Decreased FOXO1 Expression Is Correlated with Poor Prognosis in Myelodysplastic Syndromes

Round 1

Reviewer 1 Report

In the present study, Zhang et al, address the role of FOXO1 gene as an independent prognostic marker in MDS and it could be a potential candidate target therapy. This is an interesting manuscript that shows FOXO1 as an independent prognostic marker in MDS. Some points need to be addressed:

1.      Author should add the significance of FOXO1 gene with respect to immune environment in general (in cancers) in the introduction section

2.      Is there any specific rationale to study the role of FOXO1's in MDS? In addition to FOXO1, other FOXOs such as FOXO3 and FOXO4 are also involved in cancer.

3.      In the materials and methods section, clearly define the difference between PFS and OS like both are calculated from the date of diagnosis to the date of death, which is confusing.

4.      It would be better to include the criteria for stratifying FOXO1 into low and high expression in the statistical analysis section rather than the results section.

5.      Every table should include the abbreviations below, such as Neu, Plt, IST, HAM, WPSS, etc.

6.      The authors mention in the introduction that tFOXO1 has anti-tumor effects on a variety of tumors, including hematological tumors, digestive system tumors, prostate tumors and breast. However, some studies suggest the opposite.  Thus, I strongly encourage the authors to comment this discrepancy in the literature and discuss the possible explanations.

Author Response

1. Based on your suggestion, we have added the regulatory role of FOXO1 gene on the tumor immune microenvironment in the introduction. New additions to the introduction are marked in red.
2. The reason for choosing FOXO1 is that according to previous literature reports, FOXO1 is considered to be a representative anti-tumor gene in the FOXO family, and has also a role in regulating immunity, which is explained in red font in the introduction.
3. I'm very sorry for confusing you in defining PFS and OS. According to your suggestion, referring to the definition standard of OS and PFS in Eberhardt’s article (Phase III Study of Surgery Versus Definitive Concurrent Chemoradiotherapy Boost in Patients With Resectable Stage IIIA(N2) and Selected IIIB Non–Small-Cell Lung Cancer After Induction Chemotherapy and Concurrent Chemoradiotherapy (ESPATUE), J Clin Oncol, 2015; 33(35):4194-201). We re-described the definitions of PFS and OS, which are marked in red font in the text.
4. According to your suggestion, we put the FOXO1 classification criteria in the statistical analysis section.
5. Thank you very much for the reminder, we have added abbreviations below all tables. See section in red font.
6. Thank you very much for your constructive comments. Although this question is a bit difficult for me，I discuss the differences roles of FOXO1 in different tumors in the discussion section. See section in red font. Correspondingly, I also update the references in the text.

Reviewer 2 Report

Interesting article on a potential prognostic marker in MDS.

Some section need to be improoved as follow:

Section 3.2: it should be stated how FOXO1 IHC was evaluated (median % of positive cells, intensity) and how the patients were divided into two groups.

The authors should also include a section describing the FOXO1 staining in normal bone marrow (at least with IHC) and compare it with MDS.

Finally in section 3.4 the author should describe the cell lineage expressing FOXO1 in IHC.

Author Response

Thank you very much for your review.

1. In section 3.2, according to the median value of FOXO1 mRNA expression, patients were divided into FOXO1 low-expression and high-expression groups, and the clinical data were analyzed and compared. In order to avoid confusion, the grouping criteria of FOXO1 low and high expression groups are explained in the statistics section and section 3.2. In section 3.4, the expression of FOXO1 protein was detected in 3 matched patients, that is, the expression of FOXO1 protein before and after disease progression. At your request, the FOXO1 IHC staining scoring method is detailed in the methods ICH section, and the FOXO1 IHC staining scoring statistics for 3 paired cases are supplemented in section 3.4. And add it in the corresponding position with red font.
2. According to your request, we supplement the results of FOXO1 staining index of normal human bone marrow, which are added in section 3.4, described in red font.
3. Thank you very much for your question, but I am confused about it. First, the main purpose of this study was to determine whether FOXO1 expression could be used as a monitoring indicator for MDS disease progression. Therefore, RT-PCR and IHC were used to monitor FOXO1 expression in 3 matched MDS patients during disease progression. As the disease progressed, FOXO1 expression decreased significantly. Second, is the IHC performed in the bone marrow biopsy of the MDS patient in this paper comparable with the IHC performed in the cell line? Thirdly, in the introduction part, our previous article has studied the expression of FOXO1 in MDS cell lines. Therefore, I implore you not to ask me to supplement the expression of the FOXO1 protein in the cell line. Thanks again very much!