Table 1.
Clinical and molecular data of the patients included in the study
| Cancer type | MMR deficiency type | MSI/IHC status | ID number | Sex | Age | Tumor localization* | Affected gene | DNA (protein) change** | Molecular defect | MLH1 methylation | ÌSI-PCR | IHC | MSI-NGS | |||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| MLH1 | MSH2 | MSH6 | PMS2 | |||||||||||||
| Hereditary | Classical MMR deficiency (n=16) | MSI+ IHC+ | H1 | F | 43 | proximal CRC | MLH1 | c.896_897insC (p.Pro300SerfsTer7) | deletion out-of-frame | − | + | X | X | |||
| H2 | F | 60 | proximal CRC | MLH1 | c.392C>G (p.Ser131Ter) | nonsense mutation | − | + | X | X | ||||||
| H3 | F | 57 | proximal CRC | MLH1 | c.683T>C (p.Leu228Pro) | missense mutation | − | + | X | X | 49,3 | |||||
| H4 | F | 41 | proximal CRC | MLH1 | c.1667+1del (p.?) | splice site mutation | − | + | X | X | 74 | |||||
| H5 | M | 15 | proximal CRC | MSH2 | c.2211-2A>C (p.?) | splice site mutation | − | + | X | X | ||||||
| H6 | M | 50 | proximal CRC | MSH2 | c.2211-2A>C (p.?) | splice site mutation | − | + | X | X | ||||||
| H7 | F | 63 | proximal CRC | MSH2 | c.1786_1788delAAT (p.Asn596del) | deletion in-frame | − | + | X | X | ||||||
| H8 | M | 46 | distal CRC | MSH2 | c.1786_1788delAAT (p.Asn596del) | deletion in-frame | − | + | X | X | ||||||
| H9 | M | 41 | distal CRC | MSH2 | c.209_211+11del (p.?) | deletion in-frame | − | + | X | X | ||||||
| H10 | F | 27 | proximal CRC | MSH2 | c.1012 G>A (p.Gly338Arg) | missense mutation | − | + | X | X | 56,5 | |||||
| H11 | F | 52 | distal CRC | MSH6 | c.3991C>T, (p.Arg1331Ter) | nonsense mutation | − | + | X | 50 | ||||||
| H12 | F | 48 | endometrial | MSH6 | :c.3172G>C (p.Asp1058His) | missense mutation | − | + | X | 36 | ||||||
| H13 | M | 40 | distal CRC | MSH6 | c.3263delT (p.Phe1088SerfsTer2) | deletion out-of-frame | − | + | X | 42,9 | ||||||
| H14 | M | 41 | proximal CRC | PMS2 | g.(5984924_5987848)_ (6015520_?)del (p.?) | large deletion | − | + | X | |||||||
| H15 | M | 68 | distal CRC | PMS2 | c.2192_2196delTAACT (p.Leu731CysfsTer3) | deletion out-of-frame | − | + | X | 73,5 | ||||||
| H16 | M | 65 | proximal CRC | PMS2 | c.1321_1322delA (p.Arg443GlufsTer5) | deletion out-of-frame | + | + | X | X | ||||||
| Unusual MMR deficiency (n=9) | Type1 MSI+IHC- | H17 | M | 55 | proximal CRC | MLH1 | c.244A>G (p.Thr82Ala) | missense mutation | − | + | 54 | |||||
| H18 | M | 51 | proximal CRC | MLH1 | c.244A>G (p.Thr82Ala) | missense mutation | − | + | 53,8 | |||||||
| H19 | M | 38 | proximal CRC | MLH1 | c.62C>T (p.Ala21Val) | missense mutation | − | + | 43 | |||||||
| H20 | F | 37 | distal CRC | MSH6 | c.2927G>C (p.Arg976Pro) | missense mutation | − | + | 40 | |||||||
| Type2 MSI-IHC+ | H21 | F | 52 | endometrial | MSH6 | c.900_901insTC (p.Lys301SerfsTer5) | insertion out of frame | + | − | / X | / X | X | / X | 12 | ||
| H22 | F | 41 | endometrial | MSH6 | c.3514dupA (p.Arg1172LysfsTer5) | deletion out-of-frame | − | − | X | 4,3 | ||||||
| H23 | F | 45 | distal CRC | PMS2 | c.2437C>T (p.Arg813Trp) | missense mutation-VUS | − | − | X | 1,7 | ||||||
| H24 | F | 44 | distal CRC | MSH6 | c.2384T>C (p.Ile795Thr) | missense mutation-VUS | − | − | X | 1,16 | ||||||
| H25 | M | 51 | proximal CRC | MSH6 | c.1151_1156dupGGAGGC (p.Arg384_385dup) | Insertion in-frame - VUS | − | − | X | 2 | ||||||
