Phenotype of Very-Early-Onset In ammatory Bowel Disease with Interleukin-10 Receptor A Mutations in 22 Chinese Children

Dexiu Guan Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Jing Zhang Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Shu Guo Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Feihong Yu Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Jin Zhou Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Guoli Wang Beijing Children's Hospital, Capital Medical University, National Center for Children's Health Xiwei Xu (  xuxiweibch@163.com ) Beijing Children's Hospital, Capital Medical University, National Center For Children's Health

Statistics were analyzed using SPSS 22. Continuous data were presented as mean and SD or median and interquartile range (IQR). Categorical variables were reported as frequency and percentage. P < 0.05 was considered statistically signi cant.

Ethical approval
This study was approved by the Ethics Committee of Beijing Children's Hospital, Capital Medical University.

Demographic Features
In this study, 22

Characteristics of the IL10RA Mutations
Among these patients, 81.8% (18/22) of patients had onset of disease within 1 month after birth, all of these patients showed symptoms within 6 months of life, the median age of onset of symptoms, including diarrhea, fever and oral ulcer, was 8.5 (IQR: 3.0-24.0) days. Regarding the rst symptoms experienced by the patients, diarrhea, fever and oral ulcers were experienced in 81.8% (18/22) The average weight-for-age Z score and height-for-age Z score on initial diagnosis were -2.07(IQR: -2.61~-1.78) and -1.74±1.34 respectively.
Colonoscopy revealed that colonic aphthous and deep ulcers and deep were common (85%, 17/20), and cobblestone change was found in 5 patients (25%, 5/20). Typical histopathological evaluation showed unspeci c acute or chronic colitis with multiple ulcerations, crypt structural deformation, cryptitis, and crypt abscess. As of April 1, 2020, 2 patients lost to follow-up, 9 patients were dead, the mortality rate was 45% (9/20), and 11 patients (55%, 11/20) were alive. Among the 11 patients, 2 patients remained stable and showed no gastrointestinal symptoms after UCBT, 1 patient achieved sustained clinical remission receiving thalidomide, 2 patients were clinically improved with thalidomide, and clinical improvement was also achieved in 1 patient with exclusive enteral nutrition (EEN). The remaining 5 patients still had recurrent disease activity. None of the patients developed lymphoma.

