Long term outcomes in children with trichohepatoenteric syndrome

Trichohepatoenteric syndrome (THES) is a rare autosomal recessive disorder caused by mutations in either TTC37 or SKIV2L, usually leading to congenital diarrhea as part of a multisystem disease. Here, we report on the natural history of the disease for the largest UK cohort of patients with THES from 1996 to 2020. We systematically reviewed the clinical records and pathological specimens of patients diagnosed with THES managed in a single tertiary pediatric gastroenterology unit. Between 1996 and 2020, 13 patients (7 female and 6 male) were diagnosed with THES either by mutation analysis or by clinical phenotype. Two patients died from complications of infection. All patients received parenteral nutrition (PN) of which six patients were weaned off PN. All patients had gastrointestinal tract inflammation on endoscopy. Almost half of the cohort were diagnosed with monogenic inflammatory bowel disease (IBD) by the age of 11 years, confirmed by endoscopic and histological findings. Protracted diarrhea causing intestinal failure improves with time in all patients with THES, but monogenic IBD develops in later childhood that is refractory to conventional IBD treatments. Respiratory issues contribute to significant morbidity and mortality, and good respiratory care is crucial to prevent comorbidity.


| INTRODUCTION
Trichohepatoenteric syndrome (THES) is a rare autosomal recessive disorder that usually presents with protracted diarrhea during infancy.
Global prevalence is estimated at 1 in 1,000,000 live births (Fabre et al., 2013).
Mutations in two genes-tetratricopeptide repeat domain 37 (TTC37) and SKIV2L have been identified as the cause of THES (Fabre et al., 2012;Hartley et al., 2010).These genes encode cofactors of the human Ski complex, a multiprotein complex required for cytoplasmic exosome-mediated RNA surveillance (Tuck et al., 2020).
During infancy, patients often require parenteral nutrition (PN) to support growth and development.Some patients subsequently develop monogenic inflammatory bowel disease (IBD) (Busoni et al., 2017;Kammermeier et al., 2017;Uhlig et al., 2021) that is often refractory to conventional IBD treatments.
Clinical presentation however varies significantly; some patients do not develop intractable diarrhea, previously thought to be a hallmark of THES (Karaca et al., 2019;Poulton et al., 2019;Rider et al., 2015).Patients with SKIV2L mutations may be more severely affected than patients with TTC37 mutations (Bourgeois et al., 2018).
Our study reports on the long-term outcomes of the largest UK cohort of patients with THES.

| Editorial policies and ethical considerations
This study was registered as a service evaluation project under registration number CARMS-30202 at Birmingham Children's Hospital.
Consent for anonymized data usage was granted by our governance service unit.Local research ethics committee approval was not required.

| Data collection and analysis
We systematically reviewed the case notes and electronic records for all patients with THES (n = 13) in our center between 1996 and 2020.
We recorded the following data: anthropometry, clinical features, laboratory findings including baseline immunology (lymphocyte subsets, immunoglobulins, and functional antibodies), mutation analysis, histopathology results, endoscopic findings, mode of nutritional support and treatment modalities for diarrhea, and bowel inflammation.Weight-for-age and height-for-age Z-scores were calculated using World Health Organization child growth standards for 0-5 years and growth reference data for 5-19 years.
All patients had characteristic facial features of THES.Trichorrhexis nodosa was found in 11/13 (85%) of patients.Two patients had earlier hair samples that were negative for trichorrhexis nodosa, before later having positive samples.Two patients have to date had negative microscopy for trichorrhexis nodosa; both have genetically confirmed THES.
Seven patients had duodenal disaccharidase measurement.Lactase levels were low in 5/7 (71.4%), while sucrase levels were normal in all 7 patients.
All patients had evidence of gastrointestinal (GI) tract inflammation, with most patients having both upper and lower GI involvement.
Table 2 describes the endoscopic and histologic findings in our patients.To compare the changes over time, we separated our findings into two epochs-early childhood (<6 years) and later childhood (6-17 years).
Graft-versus-host disease (GvHD)-like changes, including increased apoptosis, villous atrophy and eosinophilia were seen more frequently in early biopsies compared to biopsies in later childhood.
Six patients developed a secondary GI phenotype of IBD in later childhood, confirmed by endoscopic and histologic findings.Symptoms included vomiting, diarrhea, abdominal pain, oral and anal ulceration, and GI bleeding.Endoscopic findings leading to a diagnosis of IBD included mucosal inflammation, ulceration and lower GI strictures.

