## Introduction

The advent of next-generation sequencing—first in the research environment and recently in diagnostic laboratories—provides a powerful tool to interrogate the exome, or even the entire genome, without a priori knowledge in pediatric patients with a disorder that has a suspected underlying genetic cause.1,2 The burden of genetic disease in child health is increasingly recognized, with an estimated 34% of inpatient children admissions having a clear genetic underlying cause, of which a significant and expanding proportion are identified as Mendelian disorders.3,4 Whole-exome sequencing (WES) enables examination of the coding part of the genome for variants in genes linked to Mendelian disorders. Trio-based WES of patients with suspected rare, monogenic disorders and their healthy parents has previously been demonstrated as a particularly effective method to find the causal variant in cases suspected to be caused by a de novo variant or due to autosomal recessive inheritance. WES gives value to parents by allowing timely interventions and altered management and providing information necessary to make reproductive choices.5,6,7,8,9 WES has a diagnostic advantage in situations where conventional single-gene or gene-panel tests may not be appropriate because a relevant genetic test has not yet been developed or because of genetic heterogeneity, incomplete or atypical clinical presentation, or lack of knowledge of the causal gene. In a research setting at the Sylvia Tóth Center (STC; Utrecht, The Netherlands), we performed trio WES in a particularly difficult-to-diagnose patient population that could especially benefit from this emerging technology. The STC is a multidisciplinary center that specializes in diagnosing children with intellectual disability, and a large proportion of the diagnoses are attributed to genetic causes.10

Prior to admittance to the STC, patients are traditionally evaluated at peripheral hospitals in an iterative approach in which subsequent tests are introduced after initial diagnostic tests have negative results. This approach is effective in solving cases with an easily recognizable etiology, but other patients must undergo a “diagnostic odyssey,” subjected to numerous hospitalizations, diagnostic tests, and procedures over the course of many years—possibly with no diagnosis. In the Netherlands, these patients eventually end up in tertiary-level diagnostic facilities such as the STC.

The patients referred to the STC with intellectual disability seem highly suitable for a WES-based diagnostic approach because single-gene disorders account for at least one-quarter of intellectual-disability cases.11 The present study compared the effectiveness of WES with that of traditional diagnostic investigations. We provide an extensive overview of the costs of the traditional diagnostic trajectory in this patient group. We also examine the cost savings that early implementation of WES would enable by rendering various genetic or metabolic tests unnecessary. The findings of this research can be used as input in the discussion regarding the societal, individual, and monetary value of next-generation sequencing.12 A recent study assessed comprehensive costs of the traditional diagnostic pathway in a different group of patients, with complex pediatric neurological disorders.13 However, a clear determination of the diagnostic yield when actually performing WES compared with traditional diagnostic procedures and costs was not made. Reliable information on all resources used in the current clinical and diagnostic pathway is crucial for clinical diagnostic centers considering implementing WES as a diagnostic tool. In addition, this information could serve as input for future value-based research for the diagnostic trajectory.

### Cost scenario analyses

In three hypothetical scenarios, we evaluated the impact of WES on cost savings with the assumption that WES would render certain diagnostic investigations unnecessary. If WES was performed as a first diagnostic approach for patients in our group who ultimately received a genetic diagnosis, cost savings of genetic testing and metabolic testing would average $4,986 (median:$5,342; range: $0–$10,684) and $2,533 (median:$2,446; range: $2,204–$2,866), respectively. For patients who did not receive a diagnosis following WES but for whom WES would replace genetic testing, savings would average $5,699 (median:$4,854; range: $890–$18,696).

In the final scenario, we assumed that WES would result in a 50% reduction of number and cost in the categories of health-care visits, imaging, biochemical tests, and patient day admission. For the diagnosed patients we calculated mean total savings of $1,660 and for the undiagnosed patients$4,269.

## Discussion

Implementation of WES in clinical practice is at a turning point. The application of WES in diagnostics has transformed the clinic and allowed massive interrogation of the entire coding region of the genome. The diagnostic yield has been proven in numerous studies, and the cost continues to fall. However, information is lacking on the prospective yield using WES compared with the cost of patient diagnostics retrospectively using the traditional diagnostic pathway. An initial study focused on retrospective cost analysis of patient resource use in a cohort of neurological patients and the potential application of WES.13 The current study not only links the costs of patient diagnostics using traditional means but also reveals the higher yield and lower costs when actually implementing WES in a heterogeneous, difficult-to-diagnose patient group. This knowledge is essential for clinical diagnostic centers considering WES implementation so that the cost for increased diagnostic yield can be ascertained and considered if WES is advantageous.

