EMX2 inhibits clear cell renal cell carcinoma progress via modulating Akt/FOXO3a pathway.

Empty spiracles homeobox 2 (EMX2) is initially identified as a key transcription factor that plays an essential role in the regulation of neuronal development and some brain disorders. Recently, several studies emphasized that EMX2 could as a tumor suppressor, but its role in human clear cell renal cell carcinoma (ccRCC) remains unclear. In the present study, we investigated the role and underlying mechanism of EMX2 in the regulation of ccRCC progress. Our results demonstrated that EMX2 expression was markedly decreased in ccRCC tissues and cell lines, and low EMX2 expression predicted the poor prognosis of ccRCC patients. In addition, forced expression of EMX2 significantly inhibited the cell growth, migration, and invasion in vitro, as well as ccRCC tumor growth in nude mice, via, at least in part, regulating Akt/FOXO3a pathway. In detail, EMX2 could attenuate the phosphorylation levels of Akt and FOXO3a, and increase FOXO3a expression without affecting total Akt expression in vivo and in vitro. Meanwhile, shRNA-mediated knockdown of FOXO3a expression could obviously attenuate the effects of EMX2 on cell growth, migration, invasion, and tumor growth. Furthermore, EMX2 could significantly attenuate the interaction between Akt and FOXO3a. Taken together, our results demonstrated that EMX2 could inhibit ccRCC progress through, at least in part, modulating Akt/FOXO3a signaling pathway, thus representing a novel role and underlying mechanism of EMX2 in the regulation of ccRCC progress.

ß-Arrestin2 promotes docetaxel resistance of castration-resistant prostate cancer via promoting hnRNP A1-mediated PKM2 alternative splicing.

PURPOSE: To investigate the influence of ß-arrestin2 on the docetaxel resistance in castration-resistant prostate cancer (CRPC) and elucidate the underlying molecular mechanisms. METHODS: PC3 and DU145 cells with stable ß-arrestin2 overexpression and C4-2 cells with stable ß-arrestin2 knockdown, were constructed via using lentivirus and puromycin selection. MTT and colony formation assays were carried out to investigate the effect of ß-arrestin2 expression on the docetaxel resistance of CRPC cells. Glycolysis analysis was used to assess the glycolytic capacity modulated by ß-arrestin2. GO enrichment analysis, gene set enrichment analysis and Spearman correlation test were carried out to explore the potential biological function and mechanism via using public data from GEO and TCGA. The expressions of PKM2, Phospho-PKM2, Phospho-ERK1/2 and hnRNP A1 were detected by western blot. Functional blocking experiments were carried out to confirm the roles of PKM2 and hnRNP A1 in the regulation of ß-arrestin2's biological functions via silencing PKM2 or hnRNP A1 expression in cells with stable ß-arrestin2 overexpression. Finally, nude mice xenograft models were established to confirm the experimental results of cell experiments. RESULTS: ß-Arrestin2 significantly decreased the sensitivity of CRPC cells to docetaxel stimulation, through enhancing the phosphorylation and expression of PKM2. Additionally, ß-arrestin2 increased PKM2 phosphorylation via the ERK1/2 signaling pathway and induced PKM2 expression in a post-transcriptional manner through an hnRNP A1-dependent PKM alternative splicing mechanism, rather than by inhibiting its ubiquitination degradation. CONCLUSION: Our findings indicate that the ß-arrestin2/hnRNP A1/PKM2 pathway could be a promising target for treating docetaxel-resistant CRPC.

New hope for tumor immunotherapy: the macrophage-related "do not eat me" signaling pathway.

The "do not eat me" signaling pathway is extremely active in tumor cells, providing a means for these cells to elude macrophage phagocytosis and escape immune surveillance. Representative markers of this pathway, such as CD47 and CD24, are highly expressed in numerous tumors. The interaction of SIRPα with CD47 reduces the accumulation of non-myosin ⅡA on the cell membrane. The combination of CD24 and Siglec10 ultimately leads to the recruitment of SHP-1 or SHP-2 to reduce signal transduction. Both of them weaken the ability of macrophages to engulf tumor cells. Blocking the mutual recognition between CD47-SIRPα or CD24-Siglec10 using large molecular proteins or small molecular drugs represents a promising avenue for tumor immunotherapy. Doing so can inhibit signal transduction and enhance macrophage clearance rates of cancer cells. In this paper, we summarize the characteristics of the drugs that affect the "do not eat me" signaling pathway via classical large molecular proteins and small molecule drugs, which target the CD47-SIRPα and CD24-Siglec10 signaling pathways, which target the CD47-SIRPα and CD24-Siglec10 signaling pathways. We expect it will offer insight into the development of new drugs centered on blocking the "do not eat me" signaling pathway.

