Adverse childhood experiences and the development of Down syndrome regression disorder

Down syndrome regression disorder (DSRD) is a clinical symptom cluster of acute or subacute neurocognitive regression in otherwise health persons with Down syndrome. The objective of this study was to evaluate if adverse childhood experiences (ACEs) were more prevalent in children with DSRD than those with DS alone. A survey‐based, cohort‐based study was performed. Caregivers of individuals with DSRD with onset of symptoms between age 10 and 30 years and DS alone were administered the ACEs questionnaire via an online REDCap survey. A total of 159 responses were collected after excluding incomplete surveys and those not meeting criteria for DSRD. Individuals with DSRD were not more likely to experience ACEs (p = 0.18, 95% confidence interval [CI]: 0.43–1.17). In those with ACEs prior to the onset of symptoms, the median time prior was 7 months (interquartile range: 5–10). Individuals with DSRD were more likely to report three or more ACEs (52, 33%) compared to those with DS alone (39, 22%) (p = 0.02, 95% CI: 1.08–2.87). Exposure to ACEs were not predictive of response to particular therapeutic interventions although those with multiple ACEs 3 months prior to the onset of symptoms was associated with lower response rates to benzodiazepines and immunotherapy (p = 0.02, 95% CI: −3.64–−1.13). This study provides preliminary data that individuals with DSRD experience ACEs at a similar rate to individuals with only DS alone, although three or more ACEs, often preceding the onset of symptoms, was more prevalent in individuals with DSRD.


| INTRODUCTION
Down syndrome regression disorder (DSRD) is a clinical symptom cluster of acute or subacute developmental regression, catatonia, bradykinesia, cognitive decline, neuropsychiatric disturbance, and language deterioration in persons with Down syndrome (DS) (Mircher et al., 2017;Rosso et al., 2020;Santoro et al., 2020;Worley et al., 2015). The symptoms are severe and disabling and greatly impacts the quality of life in otherwise healthy persons with DS.
The etiology of DSRD remains unclear although emerging data has supported psychiatric, immunologic, and systemic causes Santoro, Baumer, et al., 2022;. Preceding environmental stressors have been reported in between 29% and 100% of individuals with DSRD Sargado et al., 2022). A variety of stressors including abuse, medical hospitalization, adolescent transitions, and changes in the home environment (moving, divorce, etc.) have been reported preceding the onset of DSRD, often occurring within months of symptom onset (Santoro, Baumer, et al., 2022;Sargado et al., 2022). Adverse childhood experiences (ACEs) are amongst the stressors that have been surmised to be related to changes in stable and consistent life structures. The temporal association between stressors and symptom onset in many individuals with DSRD is a justification for primary psychiatric/psychologic diagnosis and trial of therapeutics such as antidepressants, antipsychotics, and in severe cases, electroconvulsive therapy (Santoro, Baumer, et al., 2022;. However, the interface of stressors and immune dysregulation, an emerging cause of DSRD  is under-recognized and may be a unifying link among individuals with this symptom cluster (Dube et al., 2009;Eid et al., 2022).
To date, there have been no large-scale, controlled, studies evaluating the role of environmental stressors in the development of DSRD.
The specific aims of this study were to (1) evaluate the prevalence of ACEs in individuals with DSRD prior to the onset of symptoms and (2) to determine if the rates of ACEs in persons with DSRD are higher than in individuals with DS only.

| Approval and data availability
This study was approved by the institutional review board at the University of Southern California. Virtual consent was obtained by caregivers or guardians of all individuals. Anonymized data is available upon request to qualified researchers. The first page of this survey was a consent with viewing of questionnaire only after completion of consent/assent.

| Study population
Individuals were recruited from the Children's Hospital Los Angeles (CHLA) Down syndrome neurology clinic and via virtual recruitment on a Facebook support group for individuals and caregivers of persons with DSRD. The recruitment period lasted 8 weeks (August 1, 2022 to September 30, 2022). The only presurvey exclusion criteria in this study was age of onset <10 years or > 30 years.

| Study design
This study utilized a retrospective case-control design.

