High Degree Of Synergism Research Articles

P1278: ACTIVATION OF THE NECROPTOSIS PATHWAY PLAYS A ROLE IN SYNERGISM WITH COMBINATION OF THE IAP ANTAGONIST, TOLINAPANT AND HYPOMETHYLATING AGENTS (HMA) IN IN VITRO MODELS OF T-CELL LYMPHOMA (TCL).

Background: Peripheral T-cell lymphoma (PTCL) are a highly aggressive and heterogeneous group of non-Hodgkin lymphomas that often carry a poor prognosis with standard chemotherapy. Tolinapant is a potent, non-peptidomimetic antagonist of cIAP1, cIAP2 and XIAP. In an ongoing Phase 2 trial (NCT02503423), tolinapant has shown activity against highly pre-treated peripheral and cutaneous T-cell lymphoma. HMAs have also shown clinical responses in some subsets of PTCL. Both HMAs and IAP antagonists show immunomodulatory anti-cancer potential in pre-clinical studies. We have also shown that decitabine and tolinapant as single agent induced necroptosis in mouse TCL cell lines with the activation of the RIPK1/RIPK3/MLKL necroptosis pathway. In addition, re-expression of RIPK3 in CT26 cells by CRISPR increased lytic cell death on treatment with tolinapant indicating its essential role in the necroptosis pathway. Aims: We explored the sensitivity of a range of TCL lines to tolinapant, established the synergy coefficient and interrogated the role of necroptosis in the mechanism of synergy between tolinapant and drugs active against PTCL, the HMAs; azacytidine and decitabine Methods: A panel of 10 human TCL lines were tested in proliferation assays (CellTiterGlo) for sensitivity to tolinapant in the presence or absence of 10 ng/ml of TNFa. For combination studies, each drug was tested at the IC10, IC20 and IC40 concentrations, in the presence or absence of TNFa. Synergy coefficient was calculated using the Excess over Bliss model with SynergyFinder. Additionally, the on-target effects of the drugs were measured by analyzing levels of the IAPs, DNMTs and key apoptosis and necroptosis markers, such as PARP and RIPK3 by Western blotting. Results: TCL Lines demonstrate varying sensitivities to tolinapant with the most sensitive cell line, ALK+ ALCL SUP-M2, having an IC50 as low as 20 nM ± 1 nM while a resistant CTCL cell line HH had an IC50 of over 20 µM. In combination experiments using both cell lines, tolinapant plus azacytidine or decitabine displayed a very high degree of synergism; Average synergy score of 50.9 and 49.8 with azacytidine and 22.6 and 31.3 with decitabine for HH and SUP-M2, respectively. Of note, a high degree of synergism was also seen with concentrations 10 fold lower of azacytidine or 100 fold lower of decitabine, added daily (Figure 1). The combination of tolinapant and the HMAs led to a decrease in the levels of cIAP1 and DNMT3a in TCL lines, demonstrating on-target activity of tolinapant and the HMAs, respectively. In addition, both azacytidine and decitabine increased the levels of RIPK3 up to 3-4 fold and 6-8 fold respectively, in a concentration dependent manner at 72Hrs by western blot analysis, indicating activation of the necroptosis pathway. Image:Figure 1: 3D synergy plots showing average synergy scores for decitabine in combination with tolinapant in the CTCL line HH.Summary/Conclusion: Tolinapant has demonstrated varying cytotoxic effects against a range of TCL lines as a single agent. Tolinapant, in combination with the HMAs, azacytidine and decitabine, displays a very high degree of synergism, in vitro, even at very low concentrations of both HMA drugs. The HMAs were able to activate the necroptosis pathway, as shown by the increase in RIPK3. Thus, activation of the necroptosis pathway, is a possible mechanism for the high degree of synergism displayed when HMAs are used in combination with tolinapant in the TCL lines in vitro. These data provide the rationale to initiate clinical trials with the combination of tolinapant and decitabine.

