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. 2022 Apr 28;14(9):2204.
doi: 10.3390/cancers14092204.

Microsatellite Status Detection in Gastrointestinal Cancers: PCR/NGS Is Mandatory in Negative/Patchy MMR Immunohistochemistry

Federica Zito Marino  1 , Martina Amato  1 , Andrea Ronchi  1 , Iacopo Panarese  1 , Franca Ferraraccio  1 , Ferdinando De Vita  2 , Giuseppe Tirino  2 , Erika Martinelli  2 , Teresa Troiani  2 , Gaetano Facchini  3 , Felice Pirozzi  4 , Michele Perrotta  5 , Pasquale Incoronato  6 , Raffaele Addeo  7 , Francesco Selvaggi  8 , Francesco Saverio Lucido  8 , Michele Caraglia  2 , Giovanni Savarese  9 , Roberto Sirica  9 , Marika Casillo  9 , Eva Lieto  10 , Annamaria Auricchio  10 , Francesca Cardella  10 , Ludovico Docimo  8 , Gennaro Galizia  11 , Renato Franco  1
Affiliations

Microsatellite Status Detection in Gastrointestinal Cancers: PCR/NGS Is Mandatory in Negative/Patchy MMR Immunohistochemistry

Federica Zito Marino et al. Cancers (Basel). .

Abstract

Background: Microsatellite instability (MSI) is a predictive biomarker for immune checkpoint inhibitors. The main goal was to investigate the discordance between IHC and PCR/NGS for MSI testing in gastrointestinal cancers.

Methods: Two series were analyzed through IHC for mismatch-repair-system proteins (MMRP) and PCR, with one series of 444 colorectal cancers (CRC) and the other of 176 gastric cancers (GC). All cases with discordant results between IHC and PCR were analyzed by NGS. IHC staining was evaluated as follows: proficient MMR (pMMR), with all MMR positive; deficient MMR (dMMR), with the loss of one heterodimer; and cases with the loss/patchy expression of one MMR (lo-paMMR). Cases with instability in at least two markers by PCR were MSI-high (MSI-H) and with instability in one marker, MSI-low (MSI-L). Cases without instability were evaluated as microsatellite-stable (MSS).

Results: In the CRC cohort, 15 out of 444 cases were dMMR and 46 lo-paMMR. Among the 15 dMMR, 13 were MSI-H and 2 MSS. Among the 46 lo-paMMR, 13 were MSI-H and 33 were MSS. In the GC cohort, 13 out of 176 cases were dMMR and 6 cases lo-paMMR. Among the 13 dMMR, 12 were MSI-H and only 1 was MSS. All six lo-paMMR cases were MSS. All NGS results were in agreement with PCR.

Conclusions: In clinical practice, MMR-IHC could be used as a screening test and additional molecular analysis is mandatory exclusively in cases carrying loss/patchy MMR-IHC.

Keywords: PCR; gastrointestinal cancers; microsatellite instability; mismatch-repair-system-protein deficient; mismatch-repair-system-protein patchy.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Representative results of CRC case showing dMMR-IHC and MSI-H by PCR. (A) MMR-IHC results: loss expression of MLH1 and PMS2 in tumor cells with positive internal control (original magnification 10×); intact expression of MSH2 and MSH6 in tumor cells (original magnification 10×). (B) MSI-PCR results: stability of NR22, NR24, CAT25 and MONO27; instability of BAT25, BAT26, NR21 and NR27 (red lines indicate samples while blue lines indicate MSS controls).
Figure 2
Figure 2
Representative results of CRC case showing dMMR-IHC and MSI-H by PCR. (A) MMR-IHC results: intact expression of MLH1 and PMS2 in tumor cells (original magnification 10×); loss expression of MSH2 and MSH6 in tumor cells with positive internal control (original magnification 10×). (B) MSI-PCR results: stability of NR21, NR22 and MONO27; instability of BAT25, BAT26, NR21, NR24, NR27 and CAT25 (red lines indicate samples while blue lines indicate MSS controls).
Figure 3
Figure 3
Representative results of CRC case showing loMMR-IHC and MSI-H by PCR. (A) MMR-IHC results: intact expression of MLH1, PMS2 and MSH2 in tumor cells (original magnification 10×); loss expression of MSH6 in tumor cells with positive internal control (original magnification 10×). (B) MSI-PCR results: stability of BAT26, NR22, NR24 and MONO27; instability of BAT25, NR21, NR27 and CAT25 (red lines indicate samples while blue lines indicate MSS controls).
Figure 4
Figure 4
Representative results of CRC case showing paMMR-IHC and MSS by PCR. (A) MMR-IHC results: patchy expression of MLH1 and PMS2 in tumor cells with positive internal control (original magnification 10×); intact expression of MSH2 and MSH6 in tumor cells (original magnification 10×). (B) MSS-PCR results: stability of BAT25, BAT26, NR21, NR22, NR24, NR27, CAT25 and MONO27 (red lines indicate samples while blue lines indicate MSS controls).
Figure 5
Figure 5
Representative results of CRC case showing paMMR-IHC and MSI-H by PCR. (A) MMR-IHC results: intact expression of MLH1, MSH2 and MSH6 in tumor cells (original magnification 10×); patchy expression of PMS2 in tumor cells with positive internal control (original magnification 10×). (B) MSI-PCR results: stability of BAT25 and NR22; instability of BAT26, NR21, NR24, NR27, CAT25 and MONO27 (red lines indicate samples while blue lines indicate MSS controls).
Figure 6
Figure 6
Representative results of GC case showing dMMR-IHC and MSS by PCR. (A) MMR-IHC results: loss expression of MLH1 and PMS2 in tumor cells with positive internal control (original magnification 10×); intact expression of MSH2 and MSH6 in tumor cells (original magnification 10×). (B) MSS-PCR results: stability of BAT25, BAT26, NR21, NR22, NR24, NR27, CAT25 and MONO27 (red lines indicate samples while blue lines indicate MSS controls).
Figure 7
Figure 7
Representative results of GC case showing paMMR-IHC and MSS by PCR. (A) MMR-IHC results: intact expression of MLH1, MSH2 and MSH6 in tumor cells (original magnification 10×); patchy expression of PMS2 in tumor cells with positive internal control (original magnification 10×). (B) MSS-PCR results: stability of BAT25, BAT26, NR21, NR22, NR24, NR27, CAT25 and MONO27 (red lines indicate samples while blue lines indicate MSS controls).
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