Results of a Pilot External Quality Assessment Scheme for Genetic Testing of Newborns with Spinal Muscular Atrophy.

BACKGROUND: This study aims to evaluate the ability of laboratories to perform spinal muscular atrophy (SMA) genetic testing in newborns based on dried blood spot (DBS) samples, and to provide reference data and advance preparation for establishing the pilot external quality assessment (EQA) scheme for SMA genetic testing of newborns in China. METHODS: The pilot EQA scheme contents and evaluation principles of this project were designed by National Center for Clinical Laboratories (NCCL), National Health Commission. Two surveys were carried out in 2022, and 5 batches of blood spots were submitted to the participating laboratory each time. All participating laboratories conducted testing upon receiving samples, and test results were submitted to NCCL within the specified date. RESULTS: The return rates were 75.0% (21/28) and 95.2% (20/21) in the first and second surveys, respectively. The total return rate of the two examinations was 83.7% (41/49). Nineteen laboratories (19/21, 90.5%) had a full score passing on the first survey, while in the second survey twenty laboratories (20/20, 100%) scored full. CONCLUSIONS: This pilot EQA survey provides a preliminary understanding of the capability of SMA genetic testing for newborns across laboratories in China. A few laboratories had technical or operational problems in testing. It is, therefore, of importance to strengthen laboratory management and to improve testing capacity for the establishment of a national EQA scheme for newborn SMA genetic testing.

Result of a Pilot External Quality Assessment Scheme for Clinical Diagnosis of Inherited Metabolic Disorders in China.

BACKGROUND: We aimed to evaluate the diagnostic capabilities of Chinese laboratories for inherited metabolic disorders (IMDs) using gas chromatography-mass spectrometry (GC-MS) on urine samples. Meanwhile, based on the result of the pilot external quality assessment (EQA) scheme, we hope to establish a standardized and reliable procedure for future EQA practice. METHODS: We recruited laboratories that participated in the EQA of quantitative analysis of urinary organic acids with GC-MS before joining the surveys. In each survey, a set of five real urine samples was distributed to each participant. The participants should analyze the sample by GC-MS and report the "analytical result", "the most likely diagnosis", and "recommendation for further tests" to the NCCL before the deadline. RESULTS: A total of 21 laboratories participated in the scheme. The pass rates were 94.4% in 2020 and 89.5% in 2021. For all eight IMDs tested, the analytical proficiency rates ranged from 84.7% - 100%, and the interpretational performance rate ranged from 88.2% - 97.0%. The performance on hyperphenylalaninemia (HPA), 3-methylcrotonyl-CoA carboxylase deficiency (MCCD), and ethylmalonic encephalopathy (EE) samples were not satisfactory. CONCLUSIONS: In general, the participants of this pilot EQA scheme are equipped with the basic capability for qualitative organic acid analysis and interpretation of the results. Limited by the small size of laboratories and samples involved, this activity could not fully reflect the state of clinical practice of Chinese laboratories. NCCL will improve the EQA scheme and implement more EQA activities in the future.

[Clinical phenotype and genetic analysis of a patient with a heterozygous 6p25.3 deletion and partial trisomy 15q].

OBJECTIVE: To investigate the clinical phenotype and genetic characteristics of a patient with a heterozygous 6p25.3 deletion and partial trisomy 15q. METHODS: A patient who had presented at the Genetics and Prenatal Diagnosis Center of the First Affiliated Hospital of Zhengzhou University on May 14, 2021 was selected as the study subject. Clinical data of the patient was collected, and G-banded chromosomal karyotyping and copy number variation sequencing (CNV-seq) were carried out. RESULTS: The patient's main clinical features have included complete uterine septum, vaginal septum, atrophy of left eyeball, abnormal fingers and toes, and mental retardation. The karyotype of the patient was 46,XX,der(6)t(6;15)(p25.3;q26.1). CNV-seq result has indicated a 1.20 Mb heterozygous deletion in the 6p25.3 region and a 10.20 Mb duplication in the 15q26.1q26.3 region. The deletion segment has included the FOXQ1 gene, which may be related with the abnormal development of the left eye. The duplication segment has a 96.16% overlap with the region associated with 15q26 overgrowth syndrome (including the IGF1R gene), which may be related to the patient' s abnormal development of the Müllerian duct, abnormal fingers and toes, and mental developmental delay. CONCLUSION: The heterozygous deletion of the 6p25.3 region and duplication of the 15q26.1q26.3 region probably underlay the abnormal clinical phenotype in this patient.

[Results of combined newborn hearing and deafness gene screening in Yuncheng area of Shanxi Province].