IL-10 Promotes CXCL13 Expression in Macrophages Following Foot-and-Mouth Disease Virus Infection.

Foot-and-mouth disease (FMD) is one of the most contagious livestock diseases in the world, posing a constant global threat to the animal trade and national economies. The chemokine C-X-C motif chemokine ligand 13 (CXCL13), a biomarker for predicting disease progression in some diseases, was recently found to be increased in sera from mice infected with FMD virus (FMDV) and to be associated with the progression and severity of the disease. However, it has not yet been determined which cells are involved in producing CXCL13 and the signaling pathways controlling CXCL13 expression in these cells. In this study, the expression of CXCL13 was found in macrophages and T cells from mice infected with FMDV, and CXCL13 was produced in bone-marrow-derived macrophages (BMDMs) by activating the nuclear factor-kappaB (NF-κB) and JAK/STAT pathways following FMDV infection. Interestingly, CXCL13 concentration was decreased in sera from interleukin-10 knock out (IL-10-/-) mice or mice blocked IL-10/IL-10R signaling in vivo after FMDV infection. Furthermore, CXCL13 was also decreased in IL-10-/- BMDMs and BMDMs treated with anti-IL-10R antibody following FMDV infection in vitro. Lastly, it was demonstrated that IL-10 regulated CXCL13 expression via JAK/STAT rather than the NF-κB pathway. In conclusion, the study demonstrated for the first time that macrophages and T cells were the cellular sources of CXCL13 in mice infected with FMDV; CXCL13 was produced in BMDMs via NF-κB and JAK/STAT pathways; and IL-10 promoted CXCL13 expression in BMDMs via the JAK/STAT pathway.

HELLPAR/RRM2 axis related to HMMR as novel prognostic biomarker in gliomas.

BACKGROUND: Gliomas are the most frequent type of central nervous system tumor, accounting for more than 70% of all malignant CNS tumors. Recent research suggests that the hyaluronan-mediated motility receptor (HMMR) could be a novel potential tumor prognostic marker. Furthermore, mounting data has highlighted the important role of ceRNA regulatory networks in a variety of human malignancies. The complexity and behavioural characteristics of HMMR and the ceRNA network in gliomas, on the other hand, remained unknown. METHODS: Transcriptomic expression data were collected from TCGA, GTEx, GEO, and CGGA database.The relationship between clinical variables and HMMR was analyzed with the univariate and multivariate Cox regression. Kaplan-Meier method was used to assess OS. TCGA data are analyzed and processed, and the correlation results obtained were used to perform GO, GSEA, and ssGSEA. Potentially interacting miRNAs and lncRNAs were predicted by miRWalk and StarBase. RESULTS: HMMR was substantially expressed in gliomas tissues compared to normal tissues. Multivariate analysis revealed that high HMMR expression was an independent predictive predictor of OS in TCGA and CGGA. Functional enrichment analysis found that HMMR expression was associated with nuclear division and cell cycle. Base on ssGSEA analysis, The levels of HMMR expression in various types of immune cells differed significantly. Bioinformatics investigation revealed the HEELPAR-hsa-let-7i-5p-RRM2 ceRNA network, which was linked to gliomas prognosis. And through multiple analysis, the good predictive performance of HELLPAR/RRM2 axis for gliomas patients was confirmed. CONCLUSION: This study provides multi-layered and multifaceted evidence for the importance of HMMR and establishes a HMMR-related ceRNA (HEELPAR-hsa-let-7i-5p-RRM2) overexpressed network related to the prognosis of gliomas.

FTL004, an anti-CD38 mAb with negligible RBC binding and enhanced pro-apoptotic activity, is a novel candidate for treatments of multiple myeloma and non-Hodgkin lymphoma.

