarcoma, evolved to AML 246 TET2 p.r1354fs 9 30 F Sarcoma Y Y MDS 16 DNMT3A p.r882h VAF blood VAF tumor Hematologic cancer clonally related to prior CH? Yes Died/died from hematologic cancer? No Yes No Unknown a No Yes Yes Unknown b Yes No Yes/Yes No No No c Yes/Yes Yes Yes/No died from respiratory failure related to lymphangitic spread of sarcoma Yes No M Lung- Carcinoid N N ET 119 CALR p.e364fs F Endometrial N N MDS-RARS 467 None Yes/No Patient died from complications of progressive endometrial cancer F Ovarian Y N CLL/SLL 108 None No M Colon Y N AML 152 None Yes/Yes M Urothelial Y Y CLL/SLL 253 None Yes/No Patient died from progressive urothelial cancer M Sarcoma N N Marginal zone lymphoma 43 None Yes/No Patient died from progressive sarcoma

12 16 10 F Glioblastoma Y Y AML 190 None Yes /Yes Patient s immediate cause of death was related to AML though admitting GBM also complicating factor M Renal Y N Follicular lymphoma 391 None No F Lung- Carcinoid F Lung-SCLC Y Y Acute leukemia- NOS Y N AML 8 None Yes/Yes from recurrent AML with inversion None Yes /Yes placed on palliative care due to poor prognosis from lung cancer with new pancytopenia and >20% circulating blasts suggestive of new acute leukemia

13 Table S4. Related to Figure 1. A. List of genes sequenced on MSK-IMPACT assay. The first 1,912 patients in the cohort underwent sequencing with a 341-gene assay and the remaining patients underwent sequencing with a 410-gene assay. A subset of genes, including PPM1D, SRSF2, SH2B3, CALR, and ZRSR2, among others (additional genes listed as Version 2) were only included in the 410 gene version of the panel, which became our standard profiling assay after the first 1,912 patients were sequenced such that associations related to these genes described in Figure 1 and Table S2 only reflects data from the last 6,898 and 3,737 patients, respectively. Gene Version Gene Version Gene Version Gene Version Gene Version ABL1 1 CXCR4 2 HIST1H3D 2 MYCL1 1 RECQL4 1 ACVR1 2 DAXX 1 HIST1H3E 2 MYCN 1 REL 1 AKT1 1 DCUN1D1 1 HIST1H3F 2 MYD88 1 RET 1 AKT2 1 DDR2 1 HIST1H3G 2 MYOD1 1 RFWD2 1 AKT3 1 DICER1 1 HIST1H3H 2 NBN 1 RHEB 2 ALK 1 DIS3 1 HIST1H3I 2 NCOA3 2 RHOA 1 ALOX12B 1 DNAJB1 2 HIST1H3J 2 NCOR1 1 RICTOR 1 ANKRD11 2 DNMT1 1 HIST2H3C 2 NEGR1 2 RIT1 1 APC 1 DNMT3A 1 HIST2H3D 2 NF1 1 RNF43 1 AR 1 DNMT3B 1 HIST3H3 2 NF2 1 ROS1 1 ARAF 1 DOT1L 1 HLA-A 2 NFE2L2 1 RPS6KA4 1 ARID1A 1 E2F3 1 HNF1A 1 NFKBIA 2 RPS6KB2 1 ARID1B 1 EED 1 HOXB13 2 NKX2-1 1 RPTOR 1 ARID2 1 EGFL7 1 HRAS 1 NKX3-1 1 RUNX1 1 ARID5B 1 EGFR 1 ICOSLG 1 NOTCH1 1 RYBP 1 ASXL1 1 EIF1AX 1 ID3 2 NOTCH2 1 SDHA 1 ASXL2 1 EIF4A2 2 IDH1 1 NOTCH3 1 SDHAF2 1 ATM 1 EIF4E 2 IDH2 1 NOTCH4 1 SDHB 1 ATR 1 EP300 1 IFNGR1 1 NPM1 1 SDHC 1 ATRX 1 EPCAM 1 IGF1 1 NRAS 1 SDHD 1 AURKA 1 EPHA3 1 IGF1R 1 NSD1 1 SETD2 1 AURKB 1 EPHA5 1 IGF2 1 NTRK1 1 SF3B1 1 AXIN1 1 EPHA7 2 IKBKE 1 NTRK2 1 SH2B3 2 AXIN2 1 EPHB1 1 IKZF1 1 NTRK3 1 SH2D1A 1 AXL 1 ERBB2 1 IL10 1 NUP93 2 SHQ1 1 B2M 1 ERBB3 1 IL7R 1 PAK1 1 SMAD2 1 BAP1 1 ERBB4 1 INHA 2 PAK7 1 SMAD3 1 BARD1 1 ERCC2 1 INHBA 2 PALB2 1 SMAD4 1 BBC3 1 ERCC3 1 INPP4A 1 PARK2 1 SMARCA 4 1 BCL10 2 ERCC4 1 INPP4B 1 PARP1 1 SMARCB 1 1 BCL2 1 ERCC5 1 INSR 1 PAX5 1 SMARCD 1 1 BCL2L1 1 ERG 1 IRF4 1 PBRM1 1 SMO 1

