Cross-sectional network analysis of plasma proteins/metabolites correlated with pathogenesis and therapeutic response in acute promyelocytic leukemia

The treatment of PML/RARA+acute promyelocytic leukemia(APL)with all-trans-retinoic acid and arsenic trioxide(ATRA/ATO)has been recognized as a model for translational medicine research.Though an altered microenvironment is a general cancer hallmark,how APL blasts shape their plasma composition is poorly understood.Here,we reported a cross-sectional correlation network to interpret multilayered datasets on clinical parameters,proteomes,and metabolomes of paired plasma samples from patients with APL before or after ATRA/ATO induction therapy.Our study revealed the two prominent features of the APL plasma,suggesting a possible involvement of APL blasts in modulating plasma composition.One was characterized by altered secretory protein and metabolite profiles correlating with heightened proliferation and energy consumption in APL blasts,and the other featured APL plasma-enriched proteins or enzymes catalyzing plasma-altered metabolites that were potential trans-regulatory targets of PML/RARA.Furthermore,results indicated heightened interferon-gamma signaling characterizing a tumor-suppressing function of the immune system at the first hematological complete remission stage,which likely resulted from therapy-induced cell death or senescence and ensuing supraphysiological levels of intracellular proteins.Overall,our work sheds new light on the pathophysiology and treatment of APL and provides an information-rich reference data cohort for the exploratory and translational study of leukemia microenvironment.

Catalytic activity of Setd2 is essential for embryonic development in mice: establishment of a mouse model harboring patient-derived Setd2 mutation

SETD2 is the only enzyme responsible for transcription-coupled histone H3 lysine 36 trimethylation(H3K36me3).Mutations in SETD2 cause human diseases including cancer and developmental defects.In mice,Setd2 is essential for embryonic vascular remodeling.Given that many epigenetic modifiers have recently been found to possess noncatalytic functions,it is unknown whether the major function(s)of Setd2 is dependent on its catalytic activity or not.Here,we established a site-specific knockin mouse model harboring a cancer patientderived catalytically dead Setd2(Setd2-CD).We found that the essentiality of Setd2 in mouse development is dependent on its methyltransferase activity,as the Setd2 CD/CD and Setd2^(−/−)mice showed similar embryonic lethal phenotypes and largely comparable gene expression patterns.However,compared with Setd2^(−/−),the Setd2 CD/CD mice showed less severe defects in allantois development,and single-cell RNA-seq analysis revealed differentially regulated allantois-specific 5′Hoxa cluster genes in these two models.Collectively,this study clarifies the importance of Setd2 catalytic activity in mouse development and provides a new model for comparative study of previously unrecognized Setd2 functions.

Spatial metabolomics highlights metabolic reprogramming in acute myeloid leukemia mice through creatine pathway

Acute myeloid leukemia(AML)is recognized as an aggressive cancer that is characterized by significant metabolic reprogramming.Here,we applied spatial metabolomics to achieve high-throughput,in situ identification of metabolites within the liver metastases of AML mice.Alterations at metabolite and protein levels were further mapped out and validated by integrating untargeted metabolomics and proteomics.This study showed a downregulation in arginine's contribution to polyamine biosynthesis and urea cycle,coupled with an upregulation of the creatine metabolism.The upregulation of creatine synthetases Gatm and Gamt,as well as the creatine transporter Slc6a8,resulted in a marked accumulation of creatine within tumor foci.This process further enhances oxidative phosphorylation and glycolysis of leukemia cells,thereby boosting ATP production to foster proliferation and infiltration.Importantly,we discovered that inhibiting Slc6a8 can counter these detrimental effects,offering a new strategy for treating AML by targeting metabolic pathways.

Systemic and dynamic immune landscape of Omicron-infected subjects treated with Lianhua Qingwen capsules

To the Editor:The COVID-19 has represented an unexpected global public health challenge due to the profound immune evasion and transmission capabilities of the responsible virus SARS-CoV-21,being in great short of effective prophylactic and curative methods.Lianhua Qingwen(LHQW)capsule,a traditional Chinese medicine(TCM)formula,is officially included in China's COVID-19 treatment guidelines and has shown antiviral activity in vitro and in vivo.Recent clinical trials have shown that LHQW can alleviate symptoms and accelerate recovery in Omicron-infected patients2.The immune pathology of COVID-19 involves a complex interplay between virus-induced injuries and multifactorial immune responses.However,a comprehensive analysis of the immunological basis of LHQW's effects in Omicron-infected subjects is still lacking.

