A Novel Multi-Functional Fluorescence Probe for the Detection of Al3+/Zn2+/Cd2+ and its Practical Applications.

A novel multi-functional fluorescence probe HMIC based on hydrazide Schiff base has been successfully synthesized and characterized. It can distinguish Al3+/Zn2+/Cd2+ in ethanol, in which fluorescence emission with different colors (blue for Al3+, orange for Zn2+, and green for Cd2+) were presented. The limits of detection of HMIC towards three ions were calculated from the titration curve as 7.70 × 10- 9 M, 4.64 × 10- 9 M, and 1.35 × 10- 8 M, respectively. The structures of HMIC and its complexes were investigated using UV-Vis spectra, Job's plot, infrared spectra, mass spectrometry, 1H-NMR and DFT calculations. Practical application studies have also demonstrated that HMIC can be applied to real samples with a low impact of potential interferents. Cytotoxicity and cellular imaging assays have shown that HMIC has good cellular permeability and potential antitumor effects. Interestingly, HMIC can image Al3+, Zn2+ and Cd2+ in the cells with different fluorescence signals.

Combination of DNA Damage, Autophagy, and ERK Inhibition: Novel Evodiamine-Inspired Multi-Action Pt(IV) Prodrugs with High-Efficiency and Low-Toxicity Antitumor Activity.

Exploring multi-targeting chemotherapeutants with advantages over single-targeting agents and drug combinations is of great significance in drug discovery. Herein, we employed phytogenic evodiamine (EVO) and conventional Pt(II) drugs to design and synthesize multi-target EVO-Pt(IV) anticancer prodrugs (4-14). Among them, compound 10 exhibited a 118-fold enhancement in the IC50 value compared to cisplatin and low toxicity to normal cells. Further studies proved that 10 significantly enhanced intracellular Pt accumulation and DNA damage, perturbed mitochondrial membrane potential, inhibited cell migration and invasion, upregulated reactive oxygen species levels, and induced apoptosis and autophagic cell death. Molecular docking assay revealed that 10 fits perfectly into the extracellular signal-regulated protein kinase (ERK)-1 pocket, which was verified to produce profound ERK suppression. Most strikingly, compound 10 exhibited superior in vivo antitumor efficiency and effectively attenuated systemic toxicity. Our results emphasize that functionalizing platinum drugs with the multi-target EVO could generate synergistically excellent anticancer activity with low toxicity and decreased resistance, which may represent a brand-new cancer therapy modality.

Rapid GSH detection and versatile peptide/protein labelling to track cell penetration using coumarin-based probes.

Biothiols play essential roles in balancing the redox state and modulating cellular functions. Fluorescent probes for monitoring/labelling biothiols often suffer from slow reaction rates, strong background fluorescence and cytotoxic byproduct release. Thus, developing facile and versatile probes to overcome the challenges is still in high demand. Here, we report four coumarin-maleimides as fast responding and fluorogenic probes to detect GSH or label peptides/proteins. The probes quantitatively and selectively react with GSH via Michael addition within 1-2 min, achieving an 11-196-fold increase in fluorescence quantum yield via blockage of the photoinduced electron transfer (PET) process. Optimized probe 4 is applied for the detection of GSH in vitro (A549 cells) and in vivo (zebrafish embryos). Taking advantage of the fast Michael addition between the maleimide moiety and the sulfhydryl group, we expand the application of our method for fluorescent labelling of peptides/proteins and for tracking their cellular uptake process. The labelling strategy works for both Cys-bearing and Cys-free proteins after the introduction of a sulfhydryl group using Traut's reagent. Fluorescence assay reveals that the TAT-peptide can efficiently enter cells, but H3 protein, part of nucleosomes, prefers to bind on the cell membrane by electrostatic interactions, shedding light on the cellular uptake activity of nucleosomes and affording a potential membrane staining strategy. Overall, our study illustrates the broad potential of coumarin-maleimide based dual-functional probes for GSH detection and versatile protein labelling in biochemical research.

