Target For Cancer Drug Development Research Articles

Transcription factor EHF interacting with coactivator AJUBA aggravates malignancy and acts as a therapeutic target for gastroesophageal adenocarcinoma

Transcriptional dysregulation of genes is a hallmark of tumors and can serve as targets for cancer drug development. However, it is extremely challenging to develop small-molecule inhibitors to target abnormally expressed transcription factors (TFs) except for the nuclear receptor family of TFs. Little is known about the interaction between TFs and transcription cofactors in gastroesophageal adenocarcinoma (GEA) or the therapeutic effects of targeting TF and transcription cofactor complexes. In this study, we found that ETS homologous factor (EHF) expression is promoted by a core transcriptional regulatory circuitry (CRC), specifically ELF3-KLF5-GATA6, and interference with its expression suppressed the malignant biological behavior of GEA cells. Importantly, we identified Ajuba LIM protein (AJUBA) as a new coactivator of EHF that cooperatively orchestrates transcriptional network activity in GEA. Furthermore, we identified KRAS signaling as a common pathway downstream of EHF and AJUBA. Applicably, dual targeting of EHF and AJUBA by lipid nanoparticles cooperatively attenuated the malignant biological behaviors of GEA in vitro and in vivo. In conclusion, EHF is upregulated by the CRC and promotes GEA malignancy by interacting with AJUBA through the KRAS pathway. Targeting of both EHF and its coactivator AJUBA through lipid nanoparticles is a novel potential therapeutic strategy.

Identification of 1,3,4-oxadiazoles as tubulin-targeted anticancer agents: a combined field-based 3D-QSAR, pharmacophore model-based virtual screening, molecular docking, molecular dynamics simulation, and density functional theory calculation approach

Cancer is one of the most prominent causes of death worldwide and tubulin is a crucial protein of cytoskeleton that maintains essential cellular functions including cell division as well as cell signalling, that makes an attractive drug target for cancer drug development. 1,3,4-oxadiazoles disrupt microtubule causing G2-M phase cell cycle arrest and provide anti-proliferative effect. In this study, field-based 3D-QSAR models were developed using 62 bioactive anti-tubulin 1,3,4-oxadiazoles. The best model characterized by PLS factor 7 was rigorously validated using various statistical parameters. Generated 3D-QSAR model having high degree of confidence showed favourable and unfavourable contours around 1,3,4-oxadiazole core that assisted in defining proper spatial positioning of desired functional groups for better bioactivity. A five featured pharmacophore model (AAHHR_1) was developed using same ligand library and validated through enrichment analysis (BEDROC160.9 value = 0.59, Average EF 1% = 27.05, and AUC = 0.74). Total 30,212 derivatives of 1,3,4-oxadiazole obtained from PubChem database was prefiltered through validated pharmacophore model and docked in XP mode on binding cavity of tubulin protein (PDB code: 1SA0) which led into the identification of 11 HITs having docking scores between −7.530 and −9.719 kcal/mol while the reference compound Colchicine exerted docking score of −7.046 kcal/mol. Following the analysis of MM-GBSA and ADME studies, HIT1 and HIT4 emerged as the two promising hits. To verify their thermodynamic stability at the target site, molecular dynamic simulations were carried out. Both HITs were further subjected to DFT analysis to determine their HOMO-LUMO energy gap for ensuring their biological feasibility. Finally, molecular docking based structural exploration for 1,3,4-oxadiazoles to set up a lead of Formula I for further advancements of tubulin polymerization inhibitors as anti-cancer agents. Communicated by Ramaswamy H. Sarma

Design, synthesis, and biological evaluation of novel bivalent PI3K inhibitors for the potential treatment of cancer

Phosphatidylinositol 3-kinase (PI3K) signaling is among the most common alterations in cancer and has become a key target for cancer drug development. Based on a 4-methyl quinazoline scaffold, we designed and synthesized a novel series of bivalent PI3K inhibitors with different linker lengths and types. Bivalent PI3K inhibitor 27 demonstrates improved PI3K potency and antiproliferative cell activity, relative to the corresponding monovalent inhibitor 11. Compound 27 also significantly blocks the PI3K signal pathway, induces cell cycle arrest in G1 phase, and inhibits colony formation and cell migration. Furthermore, compound 27 shows dose-dependent anticancer efficacies in a HGC-27 xenograft mice model. Overall, this work provides a possible strategy to discover novel PI3K inhibitors for the treatment of cancers.