Anti-CD38 monoclonal antibodies (mAbs), daratumumab, and isatuximab have represented a breakthrough in the treatment of multiple myeloma (MM). Recently, CD38-based mAbs were expected to achieve increasing potential beyond MM, which encouraged us to develop new anti-CD38 mAbs to meet clinical needs. In this study, we developed a novel humanized anti-CD38 antibody, FTL004, which exhibited enhanced pro-apoptotic ability and negligible binding to red blood cells (RBCs). FTL004 presented a better ability to induce direct apoptosis independent of Fc-mediated cross-linking against lymphoma and MM cell lines as well as primary myeloma cells derived from MM patients. For instance, FTL004 induced RPMI 8226 cells with 55% early apoptosis cells compared with 20% in the isatuximab-treated group. Of interest, FTL004 showed ignorable binding to CD38 on human RBCs in contrast to tumor cells, even at concentrations up to 30 µg/mL. Furthermore, with an engineered Fc domain, FTL004 displayed stronger antibody-dependent cellular cytotoxicity (ADCC) against CD38+ malignant cells. In vivo MM and non-Hodgkin lymphoma tumor xenograft models showed that FTL004 possessed an effective anti-tumor effect. Cryo-electron microscopy structure resolved two epitope centers of FTL004 on CD38: one of which was unique while the other partly overlapped with that of isatuximab. Taken together, FTL004 distinguishes it from other CD38 targeting mAbs and represents a potential candidate for the treatment of MM and non-Hodgkin lymphoma.

Peste des Petits Ruminants Virus Upregulates STING to Activate ATF6-Mediated Autophagy.

Peste des petits ruminants virus (PPRV) infection leads to autophagy, and the molecular mechanisms behind this phenomenon are unclear. Here, we demonstrate that PPRV infection results in morphological changes of the endoplasmic reticulum (ER) and activation of activating transcription factor 6 (ATF6) of the ER stress unfolded protein response (UPR). Knockdown of ATF6 blocked the autophagy process, suggesting ATF6 is necessary for PPRV-mediated autophagy induction. Further study showed that PPRV infection upregulates expression of the ER-anchored adaptor protein stimulator of interferon genes (STING), which is well-known for its pivotal roles in restricting DNA viruses. Knockdown of STING suppressed ATF6 activation and autophagy induction, implying that STING functions upstream of ATF6 to induce autophagy. Moreover, the STING-mediated autophagy response originated from the cellular pattern recognition receptor melanoma differentiation-associated gene 5 (MDA5). The absence of MDA5 abolished the upregulation of STING and the activation of autophagy. The deficiency of autophagy-related genes (ATG) repressed the autophagy process and PPRV replication, while it had no effect on MDA5 or STING expression. Overall, our work revealed that MDA5 works upstream of STING to activate ATF6 to induce autophagy. IMPORTANCEPPRV infection induces cellular autophagy; however, the intracellular responses and signaling mechanisms that occur upon PPRV infection are obscure, and whether innate immune responses are linked with autophagy to regulate viral replication is largely unknown. Here, we uncovered that the innate immune sensor MDA5 initiated the signaling cascade by upregulating STING, which is best known for its role in anti-DNA virus infection by inducing interferon expression. We first provide evidence that STING regulates PPRV replication by activating the ATF6 pathway of unfolded protein responses (UPRs) to induce autophagy. Our results revealed that in addition to mediating responses to foreign DNA, STING can cross talk with MDA5 to regulate the cellular stress response and autophagy induced by RNA viruses; thus, STING works as an adaptor protein for cellular stress responses and innate immune responses. Modulation of STING represents a promising approach to control both DNA and RNA viruses.

p53 Inhibits Bmi-1-driven Self-Renewal and Defines Salivary Gland Cancer Stemness.

