Background
Methods
Study population
Targeted sequencing
Sequencing data analysis
Validation by cycling temperature capillary electrophoresis
Genetic variants nomenclature and classification
In silico analyses of VUS
Cell-based minigene splicing assays
Results
Family history and clinical characteristics
Patient_ID | Institution | Familial path_BRCA1 or path_BRCA2 variantFamilial path_BRCA1 or path_BRCA2 variant | ICD9 diagnosis (age) | Pathogenic variant identified in the current study |
---|---|---|---|---|
17,161 | HCBNRH |
BRCA2 c.5217_5223delTTTAAGT (p.Tyr1739Terfs)BRCA2 c.5217_5223delTTTAAGT (p.Tyr1739Terfs)
| OC (67) |
ATM c.468G > A (p.Trp156Ter)*ATM c.468G > A (p.Trp156Ter)*
|
6475 | HCBNRH |
BRCA1 c.1011dupA (p.Val340Glyfs)BRCA1 c.1011dupA (p.Val340Glyfs)
| BC (52) |
ATM c.9139C > T (p.Arg3047Ter)ATM c.9139C > T (p.Arg3047Ter)
|
13,141 | HCBNRH |
BRCA1 c.1072delC (p.Leu358Cysfs)BRCA1 c.1072delC (p.Leu358Cysfs)
| EC (57) |
MSH6 c.2864delC (p.Thr955fs)*MSH6 c.2864delC (p.Thr955fs)*
|
1873 | HCBNRH |
BRCA1 c.1556delA (p.Lys519Argfs)BRCA1 c.1556delA (p.Lys519Argfs)
| MTHM (56), BC (70) | Not |
5378 | HCBNRH |
BRCA1 c.697_698delGT (p.Val233Asnfs)BRCA1 c.697_698delGT (p.Val233Asnfs)
| BC (52) | Not |
5180 | HCBNRH |
BRCA1 c.5194-2A > CBRCA1 c.5194-2A > C
| BC (39) | Not |
22 | HCBNRH |
BRCA2 c.3847_3848delGT (p.Val1283Lysfs)BRCA2 c.3847_3848delGT (p.Val1283Lysfs)
| BC (63) | Not |
243 | HCBNRH |
BRCA2 c.3847_3848delGT (p.Val1283Lysfs)BRCA2 c.3847_3848delGT (p.Val1283Lysfs)
| CVC (41) | Not |
5348 | HCBNRH |
BRCA1 c.1556delA (p.Lys519Argfs)BRCA1 c.1556delA (p.Lys519Argfs)
| BC (68) | Not |
6031 | HCBNRH |
BRCA1 c.1556delA (p.Lys519Argfs)BRCA1 c.1556delA (p.Lys519Argfs)
| BC (66) | Not |
6032 | HCBNRH |
BRCA1 c.3228_3229delAG (p.Gly1077Alafs)BRCA1 c.3228_3229delAG (p.Gly1077Alafs)
| OC (55) | Not |
6207 | HCBNRH |
BRCA1 c.697_698delGT (p.Val233Asnfs)BRCA1 c.697_698delGT (p.Val233Asnfs)
| BC (47) | Not |
8085 | HCBNRH | BRCA1 c.3228_3229delAG (p.Gly1077Alafs)BRCA1 c.3228_3229delAG (p.Gly1077Alafs) | BC (55), CC (66) | Not |
11,717 | HCBNRH | BRCA1 c.1556delA (p.Lys519Argfs)BRCA1 c.1556delA (p.Lys519Argfs) | BC(42,57) | Not |
12,470 | HCBNRH |
BRCA1 c.3178G > T (p.Glu1060Ter)
| BC (39) | Not |
13,023 | HCBNRH |
BRCA2 c.5217_5223delTTTAAGT (p.Tyr1739Terfs)
| BC (59) | Not |
15,529 | HCBNRH |
BRCA2 c.4821_4823delTGAins
| BC (48) | Not |
22,325 | HCBNRH |
BRCA1 c.5047G > T (p.Glu1683Ter)
| BC (45) | Not |
1,100,948 | UM |
BRCA2 c.6591_6592delTG (p.Glu2198Asnfs)
| BC (44) |
BRCA2 c.9382C > T (p.Arg3128Ter)
|
12,010,643 | UM |
BRCA2 c.7360delA (p.Ile2454Phefs)
| BC (56) |
MUTYH c.1178G > A (p.Gly393Asp)
|
75,443 | UM |
BRCA2 c.5909C > A (p.Ser1970Ter)
| BC (55) | Not |
