Objective:To reassess the pathogenicity of copy number variants (CNVs) involving chromosomal microdeletions and microduplications classified as variants of uncertain significance (VUS).Methods:This retrospective study analyzed 1 882 pregnant women who underwent invasive prenatal diagnosis for chromosomal microarray analysis (CMA) at the First Medical Center, Chinese PLA General Hospital between January 1, 2018, and December 31, 2022. The results were classified according to the American College of Medical Genetics and Genomics guidelines, with 82 fetuses rated as VUS selected for the study. We analyzed invasive prenatal diagnostic indications, followed up on fetal ultrasound findings, parental origin identification results, and pregnancy outcomes, and reclassified VUS CNVs based on the latest evidence. Descriptive statistical analysis was applied to the data.Results:(1) Among the 82 fetuses with VUS CNVs, prenatal diagnostic indications included fetal structural abnormalities detected by ultrasound (21 cases, 25.6%), abnormal non-invasive prenatal testing (NIPT) results (12 cases, 14.6%), high-risk serum screening (seven cases, 8.5%), advanced maternal age (≥35 years at expected delivery, 28 cases, 34.1%), and other indications (14 cases, 17.1%). Sixteen cases (19.5%) exhibited abnormal phenotypes, with seven pregnancies terminated due to severe structural abnormalities detected by prenatal ultrasound. Seventy-five live births were followed up for 25 (13-66) months. (2) Among the 82 cases, five fetuses had two VUS CNVs detected by CMA, while the remaining 77 had only one, totaling 87 VUS CNVs. Of these, 63 (72.4%) were chromosomal microduplications and 24 (27.6%) were chromosomal microdeletions. The size of the CNV segments ranged from 0.85 (0.05-5.61) Mb, with 82 segments less than 2 Mb. Parental origin identification was refused by 44 cases (53.7%), while 38 (46.3%) underwent the test, revealing eight (21.0%) de novo variants and 30 (78.9%) inherited from either parent (12 maternal and 18 paternal). (3) Among the 87 VUS CNVs, the ratings of 11 CNVs (12.6%) changed after re-evaluation. This included one 4p16.2 microdeletion and two 15q11.2 microdeletions being upgraded to pathogenic, one 16p13.11 microduplication being upgraded to likely pathogenic, one Xp22.31 microduplication and two 2q13 microdeletions being downgraded to likely benign, and four Xp22.31 microduplications being downgraded to benign. (4) Among the 16 fetuses with abnormal phenotypes, seven with prenatal abnormalities terminated pregnancies, including six with structural abnormalities and one with severe fetal growth restriction. After re-evaluation, one case was upgraded to pathogenic, while six remained VUS. Nine live births with postnatal abnormal phenotypes showed no change in classification after re-evaluation. Among the 66 cases (80.5%) without abnormal phenotypes, 10 had their classifications changed after re-evaluation. Conclusions:Fetuses with VUS CNVs often exhibit no significant abnormal phenotypes and have a relatively favorable prognosis, however, further floow-up is still needed. Parental origin identification can provide valuable insights for genetic counseling.

Objective:To investigate the distribution of CGG repeat numbers in the FMR1 gene among reproductive-age women in China, providing data reference for carrier screening and genetic counseling of Fragile X syndrome. Methods:This cross-sectional study recruited 16 610 reproductive-age women from 12 medical institutions between July 2022 and October 2023. Peripheral venous blood samples (3 mL) were collected, and genomic DNA was extracted. The number of CGG repeats in the FMR1 gene was determined using the triplet-primed polymerase chain reaction (TP-PCR) combined with capillary electrophoresis technology. Statistical analyses were performed to assess the prevalence and distribution of CGG repeat expansions. Results:Among 16 610 women of childbearing age, 5 684 (34.220%) women had the same number of CGG repeats in the two alleles of FMR1 gene, and 10 926 (65.780%) women had different numbers of repeats in the two alleles. Among the 33 220 FMR1 alleles in 16 610 women of reproductive age, the most common CGG repeat numbers were 29 [48.645% (16 160/33 220)] and 30 [26.276% (8 729/33 220)], while the most frequent CGG genotype was CGG 29/29 [24.726% (4 107/16 610)]. The CGG repeat numbers of FMR1 gene were normal in 16 498 women (99.326%). Among the 112 women (0.674%) with CGG repeat abnormities, 96 (0.578%) women were classified as intermediate carriers, 15 (0.090%) as premutation carriers, and 1 (0.006%) as a full mutation carrier, whose CGG genotype was (36, >200). Conclusion:In the general reproductive-age female population in China, the normal CGG repeat numbers of the FMR1 gene account for 99.326%, while the intermediate carrier rate is 0.578%, and the combined carrier rate of the premutation and full mutation types is 0.096%.

