Diagnostic Accuracy of Methods for Detection of Antibodies against Type I Interferons in Patients with Endocrine Disorders

Comment: Reviewer #1.
The authors thank the Reviewer for the evaluation of the manuscript
Your work is scientifically interesting in the field of personalised medicine.
1. In the patient groups with low patient number ( n=3 and n=4 ) you could not made correct conclusions
The following groups were analyzed: 18 patients with APS-1 (group 1), 89 patients with autoimmune endocrine pathology (group 2), 71 patients with non-autoimmune endocrine pathology (group 3) and 28 healthy individuals (group 4).We did not conduct a subgroup analysis, for example, patients with hypergonadotropic hypogonadism (HH) of autoimmune origin (n=4) or patients with autoimmune adrenal insufficiency (AAI) (n=3).
2. Please correct references 5; 10; 12; 21 - use small letters for article titles text flow.
These references were corrected in the revised version of the manuscript.


Reviewer #2.
This is an excellent study in which the author has evaluated the sensitivity and specificity of anti-IFN-I auto-Abs detection using various methods: multi- 87 plex microarray analysis, cell based autoantibodies, and a commercially available ELISA.
The authors thank the Reviewer for the evaluation of the manuscript.
1. The abstract is written well and precisely. However, please consider adding a sentence about the clinical utility of this study for general clinicians and endocrinologists.
The following sentence was added to the abstract:Moreover, determination of antibodies to IFN-ω and IFN-α2 allows a short-term diagnosis in patients with isolated and atypical forms of APS-1.
2. The role of autoantibodies in COVID-19 ( lines 40-52 ) adds little value and could be presented in a precise manner.
This paragraph was corrected as follows:Lines 42-47 were removed.The following sentence was added:Thus, the long-standing opinion that there is no increased susceptibility to viral infections in patients with APS-1, despite the presence of auto-Abs against IFN-I [13], is refuted.
New reference #13: Kisand, K.; Link, M.; Wolff A.S.B.; Meager A.; Tserel L.; Org T.; Murumägi A.; Uibo R.; Willcox N.; Podkrajšek K.T.; et al. Interferon autoantibodies associated with AIRE deficiency decrease the expression of IFN-stimulated genes. Blood 2008, 112, 2657–2666. doi: 10.1182/blood-2008-03-144634.
Therefore, this paragraph now re-written as follows:
Nevertheless, recent data indicate that anti-interferon auto-Abs are more common in the population than previously thought. Over the past two years, the role of auto-Abs against IFN-I in SARS-CoV-2 infection has been demonstrated [7], moreover, they have been shown to underlie severe side effects after vaccination with a live attenuated yellow fever virus vaccine [8]. Multiple studies showed that more than 10% of patients with neutralizing auto-Abs against IFN-I had life-threatening COVID-19 pneumonia [9-11]. The presence of auto-Abs against IFN-I appears to remain clinically asymptomatic in people prior to their infection with SARS-CoV-2. To date, a case of APS-1 was diagnosed in two brothers, 7-year-old and 13-year-old, with life-threatening pneumonia caused by COVID-19. Due to the severity of COVID-19 and their medical history, APS-1 was suspected and the patients underwent appropriate molecular testing. In addition to a mutation in the AIRE gene, the patients were found to carry auto-Abs against IFN-I [12]. Thus, the long-standing opinion that there is no increased susceptibility to viral infections in patients with APS-1, despite the presence of auto-Abs against IFN-I [13], is refuted. Currently, there are also suggestions about the possible role of autoantibodies against IFN-I in other severe viral and malignant diseases, especially in the elderly [14].
3. Table 1 is beneficial to understand this study and its results at a glance. Still, the author should consider adding another table to discuss the sensitivity and specificity of the three tests to make them more reader-friendly.
We agree. Values of specificity and sensitivity of the assays used in this study are presented in Table 2.The title of the Table 2 was changed as follows:
Table 2. Anti -IFN-I positive patients by group (n/n total), sensitivity and specificity of assays.
4. The results and discussion section are good. Please consider adding a conclusion at the end, along with the clinical utility of this study.
The ‘Conclusions’ section was added to the manuscript:
Microarray-based assay showed almost 100% sensitivity and specificity in detection of anti- IFN-I antibodies in patients with endocrine disorders. Determination of antibodies to IFN-ω and IFN-α-2 can be used as an inexpensive and rapid way to diagnose APS-1. Thanks to these antibodies, the number of people who need to undergo genetic analysis can be reduced, i.e. in patients who do not have antibodies to IFN-ω and IFN-α-2, the diagnosis of APS-1 can be ruled out without testing for the AIRE gene. Also, this analysis may be recommended for individuals who do not fully meet the diagnostic criteria: patients who failed to detect a second mutation in the AIRE gene, or patients with dominant-negative mutations.