SUPPLEMENTAL MATERIAL

Supplemental Table 1. Guardant360 54 gene cell-free DNA NGS panel. Complete exon coverage for genes in bold. Unbolded genes had critical exon coverage.

Point mutations and amplifications in EGFR, ERBB2, and MET:

ABL1 / AKT1 / ALK / APC / AR / ATM
BRAF / CDH1 / CDKN2A / CSF1R / CTBBB1 / EGFR*
ERBB2* / ERBB4 / EZH2 / FBXW7 / FGFR1 / FGFR2
FGFR3 / FLT3 / GNA11 / GNAQ / GNAS / HNF1A
HRAS / IDH1 / IDH2 / JAK2 / JAK3 / KDR
KIT / KRAS / MET* / MLH1 / MPL / MYC
NOTCH1 / NPM1 / NRAS / PDGFRA / PIK3CA / PTPN11
PTEN / PROC / RB1 / RET / SMAD4 / SMARCB1
SMO / SRC / STK11 / TERT / TP53 / VHL

*Copy number amplification of EGFR, ERBB2 (HER2) and MET genes was done (in addition of mutations (complete exon coverage)).

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Supplemental Table 2. Frequency of patients with metastatic/recurrent/advanced unresectable disease

Number (%) of patients with metastatic/recurrent/advanced unresectable disease at the time of ctDNA testing
Overall (N = 168) / 143 (85.1%)
Tumor type:
Brain (N = 56) / 38 (67.9%)
Lung (N = 47) / 44 (93.6%)
Breast (N = 36) / 34 (94.4%)
Gastrointestinal (N = 13) / 13 (100%)
Genitorurinary (N = 8) / 8 (100%)
Gynecologic (N = 2) / 2 (100%)
Other (N = 6) / 4 (66.7%)
% of ctDNA:
At least one alteration with ctDNA percentage ≥ 5% (N=27) / 22 (81.5%) P=0.029 (Fisher’s exact test)
All alterations with ctDNA percentage < 5% (N=141) / 82 (58.2%)
Matching status:
Patients received matched therapy* (N=15) (12 evaluable) / 13 (86.7%) P=0.182 (Fisher’s exact test)
Patients received an unmatched therapy (N=39) (28 evaluable) / 25 (64.1%)

*Informed by the ctDNA test; some patients were inevaluable because they were not evaluated for response.

Supplemental Table 3. List of the 12 evaluable patients treated with a matched therapy targeting ≥ 1 alteration detected by the ctDNA test.

# / Tumor type / Relevant targeted alterations(s) in ctDNA / Targeted alteration(s) also identified in tissue / Matched drug(s) / Outcome / PFSa / Line of therapy
1 / Lung / EGFR T790M (also EGFR amplification) / YES / AZD9291 trial
(Irreversible EGFR inhibitor) / PR / 8.4 / 3
2 / Lung / EGFR mutations (E746K and M881V) and amplification / YES / erlotinib / PR / 7.6+ / 1
3 / Breast (ER+) / PTEN and PIK3CA mutations / YES / exemestane/everolimus / PR / 3.3+ / 2
4 / Lung / EGFR T790M (also EGFR L861Q and EGFR G719S detected) / YES / AZD9291 trial
(Irreversible EGFR inhibitor) / SD ≥ 6 months / 8.6+ / 2
5 / Lung / EGFR T790M (also EGFR A750E) / YES / AZD9291 trial
(Irreversible EGFR inhibitor) / SD ≥ 6 months / 7.0 / 2
6 / Brain / TP53* mutation / YES / bevacizumab* / SD / 3.6 / 4
7 / Breast (ER+) / PIK3CA mutation / NO / exemestane/everolimus / PD / 1.0 / 3
8 / Breast (ER+) / PIK3CA mutation / No other testing performed other than ctDNA / exemestane/everolimus / PD / 2.0 / 3
9 / Gastrointestinal / ERBB2 amplification / YES / pertuzumab + trastuzumab / PD / 1.3 / 5
10 / Gastrointestinal / TP53* mutation / No other testing performed other than ctDNA / bevacizumab* / PD / 0.8 / 1
11 / Genitourinary / MET amplification / No other testing performed other than ctDNA / cabozantinib / PD / 2.1 / 5
12 / Lung / EGFR L858R, ERBB2 mutation / YES / afatinib / PD / 0.3 / 2

aPFS = Progression-free survival: a “+” sign indicated that the therapy was still ongoing; PD = Progressive disease; PR = Partial response; SD = Stable disease.

*TP53 alterations were considered relevant for antiangiogenic therapy (such as bevacizumab)(42,43).

Supplemental Figure 1. Diagram of the population analyzed for treatment outcome

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