Objective:To carry out carrier screening among people of childbearing age, detect the pathogenic genes of monogenic genetic diseases and analyze the carrier status of pathogenic variants, so as to provide fertility guidance and intervention measures for high-risk families.Methods:From August 2022 to August 2023, 1 533 families of childbearing age who met the criteria were recruited in the Chinese PLA General Hospital, including a total of 3 044 subjects. According to the standard enrollment procedure, 223 genes (197 autosomal recessive genes and 26 X-linked genes) of the subjects were tested. According to the screening results, genetic counseling and fertility guidance were provided to the subjects. Invasive prenatal diagnosis was performed for high-risk couples (both couples being carriers of the same autosomal recessive disease gene or the woman was a carrier of X-linked disease gene), and their pregnancy pattern, outcome and offspring phenotype were followed up.Results:(1) A total of 3 044 cases from 1 511 couples and women of childbearing age from 22 families were included for carrier screening. Totally 1 503 families chose simultaneous screening and 30 families chose sequential screening out of the 1 533 families. Among the 3 044 subjects, 1 603 individuals carried at least one pathogenic or likely pathogenic variant, and the overall carrier rate was 52.66% (1 603/3 044). A total of 2 292 pathogenic or likely pathogenic variants were detected, and 0.75 variants (2 292/3 044) were detected per capita. (2) The three genes with the highest carrier rates were GJB2 (8.67%, 264/3 044), CYP21A2 (3.19%, 97/3 044) and PAH (3.09%, 94/3 044). There were 32 genes with a carrier rate ≥1/200, 17 genes with a carrier rate ≥1/100, and 7 genes with a carrier rate ≥1/50. (3) Thirty-eight high-risk families were identified. After excluding G6PD gene mutation, there were 33 high-risk families, of which 25 couples were carriers of the same autosomal recessive gene, 9 women were carriers of X-linked gene, and 1 family was double high-risk couple with both autosomal recessive and X-linked gene. After further excluding the GJB2 c.109G>A mutation, 21 high-risk families were identified. Preimplantation genetic testing for monogenic disease was performed in 12 families after genetic counseling. Prenatal diagnosis was completed in 4 out of 5 high-risk families who conceived naturally. Two fetuses carried the parental variants and terminated the pregnancy, one fetus did not carry the parental variants but was induced due to trisomy 21 syndrome, and one fetus was a carrier of congenital disorders of glycosylation type 1a.Conclusions:Carrier screening effectively identifies high-risk genetic disease families and provides reproductive guidance to prevent the birth of affected children. However, establishing multidisciplinary team is essential for managing complex cases. Implementation should prioritize prenatal institutions with genetic counseling or diagnostic expertise for monogenic disorders or established referral networks.

