OBJECTIVES:Non-syndromic cleft lip with or without cleft palate (NSCL/P) is a common birth defect influenced by genetic and environmental factors, with genetic factors playing a major role. This study aims to investigate the association between the dystrophin (DMD) gene and NSCL/P in a Chinese Han population. METHODS:Four tag single nucleotide polymorphisms (SNPs) in the DMD gene were selected and allelic and genotype-based association analyses were performed on 1 780 patients with NSCL/P and 1 823 normal controls. RESULTS:Comparison with the controls showed that patients with NSCL/P presented three SNPs (rs5971698, rs5928208, and rs5972815) with significant associations with NSCL/P or its subphenotypes. Allelic association analysis revealed that rs5971698 was associated with NSCL/P, unilateral cleft lip with or without cleft palate (UCL/P), left cleft lip with or without cleft palate (LCL/P), unilateral cleft lip and cleft palate (UCLP), left cleft lip and cleft palate (LCLP), unilateral cleft lip (UCL), and left cleft lip only (LCL) (P<0.05); rs5928208 was associated with non-syndromic cleft lip and cleft palate (NSCLP), bilateral cleft lip with or without cleft palate (BCL/P), LCL/P, bilateral cleft lip and cleft palate (BCLP), and LCL (P<0.05); and rs5972815 was associated with UCLP, UCL, and right cleft lip only (RCL)(P<0.05), exhibiting significant laterality bias. Genotype analysis further confirmed these associations. Functional predictions suggested that different alleles at rs5928208 and rs5972815 may influence transcription factor binding affinity. CONCLUSIONS:This study identified associations between SNPs in the DMD gene and NSCL/P in a Western Chinese Han population, providing new evidence for distinct genetic susceptibility loci among NSCL/P subtypes.

ObjectiveGenome-wide association studies have identified over 80 loci associated with nonsyndromic orofacial cleft (NSOC), yet substantial heritability remains unexplained. Insights from syndromic orofacial cleft (SOC) implicated genes could help bridge this gap.DesignA case-control association study in a Han Chinese cohort was conducted to evaluate the association between SOC-implicated genes and NSOC subtypes using association, linkage disequilibrium (LD), and haplotype analyses.SettingTertiary medical center.Patients, ParticipantsThe study included 1626 cases of non-syndromic cleft lip with or without cleft palate, 930 cases of non-syndromic cleft palate only, and 2255 controls.InterventionsPeripheral blood (cases) and umbilical cord blood (controls) were collected for DNA extraction.Main Outcome MeasuresAllelic (Pearson' s χ2, 1 df) and genotypic (Pearson' s χ2, 2 df) associations between SNPs and NSOC subtypes were evaluated, with odds ratios (ORs) and 95% confidence intervals (CIs). LD and sliding-window haplotype association analyses were performed in Haploview. SNPs with minor allele frequency (MAF) >0.05 and in Hardy-Weinberg equilibrium in controls were analyzed. The significance threshold was P < 1.27 × 10-5 after Bonferroni correction.ResultsAllelic analysis identified 23 SNPs that were significantly associated with NSOC subtypes (lowest P = 2.02 × 10-22). Genotypic analysis identified 39 significant SNPs (lowest P = 1.09 × 10-36). Signals mapped to 7 genes. Haplotype analyses revealed a shared causal variant block at MYMK and TWIST2, and allelic heterogeneity at NEDD4L.ConclusionsWe identified MYMK, TWIST2, and NEDD4L as NSOC-associated genes. Using SOC genes as prior knowledge reveals loci missed by standard GWAS, offering key insights into NSOC pathogenesis.

Objectives Orofacial cleft (OC) can be classified into syndromic orofacial cleft (SOC) and non-syndromic orofacial cleft (NSOC), depending on whether there are other congenital deformities. Craniosynostosis, the premature closure of cranial sutures, is a common phenotype of SOC resulting in abnormal ossification of skull and brain development disorders. Its correlation with OC offers a promising approach to identify susceptibility genes for NSOC by examining causative genes of SOCs with craniosynostosis. Materials and Methods This study included 2556 patients with NSOC and 2255 normal controls from western Han Chinese with their genomic DNA samples. We selected 31 causative genes of 34 syndromes with both craniosynostosis and OC as candidate genes and performed quality control. Allelic and genotypic association analyses and haplotype analysis were performed to identify statistically significant single nucleotide polymorphisms (SNPs). Results In allelic association analysis performed with 1265 qualified SNPs in 20 genes, only rs2239936, located in MYH3 gene, was statistically associated with non-syndromic cleft lip only (NSCLO) ( P = 1.70×10−07, OR = 1.33, 95%CI: 1.17-1.52) and non-syndromic cleft palate only (NSCPO) ( P = 6.43×10−05, OR = 1.33, 95%CI: 1.16-1.52). The higher frequency of allele G in NSCPO suggesting that minor allele G at rs2239936 will result in an elevated risk of NSCPO. However, rs2239936 only exhibited a statistical association with NSCPO in genotypic association analysis ( P = 8.06×10−06) and haplotype analysis ( P = 1.43×10−05). Conclusion This study identified that allele G at rs2239936 in MYH3 gene was significantly associated with NSCPO as a risk factor and MYH3 was a new susceptibility gene for NSCPO in western Han Chinese population.