88,295 | UM |
BRCA2 c.7977-1G > C
| BC (44) | Not |
64,949 | UM |
BRCA2 c.5909C > A (p.Ser1970Ter)
| BC (55) | Not |
67,723 | UM |
BRCA2 c.4866delA p.(Arg1622Serfs*14)
| BC (46) | Not |
84,510 | UM |
BRCA2 c.5946delT (p.Ser1982Argfs)
| BC (67) | Not |
13,007,862 | UM | BRCA2 c.5909C > A (p.Ser1970Ter) | BC (31) | Not |
9,009,462 | UM | BRCA2 c.6535_6536insA (p.Val2179Aspfs) | BC (67) | Not |
900,178 | UM | BRCA2 c.1889delC (p.Thr630Asnfs) | BC (49,77) | Not |
10,005,829 | UM |
BRCA2 c.9541_9554del p.(Met318CysfsTer13)
| BC (38) | Not |
10,007,016 | UM | BRCA2 c.632-1G > A | BC (51) | Not |
10,003,959 | UM |
BRCA2 c.6275_6276delTT (p.Leu2092Profs)
| BC (55) | Not |
12,852 | UM |
BRCA2 c.1929delG (p.Arg645Glufs)
| BC (56) | Not |
12,001,161 | UM | BRCA2 c.7958 T > C (p.Leu2653Pro) | BC (67) | Not |
13,017,067 | UM | BRCA2 c.755_758delACAG (p.Asp252Valfs) | BC (74) | Not |
688 | UM | BRCA2 c.1929delG (p.Arg645Glufs) | BC (32) | Not |
40,540 | UM | BRCA2 c.8535_8538delAGAG p.(Glu2846LysfsTer16) | BC (69) | Not |
9,001,644 | UM | BRCA2 c.4965C > G (p.Tyr1655Ter) | BC (39, 45) | Not |
89,205 | UM | BRCA2 c.5946delT (p.Ser1982Argfs) | BC (77) | Not |
10,002,068 | UM | BRCA2 del exons 14–16 | BC (37) | Not |
10,004,590 | UM | BRCA2 c.2672dupT | BC (67,67) | Not |
40,286 | UM | BRCA2 c.7069_7070delCT p.(Leu2357ValfsTer2) | BC (36,53) | Not |
76,618 | UM | BRCA2 c.4478_4481delAAAG (p.Glu1493Valfs) | BC (51) | Not |
12,015,576 | UM | BRCA2 c.9382C > T (p.Arg3128Ter) | BC (45) | Not |
61,420 | UM | BRCA2 c.5350_5351delAA p.(Asn1784HisfsTer2) | BC (59) | Not |
960,579 | UM | BRCA2 c.2808_2811del4 (p.Ala938Profs) | BC (39) | Not |
14,965 | UM | BRCA2 c.5682C > G p.(Tyr1894Ter) | BC (59) | Not |
20,468 | UM |
BRCA2 c.6275_6276delTT (p.Leu2092Profs)
| BC (38) | Not |
56,193 | UM |
BRCA2 c.7884dupA (p.Trp2629Metfs)
| BC (79) | Not |
Germline findings
Validation of the cancer gene panel output
Variants of unknown significance (VUS) and predicted protein alterations
Patient ID | Genomic position (GRCh37) | Gene | Exon | Nucleotide change (cNomen) | Predicted protein change (pNomen) | dbSNPrsID | Non-Finnish European population frequency* | Reference splice site-dedicated analyses | Cryptic splice site-dedicated analyses | ESR-dedicated analyses | |||||||||||
---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
Nearest reference | MES scores | SSFL scores | Potential local splice effect | Local MES scores | ∆tESRseq | ∆Hzei | ΔΨ | ||||||||||||||
Distance | Type | WT | Var | VAR vs WT | WT | Var | VAR vs WT | WT | Var | ||||||||||||
(nt) | (3′ or 5’ss) | ∆ (%) | ∆ (%) | ||||||||||||||||||
688 | chr_16_68835593_G_A |
CDH1
