Global characterization of modifications to the charge isomers of IgG antibody

Posttranslational modifications of antibody products affect their stability,charge distribution,and drug activity and are thus a critical quality attribute.The comprehensive mapping of antibody modifications and different charge isomers(CIs)is of utmost importance,but is challenging.We intended to quantitatively characterize the posttranslational modification status of CIs of antibody drugs and explore the impact of posttranslational modifications on charge heterogeneity.The CIs of antibodies were fractionated by strong cation exchange chromatography and verified by capillary isoelectric focusing-whole column imaging detection,followed by stepwise structural characterization at three levels.First,the differences between CIs were explored at the intact protein level using a top-down mass spectrometry approach;this showed differences in glycoforms and deamidation status.Second,at the peptide level,common modifications of oxidation,deamidation,and glycosylation were identified.Peptide mapping showed nonuniform deamidation and glycoform distribution among CIs.In total,10 N-glycoforms were detected by peptide mapping.Finally,an in-depth analysis of glycan variants of CIs was performed through the detection of enriched glycopeptides.Qualitative and quantitative analyses demonstrated the dynamics of 24 N-glycoforms.The results revealed that sialic acid modification is a critical factor accounting for charge heterogeneity,which is otherwise missed in peptide mapping and intact molecular weight analyses.This study demonstrated the importance of the comprehensive analyses of antibody CIs and provides a reference method for the quality control of biopharmaceutical analysis.

Identification of hedgehog signaling as a potential oncogenic driver in an aggressive subclass of human hepatocellular carcinoma: A reanalysis of the TCGA cohort

Hepatocellular carcinoma(HCC)is a heterogeneous disease and the second most common cause of cancer-related death worldwide.Marked developments in genomic technologies helped scientists to understand the heterogeneity of HCC and identified multiple HCC-related molecular subclasses.An integrative analysis of genomic datasets including 196 patients from The Cancer Genome Atlas(TCGA)group has recently reported a new HCC subclass,which contains three subgroups(iCluster1,iCluster2,and iCluster3).However,the transcriptional molecular characteristics underlying the iClusters have not been thoroughly investigated.Herein,we identified a more aggressive subset of HCC patients in the iCluster1,and re-clustered the TCGA samples into novel HCC subclasses referred to as aggressive(Ag),moderate-aggressive(M-Ag),and less-aggressive(L-Ag)subclasses.The Ag subclass had a greater predictive power than the TCGA iCluster1,and a higher level of alpha fetoprotein,microscopic vascular invasion,immune infiltration,isocitrate dehydrogenase 1/2 mutation status,and a worse survival than MAg and L-Ag subclasses.Global transcriptomic analysis showed that activation of hedgehog signaling in the Ag subclass may play key roles in tumor development of aggressive HCC.GLI1,a key transcriptional regulator of hedgehog signaling upregulated in the Ag subclass,was correlated with poor prognosis of HCC,and may be a potential prognostic biomarker and therapeutic target for Ag subclass HCC patients.

A time-resolved proteotranscriptomics atlas of the human placenta reveals pan-cancer immunomodulators

Dear Editor,The placenta separates fetus and mother,with the trophoblast playing a most important role in defense against potential harm from the maternal immune system.Although the placenta is normal tissue,it shares several common features with malignant cells.1 Based on similar strategies,these cells are able to successfully coexist in an immunologically hostile environment.2 A detailed analysis and comparison of different placental model systems will not only contribute to our in-depth understanding of the exciting field of placental development research but also may offer valuable insights on the broad field of cancer studies.