Background Limb-girdle muscular dystrophy R8 (LGMD R8) is a rare autosomal recessive muscle disease caused by TRIM32 gene biallelic defects. The genotype–phenotype correlation of this disease has been reported poorly. Here, we report a Chinese family with two female LGMD R8 patients. Methods We performed whole-genome sequencing (WGS) and Sanger sequencing on the proband. Meanwhile, the function of mutant TRIM32 protein was analyzed by bioinformatics and experimental analysis. In addition, a summary of the reported TRIM32 deletions and point mutations and an investigation of genotype–phenotype correlation were performed through a combined analysis of the two patients and other cases reported in previous literature. Results The two patients displayed typical symptoms of LGMD R8, which worsened during pregnancy. Genetic analysis by whole-genome sequencing (WGS) and Sanger sequencing showed that the patients were compound heterozygotes of a novel deletion (chr9.hg19:g.119431290_119474250del) and a novel missense mutation (TRIM32:c.1700A > G, p.H567R). The deletion encompassed 43 kb and resulted in the removal of the entire TRIM32 gene. The missense mutation altered the structure and further affected function by interfering with the self-association of the TRIM32 protein. Females with LGMD R8 showed less severe symptoms than males, and patients carrying two mutations in NHL repeats of the TRIM32 protein had earlier disease onset and more severe symptoms than other patients. Conclusions This research extended the spectrum of TRIM32 mutations and firstly provided useful data on the genotype–phenotype correlation, which is valuable for the accurate diagnosis and genetic counseling of LGMD R8.

SummaryHaemoglobin H (Hb H) disease (intermediate status of α‐thalassemia) shows marked phenotypic variability from asymptomatic to severe anaemia. Apart from the combined β‐thalassemia allele ameliorating clinical severity, reports of genetic modifier genes affecting the phenotype of Hb H disease are scarce which bring inconvenience to precise diagnosis and genetic counselling of the patients. Here, we present a novel mutation (c.948C>A, p.S316R) in the PIP4K2A gene in a female Hb H disease patient who displayed moderate anaemia and a relatively high Hb H level. Haematological analysis in her family members revealed that individuals carrying this mutation have upregulated β‐globin expression, leading to a more imbalanced β/α‐globin ratio and more Hb H inclusion bodies in peripheral red blood cells. According to functional experiments, the mutant PIP4K2A protein exhibits enhanced protein stability, increased kinase activity and a stronger regulatory effect on downstream proteins, suggesting a gain‐of‐function mutation. Moreover, introduction of the S316R mutation into HUDEP‐2 cells increased expression of β‐globin, further inhibiting erythroid differentiation and terminal enucleation. Thus, the S316R mutation is a novel genetic factor associated with β‐globin expression, and the PIP4K2A gene is a new potential modifier gene affecting the α‐thalassemia phenotype.

PurposeShiliao Decoction (SLD) was developed for treatment and prevention of cancer-associated malnutrition (CAM) in China. In this study, we aim to discover SLD’s active compounds and demonstrate the mechanisms of SLD that combat CAM through network pharmacology and molecular docking techniques.MethodsAll components of SLD were retrieved from the pharmacology database of Traditional Chinese Medicine Systems Pharmacology (TCMSP). The GeneCards database and the Online Mendelian Inheritance in Man database (OMIM) were used to identify gene encoding target compounds, and Cytoscape was used to construct the drug compound–target network. The network of target protein-protein interactions (PPI) was constructed using the STRING database, while gene ontology (GO) functional terms and the Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathways associated with potential targets were analyzed using a program in R language (version 4.2.0). Core genes linked with survival and the tumor microenvironment were analyzed using the Kaplan–Meier plotter and TIMER 2.0 databases, respectively. Protein expression and transcriptome expression levels of core gene were viewed using the Human Protein Atlas (HPA) and the Cancer Genome Atlas (TCGA). A component-target-pathway (C-T-P) network was created using Cytoscape, and Autodock Vina software was used to verify the molecular docking of SLD components and key targets.ResultsThe assembled compound–target network primarily contained 134 compounds and 147 targets of the SLD associated with JUN, TP53, MAPK3, MAPK1, MAPK14, STAT3, AKT1, HSP90AA1, FOS, and MYC, which were identified as core targets by the PPI network. KEGG pathway analysis revealed pathways involved in lipid and atherosclerosis, the PI3K/Akt signaling pathway, and immune-related pathways among others. JUN is expressed at different levels in normal and cancerous tissues, it is closely associated with the recruitment of different immune cells and has been shown to have a significant impact on prognosis. The C-T-P network suggests that the active component of SLD is capable of regulating target genes affecting these related pathways. Finally, the reliability of the core targets was evaluated using molecular docking technology.ConclusionThis study revealed insights into SLD’s active components, potential targets, and possible molecular mechanisms, thereby demonstrating a potential method for examining the scientific basis and therapeutic mechanisms of TCM formulae.