Synthesis of 4-styrylpyrazoles and Evaluation of their Inhibitory Effects on Cyclin-dependent Kinases.

Cycle-regulating and transcriptional cyclin-dependent kinases (CDKs) are attractive targets in cancer drug development. Several CDK inhibitors have already been obtained or are close to regulatory approval for clinical applications. Phenylazopyrazole CAN508 has been described as the first selective CDK9 inhibitor with an IC50 of 350 nM. Since the azo-moiety is not a suitable functionality for drugs due to pharmacological reasons, the preparation of carbo-analogues of CAN508 with similar biological activities is desirable. The present work is focused on the synthesis of carbo-analogues similar to CAN508 and their CDK inhibition activity. Herein, the synthesis of 21 novel carbo analogues of CAN508 and their intermediates is reported. Subsequently, target compounds 8a - 8u were evaluated for protein kinase inhibition (CDK2/cyclin E, CDK4/cyclin D, CDK9/cyclin T) and antiproliferative activities in cell lines (K562, MCF-7, MV4-11). Moreover, the binding mode of derivative 8s in the active site of CDK9 was modelled. Compounds 8a - 8u were obtained from key intermediate 7, which was prepared by linear synthesis involving Vilsmeier-Haack, Knoevenagel, Hunsdiecker, and Suzuki-Miyaura reactions. Styrylpyrazoles 8t and 8u were the most potent CDK9 inhibitors with IC50 values of approximately 1 μM. Molecular modelling suggested binding in the active site of CDK9. The flow cytometric analysis of MV4-11 cells treated with the most active styrylpyrazoles showed a significant G1-arrest. The prepared styrylpyrazoles showed inhibition activity towards CDKs and can provide a novel chemotype of kinase inhibitors.

A phase I clinical trial to evaluate the safety, tolerability, and pharmacokinetics of TST001 in patients with locally advanced or metastatic solid tumors.

TPS375 Background: In normal conditions, Claudin (CLDN)18.2 is a tight junction protein with expression strictly confined to differentiated epithelial cells in gastric mucosa. CLDN18.2 has been found to be upregulated, and involved in tumor development and progression in a variety of tumor types such as gastric, pancreatic, and bile duct cancer (BTC). The biological characteristics of CLDN18.2 suggest it is an ideal therapeutic target for cancer drug development. IMAB362 is the first anti-CLDN18.2 monoclonal antibody (mAb) of high potency to have been tested in humans and it revealed clinical efficacy in gastric cancer in a phase II study. TST001, a humanized IgG1 mAb, binds to a distinct epitope of CLDN18.2 with higher affinity and mediates CLDN18.2 expressing cancer cell death through antibody-dependent cellular cytotoxicity (ADCC) in comparison with IMAB362; Furthermore, TST001 is produced using an optimized glycoengineering process to increase affinity to FcR. The enhanced binding to CLDN18.2 on tumor cells and FcR on NK cells results in more efficient engagement of the tumor cells with NK cells and antibody mediated cellular cytotoxicity. In preclinical xenograft studies, TST001 displayed potent anti-tumor activities in the tumor models with medium to high level of CLDN18.2 expression and synergy anti-cancer effect with checkpoint inhibitor. A mAb specific for CLDN18.2 was also developed as an IHC based biomarker for patient enrollment in the clinical trials. Methods: This is an open-label, multi-center, phase I clinical trial to evaluate the safety, maximum tolerated dose (MTD), pharmacokinetics (PK) profile and preliminary anti-cancer effect of TST001 in patients with locally advanced or metastatic solid tumors. (NCT04396821) The study consists of two parts: Part A is a 3+3 dose escalation design with sequential dose cohorts of 1, 3, 6, 10mg/kg in Q2W and Q3W schedules. Based on the emerging safety data, higher doses may be proposed for testing. About 27-54 patients will be enrolled. Dose expansion (Part B) will utilize doses of TST001 based on the emerging data from Part A. In Part B, up to 20 patients with CLDN18.2 overexpression per tumor specific cohort will be enrolled to 3 cohorts: A: TST001 single agent in gastric/gastroesophageal junction (G/GEJ) cancer; B: TST001 + nivolumab in G/GEJ cancer; C: TST001 single agent in pancreatic cancer or BTC. All patients in Part B will be selected by CLDN18.2 expression by central lab testing. The safety, anti-tumor activity, and PK will be further assessed in Part B. Enrolment began in July 2020 in the USA and is ongoing in multiple sites. As of 20 September, 2021, 23 subjects were dosed in Part A and the dose of 10mg/kg is being tested. Another phase I study of TST001 single agent and in combination with chemotherapy in patients with metastatic solid tumor is also ongoing in China (NCT04495296).