Discussion
Since the initial report patients with IL10R signaling defects in 2009 by Glocker et al [8], more than 130 VEO-IBD patients with IL-10 or IL-10R gene mutations have been reported [10,11]. Most patients were from Europe and Asian, where IL-10R mutations were more frequent than IL-10 mutations. In this study, we identi ed 22 VEO-IBD patients with IL10RA gene mutations. In general, VEO-IBD patients with IL-10RA mutations suffered from even more complicated, severe, and intractable disease. They were characterized by very early onset symptoms, common perianal diseases (abscess, stulas, ssure, skin tag) and extra-intestinal manifestations, such as skin rash and oral ulcers, respond poorly to traditional treatment, and high surgery rate and mortality [11].
In this study, we identi ed the c.C301T (p.R101W) mutation in exon 3 and the c.G537A (p.T179T) mutation in exon 4 were the most common mutations in the IL10RA gene in Chinese VEO-IBD patients, with a mutation rate of 86.4% and 36.4%, respectively. This was consistent with previous reports from East Asia, but signi cantly higher than other regions [11][12][13][14]. These mutations may disrupt signaling after activation of the IL-10 receptor, and therefore, STAT3 was not phosphorylated and intractable in ammatory responses developed in the gut of pediatric patients [15,16]. In addition, we found that 6 patients (27.3%, 6/22) had same compound heterozygote IL10RA p.R101W/p. T179T mutations, and these patients were born in unrelated families. This was similar to study from China before [17]. In this study, 1 novel mutation of IL-10RA was identi ed c.635G>C (p.R212P), we did not perform a functional study on this mutation due to conditional constraints. This novel mutant site was not found in HGMD or the other databases and was predicted to be deleterious. Therefore, we speculated that this novel mutation formed a compound heterozygous mutation with the c.C301T (p.R101W) mutation, thereby causing the patient 20 to develop clinical symptoms.
In our study, we did not nd mutations in IL10RB or IL10 in our patients, which was consistent with previous reports from East Asia. IL10 RA mutation was more common in East Asian patients than in Western countries, where comparable numbers of IL 10RA and IL 10RB mutation cases exist [18,19]. Furthermore, 81.8% of our patients (18/22) were compound heterozygous mutations, as they were born to unrelated nonconsanguineous families. Which was in contrast to North America and Europe, where the patients were born to consanguineous parents and had homozygous mutations. The high rate of consanguinity in the Arabic population resulted in a high incidence of IL10R homozygous mutations and that associated with the prevalence of VEO-IBD [20]. The genetic architecture of East Asia may be different compared to other regions.
VEO-IBD patients with IL-10RA mutations had extremely early onset of symptoms, all of these patients showed symptoms within 6 months after birth, the median age of onset time was 8.5 days. These patients had refractory diarrhea, hematochezia, fever and growth retardation. In addition, these patients might have recurrent perianal lesions such as abscesses, stulas, skin tag and ssures, which was consistent with previous researches that patients had extremely early onset of symptoms and multiple perianal lesions [21,22]. In our study, 77.3% (17/22) of our patients suffered from multiple perianal lesions, only 5 patients did not suffer from perianal disease. For extra-intestinal manifestations, oral ulcers and skin rash were very common in these patients, accounting for 72.7% and 63.6%, respectively. Failure to thrive was relatively common in these patients because of the earlier onset, more severe disease course, di culty in achieving remission, reduced food intake and increased nutritional requirements [23]. In this study, 3 patients had a positive family history that included their siblings dying from similar symptoms.
All the patients in this study were clinically and endoscopically diagnosed with CD. It was interesting to note that VEO-IBD patients with IL-10RA mutations predominantly presented as isolated colitis with deep ulcerations and cobblestone change. However, only 9.1% (2/22) of the patients had upper gastrointestinal or small bowel involvement. This suggested that the colon was particularly prone for immune disorders, which was consistent with other studies involving Turkish and whites [4,18]. Moran CJ et al. reported that the IL-10RA polymorphisms were associated with early-onset UC [24]. These ndings suggested that identifying IL-10R mutations may be more important than classifying the disease types (CD or UC) in VEO-IBD patients.
Current therapeutic strategies for pediatric IBD include the use of mesalazine, sulfasalazine, EEN, corticosteroids, immunomodulators (azathioprine, 6-mercaptopurine, methotrexate), and anti-TNFantibodies. In this study, the effects of mesalazine, steroids, thalidomide, immunosuppressive agents and biological agents were limited. These were consistent with reports from Western countries [21,25]. In our study, 1 patient achieved sustained clinical remission and 2 patients were clinically improved with thalidomide. This was consistent with Veenbergen S et al. report that thalidomide could promote clinical remission of intestinal symptoms in patients with IL10RA mutation [26]. It was interesting that in our study 1 patient with EEN also achieved clinical improvement, this was consistent with Talya L. Miller et al who described two infants presenting with VEO-IBD achieved clinical remission using EEN [27]. However, thalidomide was currently considered to be a potentially effective treatment for the VEO-IBD patients with IL10RA mutations, but long-term follow-up assessment was still required. In our study, 2 patients underwent UCBT and remained stable after UCBT. As reported before, Hematopoietic stem cell transplantation (HSCT) was a curative treatment for VEO-IBD patients with IL-10 or IL-10R mutations [11,13]. However, high risks and high costs should be taken into considerations, and long-term follow-up observations were required. Therefore, some patients from our study refused transplantation.
Our results showed that 18.2% (4/22) of the patients had surgical intervention. Due to poor treatment e cacy, some patients with IL-10RA mutations have had to undergo intestinal resection and ileostomy or colostomy [21]. Shim JO et al also reported that patients with IL-10RA mutations were associated with infantile onset, perianal stulae and early surgical interventions [15].
VEO-IBD patients with IL10RA mutations responded poorly to conventional therapy and had a higher mortality rate. Our study showed a high mortality rate of 45% (9/20). Huang ZH et al reported an overall mortality of 21.4% in patients with IL10RA mutations in China [17]. The study by Mathurin Fumery et al [28] showed that the mortality rate of pediatric CD patients was 0.93%, with a median follow-up time of 11.1 years [IQR, 7.3-15.0]. Despite aggressive treatment, VEO-IBD patients with IL10RA mutations still had a higher mortality rate than pediatric IBD. The high mortality rate in our study was due to severe disease course and several patients gave up treatment.

Conclusions
VEO-IBD patients with IL10RA mutations were phenotypically and genetically distinct from pediatric IBD. We provided the clinical phenotype, endoscopic results, medical therapy, surgical interventions and genetic characteristics of VEO-IBD patients with IL10RA mutations. Therefore, we should consider genetic defects in the IL-10 signaling pathway including IL-10RA in VEO-IBD patients, particularly when they had early onset of symptoms, perianal stulae and severe colitis.

Declarations
Ethics approval and consent to participate This study was reviewed and approved by the Ethics Committee of Beijing Children's Hospital, Capital Medical University.

Consent for publication
Not applicable.

Availability of data and materials
The datasets generated during and/or analysed during the current study are available from the corresponding author on reasonable request.

Competing interests
The authors declare no con icts of interest.

This work was supported by Beijing Municipal Administration of Hospitals Gastroenterology Discipline
Collaborative Development Center Digestive Subproject of major project (XXZ0504).

Authors' contributions
Xiwei Xu designed the research and reviewed the paper; Dexiu Guan collected the data and wrote the paper; Jing Zhang and Shu Guo were responsible for follow-up; Feihong Yu, Jin Zhou, Guoli Wang collected and analyzed the data. All authors read and approved the nal manuscript.