Objective:To investigate the distribution of CGG repeat numbers in the FMR1 gene among reproductive-age women in China, providing data reference for carrier screening and genetic counseling of Fragile X syndrome. Methods:This cross-sectional study recruited 16 610 reproductive-age women from 12 medical institutions between July 2022 and October 2023. Peripheral venous blood samples (3 mL) were collected, and genomic DNA was extracted. The number of CGG repeats in the FMR1 gene was determined using the triplet-primed polymerase chain reaction (TP-PCR) combined with capillary electrophoresis technology. Statistical analyses were performed to assess the prevalence and distribution of CGG repeat expansions. Results:Among 16 610 women of childbearing age, 5 684 (34.220%) women had the same number of CGG repeats in the two alleles of FMR1 gene, and 10 926 (65.780%) women had different numbers of repeats in the two alleles. Among the 33 220 FMR1 alleles in 16 610 women of reproductive age, the most common CGG repeat numbers were 29 [48.645% (16 160/33 220)] and 30 [26.276% (8 729/33 220)], while the most frequent CGG genotype was CGG 29/29 [24.726% (4 107/16 610)]. The CGG repeat numbers of FMR1 gene were normal in 16 498 women (99.326%). Among the 112 women (0.674%) with CGG repeat abnormities, 96 (0.578%) women were classified as intermediate carriers, 15 (0.090%) as premutation carriers, and 1 (0.006%) as a full mutation carrier, whose CGG genotype was (36, >200). Conclusion:In the general reproductive-age female population in China, the normal CGG repeat numbers of the FMR1 gene account for 99.326%, while the intermediate carrier rate is 0.578%, and the combined carrier rate of the premutation and full mutation types is 0.096%.

Objective:To carry out carrier screening among people of childbearing age, detect the pathogenic genes of monogenic genetic diseases and analyze the carrier status of pathogenic variants, so as to provide fertility guidance and intervention measures for high-risk families.Methods:From August 2022 to August 2023, 1 533 families of childbearing age who met the criteria were recruited in the Chinese PLA General Hospital, including a total of 3 044 subjects. According to the standard enrollment procedure, 223 genes (197 autosomal recessive genes and 26 X-linked genes) of the subjects were tested. According to the screening results, genetic counseling and fertility guidance were provided to the subjects. Invasive prenatal diagnosis was performed for high-risk couples (both couples being carriers of the same autosomal recessive disease gene or the woman was a carrier of X-linked disease gene), and their pregnancy pattern, outcome and offspring phenotype were followed up.Results:(1) A total of 3 044 cases from 1 511 couples and women of childbearing age from 22 families were included for carrier screening. Totally 1 503 families chose simultaneous screening and 30 families chose sequential screening out of the 1 533 families. Among the 3 044 subjects, 1 603 individuals carried at least one pathogenic or likely pathogenic variant, and the overall carrier rate was 52.66% (1 603/3 044). A total of 2 292 pathogenic or likely pathogenic variants were detected, and 0.75 variants (2 292/3 044) were detected per capita. (2) The three genes with the highest carrier rates were GJB2 (8.67%, 264/3 044), CYP21A2 (3.19%, 97/3 044) and PAH (3.09%, 94/3 044). There were 32 genes with a carrier rate ≥1/200, 17 genes with a carrier rate ≥1/100, and 7 genes with a carrier rate ≥1/50. (3) Thirty-eight high-risk families were identified. After excluding G6PD gene mutation, there were 33 high-risk families, of which 25 couples were carriers of the same autosomal recessive gene, 9 women were carriers of X-linked gene, and 1 family was double high-risk couple with both autosomal recessive and X-linked gene. After further excluding the GJB2 c.109G>A mutation, 21 high-risk families were identified. Preimplantation genetic testing for monogenic disease was performed in 12 families after genetic counseling. Prenatal diagnosis was completed in 4 out of 5 high-risk families who conceived naturally. Two fetuses carried the parental variants and terminated the pregnancy, one fetus did not carry the parental variants but was induced due to trisomy 21 syndrome, and one fetus was a carrier of congenital disorders of glycosylation type 1a.Conclusions:Carrier screening effectively identifies high-risk genetic disease families and provides reproductive guidance to prevent the birth of affected children. However, establishing multidisciplinary team is essential for managing complex cases. Implementation should prioritize prenatal institutions with genetic counseling or diagnostic expertise for monogenic disorders or established referral networks.