Thalidomide-based Pt(IV) prodrugs designed to exert synergistic effect of immunomodulation and chemotherapy.

Combination of immune- and chemo-therapy has become a new trend in cancer treatment. Food and Drug Administration (FDA)-approved immune-modulatory agent, thalidomide, can modulate the related proteins of upstream signaling pathway of programmed cell death-ligand 1 (PD-L1), including nuclear transcription factor κB (NF-κB), hypoxia inducible factor-1α (HIF-1α), epidermal growth factor receptor (EGFR), and signal transducer and activator of transcription 3 (STAT3), all acting as key antitumor target proteins. In this work, we conjugated thalidomide with oxidized cisplatin to construct multi-functional Pt(IV) prodrugs, named thaliplatins 4-6, to investigate the anti-tumor effect of immuno- and chemo-therapy. Among them, thaliplatin 6 exerted remarkable cytotoxicity against the tested cancer cell lines, showing 15-26 and 9-20 times higher IC50 values than those of single cisplatin or the combination of cisplatin + thalidomide, respectively. Moreover, thaliplatin 6 could rapidly accumulated into cells, markedly triggered DNA damage, and induced cell S phase arrest and apoptosis, as well as inhibited cell migration and invasion in breast carcinoma cell line (MCF-7). Fluorescent confocal and western blotting experiments proved that 6 significantly regulated NF-κB, EGFR, HIF-1α and phosphor-signal transducer and activator of transcription 3 (p-STAT3), and simultaneously inhibited PD-L1 expression to interrupt programmed cell death 1 (PD-1)/PD-L1 signaling pathway, suggesting a synergistic action of cisplatin and thalidomide. Most strikingly, in vivo tests indicated that 6 effectively decreased tumor growth with no observable systemic toxicity, being superior to the anticancer efficacy of cisplatin.

A novel double target fluorescence probe for Al3+/Mg2+ detection with distinctively different responses and its applications in cell imaging.

The metal cations, Al3+ and Mg2+, could affect human health and cell biological processes. Their fast and selective detection using one probe remains a challenge. A novel fluorescence probe, N'-((1-hydroxynaphthalen-2-yl)methylene)isoquinoline-3-carbohydrazide (NHMI), was developed for selectively monitoring Al3+ and Mg2+. The probe NHMI showed a distinctive "turn-on" fluorescence signal towards Al3+ and Mg2+ (cyan for Al3+ with 2556-folds enhancement and yellow for Mg2+ with 88-folds enhancement), which is quite distinct from other metal cations and allows for naked-eye detection. This interesting response was attributed to the influence of PET, ESIPT process and CHEF effect, when Al3+ or Mg2+ chelated with NHMI. Furthermore, the fluorescence titration experiments manifested that the detection limit of probe NHMI for Al3+/Mg2+ was as low as 1.20 × 10-8 M and 7.69 × 10-8 M, respectively. The formed complexes NHMI-Al3+ and NHMI-Mg2+ were analyzed by Job's plot, ESI-MS, 1H NMR and FT-IR. The coordination pockets and fluorescence mechanisms of two metal complexes were explored by density functional theory calculation. Moreover, NHMI showed low cytotoxicity and good cell permeability. Fluorescence bioimaging of Al3+/Mg2+ in MCF-7 cells with NHMI indicated its potential application in biological diagnostic analysis.

A dual-functional fluorescent probe for sequential determination of Cu2+/S2- and its applications in biological systems.

A new acylhydrazine-derived Schiff base fluorescence probe DMI based on "ON-OFF-ON" fluorescence strategy was presented in this paper. Probe DMI could detect Cu2+ selectively and sensitively with dramatic fluorescence quenching in CH3OH-PBS (v/v = 3:7) mixed solution. Once the complex DMI-Cu2+ interacted with S2-, 10.67-folds fluorescence increase was induced via a displacement mechanism under the same experimental conditions. The corresponding detection limits for Cu2+ and S2- were calculated to be 1.52 × 10-8 M and 1.79 × 10-8 M, respectively. The structures of DMI and DMI-Cu2+ were systematically characterized by Job's plot analysis, ESI-MS, IR, X-ray diffraction and density functional theory calculations. Furthermore, fluorescence imaging in MCF-7 cells and zebrafish demonstrated the probe DMI could act as a useful tool to monitor and track intracellular Cu2+ and S2-, which was encouraged by remarkable fluorescence performance and low cytotoxicity. Importantly, the complex DMI-Cu2+ could be applied to detect corrupt blood samples, which could estimate the time of death.