Therapeutic targeting of "undruggable" MYC.

Summaryc-MYC controls global gene expression and regulates cell proliferation, cell differentiation, cell cycle, metabolism and apoptosis. According to some estimates, MYC is dysregulated in ≈70% of human cancers and strong evidence implicates aberrantly expressed MYC in both tumor initiation and maintenance. In vivo studies show that MYC inhibition elicits a prominent anti-proliferative effect and sustained tumor regression while any alteration on healthy tissue remains reversible. This opens an exploitable window for treatment that makes MYC one of the most appealing therapeutic targets for cancer drug development. This review describes the main functional and structural features of the protein structure of MYC and provides a general overview of the most relevant or recently identified interactors that modulate MYC oncogenic activity. This review also summarizes the different approaches aiming to abrogate MYC oncogenic function, with a particular focus on the prototype inhibitors designed for the direct and indirect targeting of MYC.

N-Propargylglycine: a unique suicide inhibitor of proline dehydrogenase with anticancer activity and brain-enhancing mitohormesis properties

Proline dehydrogenase (PRODH) is a mitochondrial inner membrane flavoprotein critical for cancer cell survival under stress conditions and newly recognized as a potential target for cancer drug development. Reversible (competitive) and irreversible (suicide) inhibitors of PRODH have been shown in vivo to inhibit cancer cell growth with excellent host tolerance. Surprisingly, the PRODH suicide inhibitor N-propargylglycine (N-PPG) also induces rapid decay of PRODH with concordant upregulation of mitochondrial chaperones (HSP-60, GRP-75) and the inner membrane protease YME1L1, signifying activation of the mitochondrial unfolded protein response (UPRmt) independent of anticancer activity. The present study was undertaken to address two aims: (i) use PRODH overexpressing human cancer cells (ZR-75-1) to confirm the UPRmt inducing properties of N-PPG relative to another equipotent irreversible PRODH inhibitor, thiazolidine-2-carboxylate (T2C); and (ii) employ biochemical and transcriptomic approaches to determine if orally administered N-PPG can penetrate the blood–brain barrier, essential for its future use as a brain cancer therapeutic, and also potentially protect normal brain tissue by inducing mitohormesis. Oral daily treatments of N-PPG produced a dose-dependent decline in brain mitochondrial PRODH protein without detectable impairment in mouse health; furthermore, mice repeatedly dosed with 50 mg/kg N-PPG showed increased brain expression of the mitohormesis associated protease, YME1L1. Whole brain transcriptome (RNAseq) analyses of these mice revealed significant gene set enrichment in N-PPG stimulated neural processes (FDR p < 0.05). Given this in vivo evidence of brain bioavailability and neural mitohormesis induction, N-PPG appears to be unique among anticancer agents and should be evaluated for repurposing as a pharmaceutical capable of mitigating the proteotoxic mechanisms driving neurodegenerative disorders.

Carcinogen-Induced Model of Proangiogenesis in Zebrafish Embryo-Larvae.