Objective:To reassess the pathogenicity of copy number variants (CNVs) involving chromosomal microdeletions and microduplications classified as variants of uncertain significance (VUS).Methods:This retrospective study analyzed 1 882 pregnant women who underwent invasive prenatal diagnosis for chromosomal microarray analysis (CMA) at the First Medical Center, Chinese PLA General Hospital between January 1, 2018, and December 31, 2022. The results were classified according to the American College of Medical Genetics and Genomics guidelines, with 82 fetuses rated as VUS selected for the study. We analyzed invasive prenatal diagnostic indications, followed up on fetal ultrasound findings, parental origin identification results, and pregnancy outcomes, and reclassified VUS CNVs based on the latest evidence. Descriptive statistical analysis was applied to the data.Results:(1) Among the 82 fetuses with VUS CNVs, prenatal diagnostic indications included fetal structural abnormalities detected by ultrasound (21 cases, 25.6%), abnormal non-invasive prenatal testing (NIPT) results (12 cases, 14.6%), high-risk serum screening (seven cases, 8.5%), advanced maternal age (≥35 years at expected delivery, 28 cases, 34.1%), and other indications (14 cases, 17.1%). Sixteen cases (19.5%) exhibited abnormal phenotypes, with seven pregnancies terminated due to severe structural abnormalities detected by prenatal ultrasound. Seventy-five live births were followed up for 25 (13-66) months. (2) Among the 82 cases, five fetuses had two VUS CNVs detected by CMA, while the remaining 77 had only one, totaling 87 VUS CNVs. Of these, 63 (72.4%) were chromosomal microduplications and 24 (27.6%) were chromosomal microdeletions. The size of the CNV segments ranged from 0.85 (0.05-5.61) Mb, with 82 segments less than 2 Mb. Parental origin identification was refused by 44 cases (53.7%), while 38 (46.3%) underwent the test, revealing eight (21.0%) de novo variants and 30 (78.9%) inherited from either parent (12 maternal and 18 paternal). (3) Among the 87 VUS CNVs, the ratings of 11 CNVs (12.6%) changed after re-evaluation. This included one 4p16.2 microdeletion and two 15q11.2 microdeletions being upgraded to pathogenic, one 16p13.11 microduplication being upgraded to likely pathogenic, one Xp22.31 microduplication and two 2q13 microdeletions being downgraded to likely benign, and four Xp22.31 microduplications being downgraded to benign. (4) Among the 16 fetuses with abnormal phenotypes, seven with prenatal abnormalities terminated pregnancies, including six with structural abnormalities and one with severe fetal growth restriction. After re-evaluation, one case was upgraded to pathogenic, while six remained VUS. Nine live births with postnatal abnormal phenotypes showed no change in classification after re-evaluation. Among the 66 cases (80.5%) without abnormal phenotypes, 10 had their classifications changed after re-evaluation. Conclusions:Fetuses with VUS CNVs often exhibit no significant abnormal phenotypes and have a relatively favorable prognosis, however, further floow-up is still needed. Parental origin identification can provide valuable insights for genetic counseling.

OBJECTIVE To observe the clinical efficacy of Kanglaite injection assisted with camrelizumab combined with chemotherapy in the treatment of advanced non-small cell lung cancer （NSCLC）. METHODS A total of 192 patients with advanced NSCLC and hospitalized in the TCM oncology department of our hospital from January 1st， 2018 to December 1st， 2022 were retrospectively selected as the study objects， and were divided into observation group （additional use， n＝104） and control group （without additional use， n＝88） according to whether the patients additionally received Kanglaite injection based on camrelizumab combined with chemotherapy （carboplatin+pemetrexed）. The short-term therapeutic effects of 2，4 and 6 cycles were compared between the two groups. The levels of peripheral blood immune function indexes and serum tumor markers were compared before treatment， after 3 cycles of treatment and after treatment. The long-term therapeutic effects as well as the occurrence of adverse drug reaction（ADR） during hospitalization were compared between the two groups. RESULTS After 3 treatment cycles and at the end of treatment， the CD4+ T lymphocyte ratio and CD4+/CD8+ in the observation group were notably greater than the control group （P＜0.05）； the levels of serum carcinoembryonic antigen and cytokeratin 19 fragment antigen 21-1 were significantly lower than those in the control group （P＜0.05）. The overall survival of the observation group was significantly longer than that of the control group （P＜0.05）， and the median overall survival was （185.27±38.21） d and （132.11±34.23） d， respectively. There were no significant differences in the whole ADR and grade ≥3 ADR between the two groups during hospitalization（P＞0.05）. CONCLUSIONS Based on camrelizumab combined with chemotherapy， the addition of Kanglaite injection can enhance immunological response and prolong overall survival in advanced NSCLC patients.