| Medical therapies
Medications used in THES included medications to address upper GI symptoms, diarrhea, and immunosuppressive treatment (Table 4).
5-Aminosalicylates were ineffective-all patients had discontinued it at last review.For two patients, there were improvements in bowel frequency after a steroid course and this was repeated to good effect in these patients.Conversely, diarrheal frequency was unchanged in two other patients.
Sirolimus and racecadotril were trialed in one patient; both were discontinued due to lack of response.One patient received courses of exclusive enteral polymeric nutrition for treatment of IBD with no symptomatic improvement.Hepatosplenomegaly improved in some patients-on last ultrasound scan, 6/13 (46%) had persisting hepatomegaly and 3/13 (23%) had persisting splenomegaly.
T A B L E 2 Endoscopic and histologic findings.
One patient required cardiac surgery for coarctation of aorta; the other four patients were managed conservatively.
One patient underwent a nasal ciliary brushing study due to persistent respiratory symptoms This revealed well-ciliated epithelium, however with a high degree of ciliary dyskinesia.The patient did not fit into a typical diagnosis of primary ciliary dyskinesia as there were areas of normal cilia.
One patient had bronchiolitis obliterans secondary to adenovirus, resulting in prolonged ventilation, intensive care admission, and subsequent requirement for home oxygen.
6/13 (46.2%) received prophylactic antibiotics-five for respiratory prophylaxis and one post-splenic infarction.One patient had chronic pseudomonas colonization and received nebulized colomycin.

| Immunology
All patients had serial immunoglobulin level testing.7/13 (54%) had low IgG levels on initial testing done at a median age of 4.6 months (IQR 3-7.4 months).
Three patients received immunoglobulin supplementation for a median of 1.7 years (IQR 1.2-4.4years).Reason for immunoglobulin supplementation was low IgG levels in two patients, and as adjunct treatment in severe pneumonitis in another patient.
Supplementation was discontinued at a median age of 3.8 years (IQR 3.1-5.6years) after patients were evidenced to maintain immunoglobulin levels without supplementation.
T A B L E 3 Inflammatory bowel disease in trichohepatoenteric syndrome.

| DISCUSSION
THES was first identified in 1982 (Stankler et al., 1982).Until the identification of the causative gene mutations, the diagnosis was made based on the typical clinical presentation of intractable diarrhea from infancy, characteristic facial appearances, trichorrhexis nodosa, failure to thrive, and liver disease; hence why THES has also been known as syndromic or phenotypic diarrhea.
Diagnostic genomic testing through exome and whole genome sequencing, with the utilization of virtual gene panels, is becoming increasingly available in clinical practice (Kammermeier et al., 2023).
This can aid earlier identification of THES, particularly when the clinical phenotype has not fully developed.
To our knowledge, this is the largest reported cohort of patients in the UK with THES.Given the rarity of the disease, information about the long-term outcomes for THES will be valuable in guiding medical management.
Our study highlights the heterogeneity of clinical findings and outcomes in THES, in line with previous reports (Alsaleem et al., 2021;Bourgeois et al., 2018;Fabre et al., 2014).
While patients normally present early in life, two patients presented after the age of 5 years, having survived their early years without PN, albeit with poor growth.Both patients had homozygous TTC37 c.2921-2A>G mutations, potentially reflecting a milder clinical phenotype.Only one other case with similar mutations has previously been reported (Bozzetti et al., 2013).
One patient had homozygous TTC37 c.1135-2A>G mutations-to our knowledge, this represents a novel mutation not previously reported.
In our cohort, no patients had end-stage chronic liver disease, a significant cause of mortality in previous reports (Girault et al., 1994;Hartley et al., 2010).However, deaths resulted from respiratory sepsis.Children with THES do survive into adulthood, with the oldest patient in our cohort at 24 years of age.