ObjectivesGenetic studies of nonsyndromic orofacial clefts (NSOC) focus on identifying susceptibility genes and elucidating underlying pathogenesis. A correlation between orofacial clefts and hypertelorism has been established by previous studies, hinting at shared genetic risk factors.Materials and MethodsSyndromes characterized by hypertelorism and orofacial clefts were screened, followed by analysis and selection of their causative genes, resulting in 35 candidate genes. After genotyping quality control, 340 single nucleotide polymorphisms (SNPs) were analyzed. Allelic and genotypic associations were evaluated under the additive model with Bonferroni correction. Linkage disequilibrium and haplotype analysis were performed.ResultsSingle nucleotide polymorphisms that showed significant allelic associations with NSOC subtypes are notably within TRAPPC9, TBX1, and ZIC2 genes. Single nucleotide polymorphisms within TRAPPC9 were primarily associated with nonsyndromic cleft lip and palate (NSCLP), while SNPs within TBX1 exhibited risk and protective effects across NSCLP, nonsyndromic cleft lip only (NSCLO), and nonsyndromic cleft palate only. ZIC2 SNPs were significantly associated with NSCLP and NSCLO. Although 15 SNPs in CDH1 showed allelic association with NSCLO and were in strong linkage disequilibrium, no statistically significant genotypic or haplotypic associations were identified.ConclusionTRAPPC9, TBX1, and ZIC2 are identified as novel susceptibility genes for NSOC in the western Han Chinese population, with TRAPPC9 and TBX1 showing distinct risk and protective effects among subtypes. While further studies are required to confirm their role, these findings deepen our understanding of NSOC genetics, emphasize the importance of subtype analysis, and provide valuable directions for further mechanistic studies.

Colorectal carcinoma (CRC) stands as a pressing global health issue, marked by the unbridled proliferation of immature cells influenced by multifaceted internal and external factors. Numerous studies have explored the intricate mechanisms of tumorigenesis in CRC, with a primary emphasis on signaling pathways, particularly those associated with growth factors and chemokines. However, the sheer diversity of molecular targets introduces complexity into the selection of targeted therapies, posing a significant challenge in achieving treatment precision. The quest for an effective CRC treatment is further complicated by the absence of pathological insights into the mutations or alterations occurring in tumor cells. This study reveals the transfer of signaling from the cell membrane to the nucleus, unveiling recent advancements in this crucial cellular process. By shedding light on this novel dimension, the research enhances our understanding of the molecular intricacies underlying CRC, providing a potential avenue for breakthroughs in targeted therapeutic strategies. In addition, the study comprehensively outlines the potential immune responses incited by the aberrant activation of signaling pathways, with a specific focus on immune cells, cytokines, and their collective impact on the dynamic landscape of drug development. This research not only contributes significantly to advancing CRC treatment and molecular medicine but also lays the groundwork for future breakthroughs and clinical trials, fostering optimism for improved outcomes and refined approaches in combating colorectal carcinoma.