Tumor angiogenesis is the main target in cancer drug development. Discovery of antiangiogenic agents targeting different mechanisms of action is the major area of research to control tumor growth and metastasis. Zebrafish (in the embryo-larvae stage) acts as an essential preclinical efficacy-toxicity model for antiangiogenic drug discovery. We aimed to develop a carcinogen-induced model of proangiogenesis in zebrafish embryo-larvae using the carcinogens lindane and benzo[a]pyrene. Zebrafish were randomly selected for mating. Postspawning, healthy embryos were staged, dispensed in reverse-osmosis water in a 12-well plate, and incubated at 28.5 °C, wherein 24 h postfertilization they were exposed to sublethal concentrations of the carcinogens. Three days postexposure, embryos were stained with alkaline phosphatase, and the angiogenic basket was imaged using a bright-field microscope. The number of subintestinal vessels, their length from somite to the basket, and other proangiogenic parameters were measured and analyzed. The effective concentrations causing a 30% increase in subintestinal vessels for benzo[a]pyrene and lindane were 2.69 and 2.24 µM, respectively, thus proving their proangiogenic potency. The carcinogen-induced model of proangiogenesis in zebrafish embryo-larvae can be used as an effective high-throughput screening tool to assess the proangiogenic potential of carcinogenic compounds and to screen antiangiogenic drugs for better therapeutic intervention. Environ Toxicol Chem 2021;40:447-453.© 2020 SETAC.

Frontispiece: The Thioredoxin System: A Promising Target for Cancer Drug Development

Frontispiece: The Thioredoxin System: A Promising Target for Cancer Drug Development

The Thioredoxin System: A Promising Target for Cancer Drug Development

The thioredoxin system is highly conserved system found in all living cells and comprises NADPH, thioredoxin, and thioredoxin reductase. This system plays a critical role in preserving a reduced intracellular environment, and its involvement in regulating a wide range of cellular functions makes it especially vital to cellular homeostasis. Its critical role is not limited to healthy cells, it is also involved in cancer development, and is overexpressed in many cancers. This makes the thioredoxin system a promising target for cancer drug development. As such, over the last decade, many inhibitors have been developed that target the thioredoxin system, most of which are small molecules targeting the thioredoxin reductase C-terminal redox center. A few inhibitors of thioredoxin have also been developed. We believe that more efforts should be invested in developing protein/peptide-based inhibitors against both thioredoxin reductase and/or thioredoxin.

Transcription and Translation Inhibitors in Cancer Treatment.

Transcription and translation are fundamental cellular processes that govern the protein production of cells. These processes are generally up regulated in cancer cells, to maintain the enhanced metabolism and proliferative state of these cells. As such cancerous cells can be susceptible to transcription and translation inhibitors. There are numerous druggable proteins involved in transcription and translation which make lucrative targets for cancer drug development. In addition to proteins, recent years have shown that the “undruggable” transcription factors and RNA molecules can also be targeted to hamper the transcription or translation in cancer. In this review, we summarize the properties and function of the transcription and translation inhibitors that have been tested and developed, focusing on the advances of the last 5 years. To complement this, we also discuss some of the recent advances in targeting oncogenes tightly controlling transcription including transcription factors and KRAS. In addition to natural and synthetic compounds, we review DNA and RNA based approaches to develop cancer drugs. Finally, we conclude with the outlook to the future of the development of transcription and translation inhibitors.

Abstract 2503: Predicting drug sensitivity to a novel BCL-2 and BCL-xL dual inhibitor in solid tumor PDX trials

