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Table. 3. Table 3

Significant variants with pathogenicity classification detected using the NEOseq_ACTION panel

Case No.	Gene symbol	Transcript	Nucleotide change	Amino acid change	Zygosity	Variant classification by MedyPatho	Segregation	Clinical validation	Final classification	ACMG evidences^*
P2	GALNS	NM_000512.4	c.319G >A	p.Ala107Thr	Het	LP	Trans	Serum galactose-6-sulfatase 0.37 nmol/17 hr/mg protein (ref, 18.6–61.8)	LP	PM2, PM3, PP3, PP5
	GALNS	NM_000512.4	c.451C >A	p.Pro151Thr	Het	LP			LP	PM1, PM2, PM5, PP3, PP5
P5	ACADS	NM_000017.2	c.1031A >G	p.Glu344Gly	Het	LP	Not done	Serum C4 1.995 μmol/L (cut off, 0.41)	LP	PM1, PM2, PP3, PP5
	ACADS	NM_000017.2	c.164C >T	p.Pro55Leu	Het	LP			LP	PS3, PM2, PP5
P6	PCCA	NM_000282.3	c.1846-2A >G	p?	Het	LP	Trans	Urine propionylglycine 80.0 (ref, not detected)	LP	PVS1, PM2
	PCCA	NM_000282.3	c.938G >C	p.Arg313Pro	Het	VUS^{^†}			LP^{^†}	PM2, PM3, PM5, PP3
P10	PAH	NM_000277.1	c.975C >G	p.Tyr325^*	Het	P	Not done	Serum phenylalanine 163 μmol/L (ref, 13–91)	P	PVS1, PM2, PP5
	PAH	NM_000277.1	c.158G >A	p.Arg53His	Het	VUS			VUS	BS2, PM5
P14	SLC25A13	NM_014252.3	c.852_855delTATG	p.Met285Profs^*2	Het	P	Not done	Serum citrulline 378 μmol/L (ref, 8–36)	P	PVS1, PM2, PP5
	SLC25A13	NM_014252.3	c.1177+1G >A	p?	Het	P			P	PVS1, PM2, PP5
P25	ACADM	NM_000016.4	c.1189T >A	p.Try397Asn	Het	LP	Trans	Serum C6 0.145 μmol/L (cut off < 0.1), C8 0.26 μmol/L (cut off < 0.18), C10:1 0.16 μmol/L (cut off < 0.13)	LP	PM2, PM5, PP2, PP3, PP4, PP5
	ACADM	NM_000016.4	c.1231G >T	p.Val411Leu	Het	VUS^{^†}			LP^{^†}	PM2, PM3, PP2, PP4
P36	ACADM	NM_000016.4	c.1189T >A	p.Tyr397Asn	Het	LP	Not done	Serum C6 0.64 μmol/L (cut off < 0.1), C8 1.59 μmol/L (cut off < 0.12), C10:1 0.45 μmol/L (cut off < 0.08)	LP	PM2, PM5, PP2, PP3, PP4, PP5
	ACADM	NM_000016.4	c.91C >T	p.Arg31Cys	Het	VUS^{^†}			LP^{^†}	PM2, PM5, PP2, PP4, PP5
P40	MCCC2	NM_022132.4	c.838G >T	p.Asp280Tyr	Het	LP	Not done	Urine 3-hydroxyisovaleric acid 1,731.1 mmol/mol Cr (ref, < 18)	LP	PM2, PP2, PP3, PP4, PP5
	MCCC2	NM_022132.4	c.1342G >A	p.Gly448Arg	Het	VUS^{^†}			LP^{^†}	PM2, PP2, PP3, PP4, PP5
P41	PREPL	NM_006036.4	c.1979_1980del	p.Leu660Glnfs^*9	Het	LP^{^†}	Trans	Repetitive nerve stimulation test, insignificant	P^{^†}	PVS1, PM2, PM3
	PREPL	NM_006036.4	c.1747-9_1747del	p?	Het	LP^{^†}			P^{^†}	PVS1, PM2, PM3
P45^{^‡}	HBA2	NM_000517.4	c.427T >C	p.^143Qext^31	Hem	LP	Not done	Not done	LP	PM2, PM3, PM4
P46	PHKA2	NM_000292.2	c.1246-2A >G	p?	Hem	LP	Not done	GOT/GPT 59/49 IU/L (ref, 1–40/1–40)	LP	PVS1, PM2
P72	ACADS	NM_000017.2	c.164C >T	c.Pro55Leu	Hom	LP	Not done	Serum C4 0.805 μmol/L (ref, < 0.41)	LP	PS3, PM2, PP5
P74	ACADS	NM_000017.2	c.1031A >G	p.Glu344Gly	Het	LP	Not done	Serum C4 0.905 μmol/L (ref, < 0.41)	LP	PM1, PM2, PP3, PP5
	ACADS	NM_000017.2	c.1130C >T	p.Pro377Leu	Het	LP			LP	PM1, PM2, PP3, PP5
P83	ACADS	NM_000017.2	c.1031A >G	p.Glu344Gly	Hom	LP	Not done	Serum C4 1.965 μmol/L (ref, < 0.41)	LP	PM1, PM2, PP3, PP5
P111	PAH	NM_000277.1	c.1068C >G	p.Tyr356^*	Het	P	Not done	Serum phenylalanine 173 μmol/L (ref, 25–74)	P	PVS1, PM2, PP5
	PAH	NM_000277.1	c.158G >A	p.Arg53His	Het	VUS			VUS	BS2, PM5

*All variants were assessed based on the ACMG or Association for Molecular Pathology guidelines [17]; ^†Initial classification using MedyPatho was changed at final classification; ^‡Given the high homology between HBA1 and HBA2 and that common deletions account for 85% of pathogenic variants in alpha thalassemia, Sanger sequencing and multiplex ligation probe amplification were performed to confirm the variant. This patient had a heterozygous common–^SEA deletion; thus, the zygosity of the c.427T>C variant was determined to be hemizygous.

Abbreviations: ACMG, American College of Medical Genetics and Genomics; Het, heterozygous; Hem, hemizygous; Hom, homozygous; LP, likely pathogenic; P, pathogenic; VUS, variant of unknown significance; ref, reference range.

Ann Lab Med 2023;43:280~289 https://doi.org/10.3343/alm.2023.43.3.280