To the Editor: Current therapeutic strategies against breast cancer are limited by poor prognosis and unwanted side effects, especially in triple-negative breast cancer (TNBC). Ginkgetin (Gk), a natural, non-toxic bioflavonoid extracted from Ginkgo biloba leaves, is a promising therapeutic candidate against breast cancer,[1] but the underlying mechanisms remain obscure. In this study, we evaluated the anti-tumor activity of Gk on multiple breast cancer cell lines, including both human (MDA-MB-231, MDA-MB-453, MDA-MB-436, MCF-7, and SUM-159) and mouse (4T1) cell lines in vitro. We next used high-throughput sequencing and bioinformatics analysis to identify the alterations in the expression of microRNA (miRNA) and messenger RNA (mRNA) and search for the potential regulating signaling pathways after Gk treatment. The findings were further validated using the dual luciferase reporter gene assay, Western blotting (WB), etc. Finally, the anti-tumor effect of Gk and the involvement of the miR-122-5p/polypeptide N-acetylgalactosaminyltransferase 10 (GALNT10) axis were assessed in an MDA-MB-231 xenograft mouse model. The anti-proliferative activity of Gk on breast cancer was evaluated with the cell counting kit-8 (CCK-8) assay. As shown in Supplementary Figure 1 and 2A,B, https://links.lww.com/CM9/C112, Gk exhibited dose-dependent cytotoxicity in all the tested cell lines, with the half maximal inhibitory concentration (IC50) ranging from 5.67 μmol/L to 11.5 μmol/L at 48 h and 3.7 μmol/L to 9.3 μmol/L at 72 h. The formula of Gk is C32H22O10 (molecular weight, 566.51) [Supplementary Figure 2C, https://links.lww.com/CM9/C112]. Compared to the control group, Gk treatment reduced the cell numbers and induced apparent morphology changes in breast cancer cells, including cell shrinkage, wrinkled membrane, blebbing, and chromatin condensation [Supplementary Figure 2D, https://links.lww.com/CM9/C112]. We next applied the Annexin V/PI apoptosis detection assay after Gk treatment to investigate the type of cell death. We found that Gk significantly increased the proportion of apoptotic cells (Annexin V+PI− and Annexin V+PI+) dose-dependently [Supplementary Figure 1B and 3A–C, https://links.lww.com/CM9/C112]. These results were supported by TUNEL staining [Supplementary Figure 3D, https://links.lww.com/CM9/C112]. Next, WB was used to examine the expression of apoptosis-related proteins. As shown in Supplementary Figure 1C, https://links.lww.com/CM9/C112, Gk treatment drastically increased the expression of cleaved caspase 3 (CASP3), BAX, and cytochrome C (Cyto C) but reduced the expression of BCL-2 in MDA-MB-231 cells. Results from the wound healing assay indicated that Gk inhibited the migration of MDA-MB-231 cells [Supplementary Figure 3E, https://links.lww.com/CM9/C112]. After exposure to Gk for 24 h, the number of colonies dramatically decreased compared to the untreated group, becoming almost invisible when treated with 20 μmol/L of Gk [Supplementary Figure 1D, https://links.lww.com/CM9/C112]. Therefore, Gk can effectively inhibit cell proliferation and migration and induce cell apoptosis in breast cancer. Since the most-effective apoptosis was observed in MDA-MB-231 cells upon Gk exposure, which can be divided into early- and end-stage apoptosis [Supplementary Figure 1B, https://links.lww.com/CM9/C112], we chose MDA-MB-231 cells treated with 10 μmol/L Gk for the transcriptome sequencing. RNA sequencing results indicated that over 1000 genes were differentially expressed in MDA-MB-231 cells after treatment with Gk compared to the control group [Supplementary Figure 4A, https://links.lww.com/CM9/C112]. Similarly, there are 351 miRNAs whose expression was significantly altered by Gk treatment, including 182 up-regulated and 169 down-regulated miRNAs [Supplementary Figure 1E, https://links.lww.com/CM9/C112]. KEGG and GO enrichment analyses were performed to clarify the functions of those differentially expressed genes (DEGs). The top seven altered terms of the KEGG pathways and GO plots are displayed in Supplementary Figure 4B–E, https://links.lww.com/CM9/C112. The results indicated that plenty of signaling pathways, biological processes, cellular components, and molecular functions were significantly changed by Gk treatment. For instance, as for the biological process, cytoplasmic translation and sensory perception of the light stimulus were overactivated, while other functions such as MAPK cascade, cell junction organization, and regulation of cell adhesion were inhibited by Gk. Since its first discovery in 1993, miRNA has been proven to be critical in almost all biological processes, including cell homeostasis, proliferation, differentiation, survival, and metabolism.[2,3] miRNAs are non-coding nucleic acids composed of 18–25 nucleotides, which can modulate gene expression by inhibiting or degrading their target mRNA.[3] We can see from the volcano plot that the miRNAs showing a log2 fold change of >5 or <−5 include hsa-miR-215-5p, hsa-miR-122-5p, hsa-miR-190b-5p, and hsa-miR-27a-5p, among which has-miR-122-5p changed the most significantly [Supplementary Figure 5A, https://links.lww.com/CM9/C112]. Real-time qPCR (qRT-PCR) results proved that Gk treatment significantly increased the expression of miR-122-5p in a dose-dependent manner [Supplementary Figure 5B, https://links.lww.com/CM9/C112]. To explore the function of miR-122-5p in Gk-mediated anti-tumor effects, MDA-MB-231 cells were transfected with a miRNA inhibitor against miR-122-5p (122i), followed by Gk treatment. Inhibitor NC (NCi) was used as the control. As a result, inhibition of miR-122-5p relieved the apoptosis-inducing [Supplementary Figure 5C, https://links.lww.com/CM9/C112] and anti-proliferative [Supplementary Figure 1F, https://links.lww.com/CM9/C112] activities of Gk. Meanwhile, in the presence of 122i, Gk displayed a reduced ability to prevent the migration of MDA-MB-231 