Abstract Background: BCL-2, an anti-apoptotic protein and oncogene, has been considered one of the top 10 targets for cancer drug development. The inhibition of BCL-2 and other anti-apoptotic proteins will help to restore the apoptotic pathway. Aiming to target much diverse BCL-2 family protein dependency in solid tumors, we have developed a potent BCL-2 and BCL-xL dual inhibitor APG-1252, which is currently in phase 1 trials for solid tumors in USA, China and Australia. Mechanistically, BCL-2 and BCL-xL protect cells from cell death by sequestering pro-death BH3-only proteins like BIM. Thus, BCL-2 and BCL-xL inhibitors are designed to disrupt the protein complexes and release pro-death proteins to trigger downstream apoptotic cascade. In this study, we intended to further understand the mechanism of action of APG-1252 in vivo and factors regulating its sensitivity. To this end, we conducted trials in SCID mice carrying patient-derived xenograft (PDX) tumors, and studied target engagement. Methods and Experiments: We first selected eleven gastric and esophageal cancer PDX models that bore high levels of BCL-xL (BCL-xLhigh) but normal levels of MCL-1 (MCL-1nor) based on the gene expression profiles and the consideration of that BCL-xL may play a major role in tumorigenesis in solid tumors and MCL-1 a resistant factor for the dual inhibitor. Mice bearing PDX tumors were intravenously treated with 100 mg/kg of APG-1252 twice per week for three weeks. At the end of treatments, tumor samples were collected. An advanced ELISA system (MSD, Meco Scale Discovery) and Western blotting were performed to detect BCL-2 and BCL-xL protein complex and individual proteins. Results: 1. The MSD assays revealed dominant BCL-xL:BIM complex signals in the solid tumor PDX samples in comparison with hematological malignancy cell lines, in which BCL-2:BIM was the major complex. 2. The dual inhibitor treatment disrupted BCL-xL:BIM complex, thus BCL-xL:BIM complex may serves as a pharmacodynamic (PD) marker for target engagement. 3. BCL-2 family protein profiles showed high BCL-xL and MCL-1 levels in the solid tumor PDXs. The treatment with APG-1252 led to a significant increase in BCL-2 protein levels, while there was no major change on pro-apoptotic BH3-only proteins BIM and PUMA. 4. The more efficacious responders to the treatment exhibited a more substantial decrease in BCL-xL:BIM complex signals and low MCL-1 protein levels, whereas a much less significant decrease in BCL-xL:BIM complex and higher MCL-1 levels were found in the poor or non-responders. Conclusion: Both BCL-xL protein and BCL-xL:BIM complex are dominant in the solid tumor PDX tissues. The decrease of BCL-xL:BIM complex indicates the on-target activity after the treatment with APG-1252. Thus, the complex may be used to monitor the target engagement and, potentially, a predictive biomarker for APG-1252 efficacy. These assays may be applied to patient samples to guide the clinical trials. Citation Format: Jing Deng, Xiaojing Huang, Guoqin Zhai, Tingting Mao, Yan Yin, Douglas D. Fang, Dajun Yang, Yifan Zhai. Predicting drug sensitivity to a novel BCL-2 and BCL-xL dual inhibitor in solid tumor PDX trials [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2503.

Abstract 2756: A high-resolution G-quadruplex structure involved in cmyc oncogene regulation

Abstract Gene expression needs strict regulation in human cells to prevent tumorigenesis. Among other control mechanisms, guanine-rich sequences which are clustered in promoter regions of oncogenes can form four stranded nucleic acid structures, called G-quadruplexes (G4), and regulate their transcription. The cmyc protein is the most important cellular regulator for proliferation and is often overexpressed in cancer cells. The DNA G-quadruplex formed in the cmyc promoter is a transcriptional silencer element and stabilization of this G4 by small molecules suppresses cmyc transcription, making it a promising target for cancer therapy. High-resolution structures of the highly diverse G4s provide the opportunity to understand their molecular interactions with transcription factors and to design small molecules that specifically bind a particular G4 conformer. Characteristically, multiple G4s can be formed in gene promoter regions because their G-rich sequence generally contains more than the minimum of four G-stretches. While all reported cmyc G4s share the same core with parallel oriented strands, their overall shape is individualized by distinct combinations of varying flanking and loop motifs. In this study, we present a new high-resolution NMR structure of a G4 found to form in the cmyc promoter region. The determined cmyc G4 has a 1:6:1 loop-size arrangement and its conformation is largely defined by its long central loop of 6 nt. We show that this G4 structure is the preferred target of nucleolin protein in comparison to other G4s formed in the cmyc promoter, demonstrating how the polymorphism of a G-rich sequence can modulate the interaction with transcription factors and other regulators. Our NMR structure shows that, despite the length of the 6-nt loop motif and the associated increased flexibility, the folding converges well due to a base pairing between the central loop and the 5’-flanking. Thus, the long loop is pulled towards the 5’-end while analogous loop-flanking interactions are prevented at the opposite 3’-site. In conclusion, the presented new addition to the ensemble of possible cmyc promoter G4 conformations extends the folding landscape for this promoter region, in particular due to the long central loop. This may shed light on how proteins such as nucleolin can target specific G4 and shift the equilibrium of folding to fine-tune gene expression. Finally, this new structure is another promising target for cancer drug development. Citation Format: Jonathan Dickerhoff, Luying Chen, Guanhui Wu, Buket Onel, Clement Lin, Danzhou Yang. A high-resolution G-quadruplex structure involved in cmyc oncogene regulation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2756.