PURPOSE: Mucoepidermoid carcinoma (MEC) is a poorly understood salivary gland malignancy with limited therapeutic options. Cancer stem cells (CSC) are considered drivers of cancer progression by mediating tumor recurrence and metastasis. We have shown that clinically relevant small molecule inhibitors of MDM2-p53 interaction activate p53 signaling and reduce the fraction of CSC in MEC. Here we examined the functional role of p53 in the plasticity and self-renewal of MEC CSC. EXPERIMENTAL DESIGN: Using gene silencing and therapeutic activation of p53, we analyzed the cell-cycle profiles and apoptosis levels of CSCs in MEC cell lines (UM-HMC-1, -3A, -3B) via flow cytometry and looked at the effects on survival/self-renewal of the CSCs through sphere assays. We evaluated the effect of p53 on tumor development (N = 51) and disease recurrence (N = 17) using in vivo subcutaneous and orthotopic murine models of MEC. Recurrence was followed for 250 days after tumor resection. RESULTS: Although p53 activation does not induce MEC CSC apoptosis, it reduces stemness properties such as self-renewal by regulating Bmi-1 expression and driving CSC towards differentiation. In contrast, downregulation of p53 causes expansion of the CSC population while promoting tumor growth. Remarkably, therapeutic activation of p53 prevented CSC-mediated tumor recurrence in preclinical trials. CONCLUSIONS: Collectively, these results demonstrate that p53 defines the stemness of MEC and suggest that therapeutic activation of p53 might have clinical utility in patients with salivary gland MEC.

Increased ASF1B Expression Correlates With Poor Prognosis in Patients With Gliomas.

Background: Several studies have suggested that anti-silencing function 1 B (ASF1B) can serve as a good potential marker for predicting tumor prognosis. But the values of ASF1B in gliomas have not been elucidated and further confirmation is needed. Methods: Transcriptomic and clinical data were downloaded from The Cancer Genome Atlas database (TCGA), genotypic tissue expression (GTEx), and the Chinese Gliomas Genome Atlas database (CGGA). Univariate and multivariate Cox regression analyses were used to investigate the link between clinical variables and ASF1B. Survival analysis was used to assess the association between ASF1B expression and overall survival (OS). The relationship between ASF1B expression and OS was studied using survival analysis. To investigate the probable function and immunological infiltration, researchers used gene ontology (GO) analysis, gene set enrichment analysis (GSEA), and single-sample GSEA (ssGSEA). Results: In glioma tissues, ASF1B expression was considerably higher than in normal tissues. The survival analysis found that increased ASF1B expression was linked with a poor prognosis in glioma patients. ASF1B demonstrated a high diagnostic value in glioma patients, according to a Receiver Operating Characteristic (ROC) analysis. ASF1B was found to be an independent predictive factor for OS in a Cox regression study (HR = 1.573, 95% CI: 1.053-2.350, p = 0.027). GO, KEGG, and GSEA functional enrichment analysis revealed that ASF1B was associated with nuclear division, cell cycle, m-phase, and cell cycle checkpoints. Immuno-infiltration analysis revealed that ASF1B was positively related to Th2 cells, macrophages, and aDC and was negatively related to pDC, TFH, and NK CD56 bright cells. Conclusion: A high level of ASF1B mRNA expression was correlated with a poor prognosis in glioma patients in this study, implying that it could be a reliable prognostic biomarker for glioma patients.

Back on the scene: Advances and challenges in CD3-related drugs in tumor therapy.

CD3 molecules are mainly distributed on the membrane of mature T cells. They are involved in T cell antigen recognition, signal transduction, and regulation of T cell development. CD3-related monoclonal antibodies (mAbs) are mainly used in the treatment of autoimmune diseases. Nearly half of all bispecific antibodies developed are used in tumor therapy, one of which is CD3 antigen. In this review, we discuss the importance of biological function and the crucial role of CD3 in tumor therapy. We highlight the research status of antibodies and small molecules targeting CD3 to provide guidance for future drug research.

Development of Dual Inhibitors Targeting Epidermal Growth Factor Receptor in Cancer Therapy.