The combination of the IAP antagonist, tolinapant and hypomethylating agents (HMA) is highly synergistic in in vitro models of T-cell lymphoma (TCL).

e15088 Background: Peripheral T-cell lymphoma (PTCL) are a highly aggressive and heterogeneous group of non-Hodgkin lymphomas that often carry a poor prognosis with standard chemotherapy. Tolinapant is a potent antagonist of inhibitors of apoptosis proteins (cIAP1/2 and XIAP). In a phase 2 trial (NCT02503423), tolinapant showed activity against heavily pre-treated patients with TCL. We explored the sensitivity of a range of TCL lines to tolinapant, established the synergy coefficient and interrogated the mechanism of synergy between tolinapant and drugs active against PTCL including romidepsin, pralatrexate, and the HMAs; azacytidine and decitabine. Methods: A panel of 10 human TCL lines were tested in proliferation assays (CellTiterGlo) for sensitivity to tolinapant in the presence or absence of 10 ng/ml of TNFα. For combination studies, each drug was tested, in the presence or absence of TNFα. Synergy coefficient was calculated using Excess over Bliss. Additionally, the on-target effects of the drugs were measured by analyzing levels of the IAPs, acetylated histones, DNMTs and key apoptosis and necroptosis markers by Western blotting. Results: TCL Lines demonstrated varying sensitivities to tolinapant with the most sensitive cell line, ALK+ ALCL SUP-M2, having an IC50 as low as 20 nM ± 1 nM while a resistant CTCL cell line HH had an IC50 of over 20 µM. In combination experiments using both cell lines, tolinapant plus azacytidine or decitabine displayed the highest degree of synergism, compared to romidepsin or pralatrexate. Of note, a high degree of synergism was also seen with concentrations 10 folds lower of azacytidine or 100 folds lower of decitabine, added daily (Table). The combination of tolinapant and the HMAs led to a decrease in the levels of cIAP1 and DNMT3a in TCL lines, demonstrating on-target activity of tolinapant and the HMAs, respectively. In addition, both azacytidine and decitabine increased the levels of RIPK3 and MLKL by western blot analysis, indicating activation of the necroptosis pathway. Conclusions: Tolinapant has demonstrated varying cytotoxic effects against a range of TCL lines as a monotherapy and displays a very high degree of synergism in combination with the HMAs, azacytidine and decitabine, also at low concentration, in vitro. The HMAs were also able to activate the necroptosis pathway, providing a possible mechanism for the high degree of synergism displayed when combined with tolinapant in the TCL lines in vitro. These data provide the rationale to initiate phase I trials with the combination of tolinapant and HMAs. Average synergy scores at 48 hrs for drugs used in combination with tolinapant in a CTCL line HH and ALK+ ALCL line SUP-M2. Above 10 indicates synergy.[Table: see text]

Evaluation of the in vitro synergy of polymyxin B-based combinations against polymyxin B -resistant gram-negative bacilli

ObjectivesThis study aimed to evaluate the in vitro synergy of polymyxin B (PMB) combined with 11 other antibiotics against PMB-resistant gram-negative bacilli (GNBs). MethodsThirty-six clinical isolates of PMB-resistant GNBs were used. The MICs of all the antimicrobials tested were determined by the broth microdilution method and the checkerboard assay method. Carbapenemase genes were detected by PCR. In vitro synergy results were interpreted using the fractional inhibitory concentration index (FICI). Four combinations that showed positive interactions were subsequently evaluated in a time-kill study. ResultsAmong the 36 strains, 15 harboured the carbapenemase gene, and blaKPC was the predominant carbapenemase. The resistance rates of the 36 strains to tigecycline, meropenem, ceftazidime, and cefepime were 100% (36/36), 97% (35/36), 94% (34/36), and 97% (35/36), respectively. For Enterobacteriaceae and Pseudomonas aeruginosa, the resistance rates to aztreonam and ceftazidime-avibactam (avibactam at a fixed concentration of 4 mg/L) were 95% (19/20) and 25% (5/20), respectively. The PMB-based combinations exhibited synergism to a certain degree. The most synergistic combinations were PMB plus meropenem-avibactam (avibactam at a fixed concentration of 4 mg/L) and PMB plus tigecycline, with the synergy rates of 83.3% and 80.6%, respectively. Compared to tazobactam- and sulbactam-based β-lactam–β-lactamase inhibitor combinations (BL-BLIs), PMB with avibactam-based BL-BLIs exhibited a better synergistic effect. For Acinetobacter baumannii, PMB plus sulbactam exhibited a strong synergism with a 2∼7-fold MIC reduction of PMB. In time-kill studies, the highest degree of synergism was observed for PMB with cefepime-avibactam on all the tested isolates. For isolates with low-level resistance to PMB, PMB combined with other partner antimicrobials also showed a certain degree of synergism. ConclusionsPMB in combination with tigecycline and avibactam-based BL-BLIs could be a potential clinical option for clinical treatment of infections caused by PMB -resistant GNBs.