Abstract 947: Defining UHRF1 domains that support maintenance of human colon cancer DNA methylation and tumorigenicity

Abstract Reversing the DNA methylation abnormalities by targeting the maintenance DNA methylation machinery represents a sought-after therapy paradigm in both liquid and solid tumors. UHRF1, a multi-domain protein with both chromatin reader and writer functions, is essential for targeting DNA methyltransferase 1 (DNMT1) to replicating DNA to maintain DNA methylation in both normal and cancer cells. Dysregulation of the DNMT1-UHRF1 axis is being increasingly linked to malignant transformation and progression across cancer types. Our recent study shows that UHRF1 depletion, alone or in combination with low doses of a DNMT inhibitor, effectively induces DNA demethylation and tumor suppressor genes (TSGs) reactivation. These findings collectively suggest UHRF1 represents a promising target for developing next-generation DNA demethylation agents for cancer therapy. Understanding the collective contribution of UHRF1’s domains for maintenance methylation is essential for the major translational goal of blocking maintenance of abnormal DNA methylation in established cancers and throughout cancer initiation and progression. While the UHRF1 domains responsible for de novo or re-methylation have been identified, the domain requirements specifically for maintaining normal and cancer-specific DNA methylation are poorly characterized. Herein, by developing a carefully timed genetic complementation assay, we systematically interrogate the roles for each major domain of UHRF1 for maintaining genome-wide DNA methylation in human colorectal cancer (CRC) cells. Distinct from the domain required for establishing methylation, we find that UHRF1 histone-binding and hemimethylated DNA reader domains, but not E3 ligase activity, are essential for maintaining cancer-specific DNA methylation in both HCT116 and RKO cells. Disrupting either one of the essential domains phenocopies UHRF1 depletion for global DNA demethylation and reactivation of epigenetically silenced TSGs. Supporting the essential roles of abnormal DNA methylation in sustaining the key oncogenic properties of CRC cells, we further show genetic perturbation of either histone- or hemimethylated DNA-binding activities of UHRF1 dramatically impairs CRC proliferation, invasion, and metastasis in vitro and in vivo. Moreover, for eight UHRF1 chromatin-reader domain regulated TSGs, we reveal a strong negative correlation between high UHRF1 expression, promoter hypermethylation, and their low expression with disease progression and overall survival in CRC patients in both TCGA and two independent CRC cohorts (n=363 and 390, respectively). Taken together, our data identify important differences from domains dominant for de novo DNA methylation, and provide important implications for the oncogenic functions of UHRF1 in CRC, and for credentialing UHRF1 as a desirable target for cancer drug development and means to personalize this. Citation Format: Xiangqian Kong, Jie Chen, Wenbing Xie, Stephen M. Brown, Yi Cai, Kaichun Wu, Daiming Fan, Yongzhan Nie, Yong Tao, Ray-Whay Chiu Yen, Hariharan Easwaran, Michael J. Topper, Scott B. Rothbart, Limin Xia, Stephen B. Baylin. Defining UHRF1 domains that support maintenance of human colon cancer DNA methylation and tumorigenicity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 947.

Repositioning Salirasib as a new antimalarial agent.