Chemical compound cinobufotalin potently induces FOXO1-stimulated cisplatin sensitivity by antagonizing its binding partner MYH9

In this study,we present novel molecular mechanisms by which FOXO1 functions as a tumor suppressor to prevent the pathogenesis of nasopharyngeal carcinoma(NPC).First,we observed that FOXO1 not only controlled tumor stemness and metastasis,but also sensitized NPC cells to cisplatin(DDP)in vitro and in vivo.Mechanistic studies demonstrated that FOXO1-induced miR-200b expression through the GSK3β/β-catenin/TCF4 network-mediated stimulation of ZEB1,which reduced tumor stemness and the epithelial–mesenchymal transition(EMT)signal.Furthermore,we observed FOXO1 interaction with MYH9 and suppression of MYH9 expression by modulating the PI3K/AKT/c-Myc/P53/miR-133a-3p pathway.Decreased MYH9 expression not only reduced its interactions with GSK3β,but also attenuated TRAF6 expression,which then decreased the ubiquitin-mediated degradation of GSK3βprotein.Increased GSK3βexpression stimulated theβ-catenin/TCF4/ZEB1/miR-200b network,which increased the downstream tumor stemness and EMT signals.Subsequently,we observed that chemically synthesized cinobufotalin(CB)strongly increased FOXO1-induced DDP chemosensitivity by reducing MYH9 expression,and the reduction in MYH9 modulated GSK3β/β-catenin and its downstream tumor stemness and EMT signal in NPC.In clinical samples,the combination of low FOXO1 expression and high MYH9 expression indicated the worst overall survival rates.Our studies demonstrated that CB potently induced FOXO1-mediated DDP sensitivity by antagonizing its binding partner MYH9 to modulate tumor stemness in NPC.

A systematic survey of LU domain-containing proteins reveals a novel human gene,LY6A,which encodes the candidate ortholog of mouse Ly-6A/Sca-1 and is aberrantly expressed in pituitary tumors

The Ly-6 and uPAR(LU)domain-containing proteins represent a large family of cell-surface markers.In particular,mouse Ly-6A/Sca-1 is a widely used marker for various stem cells;however,its human ortholog is missing.In this study,based on a systematic survey and comparative genomic study of mouse and human LU domain-containing proteins,we identified a previously unannotated human gene encoding the candidate ortholog of mouse Ly-6A/Sca-1.This gene,hereby named LY6A,reversely overlaps with a lncRNA gene in the majority of exonic sequences.We found that LY6A is aberrantly expressed in pituitary tumors,but not in normal pituitary tissues,and may contribute to tumorigenesis.Similar to mouse Ly-6A/Sca-1,human LY6A is also upregulated by interferon,suggesting a conserved transcriptional regulatory mechanism between humans and mice.We cloned the full-length LY6A cDNA,whose encoded protein sequence,domain architecture,and exon‒intron structures are all well conserved with mouse Ly-6A/Sca-1.Ectopic expression of the LY6A protein in cells demonstrates that it acts the same as mouse Ly-6A/Sca-1 in their processing and glycosylphosphatidylinositol anchoring to the cell membrane.Collectively,these studies unveil a novel human gene encoding a candidate biomarker and provide an interesting model gene for studying gene regulatory and evolutionary mechanisms.

iTRAQ-based quantitative proteomic analysis on differentially expressed proteins of rat mandibular condylar cartilage induced by reducing dietary loading

As muscle activity during growth is considerably important for mandible quality and morphology, reducing dietary loading directly influences the development and metabolic activity of mandibular condylar cartilage （MCC）. However, an overall investigation of changes in the protein composition of MCC has not been fully described in literature. To study the protein expression and putative signaling in vivo, we evaluated the structural changes of MCC and differentially expressed proteins induced by reducing functional loading in rat MCC at developmental stages. Isobaric tag for relative and absolute quantitation-based 2D nano-high performance liquid chromatography （HPLC） and matrix-assisted laser desorption/ionization time-of-flight/ time-of-flight （MALDI-TOF/TOF） technologies were used. Global protein profiling, KEGG and PANTHER pathways, and functional categories were analyzed. Consequently, histological and tartrate-resistant acid phosphatase staining indicated the altered histological structure of condylar cartilage and increased bone remodeling activity in hard-diet group. A total of 805 differentially expressed proteins were then identified. GO analysis revealed a significant number of proteins involved in the metabolic process, cellular process, biological regulation, localization, developmental process, and response to stimulus. KEGG pathway analysis also suggested that these proteins participated in various signaling pathways, including calcium signaling pathway, gap junction, ErbB signaling pathway, and mitogen-activated protein kinase signaling pathway. Collagen types I and II were further validated by immunohistochemical staining and Western blot analysis. Taken together, the present study provides an insight into the molecular mechanism of regulating condylar growth and remodeling induced by reducing dietary loading at the protein level.