Cytokine genes multi-locus analysis reveals synergistic influence on genetic susceptibility in Indian SLE – A multifactor-dimensionality reduction approach

Systemic lupus erythematosus (SLE) is a prototype autoimmune disease with unclear etiology. Several loci associated with genetic susceptibility for lupus have been described. However, it lacks reports on cytokine gene-gene interactions among SLE patients from Asian population. Epistasis interaction among single nucleotide polymorphisms (SNPs) of cytokine genes in Indian SLE patients was tested using multifactor-dimensionality reduction (MDR) analysis. A total of fourteen SNPs lacking linkage disequilibrium among different cytokines genes were genotyped in a cohort of 200 SLE patients and 201 healthy individuals as controls of Indian origin. A high degree of synergism among Lymphotoxin-α (LT-α), Interleukin-1β (IL-1β) and Interleukin-10 (IL-10) gene polymorphisms was detected in our SLE patients. Furthermore, by virtue of biological inter-relations among different cytokines, a high strength of interactions was observed among pro-inflammatory (IL-1β, IL-6) and anti-inflammatory (IL-10) cytokine gene SNPs. Also, among studied pro-inflammatory cytokines and chemokines, a strong synergistic effect among Tumor Necrosis Factor-α (TNF-α), LT-α and Monocyte Chemo-attractant Protein-1 (MCP-1) SNPs was occurred. A nature of strong interaction among the candidate cytokine genes may speculate a proactive role in causing genetic susceptibility to the disease in SLE patients with Indian origin.

Highly synergistic drug combination prevents vaginal HIV infection in humanized mice

The HIV-1 epidemic remains an urgent global health concern. Young women are disproportionately at risk of acquiring the virus. A range of highly effective, female-controlled, discrete vaginal products therefore is needed to help curb the epidemic. Oral tenofovir disoproxil fumarate (TDF) and emtricitabine (FTC) are effective in HIV-1 pre-exposure prophylaxis (PrEP) and form a promising basis for a vaginal product. Here, we evaluate TDF and FTC in combination with the broadly neutralizing antibody VRC01-N using a highly reproducible humanized mouse model. The agents were vaginally dosed individually and in combination, and the efficacy of HIV-1 prevention was analyzed using the established, rigorous median-effect model. Surprisingly, the triple combination showed a high degree of synergism, unprecedented for in vivo HIV-1 PrEP, leading to a possible fivefold dose reduction for some of the agents. Vaginal administration of the TDF-FTC-VRC01-N combination holds significant promise for HIV-1 PrEP.

Synergistic Enzyme Cocktail to Enhance Hydrolysis of Steam Exploded Wheat Straw at Pilot Scale

Multiple enzymes are required for efficient saccharification of lignocellulosic biomass and no wild type organism is capable of producing all enzymes in desired levels. In this study, steam explosion of wheat straw was carried out at pilot scale and a synthetic enzyme mixture (EnzMix) was developed by partially replacing the cellulase with critical dosages of commercially available accessory enzymes (β-glucosidase, xylanase and laccase) through central composite design. Highest degree of synergism (DS) was observed with β-glucosidase (1.68) followed by xylanase (1.36). Finally, benchmarking of EnzMix (Celluclast, β-glucosidase and xylanase in a protein ratio of 20.40: 38.43: 41.16, respectively) and other leading commercial enzymes was carried out. Interestingly, saccharification improved by 75% at 6h and 30% at 24h, respectively in comparison of control. By this approach, 25% reduction in enzyme dosage was observed for obtaining the same saccharification yield. Thus development of enzyme cocktail is an effective and sustainable approach for high saccharification efficiency.

Abstract 1889: Synergistic effects of glutaminase and proteasome inhibition in multiple myeloma cell lines