| High rate of respiratory infection and comorbidity in THES
Previous reports have shown infection as the leading cause of mortality in THES (Alsaleem et al., 2021;Girault et al., 1994).We highlight specifically the high burden of respiratory comorbidity and the importance of good respiratory care.Indeed, both deaths in our cohort resulted from respiratory infection and complications thereof.
In our center, THES patients with respiratory complications have regular respiratory care input, with physiotherapy, nebulizer treatment, and prophylactic antibiotics forming the bundle of care.
It is crucial that THES patients receive regular immunology input.
We recommend monitoring immunoglobulin levels and functional antibody testing to identify immune dysregulation.This should be proactively managed, with consideration of immunoglobulin infusions and revaccination where appropriate to prevent future morbidity.

| Two distinct GI phenotypes in THES
We observed two distinct, often overlapping GI phenotypes in our cohort of patients.The first phenotype presents usually in early infancy with intractable secretory diarrhea, resulting in intestinal failure and the need for PN.Histological changes are described as GVHD-like, with villous atrophy, eosinophilic infiltration, and apoptosis.
An IBD phenotype then develops in later childhood, presenting with the clinical, endoscopic and biochemical features of IBD.Patients develop typical signs and symptoms of IBD, including mucosal ulceration, GI bleeding and intestinal strictures.We hypothesize that the development of IBD represents an evolution of the ongoing immune dysregulation in these patients.
We observed that standard IBD management, including biologic therapy, has limited efficacy.Exclusive enteral nutrition, a first-line treatment modality in pediatric Crohn's disease, does not result in disease remission.
In our experience, the infantile diarrhea in THES was not steroidresponsive but the colitis that develops in older children can respond to steroids, with reductions in bloody diarrhea and inflammatory markers.

| Weaning of PN is not permanent
Consistent with previous reports, patients with THES require PN in early life, but can be weaned off in later life (Fabre et al., 2017).Being weaned off PN is not permanent, and patients may require PN temporarily again in adolescence to proceed through puberty.This phenomenon has also been observed in patients with short bowel syndrome (Poulton et al., 2023).
There are no clear feed recommendations.Many THES patients had lactase deficiency; however, this is relatively common in Asian people (Storhaug et al., 2017) and unlikely to be a specific manifestation of THES.To facilitate PN weaning, oral diet is encouraged.Continuous enteral feeding regimens can be instituted and is almost always necessary to achieve nights off PN.
LEE ET AL.

| Strengths and limitations
Our study provides valuable longitudinal information on the natural history, management strategies and clinical outcomes of THES.Most published literature report on patients at a single timepoint; our study describes in detail the evolution of the intestinal and extra-intestinal manifestations of THES over time.
Our study has several limitations.Due to its retrospective design, some data were incomplete or unavailable.One surviving patient has not completed genetic confirmation of THES.Effects of prescribed medications were often not well-described, and efficacy (or lack thereof) was inferred by whether the patient continued the medication.Last, only a small number of patients (n = 13) are reported, due to the rarity of THES.

| Future directions
The UK prevalence of THES is unknown.Most children with THES in the UK are managed in tertiary pediatric gastroenterology centers offering home PN services.In 2019, there were 25 such centers, and 39 children with congenital enteropathies received home PN (Wiskin et al., 2021).Further multidisciplinary collaboration among these centers should be encouraged to broaden our understanding of disease phenotype, prevalence and natural history, and to improve clinical outcomes.
The molecular mechanism by which loss-of-function mutations in the TTC37 and SKIV2L genes translate into a multisystem disease with diarrhea, mucosal inflammation and extra-intestinal symptoms is still poorly understood.The Ski complex plays an important role in RNA regulation and decay (Halbach et al., 2013) while SKIV2L may have a role in the development of certain autoimmune disorders (Eckard et al., 2014).Transcriptome analysis, that is, the study of the complete set of genomic RNA transcripts, may shed further light on disease mechanisms (Vancamelbeke et al., 2017).
No effective agents currently exist to ameliorate the IBDassociated mucosal inflammation in THES.Conventional immunosuppressive treatments have only a limited effect.Better understanding of the molecular mechanisms linking the dysfunction of cellular RNA machinery to immune regulation and other specific biological processes will aid the discovery of new biomarkers and therapeutic targets for new drugs.Advances in high-throughput screening for the rapid testing of new biological compounds also provide hope that effective therapies can be discovered (Jardine et al., 2020).

| CONCLUSIONS
Our study provides important information on the long-term outcomes in THES and will assist with counseling families for the future.
Although THES remains associated with significant morbidity and mortality, most patients can expect to survive into adulthood.With longer survival, monogenic IBD has been identified as an important feature of the THES phenotype during later childhood.There is a high rate of respiratory comorbidity, and therefore multidisciplinary care should ensure close respiratory and immunology support, in addition to gastroenterology, nutrition, and hepatology care.