A novel hydrazide Schiff base self-assembled nanoprobe for selective detection of human serum albumin and its applications in renal disease surveillance.

Human serum albumin (HSA) is considered as a biomarker for the early diagnosis of renal disease, therefore identifying and detecting HSA in biological fluids (especially urine) with an easy method is of great importance. Herein, we report a novel hydrazide Schiff base fluorescent probe N'-((7-(diethylamino)-2-oxo-2H-chromen-3-yl)methylene)pyrazine-2-carbohydrazide (NPC), which self-assembled into nanoparticles in aqueous solution. Based on disassembly-induced emission and the site-specific recognition mechanism, the binding of NPC with HSA resulted in a fluorescence "turn-on" response. Probe NPC exhibited superior selectivity and sensitivity toward HSA with a detection limit of 0.59 mg L-1 in PBS and 0.56 mg L-1 in the urine sample. The site-binding mechanism of NPC with HSA was explored by fluorescence quenching study, Job's plot analysis, HSA destruction, site marker displacement and molecular docking. Fluorescence imaging of HSA in MCF-7 cells was achieved by using a non-toxic NPC probe, suggesting that NPC could be applied to visualize the level of HSA in vivo. More importantly, further practical applications of probe NPC in human urine samples were achieved with satisfactory results by using a fluorometer or test paper, which could provide extensive application in clinical diagnosis.

Anticancer Melatplatin Prodrugs: High Effect and Low Toxicity, MT1-ER-Target and Immune Response In Vivo.

Multitargeted therapy could rectify various oncogenic pathways to block tumorigenesis and progression. The combination of endocrine-, immune-, and chemotherapy might exert a highly synergistic effect against certain tumors. Herein, a series of smart Pt(IV) prodrugs 3-6, named Melatplatin, were rationally designed not only to multitarget DNA, MT1, and estrogen receptor (ER) but also to activate immune response. Melatplatin, conjugating first-line chemotherapeutic Pt drugs with human endogenous melatonin (MT), significantly enhanced drug efficacy especially in ER high-expression (ER+) cells, among which 3 presented the most potent cytotoxicity toward ER+ MCF-7 with nanomolar IC50 values 100-fold lower than cisplatin. Melatplatin could bind well to melatonin receptor (MT1) according to molecular docking. Besides, 3 evidently increased intracellular accumulation and DNA damage, upregulated γH2AX and P53, and silenced NF-κB to induce massive apoptosis. Most strikingly, 3 effectively inhibited tumor growth and attenuated systemic toxicity compared to cisplatin in vivo, promoting lymphocyte proliferation in spleen to achieve immune modulation.

Electronic Cigarette Aerosol Modulates the Oral Microbiome and Increases Risk of Infection.

The trend of e-cigarette use among teens is ever increasing. Here we show the dysbiotic oral microbial ecology in e-cigarette users influencing the local host immune environment compared with non-smoker controls and cigarette smokers. Using 16S rRNA high-throughput sequencing, we evaluated 119 human participants, 40 in each of the three cohorts, and found significantly altered beta-diversity in e-cigarette users (p = 0.006) when compared with never smokers or tobacco cigarette smokers. The abundance of Porphyromonas and Veillonella (p = 0.008) was higher among vapers. Interleukin (IL)-6 and IL-1ß were highly elevated in e-cigarette users when compared with non-users. Epithelial cell-exposed e-cigarette aerosols were more susceptible for infection. In vitro infection model of premalignant Leuk-1 and malignant cell lines exposed to e-cigarette aerosol and challenged by Porphyromonas gingivalis and Fusobacterium nucleatum resulted in elevated inflammatory response. Our findings for the first time demonstrate that e-cigarette users are more prone to infection.