Abstract Background: Multiple Myeloma (MM) accounts for approximately 2.1% of all cancer deaths with a 5-year median survival rate of 49.6%. Besides chemotherapy, treatment options include monoclonal antibodies, immunomodulatory agents, and proteasome inhibitors. Carfilzomib is a selective proteasome inhibitor administered intravenously in patients with relapsed or refractory MM. With cancer cell metabolism emerging as a critical regulator of tumor progression, combining carfilzomib with an inhibitor of the metabolic machinery represents a novel therapeutic strategy to prevent MM progression. RP10107 is a potent, and selective glutaminase (GLS-1) inhibitor that demonstrated high potency against mouse (IC50=21.2 nM), rat (IC50=18.2 nM) and human (IC50=26.4 nM) enzymes with selectivity over GLS-2 (>380-fold). The objective of this study was to evaluate the effect of a combination of carfilzomib and RP10107 in MM cell lines. Methods: Glutamate concentrations in MM cell lines (MM-1S and RPMI-8226) following treatment with RP10107 was estimated by LC-MS/MS. Synergism between RP10107 and carfilzomib was determined using different concentrations of the compounds in a 5 x 5 grid. Synergism, additivity, or antagonism was calculated based on the BLISS score. Apoptosis was determined by Annexin V/7AAD staining using a MUSE® Annexin V and dead cell assay kit (Millipore) while the effect on cell cycle was estimated using Guava Cell Cycle Reagent (Millipore). Expression of cleaved PARP and cleaved caspase-8 were determined by Western Blotting. Results: Treatment of MM cell lines with RP10107 resulted in an increased ratio of glutamine to glutamate with a doubling noticed at concentrations as low as 30 nM. A high degree of synergism between RP10107 and carfilzomib was noticed in both the MM cell lines. Addition of 3 nM Carfilzomib to 30 or 500 nM RP10107 resulted in near complete inhibition of RPMI-8226 and MM-1S cell growth, respectively. Additionally, incubation with the combination (1 μM RP10107 + 15 nM Carfilzomib) for 72 h caused a G0/G1 arrest with a corresponding increase in the percent of apoptotic cells (2.5-fold) compared to the individual agents. Incubating RPMI-8226 cells with a combination of RP10107 and Carfilzomib for 48 h resulted in a 2.2- and 13.6-fold increase in expression of cleaved caspase-8 and cleaved PARP, respectively. Conclusions: Addition of RP10107, a glutaminase inhibitor, potentiated Carfilzomib activity in MM cells. Findings provide a rationale for use of the combination in future clinical trials involving MM patients thereby providing a safer and more effective alternative to currently available therapy. Citation Format: Srikant Viswanadha, Satyanarayana Eleswarapu, Kumar V. Penmetsa, Swaroop Vakkalanka. Synergistic effects of glutaminase and proteasome inhibition in multiple myeloma cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1889.

Heterologous Expression of a New Acetyl Xylan Esterase from Aspergillus niger BE-2 and its Synergistic Action with Xylan-Degrading Enzymes in the Hydrolysis of Bamboo Biomass

Efficient utilization of plant biomass by enzymatic hydrolysis is currently studied worldwide but still faces enormous challenges because of the inability to break down lignocellulosic materials with high sugar yields and low enzyme dosage. Therefore, the synergistic action between various enzymes plays an important role in reaching this goal. The synergistic cooperation between a novel acetyl xylan esterase (heterologous expressed at high levels in this study) and four other xylan-degrading enzymes (reported previously) were performed in this study. The acetyl xylan esterase (AnAxe) gene was cloned from Aspergillus niger BE-2 and expressed in Pichia pastoris GS115. The deduced amino acid (aa) sequence consisted of 304-aa and included a 23-aa signal peptide and 281-aa mature protein. The AnAxe was extracellularly expressed with a molecular weight of ca. 31 kDa. The purified AnAxe exhibited maximal specific activity of 480.2 IU/mg at pH 7.0 and 40 °C and was still thermostable below 50 °C. The metal ions used in this study and EDTA showed a slight effect on the AnAxe. A significant synergistic effect was determined between AnAxe and the other four xylan-degrading enzymes, including endo-Beta-1,4-xylanases, Beta-xylosidases, alpha-L-arabinofurano-sidases, and alpha-glucuronidases, on the degradation of bamboo biomass. The highest degree of synergism was obtained between AnAxe and endo-Beta-1,4-xylanases/Beta-xylosidases.

Abstract 2520: Synergistic combination of lurbinectedin and PARP inhibitors in breast cancer tumor cell lines