Malaria is a serious tropical disease that kills thousands of people every year, mainly in Africa, due to Plasmodium falciparum infections. Salirasib is a promising cancer drug candidate that interferes with the post-translational modification of Ras. This S-farnesyl thiosalicylate inhibits isoprenylcysteine carboxyl methyltransferase (ICMT), a validated target for cancer drug development. There is a high homology between the human and the parasite enzyme isoforms, in addition to being a druggable target. Looking to repurpose its structure as an antimalarial drug, a collection of S-substituted derivatives of thiosalicylic acid were prepared by introducing 1,2,3-triazole as a diversity entry point or by direct alkylation of the thiol. We further investigated the in vitro toxicity of FTS analogues to Plasmodium falciparum in the asexual stages and in Vero cells. An antiplasmodial activity assay was performed using a simple, high-sensitivity methodology based on nanoluciferase (NLuc)-transfected P. falciparum parasites. The results showed that some of the analogs were active at low micromolar concentration, including Salirasib. The most potent member of the series has S-farnesyl and the 1,2,3-triazole moiety substituted with phytyl. However, the compound substituted with methyl-naphthyl shows promising physicochemical and activity values. The low cytotoxicity in eukaryotic cells of the most active analogs provided good therapeutic indices, being starting-point candidates for future antimalarial drug development.

USP24 induces IL-6 in tumor-associated microenvironment by stabilizing p300 and \u03b2-TrCP and promotes cancer malignancy

We have previously demonstrated that USP24 is involved in cancer progression. Here, we found that USP24 expression is upregulated in M2 macrophages and lung cancer cells. Conditioned medium from USP24-knockdown M2 macrophages decreases the migratory and chemotactic activity of lung cancer cells and the angiogenic properties of human microvascular endothelial cell 1 (HMEC-1). IL-6 expression is significantly decreased in USP24-knockdown M2 macrophages and lung cancer cells, and IL-6-replenished conditioned medium restores the migratory, chemotactic and angiogenetic properties of the cells. USP24 stabilizes p300 and β-TrCP to increase the levels of histone-3 acetylation and NF-κB, and decreases the levels of DNMT1 and IκB, thereby increasing IL-6 transcription in M2 macrophages and lung cancer cells, results in cancer malignancy finally. IL-6 has previously been a target for cancer drug development. Here, we provide direct evidence to support that USP24 promotes IL-6 expression, which might be beneficial for cancer therapy.

\u03b3-Tocotrienol induces apoptosis in pancreatic cancer cells by upregulation of ceramide synthesis and modulation of sphingolipid transport

BackgroundCeramide synthesis and metabolism is a promising target in cancer drug development. γ-tocotrienol (GT3), a member of the vitamin E family, orchestrates multiple effects that ensure the induction of apoptosis in both, wild-type and RAS-mutated pancreatic cancer cells. Here, we investigated whether these effects involve changes in ceramide synthesis and transport.MethodsThe effects of GT3 on the synthesis of ceramide via the de novo pathway, and the hydrolysis of sphingomyelin were analyzed by the expression levels of the enzymes serine palmitoyl transferase, ceramide synthase-6, and dihydroceramide desaturase, and acid sphingomyelinase in wild-type RAS BxPC3, and RAS-mutated MIA PaCa-2 and Panc 1 pancreatic cancer cells. Quantitative changes in ceramides, dihydroceramides, and sphingomyelin at the cell membrane were detected by LCMS. Modulation of ceramide transport by GT3 was studied by immunochemistry of CERT and ARV-1, and the subsequent effects at the cell membrane was analyzed via immunofluorescence of ceramide, caveolin, and DR5.ResultsGT3 favors the upregulation of ceramide by stimulating synthesis at the ER and the plasma membrane. Additionally, the conversion of newly synthesized ceramide to sphingomyelin and glucosylceramide at the Golgi is prevented by the inhibition of CERT. Modulation ARV1 and previously observed inhibition of the HMG-CoA pathway, contribute to changes in membrane structure and signaling functions, allows the clustering of DR5, effectively initiating apoptosis.ConclusionsOur results suggest that GT3 targets ceramide synthesis and transport, and that the upregulation of ceramide and modulation of transporters CERT and ARV1 are important contributors to the apoptotic properties demonstrated by GT3 in pancreatic cancer cells.

