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DBA genotype-phenotype associated with RPL35A
Figure 1. Study population and types of RPL35A variants in patients with Diamond-Blackfan anemia (DBA). NCI: National Cancer Institute Inherited Bone Marrow Failure Study; DBAR: DBA registry; HGMD: Human Gene Mutation Database; Alabama Children’s: Arkansas Children’s Hospital; Boston Children’s: Boston Children’s Hospital; UTR: untranslated regions; ClinVar: ClinVar public data archive.
available databases and obtain in silico prediction scores.36 The St. Jude Cloud PeCan tool was used to create lollipop plots.37 For large deletions, UCSC Table Browser was used to identify all deleted genes in that region.38
Genotype-phenotype statistical analysis
The cases for the genotype-phenotype analysis were made up exclusively from the ten collaborating institutions (Figure 1). The cases identified through literature review had limited to no avail- able clinical data and were included in RPL35A variant characteri- zation. All statistical analyses used the individual case as the analysis unit. We compared 21 different clinical features between cases with large deletions versus all other pathogenic variants using Fisher exact test or Student t-test (Stata 15 software39). Bonferroni correction for 21 tests was applied. P<2.4x10-3 were considered significant.
Results
Cohort clinical characterization and genotype-phenotype relationship
We compiled a total of 45 cases of DBA caused by RPL35A variants from ten different institutions/registries after exclusion of four cases with variants of unknown sig- nificance (Figure 1). Table 1 shows the clinical features of the 45 patients from 41 different families. Fifteen of these cases have been published previously (Online Supplementary Table S3).10,14,18,40-42 Nearly half of the patients (n=22) had large deletions involving the entire RPL35A along with addi- tional genes deleted in the region. All other pathogenic vari-
ants (n=23) were either nonsense, missense, splice site, small frameshift, or inframe deletions. Six of these variants are not present in ClinVar, HGMD, or gnomAD, and are considered novel (Online Supplementary Table S3).
Forty-four percent of the cohort was male (male:female 1:1.25; P=0.14) (Table 1). Median age at identification of symptoms was at birth in patients with large deletions ver- sus two months of age in those with other pathogenic vari- ants (P=0.04) (Table 1). Age of last follow-up was similar for both groups with a median age of 10 and 12 years, respectively (P=0.37) (data not shown). All except two patients were alive at last follow-up. The cause of death was right ventricular outflow obstruction due to throm- boembolic plaque in one patient and complications related to treatment for Wilms tumor in the other patient.
Erythrocyte adenosine deaminase (eADA) values were available only for a subset of patients. Six of 11 patients with large deletion and 5 of 13 with all other RPL35A vari- ants had elevated eADA with no difference between the groups (P=0.682). Anemia was the most common present- ing symptom and was identified in all except one patient. More than 86% of patients in both groups had severe ane- mia needing treatment (Table 1), but a significant number of patients with RPL35A large deletions were steroid-resis- tant and RBC transfusion-dependent compared with patients with other pathogenic variants in RPL35A (P=0.0023) (Table 1). The number of patients who received HCT was similar in both groups.
Neutropenia was significantly associated with large dele- tions (P=0.0017). In addition, a higher proportion of patients with large deletions with severe neutropenia
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