A reliable method for the detection of BRCA1 and BRCA2 mutations in fixed tumour tissue utilising multiplex PCR-based targeted next generation sequencing

Table 2 High impact variants

Sample	Sample type	High impact variant	Class	% variant reads	NGS replicate confirmation	Sanger confirmation
AZ68⁺	Ovary	BRCA1 c.1105delG p.(Asp369MetfsTer5)	pathogenic	72	Yes	Yes
AZ75⁺	Ovary	BRCA1 c.1105delG p.(Asp369MetfsTer5)	pathogenic	68	Yes	Yes
AZ30	Ovary	BRCA1 c.4675G>A p.(Glu1559Lys)	pathogenic	58	Yes	Yes
AZ28	Ovary	BRCA1 c.5266dupC p.(Gln1756ProfsTer74)	pathogenic	52	Yes	Yes
AZ23	Ovary	BRCA2 c.7007+1G>C	pathogenic	79	Yes	Yes
AZ11	Ovary	BRCA1 c.181T >G p.(Cys61Gly)	pathogenic	84	Yes	No data*
AZ113	Breast	BRCA1 c.2253_2254delGT p.(Met751IlefsTer10)	pathogenic	52	Yes	Yes
AZ109	Breast	BRCA1 c.5095C>T p.(Arg1699Trp)	pathogenic	59	Yes	Yes
AZ17	Ovary	BRCA1 c.2060A>C p.(Gln687Pro)	VUS	90	Yes	Yes
AZ78	Ovary	BRCA2 c.1408G>C p.(Glu470Gln)	VUS	76	Yes	Yes
AZ72	Ovary	BRCA2 c.10024G>A p.(Glu3342Lys)	VUS	46	Yes	Yes
AZ39	Ovary	BRCA2 c.7788delAinsGGGT p.(Gly2596dup)	VUS	58	Yes	Yes
AZ29	Ovary	BRCA2 c.9302 T>C p.(Leu3101Pro)	VUS	82	No data	Yes
AZ10	Ovary	BRCA2 c.10095delinsGAATTATATCT p.(Ser3366AsnfsTer4)	VUS	100	Yes	Yes

These variants were confirmed by repeat NGS and validated by Sanger DNA sequencing. One sample* could not be confirmed by Sanger DNA sequencing due to PCR failure, possibly due to the poor quality of the FFPE sample. Another sample was not reanalysed by NGS, but was confirmed by Sanger DNA sequencing. ⁺Samples AZ68 and AZ75 are thought to be separate samples from the same tumour block as they were sourced from the same supplier at the same time and have exactly the same variant calls (including coding SNPs).

ISSN: 1472-6890