Epidermal growth factor receptor (EGFR) is of great significance in mediating cell signaling transduction and tumor behaviors. Currently, third-generation inhibitors of EGFR, especially osimertinib, are at the clinical frontier for the treatment of EGFR-mutant non-small-cell lung cancer (NSCLC). Regrettably, the rapidly developing drug resistance caused by EGFR mutations and the compensatory mechanism have largely limited their clinical efficacy. Given the synergistic effect between EGFR and other compensatory targets during tumorigenesis and tumor development, EGFR dual-target inhibitors are promising for their reduced risk of drug resistance, higher efficacy, lower dosage, and fewer adverse events than those of single-target inhibitors. Hence, we present the synergistic mechanism underlying the role of EGFR dual-target inhibitors against drug resistance, their structure-activity relationships, and their therapeutic potential. Most importantly, we emphasize the optimal target combinations and design strategies for EGFR dual-target inhibitors and provide some perspectives on new challenges and future directions in this field.

MiR-181a-5p Alleviates the Inflammatory Response of PC12 Cells by Inhibiting High-Mobility Group Box-1 Protein Expression.

OBJECTIVE: Neuroinflammation triggers sequelae after spinal cord injury (SCI). Inhibition of inflammation promotes recovery after SCI. MicroRNAs regulate many pathophysiological processes, including inflammation. Any role for miR-181a-5p in the inflammatory response after SCI remains unclear. Thus, we evaluated the effects of miR-181a-5p on inflammation in PC12 cells and the underlying mechanism in play. METHODS: Quantitative reverse transcription-polymerase chain reaction was used to measure the levels of miR-181a-5p and high-mobility group box-1 protein (HMGB1) in SCI tissues. Cell-counting kit-8 assays were used to assess the viability of PC12 cells treated with lipopolysaccharide (LPS). Plasmids encoding MiR-181a-5p mimics, an miR-181a-5p inhibitor, or/and the HMGB1 were transfected into PC12 cells. Quantitative reverse transcription-polymerase chain reaction or/and Western blotting were performed to assess the expression of miR-181a-5p, HMGB1, and inflammatory factors in vitro. RESULTS: MiR-181a-5p expression decreased and HMGB1 expression increased in SCI tissues and LPS-induced PC12 cells. Upregulation of miR-181a-5p (via transfection) inhibited inflammation of, and HMGB1 expression by, LPS-induced PC12 cells. HMGB1 overexpression reversed the anti-inflammatory effects of miR-181a-5p. Dual-luciferase assays confirmed that HMGB1 was a direct target of miR-181a-5p. CONCLUSIONS: miR-181a-5p attenuated the inflammatory response of LPS-induced PC12 cells by directly inhibiting HMGB1; thus, miR-181a-5p may serve as a therapeutic target in SCI.

Recent progress in DNA methyltransferase inhibitors as anticancer agents.

DNA methylation mediated by DNA methyltransferase is an important epigenetic process that regulates gene expression in mammals, which plays a key role in silencing certain genes, such as tumor suppressor genes, in cancer, and it has become a promising therapeutic target for cancer treatment. Similar to other epigenetic targets, DNA methyltransferase can also be modulated by chemical agents. Four agents have already been approved to treat hematological cancers. In order to promote the development of a DNA methyltransferase inhibitor as an anti-tumor agent, in the current review, we discuss the relationship between DNA methylation and tumor, the anti-tumor mechanism, the research progress and pharmacological properties of DNA methyltransferase inhibitors, and the future research trend of DNA methyltransferase inhibitors.

Research Advances on the Interactions between Rabies Virus Structural Proteins and Host Target Cells: Accrued Knowledge from the Application of Reverse Genetics Systems.

Rabies is a lethal zoonotic disease caused by lyssaviruses, such as rabies virus (RABV), that results in nearly 100% mortality once clinical symptoms appear. There are no curable drugs available yet. RABV contains five structural proteins that play an important role in viral replication, transcription, infection, and immune escape mechanisms. In the past decade, progress has been made in research on the pathogenicity of RABV, which plays an important role in the creation of new recombinant RABV vaccines by reverse genetic manipulation. Here, we review the latest advances on the interaction between RABV proteins in the infected host and the applied development of rabies vaccines by using a fully operational RABV reverse genetics system. This article provides a background for more in-depth research on the pathogenic mechanism of RABV and the development of therapeutic drugs and new biologics.

Combination of oncolytic adenovirus ZD55 harboring TRAIL-IETD-MnSOD and cytokine-induced killer cells against lung cancer.

