Simple IHC reveals complex MMR alternations than PCR assays: Validation by LCM and next-generation sequencing.

Evaluation of the status of mismatch repair (MMR) in tumors is crucial for determining the application of immune checkpoint inhibitors (ICIs). Conventional PCR (MSI-PCR) is the gold standard for confirming the MMR status. However, it requires visual confirmation and presents difficulties in determining MMR status. Immunohistochemistry (IHC) is a simple method and can confirming MMR protein expression in the whole tumor. We aim to investigate IHC is more suitable for evaluating MMR status in the tumor. We compared MSI-PCR and IHC by testing 319 samples from 284 patients across 14 cancer types. In discordant cases, we performed laser-capture microdissection and microsatellite instability assay by next-generation sequencing (MSI-NGS). The concordance rate between IHC and MSI-PCR testing was 98.1% (313/319). Two reasons for these discrepancies were ambiguous MSI-PCR results and heterogeneous MSI status within the tumor. Among six cases (1.9%), three were judged as MSI-H by MSI-PCR but with proficient MMR by IHC. The results of MSI-NGS revealed microsatellite stable in these three cases. The remaining three cases, two of three were MSI-H and one was MSS in whole tumor in MSI-PCR. IHC showed a "mosaic" pattern containing both proficient MMR and deficient MMR portions by IHC in all three cases. We performed microdissection and MSI-PCR and found intratumoral heterogeneity of MMR status. These results indicated the advantages of IHC and performed expanded samples (n = 1082) and two additional mosaic cases were identified. Our results clearly indicated that simple IHC is the best choice for determining MMR alterations in critical cases for ICIs treatment.

Usefulness of Five-Parameter System Reconfirmed for Cytopathology of Oral Squamous Cell Carcinoma regardless of Differentiation Degree.

BACKGROUND: We previously introduced the Five-Parameter System (FPS), which exclusively evaluates keratinized cellular findings, for use in cytology examinations of oral well-differentiated squamous cell carcinoma (SCC) and carcinoma in situ (CIS) specimens, as they occasionally lack nuclear atypia and can be challenging for categorization by The Bethesda System (TBS). This study was conducted to determine whether FPS parameters are detectable even in oral SCC/CIS specimens with apparent nuclear atypia. SUMMARY: Oral cytology specimens were obtained together with biopsy tissue samples. They were obtained from 59 malignant (HSIL and SCC) and 29 not-definitely malignant (NILM to ASC-H) specimens diagnosed using TBS. Following re-confirmation of the original TBS categorization, the specimens were re-evaluated using FPS. One or more of the FPS parameters were noted in 69 of 70 malignant specimens examined, of which 11 had been diagnosed by TBS as not-definitely malignant. The remaining one malignant specimen was diagnosed as SCC with only TBS. FPS parameters #1 (concentric arrangement), #2 (large cell number), #3 (bizarre-shaped cells), #4 (keratoglobules), and #5 (uneven filamentous cytoplasm) were observed only in malignant cases, while none were revealed in not-definitely malignant specimens. Finally, TBS supplemented with FPS achieved sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of 100%. KEY MESSAGES: FPS parameters are included in most examinations of oral cytology specimens. Thus, FPS is highly recommended for use in cytology examinations of oral SCC regardless of differentiation degree to confirm judgment based on TBS, a mandatory standard, as well as to cover its limitation of mainly evaluating nuclear atypia. FPS is considered to be an important diagnostic tool for oral cytology, especially in triage cases, which are challenging for TBS. Cytopathology should not be limited to only nuclear findings but be based on whole-cell morphology.