cells [Supplementary Figure 5D, https://links.lww.com/CM9/C112]. These results proved that the anti-tumor effects of Gk on breast cancer cells may be mediated by, at least partly, the up-regulation of miR-122-5p. To ascertain the potential downstream target of miR-122-5p, we intersected the target mRNAs of miR-122-5p with the downregulated genes after Gk treatment, which was confirmed with three databases: "mirecords," "mirtarbase", and "tarbase". Seven genes were screened out, including GALNT10, BCL2L2, UBAP2, AACS, SLC7A1, ENTPD4, and AKT3, whose expressions were highly negatively correlated with that of miR-122-5p [Supplementary Figure 5E,F, https://links.lww.com/CM9/C112]. qRT-PCR and WB assays proved a significantly reduced expression of GALNT10 after Gk treatment [Supplementary Figure 1 and 5G, https://links.lww.com/CM9/C112]. Next, we discovered that transfecting MDA-MB-231 cells with the 122i can rescue the decreased expression of GALNT10 induced by Gk [Supplementary Figure 5H, https://links.lww.com/CM9/C112]. Dual-luciferase reporter gene assay was performed to determine whether the 3′-untranslated regions (UTR) of GALNT10 can be targeted by miR-122-5p. As expected, co-transfection with mimics of miR-122-5p (mimic 122) and wild-type GALNT10 (GALNT10-WT) displayed the lowest luciferase activity, suggesting the strong interaction between miR-122-5p and GALNT10 [Supplementary Figure 1H, https://links.lww.com/CM9/C112]. Therefore, GALNT10 is a downstream target for miR-122-5p in Gk-induced cell death. This is consistent with a previous study reporting that miR-122 regulated GALNT10 expression in chronic hepatitis B virus-associated hepatocarcinogenesis.[4] GALNT10 is an oncoprotein that promotes malignant behaviors of cancer cells like proliferation, apoptosis resistance, and metastatic features.[4,5] We next investigated the function of GALNT10 in breast cancer using its specific small interfering RNA (siRNA). The siRNA significantly knocked down the expression of GALNT10 in MDA-MB-231 cells by over 70% [Supplementary Figure 5I, https://links.lww.com/CM9/C112]. Knocking down GALNT10 expression suppressed the proliferation of MDA-MB-231 cells, increased the proportion of apoptotic cells, and attenuated the invasion and migration of MDA-MB-231 cells [Supplementary Figure 1I and 5J,K, https://links.lww.com/CM9/C112]. Therefore, GALNT10 confers the malignant phenotype of breast cancer cells, whereas Gk treatment can reduce GALNT10 expression by enhancing the expression levels of its upstream regulator miR-122-5p. To find out the downstream effector molecule of GALNT10, pathways associated with GALNT10 gene expression were analyzed in the TCGA breast cancer dataset. The results of GSEA showed that GALNT10 expression was associated with the Wnt signaling pathway regulation [Supplementary Figure 6A, https://links.lww.com/CM9/C112], of which the Smad3 gene in the Wnt signaling pathway was up-regulated after Gk treatment [Supplementary Figure 6B, https://links.lww.com/CM9/C112]. In line with the downregulation of GALNT10, we discovered a dose-dependent reduction in the expression of Smad3 in MDA-MB-231 cells after Gk treatment, including total Smad3 and phosphorylated Smad3 (p-Smad3, Supplementary Figure 6C,D, https://links.lww.com/CM9/C112). Knocking down GALNT10 with siRNA resulted in decreased expression of Smad3, suggesting that Smad3 is a downstream signaling molecule of GALNT10 [Supplementary Figure 6E, https://links.lww.com/CM9/C112]. Since miR-122-5p can down-regulate GALNT10, we speculated that 122i transfection can impact Smad3 expression by regulating GALNT10. As expected, the expression of Smad3 in MDA-MB-231 cells is elevated in the presence of 122i after Gk treatment compared to pre-transfection with NCi [Supplementary Figure 6F, https://links.lww.com/CM9/C112]. These results indicated that Gk could inhibit breast cancer growth by regulating miR-122-5p/GALNT10/Smad3 signaling. To confirm the anti-tumor activity of Gk in vivo, MDA-MB-231 tumor-bearing SCID mice were administered with Gk (30 mg/kg) or vehicle intraperitoneally. As shown in Supplementary Figure 1J and 7A,B, https://links.lww.com/CM9/C112, Gk treatment significantly suppressed tumor growth compared to the vehicle group. No statistically significant changes between groups in body weight were observed after treatment [Supplementary Figure 7C, https://links.lww.com/CM9/C112], indicating the acceptable safety of Gk. To validate the involvement of the miR-122-5p/GALNT10 axis in vivo, we isolated the total miRNAs from the resected tumors after Gk treatment and assessed the expression of miR-122-5p. The expression of miR-122-5p was significantly elevated after Gk treatment [Supplementary Figure 1K, https://links.lww.com/CM9/C112]. At the same time, a downward trend was observed in the expression of GALNT10 mRNA in the tumor tissue from Gk-treated mice [Supplementary Figure 7D, https://links.lww.com/CM9/C112]. The immunofluorescence assay also revealed a dramatic decrease in the expression of GALNT10 protein in the tumor tissues in the Gk group compared to the control group [Supplementary Figure 1L, https://links.lww.com/CM9/C112]. Conclusively, Gk is a promising anti-cancer agent that inhibits tumor growth both in vitro and in vivo. High-throughput sequencing and bioinformatic analysis revealed gene alterations after Gk treatment and uncovered the critical role of miRNA-122-5p/GALNT10/Smad3 signaling, which were further supported by the results from various experiments. These results added great value for future work on Gk as a candidate agent in breast cancer therapy, including but not limited to searching for combination strategies to improve the efficacy. Funding This study was supported by the Natural Science Foundation of Sichuan Province (No. 2022NSFSC0774) and the National Natural Science Foundation of China (No. 82102896). Conflicts of interest None.