Syntheses, characterization, DNA/HSA binding ability and antitumor activities of a family of isostructural binuclear lanthanide complexes containing hydrazine Schiff base.

Three dinuclear lanthanide complexes, [Ln2(L)2(µ3-OAc)4(H2O)2]⋅2H2O (Ln = La (1), Eu (2) and Dy (3), HL = N'-(2-hydroxybenzylidene) nicotinohydrazide), have been synthesized and characterized by IR, elemental analysis and X-ray single-crystal diffraction. Crystallographic study revealed that the representative complex 1 displays a discrete dinuclear structure with a distorted tricapped trigonal prismatic geometry around La(III) ion. The interaction of complexes 1-3 with CT-DNA was investigated by absorption spectra, fluorescence quenching and viscosity, which reveals that the complexes bind to CT-DNA with a moderate intercalative mode. The complexes exhibited obvious DNA cleavage activities in the presence of H2O2. All complexes could bind to human serum albumin (HSA) with medium affinity through static mode; thus, HSA could effectively transport complexes. Furthermore, three complexes exhibited specific cytotoxicity to A549 cancer cells in micromole magnitude than other cancer cells tested and less toxicity than cisplatin for normal human cells HUVEC, in which massive cell apoptosis was induced by complexes through producing DNA damage and suppressing DNA synthesis.Communicated by Ramaswamy H. Sarma.

A highly selective colorimetric and fluorescent probe for quantitative detection of Cu2+/Co2+: The unique ON-OFF-ON fluorimetric detection strategy and applications in living cells/zebrafish.

Identifying and detecting similar target cations through combining "turn on" and "turn off" fluorescence mechanism is effective and challenging. Now a new colorimetric and ON-OFF-ON fluorescent probe N'-((7-(diethylamino)-2-oxo-2H-chromen-3-yl)methylene)-3-hydroxy-2-naphthohydrazide (L) was reported, which could detect Cu2+ and Co2+ in phosphate buffered CH3CH2OH-H2O solvent system. With the assistance of glutathione and pH adjustment, a unique ON-OFF-ON fluorescence detection strategy could be achieved for distinguishing Cu2+ and Co2+. The emission of probe could recover from the L-Cu2+ and L-Co2+ system by addition of GSH or adjusting pH value to 4, respectively, which is due to the abolishment of paramagnetic Cu2+/Co2+. Based on fluorescence titration experiments, the limit of detection was determined as 3.84 × 10-9 M and 4.55 × 10-9 M for Cu2+ and Co2+, respectively. Meanwhile, the detection limit reached 6.21 × 10-8 M for Cu2+ and 6.96 × 10-8 M for Co2+ according to absorbance signal output. Fast recognition of Cu2+/Co2+ can be achieved by obvious color changes from green to colorless under UV light, as well as from yellow to orange-red in room light. The binding mode of L toward Cu2+ and Co2+ have been systematically studied by Job's plot analysis, ESI-MS, IR and density functional theory calculations. Most strikingly, further practical applications of the probe L in fluorescence imaging were investigated in MCF-7 cells and zebrafish due to its low cytotoxicity and good optical properties, suggesting that L could serve as a fluorescent sensor for tracking Cu2+ and Co2+in vivo.

Targeting TRPV1 on cellular plasticity regulated by Ovol 2 and Zeb 1 in hepatocellular carcinoma.