| Survey
Caregivers completed an anonymous, voluntary, "open," online survey via REDCap (Appendix A) on exposure to ACEs. The 10-question ACE Questionnaire (Felitti et al., 1998) was utilized. Five additional questions written in a similar binary (yes/no) format were added on to capture other previously reported stress-related provoking factors in DSRD Sargado et al., 2022). For individuals with DSRD, only ACEs prior to the onset of symptoms were queried whereas individual with DS only were asked about ACEs at any time in their lives up to the time of the survey. Total exposure period was thus determined as months between birth and the onset of symptoms for individuals with DSRD and months between birth and the completion of the survey for individuals with DS alone. For individuals with DSRD, respondents were asked to declare how many months prior to the onset of symptoms the ACEs were using whole numbers. Responses of zero indicated a <4-week time to onset. As the study was design was to identify exposures prior to the onset of DSRD symptoms, subsequent ACEs exposure (e.g., after symptom onset) was not assessed in this study in those with DSRD. There was no established mechanism for objectively evaluating the severity of ACEs and thus this was not performed. There was no reimbursement for participation in the survey offered.
Individuals without DSRD had their survey close out after completion of ACE, demographic, and clinical questions. Medical records were not utilized in this survey and caregiver or self-report of demographics, clinical information, and ACE exposure was relied upon. Surveys that were incomplete were not considered in this study. The order with which survey ACEs-related questions were presented was random although demographic and clinical data questions were offered in the same order for each survey. All questions were presented on one page with scroll down options enabled. Caregivers and individuals did not have an option to review responses prior to completion of the study. IP addresses were referenced to ensure that multiple completions of the survey were not performed. Cookies were not used to track responses. Completion rate was determined by dividing the number of completed surveys by the total number of consents obtained.

| Confirmation of diagnosis
All caregiver responses for individuals with DSRD were evaluated post-hoc to confirm that caregiver report of symptoms was consistent with consensus guidelines on the diagnosis of DSRD . The criteria for DSRD were reproduced verbatim in the survey and respondents were asked to report if those criteria were met at the time of onset of symptoms to minimize interpretation of if individuals truly met criteria or not. This was performed by an independent reviewer (MK) who only reviewed clinical responses and was not privy to demographic or ACE question responses. Per consensus guidelines, individuals were stratified as having "possible" or "probable" DSRD by the number of criteria they met and sub-analysis were subsequently performed on these groups. This was also used as a surrogate for disease severity as individuals meeting "possible" criteria required three or more symptoms whereas individuals meeting "probable" criteria required six or more symptoms. Individuals who did not meet either possible or probable criteria for DSRD were subsequently excluded. As the survey was anonymous, confirmation of clinical symptoms or further interrogation regarding symptoms was not possible.

| Statistical analysis
Descriptive statistics were produced for demographic and clinical data. Odds ratios (ORs) with corresponding 95% confidence intervals were used to calculate the strength of association between clinical data and ACE exposures and the diagnosis of DSRD. Cohen's kappa was used to assess reliability between caregiver/self-report of DSRD and meeting existing clinical criteria for DSRD . Multivariate analysis was used to evaluate the impact of ACEs subcategories on caregiver reported responsiveness to different types of treatments and included covariate adjustment for age, sex, race, ethnicity, maternal education level, and time to diagnosis. Therapy categories included benzodiazepines, antipsychotics, antidepressants, and immunotherapy. ACEs subcategories that were used for analysis included exposure to three or more ACEs at any time point prior to symptom onset, exposure to <3 total ACEs at any time point prior to symptom onset, exposure to an ACE <3 months prior to symptom onset, exposure to an ACE three or more months prior to symptom onset, and exposure to multiple ACEs <3 months prior to symptom onset. In addition, a multivariate regression analysis was utilized to determine if statistically significant demographic factors influenced likelihood of exposure to ACEs. A p nominal value of <0.05 was considered statistically significant for each statistical test.