OBJECTIVES:The objective of this study was to explore the relationship between fibroblast growth factor/receptor (FGF/FGFR) and non-syndromic orofacial cleft (NSOC) in individuals of Han Chinese.DESIGN:Initially, we performed RNA-Seq between non-syndromic cleft lip only (NSCLO) or non-syndromic cleft palate only (NSCPO) and control groups. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were carried out to evaluate the functions of differentially expressed genes (DEGs) of FGF/FGFR. Then, we selected the most significant DEG FGFR2 and performed an association analysis in Chinese. Linkage disequilibrium (LD) and haplotype analyses were performed with HaploView and PLINK. Additional bioinformatics functional prediction for the notable single nucleotide polymorphisms was performed with HaploReg V4.1 and 3DSNP.RESULTS:Finally, we identified 32 mRNAs related to FGF/FGFR via RNA-Seq and chose FGFR2 in the subsequent association analysis. Results indicated that the single nucleotide polymorphism (SNP) rs2288336 in FGFR2 contributed significantly to both non-syndromic cleft lip with or without cleft palate (NSCL/P) and NSCLO, with p values of 5.00E-05 (OR = 0.79, 95% CI: 0.70-0.88) and 1.38E-04 (OR = 0.76, 95% CI: 0.65-0.87), respectively. In addition, rs3793893 in FGFR2 was found to be associated with NSCLO, with a p value of 1.02E-04 (OR = 0.67, 95% CI: 0.55-0.82).CONCLUSIONS:Our research demonstrated that FGFR2 is significantly more involved in NSOC than other FGF/FGFRs in Chinese and further identified rs2288336 and rs3793893 in FGFR2 associated with NSOC subtypes, which provide further evidence for the genetic etiology of NSOC in Han Chinese.