14 BCL2L11 1 ERRFI1 2 IRS1 1 PDCD1 1 SOCS1 1 BCL6 1 ESR1 1 IRS2 1 PDGFRA 1 SOX17 1 BCOR 1 ETV1 1 JAK1 1 PDGFRB 1 SOX2 1 BIRC3 2 ETV6 1 JAK2 1 PDPK1 1 SOX9 1 BLM 1 EZH2 1 JAK3 1 PGR 2 SPEN 1 BMPR1A 1 AMER1 1 JUN 1 PHOX2B 1 SPOP 1 BRAF 1 FAM175A 1 KDM5A 1 PIK3C2G 1 SRC 1 BRCA1 1 FAM46C 1 KDM5C 1 PIK3C3 1 SRSF2 2 BRCA2 1 FANCA 1 KDM6A 1 PIK3CA 1 STAG2 1 BRD4 1 FANCC 1 KDR 1 PIK3CB 1 STAT3 2 BRIP1 1 FAT1 1 KEAP1 1 PIK3CD 1 STAT5A 2 BTK 1 FBXW7 1 KIT 1 PIK3CG 1 STAT5B 2 CALR 2 FGF19 1 KLF4 1 PIK3R1 1 STK11 1 CARD11 1 FGF3 1 KRAS 1 PIK3R2 1 STK40 1 CASP8 1 FGF4 1 LATS1 1 PIK3R3 1 SUFU 1 CBFB 1 FGFR1 1 LATS2 1 PIM1 1 SUZ12 1 CBL 1 FGFR2 1 LMO1 1 PLCG2 2 SYK 1 CCND1 1 FGFR3 1 MALT1 2 PLK2 1 TBX3 1 CCND2 1 FGFR4 1 MAP2K1 1 PMAIP1 1 TCEB1 2 CCND3 1 FH 1 MAP2K2 1 PMS1 1 TCF3 2 CCNE1 1 FLCN 1 MAP2K4 1 PMS2 1 TCF7L2 2 CD274 1 FLT1 1 MAP3K1 1 PNRC1 1 TERT 1 CD276 1 FLT3 1 MAP3K13 1 POLD1 2 TET1 1 CD79A 2 FLT4 1 MAP3K14 2 POLE 1 TET2 1 CD79B 1 FOXA1 1 MAPK1 1 PPM1D 2 TGFBR1 1 CDC73 1 FOXL2 1 MAPK3 2 PPP2R1 A 1 TGFBR2 1 CDH1 1 FOXO1 2 MAX 1 PPP6C 2 TMEM127 1 CDK12 1 FOXP1 1 MCL1 1 PRDM1 1 TMPRSS2 1 CDK4 1 FUBP1 1 MDC1 1 PRKAR1 A 1 TNFAIP3 1 CDK6 1 FYN 2 MDM2 1 PTCH1 1 TNFRSF1 4 1 CDK8 1 GATA1 1 MDM4 1 PTEN 1 TOP1 1 CDKN1A 1 GATA2 1 MED12 1 PTPN11 1 TP53 1 CDKN1B 1 GATA3 1 MEF2B 1 PTPRD 1 TP63 1 CDKN2A 1 GLI1 2 MEN1 1 PTPRS 1 TRAF2 2 CDKN2B 1 GNA11 1 MET 1 PTPRT 1 TRAF7 1 CDKN2C 1 GNAQ 1 MGA 2 RAB35 2 TSC1 1 CEBPA 2 GNAS 1 MITF 1 RAC1 1 TSC2 1 CENPA 2 GPS2 2 MLH1 1 RAD21 2 TSHR 1 CHEK1 1 GREM1 1 MLL 1 RAD50 1 U2AF1 1 CHEK2 1 GRIN2A 1 MLL2 1 RAD51 1 VEGFA 2 CIC 1 GSK3B 1 MLL3 1 RAD51B 1 VHL 1 CREBBP 1 H3F3A 2 MPL 1 RAD51C 1 VTCN1 1

