Table 3.

Selected studies comparing HNSCC mutation profiles between tumour DNA (tDNA) and ctDNA

Author and publish year	Patient cohort and sample types	Methods	Results and conclusions
Perdomo et al. 2017 [115]	-Targeted mutation gene panel: fresh tumour tissue and plasma ctDNA from 36 HPV-negative HNSCC patients -Non-targeted approach using TP53 gene mutation: fresh tumour tissue, plasma ctDNA and oral rinse samples from 37 HNSCC patients	-Targeted approach: using a European cohort of patients, a 5-gene mutation panel (TP53, PTEN, NOTCH1, CASP8 and CDKN2A) -Non-targeted approach: the whole coding region of TP53 gene using a South American cohort of patients	-Targeted approach: 42% (15/36) of cases had detectable plasma ctDNA mutations, 67% were from early stages (I, II) cases. 18 mutations were detected in both plasma ctDNA the matched tDNA with allelic fractions (AF) ranging from 0.001 to 0.12 -Nontargeted: One case showed (TP53 Arg174Trp) exclusively in plasma ctDNA Four cases showed concordance in TP53 mutation between tumour tissue and oral rinse samples One case showed concordance in TP53 mutation among tumour tissue, plasma, and oral rinse
Porter et al. 2019 [118]	Tumour specimen (N = 30) and plasma ctDNA (N = 60) from patients with recurrent and metastatic HNC	-Tumour DNA: Foundation One and Caris -ctDNA: Guardant Health Inc	-TP53 (48% vs 68%) and PIK3CA (24% vs 34%) were two most common mutations in tDNA and ctDNA -Patients who had ctDNA and tDNA sequencing (N = 30), 20/30 had alterations in tDNA and ctDNA ctDNA identified a new mutation at 73% (22/30)
Mes et al. 2020 [99]	Frozen tumour tissue and matching plasma ctDNA from 27 HNSCC patients	-Illumina HiSeq -Illumina MiSeq	-Concordance rate of copy number variation (CNV) between tDNA and ctDNA was 52% (14/27) -Some mutations were only identified in ctDNA but not in matching tDNA. (CASP8, NSD1, KMT2D, CDKN2A, NOTCH1, TP53)
Wilson et al. 2020 [165]	Tumour tissue samples and plasma ctDNA from 75 HNSCC patients	-Foundation One Platform (323 genes) tDNA -Guardan360 platform in ctDNA	-13% concordance between tDNA and plasma ctDNA -TP53 was the most concordant gene -BRCA1, EGFR, KIT, BRAF, ESR1, FGFR2, FGFR3, MAP2K1 and NRAS shared similar alteration in both tDNA and ctDNA -ARID1A, ATM and MET showed more alteration in ctDNA than in tDNA -65.3% of patients had detectable actionable alteration in ctDNA
Galot et al. 2020 [43]	Plasma from 39 recurrent/metastatic HNSCC patients, and matched tumour FFPE from 18 HNSCC patients	-A 604 gene mutation panel -ddPCR	- The most frequently mutated gene in ctDNA was TP53, followed by genes in the PI3K pathway -Compared to locoregional recurrent disease, a higher probability of detecting ctDNA was found in HNSCC patients in metastatic disease (70% versus 30%) -81% of mutations identified in solid tumours were not detected in ctDNA -26% of ctDNA variants were not detected in matched tDNA - The cancer driver events identified in both tDNA and matching ctDNA were TP53, MYC, EGFR, CDKN2A and PIK3CA
Khandelwal et al. 2020 [69]	Frozen tumour tissue and plasma samples from 22 OPSCC patients	MiSeq platform	- 12/22 mutations were identified in tDNA· - 11/22 mutations were identified in plasma cfDNA· - HPV-negative OPSCC non-responder patients were more likely to have variant detected in ctDNA -Five patients carried six matching mutations (TP53 G325fs, TP53 R282W, TP53 R273C, FBXW7 R505G, FBXW7 R505L, TP53 Q331H) in both tDNA and ctDNA - A high concordance was found between TP53 mutation in ctDNA and tumour tissue in HPV-negative OPSCC
Burgener et al. 2021 [13]	Plasma ctDNA from 30 HPV-negative HNSCC patients; and TCGA data- derived tumour mutational information	CAPP-seq for ctDNA mutation analysis	-Common cancer driver mutations in TP53, PIK3CA, FAT1 and NOTCH1 were found between tDNA and ctDNA - Mutations in two genes (GRIN3A and MYC) were only specific to ctDNA
Liebs et al. 2021 [83]	HNSCC patients (N = 6) tumour tissue FFPE and matching plasma ctDNA samples	A targeted 327 cancer gene panel; The HaloPlex™ HS target enrichment system	-A relatively low (11%) mutation concordance rate was found between tDNA and ctDNA -With high input of ctDNA(> 30 ng), five genes (ACACA, ATR, LAMA2, PIK3CA and SMARCA4) were mutated both in tDNA and ctDNA - With low input DNA (< 30 ng), 52 mutations in 3 genes (FAT1, RELN and PDGFRA) were found in tDNA -With 30 ng input of tDNA and ctDNA, a total of nine mutations in four genes were found in cfDNA, (EZH2, LAMA2, PIK3CA and SMARCA4) -Analyzed ctDNA followed by their correspondent tDNA with low input (< 30 ng), 2/6 mutations (EPHA2 and FLT3) existed in both types of DNAs
Kogo et al. 2022 [72]	Frozen tumour tissue and plasma from 26 HNSCC patients	-A targeted gene panel comprising 31 genes for tDNA; -ddPCR for ctDNA	The most frequently mutated gene is TP53 (58.2%). Longitudinal positivity of ctDNA revealed prognosis
Flach et al. 2022 [37]	Tumour FFPE and plasma from HNSCC patients (N = 8)	Oncomine. Comprehensive Assay	TP53 was the most frequently mutated gene. 37.5% variants were shared between tDNA and ctDNA