Molecular target inhibitors have been regularly approved by Food and Drug Administration (FDA) for tumor treatment, and most of them intervene in tumor cell proliferation and metabolism. The RAS–RAF–MEK–ERK pathway is a conserved signaling pathway that plays vital roles in cell proliferation, survival, and differentiation. The aberrant activation of the RAS–RAF–MEK–ERK signaling pathway induces tumors. About 33% of tumors harbor RAS mutations, while 8% of tumors are driven by RAF mutations. Great efforts have been dedicated to targeting the signaling pathway for cancer treatment in the past decades. In this review, we summarized the development of inhibitors targeting the RAS–RAF–MEK–ERK pathway with an emphasis on those used in clinical treatment. Moreover, we discussed the potential combinations of inhibitors that target the RAS–RAF–MEK–ERK signaling pathway and other signaling pathways. The inhibitors targeting the RAS–RAF–MEK–ERK pathway have essentially modified the therapeutic strategy against various cancers and deserve more attention in the current cancer research and treatment.

For patients with esophageal squamous cell carcinoma (ESCC), standard therapeutic methods (cisplatin and radiotherapy) have been found to be ineffective and severely toxic. Targeted therapy emerges as a promising solution for this dilemma. It has been reported that targeted therapies are applied alone or in combination with standard conventional therapies for the treatment of a variety of cancers. To the best of our knowledge, in patients with ESCC, the combinational methods containing standard therapy and ERK-targeted therapy have yet to be explored. To analyze the prognostic role of p-ERK in ESCC patients, the Kaplan-Meier analysis and Cox regression model were used. To assess the effects of ERK-targeted therapy (GDC0994) on ESCC cells, in vitro studies including CCK-8 assay, colony formation assay, and scratch wound healing assay were conducted. In addition, the changes in cell cycle distribution and apoptosis were analyzed by flow cytometry. Besides, to assess the efficacy of different therapies in vivo, the xenograft tumor models were established by subcutaneously inoculating tumor cells into the flank/leg of mice. In patients with ESCC, a strong correlation between the high expression level of p-ERK and the poor prognosis (p < 0.01, Log-Rank test) has been identified. By analyzing the results from CCK-8 and scratch wound healing assays, we demonstrated that the ERK inhibitor repressed the viability and migration of ESCC cells. In addition, following the treatment of GDC0994, the volumes of xenograft tumors significantly decreased (p < 0.001, one-way ANOVA). Furthermore, blocking the mitogen-activated protein kinase (MAPK/ERK) pathway enhanced the therapeutic efficacy of both cisplatin and radiotherapy (p < 0.05). These findings imply the role of p-ERK in the prognosis of ESCC patients and the therapeutic value of ERK inhibitors in ESCC.

Objectives Considering limitations of previous studies and differences across populations and subtypes, this study aimed to identify new potential SNPs around IRF6 associated with non-syndromic orofacial cleft (NSOC) in Western Han Chinese. Materials and Methods We recruited 376 NSOC case-parent trios, including 125 non-syndromic cleft lip only (NSCLO) trios, 151 non-syndromic cleft lip and palate (NSCLP) trios, and 100 non-syndromic cleft palate only (NSCPO) trios. Twenty-two single-nucleotide polymorphisms (SNPs) were genotyped using MassARRAY method. Hardy-Weinberg equilibrium test, allelic transmission disequilibrium test (TDT) analysis, sliding-window haplotype TDT analysis, and tests for parent-of-origin effect were performed using the PLINK software. Pairwise linkage disequilibrium (LD) was computed using the Haploview program. Results In TDT analysis, allele A at rs17015217 (p = 0.00011, OR = 0.61 and 95% CI: 0.47-0.78) and allele T at rs12080691 (p = 0.00011, OR = 0.61 and 95% CI: 0.47-0.78) were under-transmitted among NSCLO trios but over-transmitted among NSCPO trios. Haplotypes showing evidence of under-transmission in NSCLO trios were over-transmitted in NSCPO trios. In tests for parent-of-origin effects, T allele at rs12080691 presented paternal under-transmission among NSCLO trios but over-transmission among NSCPO trios. Conclusions Allele A at rs17015217 and allele T at rs12080691 are associated with NSCLO and NSCPO with potential to have opposite effects on two subtypes in this sample from Western Han Chinese.

OBJECTIVES:This study aimed to reveal the association between single-nucleotide polymorphism of NTN1 and subtypes of non-syndromic cleft lip with or without cleft palate (NSCL/P) in Han Chinese Population.DESIGN:Initially, we selected three single-nucleotide polymorphisms (SNP) (rs4791331, rs4791774 and rs9891446) in NTN1 from previous genetic studies. Then we recruited two Han Chinese cohorts (2004 cases and 1823 controls) and divided cases into subgroups: non-syndromic right-side cleft lip, non-syndromic left-side cleft lip, non-syndromic bilateral cleft lip and non-syndromic cleft lip with palate to further evaluate the associations between the subtypes of NSCL/P and SNPs in NTN1. PLINK and Haploview program were utilized to analyze the data.RESULTS:In the association analysis under additive model, we found that G allele at rs9891446 could specifically increase the risk of right-side cleft lip (P = 0.0073, OR = 1.44, 95%CI: 1.1-1.88), which was consistent with the results of association analysis under genotypic model.CONCLUSION:This study showed that rs9891446 of NTN1 was specifically associated with right-side cleft lip in Han Chinese, which indicates that different subtypes of non-syndromic cleft lip have distinct genetic background.

Objectives Non-syndromic cleft palate only (NSCPO) is a common congenital deformity with complex etiologies. GRHL3, FAF1, and KCNJ2 have been reported to be involved in the pathogenesis of NSCPO. Up till now, there have been no replication studies based on large Han Chinese. Therefore, this study aimed to investigate associations between GRHL3, FAF1, KCNJ2, and NSCPO sub-phenotypes patients in Han Chinese. Materials and Methods Firstly, we selected 2 SNPs based on previous literatures: FAF1 (rs3827730) and GRHL3 (rs41268753). Also, we selected 8 tagSNPs in GRHL3 (rs557811, rs609352, rs10903078, rs6659209, rs12401714, rs12568599, rs3887581, rs12024148) and 2 tagSNPs in KCNJ2 (rs75855040 and rs236514). Afterward, we evaluated these SNPs among 1668 NSCPO patients and 1811 normal controls from Han Chinese. Following data were analyzed by PLINK and Haploview program. Results Association analysis under additive model showed that allele A at rs12568599 in GRHL3 gene is significantly associated with NSCPO (p = 0.0034, OR = 1.38 and 95%CI: 1.11-1.72) and its sub-phenotype incomplete cleft palate (ICP) (p = 0.0039, OR = 1.4 and 95%CI: 1.11-1.75), and it could increase the risk of both NSCPO and ICP. Conclusions This study firstly found that rs12568599 in GRHL3 is associated with NSCPO and ICP in Han Chinese, indicating that sub-phenotypes of NSCPO have different etiologies.