Objective To investigate the epidemiological characteristics and mutation spectrum of monogenic diseases in Chinese population through a large-scale,multicenter carrier screening.Methods This study was conducted among a total of 33 104 participants(16 610 females)from 12 clinical centers across China.Carrier status for 223 genes was analyzed using high-throughput sequencing and different PCR methods.Results The overall combined carrier frequency was 55.58%for 197 autosomal genes and 1.84%for 26 X-linked genes in these participants.Among the 16 669 families,874 at-risk couples(5.24%)were identified.Specifically,584 couples(3.50%)were at risk for autosomal genes,306(1.84%)for X-linked genes,and 16 for both autosomal and X-linked genes.The most frequently detected autosomal at-risk genes included GJB2(autosomal recessive deafness type 1A,393 couples),HBA1/HBA2(α-thalassemia,36 couples),PAH(phenylketonuria,14 couples),and SMN1(spinal muscular atrophy,14 couples).The most frequently detected X-linked at-risk genes were G6PD(G6PD deficiency,236 couples),DMD(Duchenne muscular dystrophy,23 couples),and FMR1(fragile X syndrome,17 couples).After excluding GJB2 c.109G>A,the detection rate of at-risk couples was 3.91%(651/16 669),which was lowered to 1.72%(287/16 669)after further excluding G6PD.The theoretical incidence rate of severe monogenic birth defects was approximately 4.35‰(72.5/16 669).Screening for a battery of the top 22 most frequent genes in the at-risk couples could detect over 95%of at-risk couples,while screening for the top 54 genes further increased the detection rate to over 99%.Conclusion This study reveals the carrier frequencies of 223 monogenic genetic disorders in the Chinese population and provides evidence for carrier screening strategy development and panel design tailored to the Chinese population.In carrier testing,genetic counseling for specific genes or gene variants can be challenging,and the couples need to be informed of these difficulties before testing and provided with options for not screening these genes or gene variants.

Objective To investigate the epidemiological characteristics and mutation spectrum of monogenic diseases in Chinese population through a large-scale,multicenter carrier screening.Methods This study was conducted among a total of 33 104 participants(16 610 females)from 12 clinical centers across China.Carrier status for 223 genes was analyzed using high-throughput sequencing and different PCR methods.Results The overall combined carrier frequency was 55.58%for 197 autosomal genes and 1.84%for 26 X-linked genes in these participants.Among the 16 669 families,874 at-risk couples(5.24%)were identified.Specifically,584 couples(3.50%)were at risk for autosomal genes,306(1.84%)for X-linked genes,and 16 for both autosomal and X-linked genes.The most frequently detected autosomal at-risk genes included GJB2(autosomal recessive deafness type 1A,393 couples),HBA1/HBA2(α-thalassemia,36 couples),PAH(phenylketonuria,14 couples),and SMN1(spinal muscular atrophy,14 couples).The most frequently detected X-linked at-risk genes were G6PD(G6PD deficiency,236 couples),DMD(Duchenne muscular dystrophy,23 couples),and FMR1(fragile X syndrome,17 couples).After excluding GJB2 c.109G>A,the detection rate of at-risk couples was 3.91%(651/16 669),which was lowered to 1.72%(287/16 669)after further excluding G6PD.The theoretical incidence rate of severe monogenic birth defects was approximately 4.35‰(72.5/16 669).Screening for a battery of the top 22 most frequent genes in the at-risk couples could detect over 95%of at-risk couples,while screening for the top 54 genes further increased the detection rate to over 99%.Conclusion This study reveals the carrier frequencies of 223 monogenic genetic disorders in the Chinese population and provides evidence for carrier screening strategy development and panel design tailored to the Chinese population.In carrier testing,genetic counseling for specific genes or gene variants can be challenging,and the couples need to be informed of these difficulties before testing and provided with options for not screening these genes or gene variants.