BACKGROUND: Our study aimed to investigate the effect of cancer-targeting gene-virotherapy and cytokine-induced killer (CIK) cell immunotherapy on lung cancer. METHODS: CIK cells were obtained from peripheral blood mononuclear cells using interferon (IFN)-γ, interleukin (IL)-2, and CD3 monoclonal antibody. The CIK cells were infected with oncolytic adenovirus ZD55 harboring tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL), manganese-containing superoxide dismutase (MnSOD), and TRAIL-isoleucine-aspartate-threonine-glutamate (IETD)-MnSOD. The cells were then cocultured with lung cancer cell lines A549 and NCI-H1650, normal cell line BEAS-2B, or injected into an A549 xenograft mouse model. RESULTS: Proliferation, colony formation, and invasion of A549 and NCI-H1650 cells were significantly inhibited by co-cultivation with CIK cells carrying oncolytic adenoviruses (in order) ZD55-TRAIL-IETD-MnSOD > ZD55-TRAIL + ZD55-MnSOD > ZD55-MnSOD > ZD55-TRAIL. Compared to BEAS-2B cells, the production of IFN-γ, TNF-α, and lactate dehydrogenase (LDH) in tumor cells was increased. Tumor volume in the xenograft model and Ki-67 expression in tumor samples were reduced after injection of CIK cells carrying oncolytic adenoviruses, in the same order as the in vivo experiments. Levels of IFN-γ, TNF-α, and LDH contents were also increased in the same order. CONCLUSIONS: Our studies confirmed the high efficacy of combined oncolytic adenovirus ZD55 harboring TRAIL-IETD-MnSOD and CIK cells against lung cancer.

C3 ester side chain plays a pivotal role in the antitumor activity of Maytansinoids.

Maytansinoids, the chemical derivatives of Maytansine, are commonly used as potent cytotoxic payloads in antibody-drug conjugates (ADC). Structure-activity-relationship studies had identified the C3 ester side chain as a critical element for antitumor activity of maytansinoids. The maytansinoids bearing the methyl group at C3 position with D configuration were about 100 to 400-fold less cytotoxic than their corresponding L-epimers toward various cell lines. The detailed mechanism of how chirality affects the anticancer activity remains elusive. In this study, we determined the high-resolution crystal structure of tubulin in complex with maytansinol, L-DM1-SMe and D-DM1-SMe. And we found the carbonyl oxygen atom of the ester moiety and the tail thiomethyl group at C3 side chain of L-DM1-SMe form strong intramolecular interaction with the hydroxyl at position 9 and the benzene ring, respectively, fixing the bioactive conformation and enhancing the binding affinity. Additionally, ligand-based and structure-based virtually screening methods were used to screen the commercially macrocyclic compounds library, and 15 macrocyclic structures were picketed out as putatively new maytansine-site inhibitors. Our study provides a possible strategy for the rational discovery of next-generation maytansine site inhibitors.

Detection of tyrosinase in living cells using an Enteromorpha Prolifera based fluorescent probe.

Melanoma, the skin cancer with the highest mortality rate, can be diagnosed at the early stage by detecting unique biomarkers. Over-expressed tyrosinase has been confirmed by dozens of clinical studies as an independent factor to evaluate the malignancy of melanoma. Using Enteromorpha Prolifera as the raw material, herein we develop a novel fluorescent probe, ECDY, which can sensitively detect the tyrosinase activity in different types of cells. More importantly, melanoma cells can be specifically distinguished through cell lysate measurements as well as the whole-cell imaging technique. Mechanically, the tyrosine groups on the surface of ECDY can be specifically recognized by tyrosinase and further converted into dopaquinone, which consequently causes the intramolecular fluorescence quenching of the probe through photoinduced electron transfer (PET). Tyrosinase can be detected within 20 min in the solution, and the detection limit is as low as 0.067 U mL-1. For the in vitro demonstration, we evaluate the fluorescence decay of ECDY in response to the intracellular tyrosinase activity within the lysate of various cell lines, including non-cancerous, non-melanoma cancerous, and mouse melanoma ones. The experimental results verify that ECDY can accurately measure the apparent tyrosinase activity in different cell lines and detect melanoma cell lysate specifically. The confocal fluorescence imaging experiments further demonstrate that ECDY can distinguish melanoma cells from others significantly. We believe that ECDY provides a new strategy for the efficient detection of tyrosinase and melanoma cells, and is expected to apply as a clinical diagnosis platform.