The purpose of this study is to analyze the clinical characteristics of a Treacher Collins syndrome (TCS) patient carrying a de novo variant of TCOF1, and briefly analyze the correlation between genetic results and clinical features. Also, the pathogenesis and clinical treatment of TCS are reviewed. A Chinese pedigree with TCS containing 8 members was enrolled. Phenotype of the proband was evaluated by a surgeon, then whole exome sequencing of the proband was performed. Then we verified the proband-derived variants by Sanger sequencing in the pedigree. Correlation between genotype and phenotype was analyzed. The study was conducted in a stomatological hospital. A Chinese pedigree with TCS containing 8 members. To ascertain the genetic variants in the Chinese pedigree with TCS. Blood samples were collected. We reported a case of typical TCS with a de novo missense variant (NM_001371623.1:c.38T>G, p.(Leu13Arg)) in exon 1 of TCOF1, who presented asymmetrical facial abnormalities, including downward slanting of the palpebral fissures, sparse eyebrows, lateral tilt of the eyeballs, bilateral external ears deformities, hypoplasia of midface, reduction of the zygomatic body, bilateral orbital invagination, right external auditory canal atresia, mandibular ramus short deformity, cleft palate and the whole face was convex. This research found a novel variant of TCS in Chinese, expanding the spectrum of TCS pathogenic variants. Genetic results combined with clinical phenotype can make a definite diagnosis and provide genetic counseling for the family.

OBJECTIVE:The role of MMP16 in lip development is unclear. This study aimed to identify nonsyndromic cleft lip with or without palate (NSCL ± P) susceptible loci of MMP16 in western Han Chinese. DESIGN:We performed targeted sequencing around MMP16 combined with a 2-phase association analysis on common variants. Phase 2 association analysis was performed with NSCL ± P specific subphenotypes (NSCL and NSCLP). Then we used rare variants burden analysis and genotyping, accompanied by motif analysis. SETTING:This study was completed in a tertiary medical center. PATIENTS, PARTICIPANTS:Phase 1 targeted sequencing included 159 patients with NSCL ± P and 542 normal controls; phase 2 included 1626 patients with NSCL ± P (1047 NSCL and 579 NSCLP) and 2255 normal controls. INTERVENTIONS:Venous blood samples were collected from patients and used to extract DNA. MAIN OUTCOME MEASURES:After Bonferroni correction, phase 1 significant threshold of p-value was 4.28 × 10-5 (0.05/1167 single nucleotide polymorphisms [SNPs]), and phase 2 was .00025 (0.05/200 SNPs). Burden analysis significant threshold p-value was .05. RESULTS:Common variants phase 1 association analysis identified 11 statistically significant SNPs (lowest p = 1.90 × 10-9, odds ratio (OR) = 0.27, 95% CI: 0.17-0.44), phase 2 replication identified 16 SNPs in NSCL ± P (lowest p = 6.26 × 10-6, OR = 0.77, 95% CI: 0.69-0.86) and 9 in NSCL (lowest p = 8.44 × 10-5, OR = 0.76, 95% CI: 0.66-0.87). Rare variants burden analysis showed no significant results, genotyping results showed they were maternally inherited. CONCLUSIONS:Our study identified MMP16 susceptible SNPs in NSCL ± P and NSCL, emphasizing its potential role in lip development. Our study also highlighted the importance to perform association analysis with subphenotypes divided.

Graphene has important application prospects in many fields, and is considered a revolutionary material in the future. The biocompatibility of graphene and its safety issues have drawn much attention. Possible toxicities of a nanomaterial might be concerned with cell deaths, releasing of damage-associated molecular patters (DAMPs), inflammation, risks of tumorigenesis etc. In the current study, we investigated the long-term pulmonary exposure model of graphene and carbon black and discovered that long-term pulmonary exposure of the materials led to lung cancer metastasis and progression. Notably, pulmonary exposure of graphene or carbon black induced cell necrosis and the release of the DAMP, namely, mitochondrial DNA (mtDNA), which is a potent stimulator for elevating the secretion of Wnt ligands in alveolar macrophages (AMs). The Wnt-activating macrophages contributed to the formation of premetastatic niches in lung tissues by enhancing the stemness potential of cancer cells and promoting their generation and proliferation. The growth of metastatic cancer induced by exposure of carbon-based nanomaterials could be effectively inhibited by treating mice with a Wnt inhibitor. Taken together, our findings elucidate the how the pulmonary cell deaths induced by graphene reshaped the tumor microenvironment by releasing DAMPs and promoting tumor metastasis, which throws the light on the relationship between nanomaterial toxicity and its tumorigenesis risk.

