Review for 10.1186/s12876-022-02175-3 (2022)

Title: The methylation of SDC2 and TFPI2 defined three methylator phenotypes of colorectal
cancer
Version: 1 Date: 21 Dec 2021
Reviewer's report:
In the present study, the authors investigated the reliability and improvement in using the
methylation level of SDC2 and TFPI2 genes to assess the methylator phenotypes of colorectal
cancer (CRC).  In order to do this, they identified the best regions of the 5’-UTR sequences of
both SDC2 and TFP12 genes and the appropriate threshold to evaluate the methylation levels
in different cohorts of colorectal cancers and normal controls derived from TCGA, two GEO
datasets and a set of patients collected at Zhongnan Hospital of Wuhan University, used as the
validation cohort.
According to the methylation status of SDC2 and TFPI2, cases were divided into 3 different
subgroups and the new subsets compared for variations in age, tumor location, genome
mutations, and microsatellite instability. Overall, the authors were able to show that the
highest methylation subset was enriched for BRAF mutations, MSI-H cases and older patients.
Accordingly, they were able to define three new clusters of differentially expressed genes with
the highest methylation group associated with transcription regulation pathways and the lower
methylation group associated with extracellular matrix interactions and cell migration pathways.
The paper is valuable because it aims to improve the methylation characterization of colorectal
cancer.  A lot of work has been carried out to improve the sensitivity and sensibility of the used
methylation markers. However, some important points are missed and the general scheme lacks
clarity.
1) The choice of   SDC2 and TFPI1 gene methylation  is at the base  of the study. However only
few information is provided regarding this point. Literature concerning the role of these two
genes is mentioned in the background, but this is a central item and it deserves more attention
and explanation. Besides, the role of these two genes can greatly affect the type of the results
(for example the obtained clusters of differentially expressed genes).
2) In general, it should be make clearer the advantage to classify colorectal cancers by using these
new methylation markers instead of the traditional MLH1 and/or MGMT etc. For example, it is
already well known that the more methylated CRC cluster (the so called CIMP) show frequent
BRAF mutations and they are MSI-H.
3) The results section should be extended and rewritten, making clear what data were obtained
from the analysis of the two explorative  cohorts (TCGA and GEO datasets) and what data were
derived from the validation cohort. Modify the Figures accordingly such as data derived from
all used cohorts are reported.
4) The validation cohort included several rectal cancers (Figure 2). This can be an important
bias for the comparison with the other two cohorts. Was this drawback taken into consideration ?
5) In the Methods section, Identification of target regions, explain why #0.3 was chosen as the
value of the most variable probes. Similarly, in the Statistical paragraph, why 0.2 was chosen
as the optimal # threshold ?
6) Several sentences should be changed because they are not understanble in the present form:
Background in the Abstract; in the Background Section lines 54-55, lines 73-75; in the Results
Section, lines 196-197 and lines 231-233. Moreover, improve the Abstract to make clear the
“take home message”
7) The manuscript should be thoroughly edited because several mispelling are reported (for
example “objects” instead of “subjects” in line 101).