The X-ray structure of tubulysin analogue TGL in complex with tubulin and three possible routes for the development of next-generation tubulysin analogues.

Microtubule-targeting agents (MTAs) are the most commonly used anti-cancer drugs. At least fourteen microtubule inhibitors and ten antibody drug conjugates (ADCs) linking MTAs are approved by FDA for clinical use in cancer therapy. In current research, we determined the crystal structure of tubulysin analogue TGL in complex with tubulin at a high resolution (2.65 Å). In addition, we summarized all of the previously published high-resolution crystal structures of ligands in the vinca site to provide structural insights for the rational design of the new vinca-site ligands. Moreover, based on the aligned results of the vinca site ligands, we provided three possible routes for designing new tubulysin analogues, namely macrocyclization between the N-14 side chain and the N-9 side chain, the hybird of tubulysin M and phomopsin A, and growing new aryl group at C-21. These designed structures will inspire the development of new MTAs or payloads in cancer therapy.

Azithromycin alleviates systemic lupus erythematosus via the promotion of M2 polarisation in lupus mice.

Our previous study demonstrated that azithromycin could promote alternatively activated (M2) macrophages under lupus conditions in vitro, which might be beneficial for lupus treatment. Thus, the aim of this study was to further confirm whether azithromycin can drive M2 polarisation in lupus and ultimately alleviate systemic lupus erythematosus (SLE) in vivo. Lymphocyte-derived DNA (ALD-DNA)-induced mice (induced lupus model) and MRL-Faslpr mice (spontaneous lupus model) were both used in the experiment. First, we observed symptoms of lupus by assessing the levels of serum anti-dsDNA antibodies and serum creatinine and renal pathology. We found that both murine models showed increased levels of serum anti-dsDNA antibodies and creatinine, enhanced glomerular fibrosis and cell infiltration, basement membrane thickening and elevated IgG deposition. After azithromycin treatment, all these medical indexes were alleviated, and kidney damage was effectively reversed. Next, macrophage polarisation was assessed in the spleen and kidneys. Macrophage infiltration in the spleen was notably decreased after azithromycin treatment in both murine models, with a remarkably elevated proportion of M2 macrophages. In addition, the expression of interleukin (IL)-1, IL-6, tumour necrosis factor (TNF)-α, inducible nitric oxide synthase (iNOS), CD86, toll-like receptor (TLR)2 and TLR4 was extremely downregulated, while the expression of transforming growth factor (TGF)-ß, arginase-1 (Arg-1), chitinase-like 3 (Ym-1), found in inflammatory zone (Fizz-1) and mannose receptor (CD206) was significantly upregulated in the kidneys after azithromycin treatment. Taken together, our results indicated for the first time that azithromycin could alleviate lupus by promoting M2 polarisation in vivo. These findings exploited the newly discovered potential of azithromycin, a conventional drug with verified safety, affordability and global availability, which could be a novel treat-to-target strategy for SLE via macrophage modulation.

A Rationale for Drug Design Provided by Co-Crystal Structure of IC261 in Complex with Tubulin.

Microtubules composed of α/ß tubulin heterodimers are an essential part of the cytoskeleton of eukaryotic cells and are widely regarded as targets for cancer chemotherapy. IC261, which is discovered as an ATP-competitive inhibitor of serine/threonine-specific casein kinase 1 (CK1), has shown its inhibitory activity on microtubule polymerization in recent studies. However, the structural information of the interaction between tubulin and IC261 is still unclear. Here, we provided a high-resolution (2.85 Å) crystal structure of tubulin and IC261 complex, revealed the intermolecular interaction between tubulin and IC261, and analyzed the structure-activity relationship (SAR). Subsequently, the structure of tubulin-IC261 complex was compared with tubulin-colchicine complex to further elucidate the novelty of IC261. Furthermore, eight optimal candidate compounds of new IC261-based microtubule inhibitors were obtained through molecular docking studies. In conclusion, the co-crystal structure of tubulin-IC261 complex paves a way for the design and development of microtubule inhibitor drugs.