Microsoft
Excel was used to tabulate data.IBM SPSS Statistics for Mac, version 26 (Armonk, NY) was used to perform statistical analysis.Continuous data are presented as medians with interquartile ranges.Categorical data are presented as rates and proportions.
-37 weeks) and median birth weight of 1440 g (IQR 1340-2035 g).Most were small for gestational age (<10th centile for weight) with median weight-for-age Z-scores of À2.4 (IQR À3.2 to À2.0).Ten patients (77%) were of Pakistani Mirpuri descent, one of Pakistani Gujrat descent, one of Nepali descent, and one of Kurdish descent.11/13 (85%) were offspring from consanguineous unions.Two patients were siblings.Eleven patients had genetically confirmed TTC37 mutations; the remaining two were diagnosed clinically.No SKIV2L mutations were detected.The two patients who did not undergo confirmatory genetics had a clinical phenotype consistent with THES and were therefore included in this study.Patient 10 unfortunately died before confirmatory genetics could be done.The second patient (Patient 5) was the sibling of a patient with genetically confirmed TTC37 (Patient 12).Two patients died.Patient 2 died at age 10 years due to respiratory failure with sepsis after co-infection with influenza and measles.Patient 10 died at age 2 years due to acute respiratory distress syndrome secondary to sepsis; no causative organisms were identified.

3. 5
| Abdominal surgery 7/13 (54%) patients had insertion of a percutaneous endoscopic gastrostomy.4/13 (31%) underwent inguinal hernia repair.Two patients underwent laparotomy for bowel obstruction-one due to meconium ileus and one due to adhesions post-splenic infarct.No patients required bowel resections.3.6 | Nutrition and growth All patients received PN.Median age at PN initiation was 7.2 months (range 5 months-7.4years).Of the 11 surviving patients, six patients have successfully stopped PN.Nine patients had their first trial off PN at a median age of 4.4 years (IQR 4.1-11.1),six successfully.Indication for stopping PN was improvement in enteral tolerance as judged by the nutritional support team.PN infusions were gradually reduced to give cyclical infusions 2-3 nights per week in all before stopping.For the six patients who have stopped PN, median age PN stopped was 8.2 years (IQR 5.3-10.4years).Median PN length was 4.3 years (IQR 3.8-6.4years).The five patients continuing PN receive cyclical infusions 3-4 nights/week.Two have never stopped PN.The other three had a period of stopping PN ranging between 0.4 and 11.4 years.Reasons for restarting PN were weight loss in one patient and optimization of pubertal growth in two patients.FI G U R E 1 Probabilities of survival based on the Kaplan-Meier curve.Almost all surviving patients (10/11) were receiving additional nutritional supplementation at last clinical contact, with eight patients on continuous overnight enteral feeds.In terms of feed formulation, five received whole protein supplements, three were on peptide-based feeds, and two received a combination of whole protein and peptide-based feeds.All patients have shown feeding aversion, despite encouragement of oral diet and timely weaning.All patients were malnourished prior to PN initiation, with median weight-for-age Z-score at PN initiation of À5.3 (IQR À6.8 to À4.8).Median change in Z-score from birth centile to PN initiation was À3 (IQR À4.8 to À2).All 11 surviving patients showed improved weight indices at last clinical encounter; median weight-for-age Z-score was À2.3 (IQR À2.8 to À2.1).One patient received growth hormone treatment for several years but did not respond.3.7 | Hepatology10/13 (77%) patients had evidence of liver involvement (transaminitis, liver fibrosis, or hepatosplenomegaly).Five patients underwent liver biopsy at a median age of 6.8 months (range 2 months-3.4years).Of these, four patients had early evidence of liver fibrosis on liver biopsy, while one had florid hepatitis with microgranulomas.Ten patients developed hepatomegaly while seven patients had splenomegaly.One patient developed massive splenomegaly and subsequently a splenic infarct.No patients developed esophageal varices or ascites.No patients progressed to end-stage liver failure.
Table 1 demonstrates the patient demographics.

Table 3
Medications used in trichohepatoenteric syndrome.