Individuals with DSRD have clinical features reported in
The median age of symptom onset was 16 years (IQR: 14-19). Nearly, all individuals (92%) had symptom onset over a period of <12 weeks.
Only 8% reported symptoms ">12 weeks" to nadir although these individuals reported multiple symptom periods yielding results that qualified as acute deterioration was during a 12-weeks window (e.g., two periods of regression more than 12 weeks apart but both with symptom nadir <12 weeks). The majority of individuals with DSRD (79%) met post-hoc criteria for probable DSRD with the remainder meeting criteria for possible DSRD. A variety of different therapies were utilized in this study with the majority of individuals (n = 143, 89%) had utilized more than one therapy.  Table 3. The majority of individuals in both groups (72% in DSRD, 78% in DS alone) reported exposure to at least one ACE although there was no statistically significant difference between groups ( p = 0.18, 95% CI: 0.43-1.17). Stratification by total number of ACEs reported was similarly not significant ( p = 0.11, 95% Response to particular therapies (benzodiazepines, antipsychotics, antidepressants, and immunotherapy) was self-reported by caregivers and sub-analyzed by the total number and timing of ACEs exposures ( Figure 2). In individuals with ≥3 ACEs at any time point (n = 52), 88% of individuals were responsive to immunotherapy compared to 72% for benzodiazepines, 59% for antipsychotics, and 45% for antidepressants ( p = 0.03, 95% CI: 1.23-3.17). In those with <3 ACEs at any time point (n = 107), these rates were 74%, 68%, 52%, and 57%, respectively ( p = 0.05, 95% CI: 1.06-2.12). Individuals with an ACEs exposure <3 months prior to presentation (n = 43), 92% of individuals were responsive to immunotherapy compared to 64% for antipsychotics, 60% for antidepressants, and 59% for benzodiazepines (p < 0.001, 95% CI: 2.14-3.77). In those with ACEs exposures >3 months prior to presentation (n = 116), these rates were 78%, 48%, 51%, and 73%, respectively (p = 0.02, 95% CI: 1.38-6.31).
Subanalysis of responses to ACEs questionnaires was performed by "possible" and "probable" diagnostic criteria for individuals with DSRD (Table 4). There were no statistically significant differences in response rates between individuals with less symptomatic "probable" DSRD compared to more symptomatic "probable" DSRD. onset of symptoms. In addition, exposure to multiple ACEs was not predictive of more clinical symptoms (defined as having "probable" vs. "possible" DSRD). While this could suggest that individuals who experience multiple ACEs within 3 months of their regression are more likely to have psychological or psychiatric etiologies for their symptoms, this study was not designed to assess this.
Exposure to environmental stressors has been linked to both the development of psychiatric disease (Carbone et al., 2019;Kasznia et al., 2021;Lakkireddy et al., 2022) and a variety of other medical disorders, including those of the brain and the immune system (Dube et al., 2009;Eid et al., 2022;Roberts et al., 2022). An emerging concept of how these stressors may be related to health outcomes, whether they be psychiatric or medical, is that ACEs may contribute . This is even more relevant when we consider that rates of response to psychiatric interventions (e.g., electroconvulsive therapy) and immunologic interventions (e.g., IVIg), both of which are >70%, and have limited overlap in therapeutic indications . In this study, immunotherapy was least effective in patients with multiple ACEs in the 3 months prior to symptom onset, which could be reflective of these cases having a more psychiatricdriven etiology. Interestingly, immunotherapy was quite effective in individuals with more than three lifetime ACEs at any time point, which is not as easily explained from a psychiatric standpoint. As we continue to learn more about the etiology of DSRD, investigation to the role of high allostatic load and the dysregulation of Percent responding to therapy Therapy type > 3 ACEs < 3 ACEs ACE < 3 mo ACE > 3 mo MulƟple ACEs < 3 mo F I G U R E 2 Percentage of individuals with Down syndrome regression disorder and multiple ACE exposures responding to a therapeutic intervention categorized by ACEs subcategory. Individuals exposed to multiple ACEs within 3 months prior to symptom onset were less likely to respond to benzodiazepines and immunotherapy. The total number of ACEs and single exposure to ACEs within or after 3 months of symptom onset did not influence response to therapy. * = p value of 0.02. ACEs, adverse childhood experiences.