ObjectiveTo explore the possible mechanism of Changweiqing （肠胃清） in the treatment of colorectal cancer. MethodsHCT 116 cancer cells were used to prepare intestinal cancer cells with silenced polypyrimidine region binding protein 3 （PTBP3） gene and stably transfected cells with overexpressed PTBP3 gene. Stably transfected cells with silenced PTBP3， stably transfected cells with overexpressed PTBP3 and untransfected cancer cells were injected into the armpit of 72 nude mice to construct three different subcutaneous transplanted tumor models of colorectal cancer cells， including the silenced model， the overexpressed model and the control model， with 24 mice per model. Mice of each transplanted tumor modelwere randomly divided into Changweiqing （CWQ） group， oxaliplatin （OXA） group and normal saline （NS） group， with 8 mice in each group. The CWQ groups were given intragastric administration of 35.9625 g/kg of Changweiqing oral liquid and were intraperitoneally injected with 0.2ml of normal saline； the NS groups were given 0.5ml of normal saline by gavage， and intraperitoneal injection of 0.2ml of normal saline； the OXA groups were intraperitoneally injected with 5 mg/kg （0.2 ml） of oxaliplatin and given 0.5ml of normal saline by gavage. Each group was given intragastric administration once a day and intraperitoneal injection three times a week. After 31 days， the weight of subcutaneous tumors in each group was measured， and the tumor inhibition rate of the groups in each model were measured. Immunohistochemistry and other methods were used to detect the expression level of cell proliferation cell nuclear antigen Ki67 and apoptosis index. Real-time PCR and Western Blot were used to detect mRNA and protein expressions of PTBP3， signal transducer and activator of transcription 3 （STAT3） splicing isoform α （STAT3α）， STAT3 splicing isoform β （STAT3β）， B-cell lymphoma/leukemia-2 （Bcl-2） splicing isoform α （Bcl-2α）， and Bcl-2 splicing isoform β （Bcl-2β） in subcutaneous tumor cells in each group. ResultsFor all three transplanted tumor models， the weight of the subcutaneous tumors and Ki67 expression level of subcutaneous tumor tissue in all CWQ groups and OXA groups were lower than those of the corresponding NS groups， while the apoptosis level were higher （P＜0.05 or P＜0.01）. The mRNA and protein expressions of PTBP3， STAT3α， and Bcl-2α in the subcutaneous tumor tissues of the silenced model CWQ group and the overexpressed model CWQ group were lower than those of the corresponding NS groups， while the mRNA and protein expression levels of STAT3β and Bcl-2β were higher （P＜0.05 or P＜0.01）. All there groups of silenced model had lower subcutaneous tumor weight， Ki67 expression level， and mRNA and protein expression levels of PTBP3， STAT3α， and Bcl-2α in subcutaneous tumor tissue， as well as higher apoptosis level and mRNA and protein expression levels of STAT3β and Bcl-2β than those in all groups of control model； all groups of overexpressed model had higher subcutaneous tumor weight， Ki67 expression level， and mRNA and protein expression levels of PTBP3， STAT3α， and Bcl-2α ， while lower apoptosis level and mRNA and protein expression levels of STAT3β and Bcl-2β than those in all control model groups （P＜0.05 or P＜0.01）. In the control model， compared with the NS group， The tumor inhibition rate of all OXA groups was higher than that of corresponding CWQ groups， respectively. Compared to that of each control model group， the tumor inhibition rate was positive value of each silenced model group， and negative value of each overexpressed model group. ConclusionPTBP3 can promote the proliferation and inhibit apoptosis of intestinal cancer cells， upregulate the expression of STAT3α and Bcl-2α， and downregulate the expression of STAT3β and Bcl-2β in intestinal cancer cells. The meachnism of action of Changweiqing in the treatment of colorectal cancer maybe related to the inhibition of PTBP3， and regulation of the expression of STAT3α， STAT3β， Bcl-2α， and Bcl-2β.