The landscape of cellular plasticity and sources with relevant niche signals in hepatocellular carcinoma is still obscure. Transient receptor potential vanilloid 1 (TRPV1), a non-selective cation channel, is involved in a variety of malignancies and overexpressed in hepatocellular carcinoma (HCC). We have investigated the role of TRPV1 in HCC from different angles by various experimental techniques, such as in vivo and in vitro experiments, and by bioinformatics analysis of data from genetic models induced by diethylnitrosamine (DEN), mice samples and human HCC samples. We find that TRPV1 knockout promotes to hepatocarcinogenesis and deconstructs the portal triad adjacent to tumor border that is contributed by originations of tumor initiating cells and biliary cells. Epithelial to mesenchymal transition (EMT) is involved and transcription factors Ovol2 and Zeb1 coordinated with Sox 10 drive gene expression in the event which is also confirmed by the expression of these proteins in human HCC samples. Treatment with TRPV1 agonist Capsaicin inhibits the growth of HCC cells in xenograft models. Our findings demonstrate that TRPV1 is a potential therapeutic target in human HCC and exerts effects on cellular plasticity with modulation of Ovol2, Zeb1 and Sox10.

A dual functional turn-on non-toxic chemosensor for highly selective and sensitive visual detection of Mg2+ and Zn2+: the solvent-controlled recognition effect and bio-imaging application.

A new dual functional turn-on chemosensor, 2,6-diformyl-4-methylphenol-di(isoquinolinyl-1-hydrazone) (HL), has been developed, which could highly selectively discriminate Mg2+ and Zn2+ in different solvent systems. The chemosensor HL exhibits rapid visual turn-on fluorescence enhancing recognition toward Mg2+/Zn2+, which is not interfered by other cations, especially for respective congeners Ca2+/Cd2+. The remarkable fluorescence enhancement (71-fold or 11-fold) was observed after adding Mg2+ in acetonitrile or Zn2+ in DMF-H2O solvent systems. Additionally such a solvent medium-controlled platform could achieve the quantitative determination of Mg2+ and Zn2+ quantitation with low detection limits of 2.97 × 10-8 M and 3.07 × 10-7 M, respectively. Furthermore, the turn-on fluorescence sensing mechanism is also investigated by 1H NMR, FT-IR and ESI-MS spectroscopy. Density functional theory (DFT) calculations derive optimized geometries of HL and its complexes. Notably, non-toxic HL also can be successfully applied as a visual probe for the practical determination of Mg2+/Zn2+ in MCF-7 cells, Zebrafish larvae, syrup and water samples, which might provide extensive application in biology and medicine fields.

Rapid induction of apoptosis in tumor cells treated with a new platinum(II) complex based on amino-thiazolidinone.

Thiazolidinone derivatives have been previously shown significant anti-cancer activities. Two amino-thiazolidinone complexes, [Pt(HTone)Cl] (1) and [Cu(HTone)Cl] (3) (HTone = (Z)-2-((E)-(1-(pyridin-2-yl)ethylidene)hydrazono)thiazolidin-4-one) and one ethyl-modified [Pt(ETone)Cl2] (2) (ETone = (Z)-3-ethyl-2-((E)-(1-(pyridin-2-yl)ethylidene) hydrazono)thiazolidin-4-one)], were designed and synthesized in order to explore novel metal-based antitumor agents. MTT assay indicated that 1 and 3 were markedly cytotoxic to MCF-7, HepG-2 and NCI-H460 tumor cells, superior to both cisplatin and the HTone ligand. Massive dead cells were observed as early as 6 h when treated with 1, indicating rapid action of 1 as compared to that of other compounds. More interestingly, Hoechst 33342 staining and flow cytometry analysis illustrated that only complex 1 could induce obvious cell apoptosis within 12 h, which was associated with the high-expression of Bax and cleavage of caspase-3 from 35 kDa to 17 kDa. By means of ICP-MS assay, we found complex 1 could largely accumulate in tumor cells in a short time. Additionally, complex 1 showed no cross resistance against the cisplatin-resistant cells.

Transient receptor potential vanilloid-type 2 targeting on stemness in liver cancer.