neuroendocrine system should be highly prioritized as optimized therapeutic interventions could be identified from this research. This is of particular importance given the established role of interferon dysregulation in persons with DS (Kong et al., 2020;Sullivan et al., 2016;Waugh et al., 2019), a system that can potentially be impacted by stress (Tripathi et al., 2021). This could be a possible explanation for why immunotherapy was particularly effective in individuals with more than three lifetime ACEs but not in those with multiple close temporal stressors. It is important to note that while exposure to mul- inclusion of an individual's data with missing reported ACEs response similarly could have created a bias toward completing ACEs questions listed earlier in the survey. In this particular cohort of individuals with DSRD, the Hawthorn effect (e.g., individuals seeking to acknowledge the role of stress in DSRD) could have also impacted study participation. This study relied on individual and caregiver responses with regards to medical history and clinical symptoms introducing recall bias into this data as well. This was partially mitigated by the use of a reviewer confirming that DSRD criteria were met in the study although the authors acknowledge that this is an imperfect post-hoc strategy. It is possible that multiple questions in the survey could have captured the same stress-related event, thus increasing the report of ACEs in both cohorts. This would not be anticipated to impact one group more than the next but may explain why the total number of ACEs was not elevated in persons with DSRD whereas simply having greater than three ACEs was. The survey tool (ACE measure) utilizes a yes/no format that was insufficient to capture severity, frequency, timing or impact of stress-related sequelae and is a major limitation of this study. Given the data obtained in this study, natural extension to investigate the features of these ACE exposures will be beneficial.
The control cohort, which represented a regional (California) sample differed from the national scope of the individuals with DSRD and is limited by being driven by a sample of convenience. For instance, lower maternal education levels (a surrogate biomarker for socioeconomic status) was present in the control group and may have yielded a comparator cohort with a higher likelihood of stressful exposures.
Another potential confounder in the data obtained on responsiveness to particular therapies is that this was not defined and was subjective for the caregiver completing the survey. This could lead to overinterpretation of response to some therapies given inability to collect objective clinical data in this study. The authors also appreciate the potential for acquiescence bias in which caregivers may have known the purpose of this study was to specifically investigate ACEs in the setting of DSRD and thus could influence the likelihood of responses.
In addition, the data presented may be influenced by attrition bias wherein caregivers are likely to search for a cause for particular symptoms associated with disease and could influence the likelihood of overreporting of the temporal relationship of stressors with symptoms. As this study did not involve direct contact with patients (e.g., phone or clinical encounter), the risk of ascertainment bias is thought to be lower. Although multivariate regression analysis did not change statistical significance regarding overall rate of exposure to ACEs or the likelihood of being exposed a cluster (three or more ACEs) this is a limitation of the study and could be reflective of differences in likelihood of endorsement ACEs at different maternal education levels. This finding is compounded by the fact that the age and sex of the caregiver completing the survey was not obtained which could have also influenced likelihood of report of ACEs as well. In addition, as individuals in the cohort group were largely recruited locally from a neurology clinic, an overrepresentation of certain medical disorders such as epilepsy was present in the comparator group.
While it is uncertain how this would affect exposure to ACEs, it should be noted that adults with neurologic disorders have been reported to have a higher number of ACEs exposures (Mendizabal et al., 2022).

| CONCLUSION
This study provides preliminary data that individuals with DSRD experience ACEs at a similar rate to individuals with only DS alone although having more than three ACEs, often preceding the onset of symptoms, was more prevalent in individuals with DSRD. Stress and ACEs could play a role in psychiatric and neuroimmunologic origins of DSRD although the exact pathogenic mechanisms remain unknown.
Future study focusing on the immediate and long-term impact of stressors on objective biomarkers of immune dysregulation (e.g., cytokines) in persons with both DS and DSRD would be ideal to link objective biochemical data to the stress response in these populations.

CONFLICT OF INTEREST STATEMENT
The authors report no conflicts of interest in the submission of this manuscript.

DATA AVAILABILITY STATEMENT
The data that support the findings of this study are available from the corresponding author upon reasonable request.