Disabled homolog 2 is required for migration and invasion of prostate cancer cells

Disabled homolog 2 （DAB2） is frequently deleted or epigenetically silenced in many human cancer cells＇ Therefore, DAB2 has always been regarded as a tumor suppressor gene. However, the role of DAB2 in tumor progression and metastasis remains unclear. In this study, DAB2 expression was upregulated along with human prostate cancer （PCa） progression. DAB2 overexpression or knockdown effects in LNCaP and PC3 cell lines were verified to address the biological functions of DAB2 in PCa progression and metastasis. LNCaP and PC3 cell lines were generated from human PCa cells with low and high metastatic potentials, respectively. The results showed that DAB2 shRNA knockdown can inhibit the migratory and invasive abilities of PC3 cells, as well as the tumorigenicity, whereas DAB2 overexpression enhanced LNCaP cell migration and invasion. Further investigation showed that DAB2 regulated the cell migration associated genes in PC3 cells, and the differential DAB2 expression between LNCaP and PC3 cells was partly regulated by histone 4 acetylation. Therefore, DAB2 may play an important role in PCa progression and metastasis.

Intra-heterogeneity in transcription and chemoresistant property of leukemia-initiating cells in murine Setd2^(-/-) acutemyeloid leukemia

Background:Heterogeneity of leukemia-initiating cells(LICs)is a major obstacle in acute myeloid leukemia(AML)therapy.Accumulated evidence indicates that the coexistence of multiple types of LICs with different pathogenicity in the same individual is a common feature in AML.However,the functional heterogeneity including the drug response of coexistent LICs remains unclear.Therefore,this study aimed to clarify the intra-heterogeneity in LICs that can help predict leukemia behavior and develop more effective treatments.Methods:Spleen cells from the primary Setd2^(-/-)-AML mouse were transplanted into C57BL/6 recipient mice to generate a transplantable model.Flow cytometry was used to analyze the immunophenotype of the leukemic mice.Whole-genome sequencing was conducted to detect secondary hits responsible for leukemia transformation.A serial transplantation assay was used to determine the self-renewal potential of Setd2^(-/-)-AML cells.A limiting-dilution assay was performed to identify the LIC frequency in different subsets of leukemia cells.Bulk and single-cell RNA sequencing were performed to analyze the transcriptional heterogeneity of LICs.Small molecular inhibitor screening and in vivo drug treatment were employed to clarify the difference in drug response between the different subsets of LICs.Results:In this study,we observed an aged Setd2^(-/-)mouse developing AML with co-mutation of Nras^(G12S) and Braf^(K520E).Further investigation identified two types of LICs residing in the c-Kit^(+)B220^(+)Mac-1^(-)and c-Kit^(+)B220^(+)Mac-1^(+)subsets,respectively.In vivo transplantation assay disclosed the heterogeneity in differentiation between the coexistent LICs.Besides,an intrinsic doxorubicinresistant transcriptional signature was uncovered in c-Kit^(+)B220^(+)Mac-1^(+)cells.Indeed,doxorubicin plus cytarabine(DA),the standard chemotherapeutic regimen used in AML treatment,could specifically kill c-Kit^(+)B220^(+)Mac-1^(−)cells,but it hardly affected c-Kit^(+)B220^(+)Mac-1^(+)cells.Transcriptome analysis unveiled a higher activation of RAS downstream signaling pathways in c-Kit^(+)B220^(+)Mac-1^(+)cells than in c-Kit^(+)B220^(+)Mac-1^(-)cells.Combined treatmentwithDAand RAS pathway inhibitors killed both c-Kit^(+)B220^(+)Mac-1^(−)and c-Kit^(+)B220^(+)Mac-1^(+)cells and attenuated disease progression.Conclusions:This study identified two cell subsets enriched for LICs inmurine Setd2^(-/-)-AML and disclosed the transcriptional and functional heterogeneity of LICs,revealing that the coexistence of different types of LICs in thismodel brings about diverse drug response.