| 3 | c.184G > A | p.Gly62Ser | 587,781,898 | 5.99e-05 | 21 | 3’ | 8.17477 | 8.17477 | 0 | 86.5179 | 86.5179 | 0 | − 1.44947 | 10.35 | −1.24 | |||
chr2_47703664_G_A |
MSH2
| 13 | c.2164G > A | p.Val722Ile | 587,781,996 | 8.99e-05 | −47 | 5’ | 10.8583 | 10.8583 | 0 | 100 | 100 | 0 | 0.59756 | 10.51 | −0.01 | ||||
chr_8_90983475_C_A |
NBN
| 6 | c.628G > T | p.Val210Phe | 61,754,796 | 0.0008158 | 44 | 3’ | 6.19815 | 6.19815 | 0 | 86.8244 | 86.8244 | 0 | − 0.782222 | −46.21 | − 0.15 | ||||
1873 | chr_5_56155672_A_G |
MAP3K1
| 3 | c.764A > G | p.Asn255Ser | 56,069,227 | 0.0269 | −71 | 5’ | 7.52484 | 7.52484 | 0 | 78.4708 | 78.4708 | 0 | New Acceptor Site? | – | 8.8 | −1.18661 | 6.7 | −0.04 |
5378 | chr 12_133244944_G_A |
POLE
| 19 | c.2171C > T | p.Ala724Val | 61,734,163 | 0.00030 | −3 | 5’ | 9.89081 | 8.73118 | −11.7 | 86.6769 | 82.5488 | −4.8 | New Donor Site? | – | 6.3 | −2.14822 | −32.05 | −0.16 |
6031 | chr17_41245621_T_C |
BRCA1
| 10 | c.1927A > G | p.Ser643Gly | 80,357,105 | NA | 1257 | 3’ | 8.86265 | 8.86265 | 0 | 87.3058 | 87.3058 | 0 | 1.44078 | 58.08 | 0.02 | |||
AXIN2
| 10 | c.2272G > A | p.Ala758Thr | 145,007,501 | 0.0039861 | 35 | 3’ | 6.34671 | 6.34671 | 0 | 86.1925 | 86.1925 | 0 | −0.942617 | 0.12 | −0.09 | |||||
chr5_112102960_C_T |
APC
| 4 | c.295C > T | p.Arg99Trp | 139,196,838 | 0.0006444 | 75 | 3’ | 7.49577 | 7.49577 | 0 | 84.8039 | 84.8039 | 0 | −2.2189 | −14.34 | − 0.08 | ||||
12,470 |
AXIN2
| 10 | c.2272G > A | p.Ala758Thr | 145,007,501 | 0.0039861 | 35 | 3’ | 6.34671 | 6.34671 | 0 | 86.1925 | 86.1925 | 0 | −0.942617 | 0.12 | − 0.09 | ||||
chr5_112128218_G_A |
APC
| 7 | c.721G > A | p.Glu241Lys | 777,603,154 | 0.0001818 | −9 | 5’ | 7.15277 | 7.15277 | 87.0697 | 87.0697 | 0 | −1.51981 | −49.76 | −0.42 | |||||
12,852 | chr_14_69061228_G_A |
RAD51B RAD51B
| 11 | c.1063G > A | p.Ala355Thr | 61,758,785 | 0.0071658 | 27 | 3’ | 11.8 | 11.8 | 0 | 80.2 | 80.2 | 0 | – | – | – | −1.24035 | −50.64 | – |
88,295 | chr10_89690828_G_A |
PTEN PTEN
| 4 | c.235G > A | p.Ala79Thr | 202,004,587 | 0.0001678 | −19 | 5’ | 9.6515 | 9.6515 | 0 | 86.8647 | 86.8647 | 0 | −1.39321 | 10.77 | 0.6 | |||
900,178 | chr11_94197365_C_T |
MRE11A MRE11A
| 11 | c.1139G > A | p.Arg380His | 587,781,646 | 4.5e-05 | 41 | 3’ | 8.9941 | 8.9941 | 0 | 95.7456 | 95.7456 | 0 | −1.57887 | −48.78 | −0.03 | |||
960,579 | chr_5_56177843_C_G |
MAP3K1
| 14 | c.2816C > G | p.Ser939Cys | 45,556,841 | 0.0221 | 447 | 3’ | 12.0063 | 12.0063 | 0 | 100 | 100 | 0 | – | – | – | −0.486881 | −16.1 | 0 |
1,000,459 | chr13_28537449_ACTT_A |
CDX2
| 3 | c.742_744del | p.Lys248delAAG | 553,066,746 | 0.0001682 | 55 | 3’ | 11.7045 | 11.7045 | 0 | 87.4307 | 87.4307 | 0 | −2.46964 | −100.08 | – | |||
1,100,948 | chr_17_7133187_A_G |
DVL2 DVL2