Abstract Lurbinectedin (PM1183) is a new tetrahydroisoquinoline derivative currently evaluated in phase II trials in BRCA1-mutated breast cancer and NSCLC patients. Two pivotal trials (in platinum-resistant ovarian cancer and in SCLC) will be launched in 2015. PM1183 shows antitumor activity against a wide variety of tumor cells with mean IC50 values in the low nanomolar range. In living cells, PM1183 inhibits trans-activated transcription and blocks NER-dependent DNA repair inducing the formation of dsDNA breaks and the collapse of replication forks that need to be repaired by homologous recombination (HR). As expected, PM1183 is more active against homologous recombination (HR)-deficient cell lines. In this work, we evaluated the in vitro antitumor effect of PM1183 when combined with the PARP inhibitors ABT-88 (Veliparib, PARP-1 and -2), AZD-2281 (Olaparib, PARP-1), BSI-201 (Iniparib, PARP-1) and BMN-673 (PARP-1 and -2) (Selleck Chemicals, Houston, TX, USA) in a panel of human breast carcinoma cells with different BRCA1 status, including MCF-7 (BRCA1 +/+), MDA-MB-231 (BRCA1 +/-), HCC-1937 (BRCA1 -/-) and MDA-MB-436 (BRCA1 -/-). The combination index (CI) of each different drug-drug combination was calculated by applying the Chou and Talalay method. Different degrees of synergism (CI < 1) were recorded when PM1183 was combined with AZD-2281 (Olaparib) and BMN-673, depending on the tumor cell line and the conditions tested. The highest degree of synergism with Olaparib was observed in MDA-MB-436 cells followed by MDA-MB-231, MCF7 and HCC-1937 cells. The highest degree of synergism with BMN-673 was observed in MCF-7 cells followed by MDA-MB-231 and HCC-1937 cells. In MDA-MB-231 cells, both combinations led to a synergistic induction of γ-H2AX expression and PARP-1 cleavage, as analyzed by western blot. Collectively, our results indicate that the combination of PM1183 and PARP inhibitors (Olaparib and BMN673) induces an artificial synthetic lethality that can be advantageously used to kill several breast cancer cells, independently from their BRCA1 status. Citation Format: Gema Santamaría, Sonia Avila, Victoria Moneo, Carmen Cuevas, Luis F. García-Fernández, Carlos M. Galmarini. Synergistic combination of lurbinectedin and PARP inhibitors in breast cancer tumor cell lines. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2520. doi:10.1158/1538-7445.AM2015-2520

Accelerated synergism along a fault: A possible indicator for an impending major earthquake

It is generally accepted that crustal earthquakes are caused by sudden displacement along faults, which rely on two primary conditions. One is that the fault has a high degree of synergism, so that once the stress threshold is reached, fault segments can be connected rapidly to facilitate fast slip of longer fault sections. The other is sufficient strain accumulated at some portions of the fault which can overcome resistance to slip of the high-strength portions of the fault. Investigations to such processes would help explore how to detect short-term and impending precursors prior to earthquakes. A simulation study on instability of a straight fault is conducted in the laboratory. From curves of stress variations, the stress state of the specimen is recognized and the meta-instability stage is identified. By comparison of the observational information from the press machine and physical parameters of the fields on the sample, this work reveals differences of temporal-spatial evolution processes of fault stress in the stages of stress deviating from linearity and meta-instability. The results show that due to interaction between distinct portions of the fault, their independent activities turn gradually into a synergetic activity, and the degree of such synergism is an indicator for the stress state of the fault. This synergetic process of fault activity includes three stages: generation, expansion and increase amount of strain release patches, and connection between them.. The first stage begins when the stress curve deviates from linearity, different strain variations occur at every portions of the fault, resulting in isolated areas of stress release and strain accumulation. The second stage is associated with quasi-static instability of the early meta-instability when isolated strain release areas of the fault increase and stable expansion proceeds. And the third stage corresponds to the late meta-instability, i.e. quasi-dynamic instability as both the expansion of strain release areas and rise of strain level of strain accumulation areas are accelerated. The synergism is accelerated when the quasi-static expansion transforms into quasi-dynamic expansion, with interaction between fault segments as its mechanism. The essence of such transformation is that the expansion mechanism has changed, i.e. expansion of isolated fault segments is replaced by linkage of the interacting segments when the fault enters the critical state of a potential earthquake. Based on the experimental results, coupled with data on the temporal-spatial evolution of earthquakes along the Laohushan-Maomaoshan fault, west of the Haiyuan fault zone in northwestern China, the synergism process of this fault before the 6 June 2000 M6.2 earthquake is analyzed.