| pMMR (n=1) | MSI-IHC- | H26 | F | 44 | distal CRC | MSH6 | c.457+1G>T (p?) | splice site mutation | − | − | 3,12 | |||||
| Sporadic | Classical MMR deficiency (n=18) | MSI+ IHC+ | S1 | M | 54 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 79,5 | ||
| S2 | M | 39 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 73,7 | |||||
| S3 | M | 72 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | ||||||
| S4 | F | 57 | endometrial | MLH1 | NA | promoter methylation | + | + | X | X | 56,5 | |||||
| S5 | F | 57 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 47,9 | |||||
| S6 | M | 73 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 76,1 | |||||
| S7 | F | 75 | endometrial | MLH1 | NA | promoter methylation | + | + | X | X | ||||||
| S8 | F | 73 | endometrial | MLH1 | NA | promoter methylation | + | + | X | X | 19,9 | |||||
| S9 | F | 53 | endometrial | MLH1 | NA | promoter methylation | + | + | X | X | 19,2 | |||||
| S10 | M | 57 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 9,1 | |||||
| S11 | M | 68 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 4,4 | |||||
| S12 | F | 60 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 41,1 | |||||
| S13 | F | 58 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 77,9 | |||||
| S14 | M | 67 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 76,7 | |||||
| S15 | M | 72 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 48,9 | |||||
| S16 | M | 77 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 64,3 | |||||
| S17 | F | 68 | proximal CRC | MLH1 | NA | promoter methylation | + | + | X | X | 52,8 | |||||
| S18 | F | 78 | distal CRC | MLH1 | NA | promoter methylation | + | + | X | X | ||||||
Figure 1.
Distribution of MSI patients by various methods for MSI detection. Classical MMR deficiency = detected by both MSI-PCR and IHC (MSI-H and loss of MMR proteins). Unusual MMR deficiency = detected by one method, MSI-PCR or IHC: type 1 (MSI-H with intact MMR protein expression) and type 2 (MSS and isolated loss of PMS2 or MSH6). *Clinically and molecularly diagnosed Lynch syndrome (pathogenic mutation in the MSH6) with MSS status, normal expression of all four MMR proteins and negative MSI-NGS result. MSI, microsatellite instability, MSS, microsatellite stability, NGS, next-generation sequencing, dMMR, deficient mismatch repair, pMMR, proficient mismatch repair, IHC, immunohistochemistry, PCR, polymerase chain reaction
Figure 2.
Quantitative MSI-NGS levels of patients with sporadic and Lynch Syndrome-associated CRC or endometrial cancers in relation to different types of MMR deficiencies, affected genes, and localization.
The threshold level for NGS-MSI positivity is set at 3.5%. MSI, microsatellite instability, MMR, mismatch repair, IHC, immunohistochemistry, VUS, variant of uncertain significance, CRC, colorectal cancer, EC, endometrial cancer
Figure 3.
Proposed strategy for more accurate cost-effective evaluation of MMR deficiency and potential subsequent NGS-MSI testing for quantitative assessment.
IHC, immunohistochemistry, LS, Lynch syndrome, dMMR, deficient mismatch repair, pMMR, proficient mismatch repair, MSI, microsatellite instability, MSS, microsatellite stability, NGS, nextgeneration sequencing