The yield from this study further confirms that WES is useful in the clinic. Current single-gene disorder analyses or chromosomal microarray analyses have a diagnostic rate of ~13–14% in these patient populations, emphasizing the need for new molecular technologies.26,27 Several previous reports using a trio-based WES strategy in large, specific patient populations reported diagnostic yields of ~25%, with some smaller studies reporting yields of up to 45–55%.7,20,28,29,30 The yield in this study is comparable to the 25% from these large patient population yields, and it demonstrates the power of WES to identify the genetic cause of disease in cohorts of intellectually disabled patients. Recent studies have reported a diagnostic yield of 37% using trio analysis via the inclusion of novel genes for which a variant was observed in only one patient.6 Our study’s yield does not include variants in entirely novel genes without the support of additional patients with similar phenotypes or functional work to establish causality, as previous reports have demonstrated that “overinterpretation” of the exome can lead to false positives.31 We expect that future essential patient data-sharing efforts and functional work will ultimately result in a higher yield for this patient cohort. Of note, the four previously known Mendelian genes that harbored diagnostic variants have only been known to be causal for these specific syndromes for less than 4 years, indicating the rapid progress in human genetics in the past few years. When this study was initiated, the underlying pathogenic mechanism for KBG (defects in ANKRD11) was still being elucidated and a diagnostic test had not yet been developed. Three of the genes—ADNP, CTNNB1, and SMARCB1—have only recently been associated with the respective syndromes. WES clearly has an advantage over traditional single-gene analysis in situations when a specific molecular test does not exist or for syndromes for which the genetic cause has only recently been discovered. For the cases that are still unsolved, the WES results can be reanalyzed periodically to determine whether a detected de novo variant can be linked to new discoveries.

Our study confirms the long traditional diagnostic trajectory and the high costs for traditional diagnostic testing in this patient population. We found a mean cost of $16,409 per patient that is comparable to other total costs recently published, albeit for a different patient group.13 The largest proportion of costs (42%) was related to previously performed gene tests. A previous study examining WES yield in a cohort of 12 patients also estimated the high amount of resources used, with a single patient’s laboratory investigations costing$22,000.32 High prices for additional diagnoses were also reported by Shashi et al.33 to be $25,000 per diagnosis if no diagnosis was obtained after a first visit. Soden et al.34 recently estimated that negative diagnostic tests for a group of neurodevelopmental disorder patients cost$19,100. In our study, patient 10 thus far has accrued a total of $47,841 in costs and still requires a diagnosis. For these patients, whole-genome sequencing is the next step in the diagnostic investigation to fully interrogate the genome. Utilizing a WES-first approach would have immediate cost savings for diagnosed as well as undiagnosed patients. With a WES cost of$3,972 compared with a mean cost of $4,986 for genetic tests and$2,533 for metabolic tests, using WES would directly save, on average, $3,547 per patient who receives a diagnosis and a mean savings of$1,727 for patients who do not receive a diagnosis using WES.

Other savings may also be realized. Before WES was introduced, the differential diagnostic workup required additional investigations (e.g., cardiac or renal ultrasounds; skeletal X-rays). If no such anomalies are reported in a genetic condition that has been diagnosed by WES, then there is no need to perform these additional investigations. These additional savings demonstrate not only that obvious genetic or metabolic tests can become redundant if WES is the first diagnostic approach but also that a proportion of other patient procedures could be omitted.

This research demonstrates that implementing WES as a first diagnostic tool for patients with intellectual disability, even in a tertiary center population, could reduce health-care costs because WES could replace a large number of genetic and metabolic investigations. Costs of WES are markedly lower than the average total traditional diagnostic trajectory costs; indeed, the cost of trio WES is already less than that of genetic investigations in all the patients in our study ($3,972 compared with$6,588). Minimally, WES would avoid genetic and metabolic tests in patients who receive a diagnosis and genetic tests in patients who do not receive a diagnosis. An additional proportion of savings would be realized, at a proportion that will vary depending on which auxiliary procedures or costs the use of WES will partially replace. Importantly, these scenarios require an initial critical stratification by a physician specialized in diagnosing patients with intellectual disability. Such a physician is able to recognize patients who have a clear clinical presentation suggesting a known underlying genetic cause that is not detectable by WES (e.g., fragile X, trinucleotide repeat, or methylation disorders) and for whom WES would not be beneficial and a waste of resources. This indicates that WES should be considered first for the majority of cases in which a genetic condition is strongly suspected.

The current study has several limitations. Because we evaluated this procedure as a pilot project, the randomly selected sample size of 17 patients is limited. Definite conclusions regarding where WES should be implemented in the diagnostic pipeline cannot be drawn on the basis of such small numbers. However, the diagnostic yield was consistent with other studies, suggesting that this yield range is what can be expected if the study is expanded to include more patients. Moreover, the small number enabled us to disentangle individual diagnostic odysseys and their economic consequences.

The cost analysis study was as comprehensive as possible, but some additional costs may exist. Only direct medical costs were taken into account in this study, and resource use collection (except genetic tests) was restricted to the UMC Utrecht.

Also, if WES were applied earlier to our patient group, there would be a clear, quantifiable cost savings of genetic and metabolic tests in patients who receive a diagnosis and savings of genetic tests in patients who do not receive a diagnosis. The reduction of these costs represents the minimum savings that would be realized with earlier WES introduction. Additionally, the contribution of the auxiliary proportion of savings in the categories of health-care visits, imaging, biochemical investigations, and day admission is debatable. For instance, the 50% cost reduction that we assume may be lower for undiagnosed patients whom further diagnostic investigations are necessary. Indeed, investigations, metabolic or otherwise, can aid in the interpretation of WES results or elucidating the disease pathogenesis.35,36 It is important for future analyses to consider these additional savings and correctly categorize diagnostic procedures that can be reduced by WES at the time of care to enable precise estimates of cost savings.

In conclusion, this study links the increase in diagnostically useful findings enabled by WES in intellectually disabled patients with the costs of traditional diagnostic investigations. The increase in causal variant detection and speed of diagnosis and the lower cost of WES compared with traditional diagnostic investigations in this patient population suggest that WES should be implemented early in clinical diagnostic centers with similar patient populations.

## Disclosure

The authors declare no conflict of interest.