| 5 | c.596 T > C | p.Met199Thr | 372,715,697 | 6.01e-05 | −61 | 5’ | 6.34467 | 6.34467 | 0 | 80.4452 | 80.4452 | 0 | 0.0509416 | −1.77 | 0.54 | |||
chr2_47641430_C_T |
MSH2
| 5 | c.815C > T | p.Ala272Val | 34,136,999 | 0.0003755 | 23 | 3’ | 10.3527 | 10.3527 | 0 | 84.3224 | 84.3224 | 0 | −2.17832 | −46.5 | −0.03 | ||||
10,002,068 | chr_17_63526198_C_T |
AXIN2
| 11 | c.2428G > A | p.Asp810Asn | 140,344,858 | 1.5e-05 | 23 | 3’ | 11.6727 | 11.6727 | 0 | 87.3948 | 87.3948 | 0 | −1.22987 | −14.33 | – | |||
10,005,829 | chr_14_69061228_G_A |
RAD51B
| 11 | c.1063G > A | p.Ala355Thr | 61,758,785 | 0.0071658 | 27 | 3’ | 11.8 | 11.8 | 0 | 80.2 | 80.2 | 0 | – | – | – | −1.24035 | −50.64 | – |
chr8_90993640_C_T |
NBN
| 3 | c.283G > A | p.Asp95Asn | 61,753,720 | 0.0030459 | −38 | 5’ | 10.7663 | 10.7663 | 0 | 94.6711 | 94.6711 | 0 | 0.318238 | 24.6 | 0.03 | ||||
chr_11_108155132_G_A |
ATM
| 26 | c.3925G > A | p.Ala1309Thr | 149,711,770 | 0.0009147 | 9.98517 | 9.98517 | 0 | 84.8076 | 84.8076 | 0 | 0.676556 | 32.96 | 0.04 | ||||||
12,001,161 | chr_14_68353893_A_G |
RAD51B
| 7 | c.728A > G | p.Lys243Arg | 34,594,234 | 0.010682 | −29 | 5’ | 9.09184 | 9.09184 | 0 | 78.9497 | 78.9497 | 0 | Cryptic 5’ss activation? | 0.9 | 7.9 | −1.48785 | −40.54 | −0.19 |
12,015,576 | chr19_15291551_C_G |
NOTCH3
| 19 | c.3083G > C | p.Trp1028Ser | rs146829488 | na | −60 | 5’ | 11.1124 | 11.1124 | 0 | 82.5954 | 82.5954 | 0 | 0.300115 | −6.4 | 0.1 | |||
11,717 | chr1_45797881_C_T |
MUTYH
| 10 | c.881G > A | p.Cys294Tyr | rs879254257 | na | −44 | 5’ | 6.31089 | 6.31089 | 0 | 72.818 | 72.818 | 0 | 1.09496 | −7.06 | 0.04 | |||
17,161 | chr_11_108139187_T_A |
ATM
| 18 | c.2689 T > A | p.Phe897Ile | 147,122,522 | 4.5e-05 | 51 | 3’ | 9.8979 | 9.8979 | 0 | 93.4253 | 93.4253 | 0 | 0.554269 | 87.95 | 0.01 | |||
22 | chr_12_133241897_A_G |
POLE
| 21 | c.2459 T > C | p.Met820Thr | 767,460,640 | 0 | −10 | 5’ | 6.58677 | 6.58677 | 0 | 77.9039 | 77.9039 | 0 | 1.28743 | −2.13 | 0.06 | |||
chr_14_68352672_A_G |
RAD51B
| 6 | c.539A > G | p.Tyr180Cys | 28,910,275 | 0.0045906 | −34 | 5’ | 9.54919 | 9.54919 | 0 | 83.7411 | 83.7411 | 0 | 0.881539 | 7.23 | −0.19 | ||||
chr_5_56155672_A_G |
MAP3K1
| 3 | c.764A > G | p.Asn255Ser | 56,069,227 | 0.0269 | −71 | 5’ | 7.52484 | 7.52484 | 0 | 78.4708 | 78.4708 | 0 | New Acceptor Site? | – | 8.8 | −1.18661 | 6.7 | −0.04 | |
6207 | chr_17_63530163_C_T |
AXIN2
| 10 | c.2272G > A | p.Ala758Thr | 145,007,501 | 0.0039861 | 35 | 3’ | 6.34671 | 6.34671 | 0 | 86.1925 | 86.1925 | 0 | −0.942617 | 0.12 | −0.09 | |||
6475 | chr_17_33433488_G_A |
RAD51D
| 6 | c.493C > T | p.Arg165Trp | 544,654,228 | 6.94e-05 | 13 | 3’ | 8.20686 | 8.20686 | 0 | 85.1161 | 85.1161 | 0 | −2.55724 | −22.32 | 1.42 |