Hypoxia-inducible factor-1 (HIF-1) inhibitors from the last decade (2007 to 2016): A "structure-activity relationship" perspective.

Tumor hypoxia is a common feature in most solid tumors and is associated with overexpression of the hypoxia response pathway. Overexpression of the hypoxia-inducible factor (HIF-1) protein leads to angiogenesis, metastasis, apoptosis resistance, and many other pro-tumorigenic responses in cancer development. HIF-1 is a promising target in cancer drug development to increase the patient's response to chemotherapy and radiotherapy as well as the survival rate of cancer patients. Since up to 1% of genes are hypoxia-sensitive, a target-specific HIF-1 inhibitor may be a better clinical candidate in cancer drug discovery. Though no HIF-1 inhibitor is clinically available to date, a lot of effort has been applied during the last decade in search of potent HIF-1 inhibitors. In this review, we will summarize the structure-activity relationship of ten different chemotypes reported to be HIF-1 inhibitors in the last decade (2007-2016), their mechanisms of action for HIF-1 inhibition, progress in the way of target-specific inhibitors, and problems associated with current inhibitors. It is anticipated that the results of these research on the medicinal chemistry of HIF-1 inhibitors will provide decent information in the design and development of future inhibitors.

Modulation of glycogen synthase kinase-3β following TRAIL combinatorial treatment in cancer cells.

Glycogen Synthase Kinase-3β (GSK3β) is a serine/threonine kinase, known to regulate various cellular processes including proliferation, differentiation, survival, apoptosis as well as TRAIL-resistance. Thus pathways that can modulate GSK3β axis are important targets for cancer drug development. Our earlier studies have shown that combinatorial treatment with Troglitazone (TZD) and TRAIL can induce apoptosis in TRAIL-resistant cancer cells. The current studies were undertaken to investigate whether GSK3β pathway was modulated during this apoptosis. Our results indicated an increase in inhibitory GSK3βSer9 phosphorylation during apoptosis, mediated via AKT. At a later time, however, TZD alone and TRAIL-TZD combination produced a dramatic reduction of GSK3β expression, which was abolished by cycloheximide. Luciferase assays with GSK3β-luc promoter reporter showed that TZD can effectively antagonize GSK3β promoter activity. Since TZD is a ligand for transcription factor PPARγ and can activate AMPK, we determined their roles on antagonism of GSK3β. Knockdown of PPARγ was unable to restore GSK3β expression or antagonize GSK3βSer9 phosphorylation. Although pretreatment with Compound C (pharmacological inhibitor of AMPK) partially rescued GSK3β expression, knockdown of AMPKα1 or α2 alone or in combination were ineffective. These studies suggested a novel PPARγ-AMPK-independent mechanism of targeting GSK3β by TZD, elucidation of which might provide newer insights to improve our understanding of TRAIL-resistance.

A specific, transmembrane interface regulates fibroblast activation protein (FAP) homodimerization, trafficking and exopeptidase activity

Fibroblast activation protein (FAP) is a cell-surface serine protease which promotes invasiveness of certain epithelial cancers and is therefore a potential target for cancer drug development and delivery. Unlike dipeptidyl peptidase IV (DPPIV), FAP exhibits prolyl endopeptidase activity and is active as a homodimer with specificity for type I collagen. The mechanism that regulates FAP homodimerization and its relation to prolyl endopeptidase activity is not completely understood. Here, we investigate key residues in the FAP TM domain that may be significant for FAP homodimerization. Mutations to predicted TM interfacial residues (G10L, S14L, and A18L) comprising a small-X3-small motif reduced FAP TM-CYTO dimerization relative to wild type as measured using the AraTM assay, whereas predicted off-interface residues showed no significant change from wild type. The results implied that the predicted small-X3-small dimer interface affect stabilization of FAP TM-CYTO homodimerization. Compared with FAPwild-type, the interfacial TM residue G10L significantly decreased FAP endopeptidase activity more than 25%, and also reduced cell-surface versus intracellular expression relative to other interfacial residues S14L and A18L. Thus, our results suggest FAP dimerization is important for both trafficking and protease activity, and is dependent on a specific TM interface.