OBJECTIVE: To analyze the clinical significance of combined newborn hearing and deafness gene screening in Yuncheng area of Shanxi Province. METHODS: Results of audiological examinations, including transient evoked otoacoustic emission and automatic discriminative auditory brainstem evoked potentials, for 6 723 newborns born in Yuncheng area from January 1, 2021 to December 31, 2021, were retrospectively analyzed. Those who failed one of the tests were considered to have failed the examination. A deafness-related gene testing kit was used to detect 15 hot spot variants of common deafness-associated genes in China including GJB2, SLC26A4, GJB3, and mtDNA12S rRNA. Neonates who had passed the audiological examinations and those who had not were compared using a chi-square test. RESULTS: Among the 6 723 neonates, 363 (5.40%) were found to carry variants. These have included 166 cases (2.47%) with GJB2 gene variants, 136 cases (2.03%) with SLC26A4 gene variants, 26 cases (0.39%) with mitochondrial 12S rRNA gene variants, and 33 cases (0.49%) with GJB3 gene variants. Among the 6 723 neonates, 267 had failed initial hearing screening, among which 244 had accepted a re-examination, for which 14 cases (5.73%) had failed again. This has yielded an approximate prevalence of hearing disorder of 0.21% (14/6 723). Among 230 newborns who had passed the re-examination, 10 (4.34%) were found to have carried a variant. By contrast, 4 out of the 14 neonates (28.57%) who had failed the re-examination had carried a variant, and there was a significant difference between the two groups (P < 0.05). CONCLUSION: Genetic screening can provide an effective supplement to newborn hearing screening, and the combined screening can provide a best model for the prevention of hearing loss, which can enable early detection of deafness risks, targeted prevention measures, and genetic counseling to provide accurate prognosis for the newborns.

Using random forest to detect multiple inherited metabolic diseases simultaneously based on GC-MS urinary metabolomics.

Inborn errors of metabolism, also known as inherited metabolic diseases (IMDs), are related to genetic mutations and cause corresponding biochemical metabolic disorder of newborns and even sudden infant death. Timely detection and diagnosis of IMDs are of great significance for improving survival of newborns. Here we propose a strategy for simultaneously detecting six types of IMDs via combining GC-MS technique with the random forest algorithm (RF). Clinical urine samples from IMD and healthy patients are analyzed using GC-MS for acquiring metabolomics data. Then, the RF model is established as a multi-classification tool for the GC-MS data. Compared with the models built by artificial neural network and support vector machine, the results demonstrated the RF model has superior performance of high specificity, sensitivity, precision, accuracy, and matthews correlation coefficients on identifying all six types of IMDs and normal samples. The proposed strategy can afford a useful method for reliable and effective identification of multiple IMDs in clinical diagnosis.

Astragaloside IV Exerts Anti-tumor Effect on Murine Colorectal Cancer by Re-educating Tumor-Associated Macrophage.

Astragaloside IV (AS-IV) has shown anti-tumorigenic properties in certain cancers for its effect of boosting the body's immune system, but its role in colorectal cancer (CRC) remains unclear. In this study, we investigated the therapeutic effect of AS-IV in CRC and explored its underlying mechanism. CT26 colon cancer cells and mouse model by injection of CT26 cells subcutaneously were used as in vitro and in vivo model. M1 and M2 macrophage-associated markers, mRNA and protein expression levels were analyzed after AS-IV treatment. Inflammatory factors and cytokines in the tumors from mouse model were detected. Repolarization effect of AS-IV in vitro on bone-marrow-derived macrophages was also detected. In vitro, AS-IV inhibited the proliferation of CT26 cells and induced cell apoptosis dose-dependently, and significantly reduced M2 macrophages and increased M1 macrophages. In mouse model, it suppressed tumor growth and decreased the production of anti-inflammatory factors such as TGF-ß, IL-10 and VEGF-A, while increased the production of pro-inflammatory factors like IFN-γ, IL-12 and TNF-α in tumor. Combination of AS-IV and checkpoint inhibitor aPD-1 exhibited synergistic antitumor effect by inhibiting tumor growth and increasing T cell infiltration. AS-IV could induce M2 macrophages polarization to the M1 phenotype. Its combination with immune checkpoint inhibitors could be expected to become a potential new strategy for the treatment of CRC.

[Digital breast tomosynthesis in diagnosis of dense breast lesions].

OBJECTIVE: To evaluate the value of digital breast tomosynthesis (DBT) in diagnosis of dense breast lesions. METHODS: Clinical and pathological data of 163 patients (58 benign lesions, 122 malignant lesions, and 180 lesions in total) with breast lesions undergoing surgical treatment in Shaoxing Central Hospital from January 2017 to December 2018 were retrospectively analyzed. The lesions were classified into non-homogeneous dense gland type and extremely dense gland type according to BI-RADS creterion. Breast MRI and DBT examinations were performed before the surgery. ROC curve was generated and the diagnostic efficacy of two examination methods for dense breast lesions was evaluated with pathological results as the gold standard. The detection rate, diagnostic accuracy of benign and malignant breast lesions were compared between two methods using chi-square test. The accuracy of lesion size preoperatively evaluated by MRI and DBT was analyzed by Pearson correlation. RESULTS: The detection rate and diagnostic accuracy for benign breast lesions by MRI were higher than those by DBT (91.4% vs. 75.9%, χ2=5.098, P<0.05 and 89.7% vs. 67.2%, χ2=8.617, P<0.01). But there were no significant differences in detection rate and accuracy for malignant lesions by MRI and DBT (98.4% vs. 95.1%, χ2=2.068, P>0.05 and 94.3% vs. 91.8%, χ2=0.569, P>0.05). The areas under the ROC curves of MRI, DBT based on BI-RADS classification were 0.910 and 0.832, respectively (Z=1.860, P>0.05). The sensitivities of MRI, DBT to breast lesions were 93.3% and 86.7%, and the specificities were 68.3% and 79.1%. DBT and MRI measurements were positively correlated with pathological measurements (r=0.887 and 0.949, all P<0.01). CONCLUSIONS: DBT can effectively diagnose benign and malignant breast lesions under dense gland background, and it has similar diagnostic efficacy with MRI for breast malignant lesions.