Abstract 1816: Blocking synthesis of oncogenic proteins in breast tumor cells through pharmacologic inhibition of the IRES

Abstract Many genes involved in the control of cell proliferation and survival, i.e. those most important to cancer biology, are now known to be regulated specifically at the translational (RNA to protein) level. An intriguing translation-regulatory element is the internal ribosomal entry site (IRES), which provides a mechanism by which oncogene-associated mRNAs can be translated independently of the global controls on protein synthesis. Tumor cells may rely on IRES-mediated translation of key oncogenic proteins to promote their own survival and proliferation under adverse conditions or exposure to cytotoxic chemotherapy. We have previously shown that the mRNA for type 1 insulin-like growth factor receptor (IGF1R) contains an IRES immediately upstream of the initiation codon. We have characterized the RNA-binding proteins that regulate the IRES, and elucidated the mechanism by which the IRES recruits the 40S ribosome. We have accumulated evidence that IGF1R IRES activity may be aberrantly upregulated in human breast cancer cells and tissues. Based on these observations, we hypothesized that IRES-mediated translation of key oncogenic proteins could be a rational target in cancer therapeutics with pharmacologic potential. Using T47D breast cancer cells genetically engineered to produce firefly luciferace under control of the IGF1R IRES, and a similar construct from which the IRES has been deleted as a control, a high throughput screen of 135000 compounds was performed. Four promising lead compounds with distinct chemical scaffolds have been identified and further validated in dose response titrations. Their specificity in blocking IRES-mediated translation was shown in reticulocyte lysates using a bicistronic construct containing the IGF1R IRES. The compounds inhibited de novo synthesis of IGF1R in T47D and SUM159 breast cancer cells with an IC50 on Western blots between <2 and 10 µg/mL. Consistent with the importance of IGF1R in tumor cell survival, the IRES inhibitors induced massive apoptosis in the same cell lines with a toxic concentration-90 in the range of <1 - 8 µg/mL. In contrast, normal human mammary epithelial cells tolerated prolonged exposure to IGF1R IRES inhibitors. A high degree of synergism (3+ to 4+) was observed when the IRES inhibitor was combined with a cytotoxic chemotherapeutic agent (doxorubicin), with combination indices of 0.153 - 0.334 and dose reduction indices for doxorubicin of 5.0-14.9. Furthermore, these compounds were shown to completely block the clonogenic survival and the formation of mammospheres in both ER positive and negative breast cancer cell lines, suggesting that cancer stem cells are sensitive to IRES inhibition. The pharmacologic inhibition of IRES-mediated translation is a novel strategy to modulate oncogene expression and could represent a new paradigm in cancer therapeutics. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1816. doi:1538-7445.AM2012-1816

Synergistic adsorption of Triton X-100 and CTAB surfactants at the toluene + water interface

Equilibrium interfacial tension measurements at 25.0 °C of the toluene + water system with two widely used surfactants, octylphenol decaethylene glycol ether (Triton X-100) and cetyl trimethyl ammonium bromide (CTAB) having concentrations much lower than their CMC were performed. According to the obtained parameters from the Szyszkowski equation, Triton has higher adsorption tendency than of CTAB. The results obtained for surfactants mixtures are analyzed by the theory of non-ideal interactions in binary mixtures (NIBMs) and the interfacial composition and the interaction parameter in the mixed adsorbed monolayer are determined. The attractive interaction shows a maximum value at nearly equal surfactants bulk mole fraction. The synergism is achieved for Triton bulk mole fractions of 0.30 and higher, and the highest degree of synergism (40.6%) is found for the bulk mole fraction of 0.52 with the lowest investigated constant interfacial tension of 28.0 mN m −1. A correlation was developed for variation of the interaction parameter with bulk mole fraction.

Trastuzumab and vinorelbine (TV) in early stages of breast cancer

e11556 Background: HER-2+ breast cancers (BCs) have a higher relapse rate, even for early stages of disease. The combination of trastuzumab and vinorelbine (TV) displays a high degree of synergism both preclinically and in metastatic BC patients that typically tolerate TV remarkably well. Occasionally, patients refuse standard treatment with chemotherapy and trastuzumab because of toxic side effects. In our institution, some of these patients accepted to receive treatment with TV. Methods: We retrospectively collected data on patients with stage I-III BCs, treated with TV as the only chemotherapy regimen. Most patients received TV on a weekly basis (one week off for V every 3–4 weeks) for ∼6 months, followed by 6 more months of T only. Results: Between May 2003 and June 2008, 23 patients were started on weekly TV. Median age was 66. Five patients received TV as preoperative treatment for BCs with the following clinical stages: IIB (1); IIIA (1); IIIB (3). The other 18 patients were pathologically staged as: stage I (11); IIA (5); IIIB (2). All cancers were HER2+; 65% of patients also received hormonal treatment for ER/PR+ disease. 3 patients had been previously treated for BC, and received TV as “adjuvant” treatment after a local relapse. Only one of these patients had previously received chemotherapy, while none had received prior T. No pathological complete response was found at surgery after preoperative TV. TV was very well tolerated, with one patient developing febrile neutropenia, 4 patients grade 3–4 uncomplicated neutropenia, and no other grade 3–4 events. During therapy, 5 patients had an asymptomatic 10–20% drop in the LVEF (Grade 1). Follow-up MUGA scans at 1 year after TV so far failed to show any significant abnormality. At an average follow-up of 26 months, one patient died for non-BC related causes, and all the other 22 patients are disease-free. Conclusions: TV is safe and very well tolerated, with no significant cardiac toxicity in our patient population. Preliminary follow-up data suggest that TV may be an acceptable alternative (neo-)adjuvant therapy for patients refusing “toxic” chemotherapy. Further studies prospectively testing TV as adjuvant treatment should be considered for patients with stage 1A BC, for which no standard treatment is clearly defined. No significant financial relationships to disclose.