唇腭裂是最常见的先天畸形之一.根据是否伴有全身其他部位的先天疾病,可分为非综合征型唇腭裂和综合征型唇腭裂.研究发现非综合征型唇腭裂为多因素遗传病,同时受多种遗传和环境危险因素的影响.而综合征型唇腭裂人群发病率相对较低,为单基因病,符合孟德尔遗传定律,呈明显的家族遗传性,且大多有明确的致病基因.本文将就5种发病率较高,较为常见的综合征型唇腭裂近年来遗传学方面的进展进行介绍,为将来综合征型唇腭裂遗传研究提供参考.

Background. Several studies have focused on the prognostic role of microRNA 222 in glioma. But different conclusions were drawn by these studies. We aimed to systematically evaluate the role of microRNA 222 in glioma by conducting a meta-analysis.Methods. A systematic literature search until January 2020 was conducted in Web of Science, EMBASE, Cochrane Library, PubMed, and China National Knowledge Infrastructure. The general characteristics and relevant data of nine articles were extracted. Hazard ratios (HRs) with 95% confidence intervals (CIs) were applied to evaluate the prognostic role of microRNA 222 in glioma. The primary outcomes were overall survival (OS) and disease-free survival (DFS).Results. Nine articles (11 data sets) with 1564 patients were included. We systematically evaluated the role of microRNA 222 for OS and DFS in glioma patients (HR forOS=1.72; 95% CI, 1.31-2.26;p=0.001; HR forDFS=1.02; 95% CI, 0.86-1.22;p=0.032). Subgroup analyses were performed according to the sources of patients, the types of the samples, the stages of the tumors, the methods for detecting the microRNA 222, and the sample size. No significant publication bias was found.Conclusion. In conclusion, our study provided evidence that a high expression of microRNA 222 was related to worse overall survival in glioma patients. However, given the limited study number, more high-quality studies are warranted in the future.

Triple-negative breast cancer (TNBC) is the most aggressive type of breast cancer and still lacks available targeted therapy options. Thioridazine hydrochloride (Thi-hyd), a common phenothiazine antipsychotic drug, has shown great tumor-suppressive potential in several types of cancer cells. In this study, we explored the anti-tumor effects of Thi-hyd on TNBC. Thi-hyd significantly inhibited the viability and migration of TNBC cells in vitro. PI3K/AKT signaling pathway was verified to be related to Thi-hyd-induced tumor inhibition. G0/G1 cell cycle arrest and apoptosis were observed after treatment with Thi-hyd. The cell cycle arrest was accompanied by down-regulation of CDK4/cyclin D1 and up-regulation of p21 and p27. The apoptosis was accompanied by mitochondrial dysfunction. In vivo, Thi-hyd significantly suppressed the growth of tumors showing a 63.73% inhibition rate in tumor weight. Spontaneous lung metastasis was also efficiently prevented by Thi-hyd treatment, with a 72.58% inhibition rate. Immunohistological analysis showed increased Ki67 expression and reduced cleaved caspase-3 expression in Thi-hyd-treated mice. No apparent side effects were observed according to blood test and H&E staining results. Collectively, our results show that Thi-hyd can be used as a potential drug for TNBC treatment and set the basis for its clinical evaluation and investigation.

Background: This study aimed to systematically assess the prognostic value of lymphocyte monocyte ratio (LMR) in patients with ovarian cancer through performing a meta-analysis. Methods: Web of Science, PubMed, EMBASE, Cochrane Library, and China National Knowledge Infrastructure databases were searched for potentially eligible studies. The baseline characteristics and relevant data were extracted. Hazard ratios with 95% confidence intervals (CIs) were combined to assess the prognostic value of LMR in patients with ovarian cancer. Results: Nine studies enrolling 2809 patients were included. The pooled hazard ratios of lower LMR for overall survival and progression free survival in patients with ovarian cancer were 1.71 (95% CI, 1.40-2.09) and 1.68 (95% CI, 1.49-1.88), respectively. Subgroup analysis and sensitivity analysis were also performed. No significant publication bias was found. Conclusion: Our results suggested that lower LMR was associated with poorer overall survival and progression free survival in patients with ovarian cancer. The findings may assist prognosis evaluation and future research on therapies based on modulating host immune response in ovarian cancer.