The malignant phenotype of the cells resulting from human liver cancer is driven by liver cancer stem-like cells (LCSLCs). Transient Receptor Potential Vanilloid-type 2 channel (TRPV2) contributes to the progression of different tumor types, including liver cancer. In the current study, the TRPV2 expression levels give rise to the effect on stemness in liver cancer cell lines. TRPV2 knockdown in HepG2 cells enhanced spheroid and colony formation, and expression levels of CD133, CD44 and ALDH1 whereas the opposite effects were observed in TRPV2 enforced expression in SMMC-7721 cells. Furthermore, TRPV2 overexpression restored inhibition of spheroid and colony formation, and stem cell markers expression in HepG2 cells with TRPV2 silencing. The addition of the TRPV2 agonist probenecid and the TRPV2 antagonist tranilast suppressed and/or increased in vitro spheroid and colony formation, and stem cell marker expression of LCSLCs and/or liver cancer cell lines, respectively. Notably, probenecid and tranilast significantly inhibited or promoted tumor growth of HepG2 xenografts in the severe combined immunodeficiency (SCID) mouse model, respectively. TRPV2 expression at protein levels revealed converse correlation with those of CD133 and CD44 in human hepatocellular carcinoma (HCC) tissue. Collectively, the data demonstrate that TRPV2 exert effects on stemness of liver cancer and is a potential target in the treatment of human liver cancer patients.

Pharmacological inhibition of TRPV4 channel suppresses malignant biological behavior of hepatocellular carcinoma via modulation of ERK signaling pathway.

TRPV4 (transient receptor potential vanilloid 4), a member of the TRP superfamily, has been reported to correlate with several different forms of cancers. However, the role of TRPV4 in human hepatocellular carcinoma (HCC) remains unclear. The present study demonstrated that elevated expression of TRPV4 was shown in HCC tumor tissues when compared with paired non-tumoral livers both in protein and mRNA levels. Furthermore, the enhanced expression of TRPV4 was highly associated with histological grade (P = 0.036) and the number of tumors (P = 0.045). Pharmacological inhibition of TRPV4 channels in HCC cells with the specific antagonist HC-067047 suppressed cell proliferation, induced apoptosis and decreased the migration capability by attenuating the epithelial-mesenchymal transition (EMT) process in vitro. The p-ERK expression was apparently repressed after treatment with the TRPV4 antagonist, further blockade of the ERK pathway with U0126 could significantly aggravate HCC cells apoptosis. In NOD-SCID mouse xenograft models, intraperitoneal injection of HC-067047 could obviously suppress tumor growth and induce apoptosis in vivo. Together, our studies showed that the antitumor effects caused by TRPV4 channel inhibition in HCC cell lines might be attributed to the suppression of EMT process and inactivation of p-ERK which induced subsequent cell apoptosis. Thus, pharmacological inhibition of TRPV4 channel may be an option for HCC treatment.

Splenectomy after partial hepatectomy accelerates liver regeneration in mice by promoting tight junction formation via polarity protein Par 3-aPKC.

AIMS: Several experimental studies have demonstrated that removal of the spleen accelerates liver regeneration after partial hepatectomy. While the mechanism of splenectomy promotes liver regeneration by the improvement of the formation of tight junction and the establishment of hepatocyte polarity is still unknown. MAIN METHODS: We analyzed the cytokines, genes and proteins expression between 70% partial hepatectomy mice (PHx) and simultaneous 70% partial hepatectomy and splenectomy mice (PHs) at predetermined timed points. KEY FINDINGS: Compared with the PHx group mice, splenectomy accelerated hepatocyte proliferation in PHs group. The expression of Zonula occludens-1 (ZO-1) indicated that splenectomy promotes the formation of tight junction during liver regeneration. TNF-α, IL-6, HGF, TSP-1 and TGF-ß1 were essential factors for the formation of tight junction and the establishment of hepatocytes polarity in liver regeneration. After splenectomy, Partitioning defective 3 homolog (Par 3) and atypical protein kinase C (aPKC) regulate hepatocyte localization and junctional structures in regeneration liver. SIGNIFICANCE: Our data suggest that the time course expression of TNF-α, IL-6, HGF, TSP-1, and TGF-ß1 and the change of platelets take part in liver regeneration. Combination with splenectomy accelerates liver regeneration by improvement of the tight junction formation which may help to establish hepatocyte polarity via Par 3-aPKC. This may provide a clue for us that splenectomy could accelerate liver regeneration after partial hepatectomy of hepatocellular carcinoma and living donor liver transplantation.