Catalytic Synergism in Physical Mixtures of Supported Iron−Cerium and Supported Noble Metal for Hydroisomerization of 1,3-Butadiene

In an effort to better understand the excellent synergism of certain physical mixtures of supported catalysts for the 1-butene double bond shift described previously, the butadiene hydroisomerization selectivity/activity of the same mixtures was studied. Limited activity/selectivity synergism for the hydroisomerization of butadiene was measured for physical mixtures of FeCe/Grafoil (alloy) and Pt/Grafoil or Pd/Grafoil (noble metal). A high degree of synergism was noted for mixtures in which the ratio of alloy to noble metal was less than about 20:1. Additions of alloy to mixtures which increased the ratio to values higher than 20 did not change the activity or selectivity. For Pd/Grafoil containing mixtures both activity and selectivity synergism could be explained completely on the basis of a hydrogen spillover model. For example, the limit in activity synergism was ascribed to the depletion of hydrogen atoms away from palladium centers (radial gradient around each noble metal particle) due to the use of...

Effect of fucoidan during activation of human plasminogen

Fucoidan [sulfated poly (L-fucopyranose)] was compared with 6-aminohexanoic acid (6-AH) or CNBr-cleaved fibrinogen (CNBr-Fbg) alone or in combination in enhancing the activation of glutamic plasminogen (Glu-Plg) or lysine plasminogen (Lys-Plg) by two-chain tissue plasminogen activator (t-PA) or LMwt-urokinase or by streptokinase. Fucoidan enhanced the t-PA activation of Glu-Plg or Lys-Plg at Plg concentrations greater than 75nM, while stimulation by CNBr-Fbg of t-PA activation followed saturation kinetics of Michaelis-Menton. During t-PA activation of Glu-Plg, a high degree of synergism was observed between 6-AH and fucoidan while the enhancement by CNBr-Fbg was not influenced by fucoidan and was reversed by 6-AH. Fucoidan alone at higher concentrations was effective in enhancing the activation of Glu-Plg by urokinase while the combination of fucoidan and 6-AH showed additive effect in enhancing the activation of Lys-Plg. The activation of Glu-Plg by streptokinase was reversed by fucoidan in a manner similar to that reported for 6-AH. The results are interpreted to suggest that CNBr-Fbg and 6-AH compete with each other for the same lysine binding sites (LBS) on the Plg molecule while fucoidan acted synergistically with 6-AH in enhancing the t-PA activation of Glu-Plg by a different mechanism. The double reciprocal plot for the interaction of Glu-Plg and urokinase also showed a significantly higher affinity between the two in presence of fucoidan.

Synergism between the antifungal agents amphotericin B and alkyl glycerol ethers

The alkyl glycerol ether rac-1-O-dodecylglycerol inhibited the growth of members of two genera of yeasts, Candida and Cryptococcus, and was strongly synergistic with amphotericin B. At one-half its MIC, dodecylglycerol decreased the MIC of amphotericin B by as much as 80-fold. This high degree of synergism between dodecylglycerol and amphotericin B was demonstrated against a number of species of yeasts including Candida albicans, Candida tropicalis, Candida parapsilosis, Cryptococcus neoformans, Cryptococcus albidus, and Cryptococcus laurentii. All fractional inhibitory concentrations (for all strains and species) were calculated to be less than 1, and most were less than 0.6, again demonstrating strong synergism. Other alkyl glycerol ethers with alkyl chain lengths ranging from 8 to 18 carbon atoms were also found to be synergistic with amphotericin B against C. neoformans and C. albicans. Electron microscopy experiments showed that C. neoformans grown in the presence of dodecylglycerol had severely abnormal, deformed capsules. Although the mechanism of action of dodecylglycerol is not known, dodecylglycerol was not simply acting as a detergent. The natural detergent sodium deoxycholate could not substitute for dodecylglycerol. At comparable and higher concentrations, sodium deoxycholate had no fungicidal effect on its own, nor did it potentiate the activity of amphotericin B. Dodecylglycerol did not interact synergistically with the water-soluble antifungal agent fluconazole. The lipid-soluble hydrophobic properties of amphotericin B appear to be important for this synergistic effect, in that alkyl glycerol ethers could promote synergism with amphotericin B by potentially increasing the interaction between membrane-bound ergosterol and amphotericin B.