15 CRKL 1 H3F3B 2 MRE11A 1 RAD51D 1 WT1 1 CRLF2 1 H3F3C 1 MSH2 1 RAD52 1 XIAP 1 CSF1R 1 HGF 1 MSH6 1 RAD54L 1 XPO1 1 CSF3R 2 HIST1H1 C 1 MST1 2 RAF1 1 XRCC2 2 CTCF 1 HIST1H2B D 1 MST1R 2 RARA 1 YAP1 1 CTLA4 1 HIST1H3A 2 MTOR 1 RASA1 1 YES1 1 CTNNB1 1 HIST1H3B 1 MUTYH 1 RB1 1 ZFHX3 2 CUL3 1 HIST1H3 C 2 MYC 1 RBM10 1 ZRSR2 2 B. Ten most recurrently mutated genes and the minimum, mean, median, and maximum coverage for the observed mutations. A recurrent mutation in ASXL1 at p.g646fsx12 which has previously been suggested to be a potential PCR artifact was observed 13 times within our cohort with sequencing depth of 232X at this position. Coverage (X) Gene Min Mean Median Max ASXL ATM CBL CHEK DNMT3A JAK PPM1D SF3B TET TP

16 Figure S1. Related to Figure 1. Top: Distribution of DNMT3A mutations observed within cohort with burgundy dots representing hotspot variants (previously reported in COSMIC database) and orange dots representing novel variants (not reported in COSMIC database). Bottom: Distribution of VAF for hotspot and novel DNMT3A CH variants, demonstrating that hotspot variants present with a higher VAF than novel variants (median VAF versus 0.039, p=<0.001).

17 Figure S2. Related to Figure 1. For all plots, Y-axis is mutation counts and X-axis depicts position of variant with relevant domains noted in inset for each gene. A. Distribution of DNMT3A mutations within our cohort (top, blue dots) and in previously published AML cohorts (bottom, yellow dots) 18. B. Distribution of TET2 mutations within our cohort (top) and in previously published AML cohorts (bottom) 18. Red dots represent protein-truncating variants and green dots represent missense variants. C. Distribution of TP53 mutations within our cohort (top, blue dots) and in previously published AML cohorts (bottom, yellow dots). 18. D. Distribution of JAK2 mutations within our cohort (top, blue dots) and in previously published AML cohorts (bottom, yellow dots) 18. E. Distribution of ASXL1 mutations within our cohort (top, blue dots) and in previously published AML cohorts (bottom, yellow dots) 18.

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19 Figure S3. A. Related to Figure 1. Distribution and variant class for mutations classified as clonal hematopoiesis in presumptive drivers (CH-PD). Note that this is in distinction to clonal hematopoiesis (CH) distribution and variant class, which is depicted in Figure 1. B. Related to Figure 1. Number of CH-PD variants seen per patient. C. Related to Figure 2. Variant class for patients with CH who were naïve to both chemotherapy and RT versus all others (prior exposure to chemotherapy, RT or both).

20 Figure S4. Related to Figure 3. Clonal hematopoiesis in presumptive drivers (CH-PD) with minimum VAF threshold of 0.2 and associations with subsequent hematologic cancers and overall survival. A. Incidence of new hematologic cancer for patients with and without CH-PD (VAF>0.2). B. OS in non-ch-pd vs. CH-PD (VAF>0.2). Differences in the incidence of new hematologic cancer were compared using Gray s test, while survival differences were assessed using the Peto & Peto modification of the Gehan- Wilcoxon test. Survival was compared both overall and stratified based on categories of age, gender, and smoking status.