Succinate and its G-protein-coupled receptor stimulates osteoclastogenesis.

The mechanism underlying bone impairment in patients with diabetes mellitus, a metabolic disorder characterized by chronic hyperglycaemia and dysregulation in metabolism, is unclear. Here we show the difference in the metabolomics of bone marrow stromal cells (BMSCs) derived from hyperglycaemic (type 2 diabetes mellitus, T2D) and normoglycaemic mice. One hundred and forty-two metabolites are substantially regulated in BMSCs from T2D mice, with the tricarboxylic acid (TCA) cycle being one of the primary metabolic pathways impaired by hyperglycaemia. Importantly, succinate, an intermediate metabolite in the TCA cycle, is increased by 24-fold in BMSCs from T2D mice. Succinate functions as an extracellular ligand through binding to its specific receptor on osteoclastic lineage cells and stimulates osteoclastogenesis in vitro and in vivo. Strategies targeting the receptor activation inhibit osteoclastogenesis. This study reveals a metabolite-mediated mechanism of osteoclastogenesis modulation that contributes to bone dysregulation in metabolic disorders.

Histone deacetylases 1 and 2 cooperate in regulating BRCA1, CHK1, and RAD51 expression in acute myeloid leukemia cells.

Resistance to chemotherapy and a high relapse rate highlight the importance of finding new therapeutic options for the treatment of acute myeloid leukemia (AML). Histone deacetylase (HDAC) inhibitors (HDACIs) are a promising class of drugs for the treatment of AML. HDACIs have limited single-agent clinical activities, but when combined with conventional or investigational drugs they have demonstrated favorable outcomes. Previous studies have shown that decreasing expression of important DNA damage repair proteins enhances standard chemotherapy drugs. In our recent studies, the pan-HDACI panobinostat has been shown to enhance conventional chemotherapy drugs cytarabine and daunorubicin in AML cells by decreasing the expression of BRCA1, CHK1, and RAD51. In this study, we utilized class- and isoform-specific HDACIs and shRNA knockdown of individual HDACs to determine which HDACs are responsible for decreased expression of BRCA1, CHK1, and RAD51 following pan-HDACI treatment in AML cells. We found that inhibition of both HDAC1 and HDAC2 was necessary to decrease the expression of BRCA1, CHK1, and RAD51, enhance cytarabine- or daunorubicin-induced DNA damage and apoptosis, and abrogate cytarabine- or daunorubicin-induced cell cycle checkpoint activation in AML cells. These findings may aid in the development of rationally designed drug combinations for the treatment of AML.

Synthesis and Antileukemic Activities of Piperlongumine and HDAC Inhibitor Hybrids against Acute Myeloid Leukemia Cells.

Synergistic-to-additive antileukemic interactions of piperlongumine (PL) and HDAC inhibitor (HDACi) SAHA (Vorinostat) provide a compelling rationale to construct PL-HDACi hybrids, such as 1-58, which recapitulated the synergism between the parental compounds in high-risk and chemoresistant AML cells. Both PL and HDACi components, either in combination or in hybrid molecules, are essential for inducing significant DNA damage and apoptosis. Introducing C2-chloro substituent to 1-58 yielded 3-35 with increased cytotoxicity but decreased selectivity in noncancerous MCF-10A cells; eliminating C7-C8 olefin of PL obtained 3-31/3-98 scaffolds which were still more active than PL or SAHA in AML and were well-tolerated by MCF-10A cells. The HDACi function was crucial for modulating expression of DNA repair and apoptosis-related proteins. Collectively, PL and SAHA hybrids are potent, multifunctional anti-AML agents, acting in part, by interfering cellular GSH defense, suppressing expression of DNA repair and pro-survival proteins, and inducing expression of pro-apoptotic proteins.