Antileukemic activity studies and cellular pharmacology of the analogues of 2-hydroxy-1H-isoindole-1,3-dione (HISD) alone and in combination with cytosine arabinoside (ara-C) against human leukemia cells CEM/0.

The 2-hydroxy-1H-isoindole-1,3-dione (HISD) derivatives are inhibitors of ribonucleotide reductase (RR). Among the seven new compounds of the PL series, three were found to be active in cell lines sensitive (CEM/0) or resistant to ara-C (CEM/ara-C/7A; CEM/dCk[-]. The compounds PL4, PL7 and PL8 exhibited IC50 values in micromolar range against the three CEM cell lines. The 3 compounds showed 100- to 1000-fold higher cytotoxicity compared to HU against all three CEM cell lines. Combination of each of the three active PL compounds with ara-C showed high degree of synergism in comparison to the combinations of HU with ara-C against both CEM/0 and CEM/ara-C/7A cell lines. The DNA synthetic capacity of CEM/0 cells treated with 10 microM PL4 for 24 or 48 h was inhibited > or = 99.8% simultaneously the cellular NTP pools were severely depleted. Treatment of CEM/0 cells with 10 microM PL4 showed steady depletion in all the dNTP pools at 1 or 2 h and complete depletion by 4 h. The enzyme activity did not recover up to 48 h in presence of the drug suggesting irreversible inhibition of RR.

Synergism between Clostridium thermocellum cellulases cloned in Escherichia coli.

We have obtained a synergistic effect during degradation of Avicel and filter paper by Clostridium thermocellum cellulases (two endoglucanases and one cellobiohydrolase) cloned in Escherichia coli. The highest degree of synergism was found at early stages of reaction, during the first 20 h: 2.5 and 2.9 on Avicel and filter paper, respectively. During combined action of all three cellulases the main product is cellobiose.

Different activation domains of Sp1 govern formation of multimers and mediate transcriptional synergism.

The process of transcriptional activation in eukaryotes by site-specific DNA-binding proteins is a key step in gene regulation. Here we have examined the properties of four distinct activator domains of the human transcription factor Sp1. In vivo transient cotransfection assays with Sp1 show that templates bearing multiple Sp1 sites activate transcription with a high degree of synergism. However, there is no evidence of cooperative binding of Sp1 to adjacent sites. Using deletion mutants of Sp1 we have determined that the glutamine-rich activation domains A and B and the previously uncharacterized carboxy-terminal domain D are all required for Sp1 to activate transcription synergistically. Gel-shift, DNase footprinting, and chemical cross-linking experiments reveal a strong correlation between the ability of Sp1 mutants to form homomultimeric complexes and their ability to activate transcription synergistically when bound to multiple sites. We have also examined the process of superactivation, in which a molecule of Sp1 tethered to DNA via its zinc fingers can be transcriptionally enhanced by interacting directly with fingerless Sp1 molecules. The domains involved in superactivation appear to be a subset of those necessary to achieve synergistic activation. These findings suggest that different domains of Sp1 carry out distinct functions and that the formation of multimeric complexes may direct synergism and superactivation.

Clinical efficacy of a synergistic combination of cefotaxime and amikacin against multiresistant Pseudomonas and Serratia infections

The synergistic activity of cefotaxime and amikacin against 21 highly resistant Pseudomonas aeruginosa and Serratia marcescens isolates was evaluated in-vitro by the checkerboard tube dilution method and in-vivo in five patients with serious infections caused by these organisms. All isolates were resistant to gentamicin, tobramycin, amikacin, and cefotaxime. Synergy was observed in 90% of isolates and occurred when the MIC of amikacin was less than 256 mg/l and of cefotaxime less than 1024 mg/l. A clinical response occurred in 100% and bacteriological cure in 80% of patients. Our results demonstrate a high degree of synergism between amikacin and cefotaxime in-vitro and clinical efficacy in the treatment of infections due to multiply-resistant Pseudomonas and Serratia species.