Clinical characterization of novel HSPA9 splice variant in a Chinese woman

Abstract

Dear Editor, HSPA9 is a highly conserved HSP70 family member that is essential in mitochondrial protein import, folding, and degradation. In human, HSPA9 mutation was thought the causative mutation for congenital sideroblastic anemia (CSA), and biallelic mutations of HSPA9 was also related to another disease called EVEN-PLUS syndrome. Currently, only these two reports described human HSPA9 variants. Here, we presented the third report of HSPA9 variant in a Chinese woman. The patient was a 50-years-old woman. She reported feeling weakness since age 18. On physical examination, she was of short stature but exhibited no dysmorphism. Pancytopenia and diverse shaped of RBC was noticed, from bone marrow smear and blood smear, it was observed that erythroid hyperplasia and granulosa hyperplasia were obviously active (Figure 1(A-D)). Iron-stained bone marrow demonstrated iron granules ringing around erythroblast nuclei (Figure 1(E)). Morphological features suggested sideroblastic anemia, most likely congenital, myelodysplastic syndrome (MDS) and acquired sideroblastic anemia were ruled out. This patient also suffered from chronic hemolytic anemia, presented some overlapping feature with MDS-RS (Figure 1(A,E)). Next-generation sequencing of the coding regions of 208 blood disease related genes (MyGenostics Company, China) identified a variant of uncertain significance, a splicing variant in HSPA9 (c.1515+1G>A). By Sanger sequencing our lab confirmed this variant, and all the other healthy family members did not carry this variant (Figure 1(F)). Decreased HSPA9 expression was observed (Figure 1(G)). Result of minigene strategy showed that this variant caused exon skipping of HSPA9 mRNA, which led to nonsense mediated decay (Figure 1(G,H)). Notably, HSPA9 is one of the genes in the proximal commonly deleted interval of 5q MDS. In mice and zebrafish models, loss of HSPA9 recapitulated ineffective hematopoiesis of MDS. This patient represented first human case and confirmed the effects of HSPA9 haploinsufficiency on cytopenia observed in MDS. CSA is a rare heterogeneous disease characterized by deposition of iron in mitochondria of erythroid precursors in the bone marrow leading to ineffective erythropoiesis. The clinical phenotype of this patient is interesting, first, it was revealed that iron overload usually appear in CSA. But this patient presented normal iron parameters. Second, CSA usually caused the erythroid dysfunction, but this patient presented multilineage cytopenia. Third, CSA usually present large proportion of ring sideroblasts, while only 6% ring sideroblasts in this patient was detected. Moreover, Mean Cell Volume of this patient was larger than normal, not consistent with previously reported phenotype of other HSPA9 variants. In summary, we described a novel and rare splicing variant in HSPA9. in vitro study showed that this variant caused exon skipping and reduced HSPA9 expression. Previous research suggested that different germline variants of HSPA9 resulted in CSA or EVEN-PLUS syndrome in human. SNP rs10117T might determine expression of the HSPA9 CSA phenotype in patients with single deleterious allele, resulting in pseudodominant inheritance. Interestingly, this patient was homozygous for rs10117T. She presented with pancytopenia and ring sideroblasts, this case represented that HSPA9 haploinsufficiency alters the hematopoietic progenitor pool and contributes to abnormal hematopoiesis in human for the first time. Maybe the severity of the phenotype correlate with the function and level of residual protein. The present case extended the clinical symptoms and variant spectrum of HSPA9 and helps to establish clinical diagnosis standards. The Ethics Committee of the First Affiliated Hospital of Nanjing Medical University approved this study and informed consent from each participant was obtained.