Synergy Coefficient Research Articles

Microwave irradiation activated persulfate and hydrogen peroxide for the treatment of mature landfill leachate effluent from a membrane bioreactor

This study systematically compared the effect of microwave irradiation activated persulfate (MW/PS) and hydrogen peroxide (MW/H2O2) on organic matter removal in mature landfill leachate effluent from a membrane bioreactor (MBR), the reactive oxygen species (ROS) produced, and the degradation and transformation mechanisms of organic substances. An obvious synergistic effect was identified in the MW/PS and MW/H2O2 processes. The synergistic effect of the MW and PS (synergy coefficient F = 23.67) was significantly higher than that of the MW and H2O2 (synergy coefficient F = 11.43). The MW/PS process had a stronger degradation effect on organic substances than the MW/ H2O2 process, even when the PS dosage was significantly lower than the H2O2 dosage. The removal rate and kobs (initial reaction rate) of the two processes could be raised by increasing the oxidant dosage. Increasing the MW output power improved the removal rate of organics by the two processes, but an excessive MW output power only improved the kobs of the two processes, while the effect on organic matter removal was not significant. The MW/H2O2 process only removed organics expeditiously under acidic conditions, while the MW/PS process resulted in a strong degradation of organic matter under a wide range of pH values. A quenching experiment using alcohols indicated that both SO4•- and •OH existed in the MW/PS process, with SO4•- playing a leading role in the degradation of organics, while •OH played a leading role in the MW/H2O2 process. A spectrum analysis based on the quenching experiment demonstrated that the two processes destroyed the characteristic structure of humus and refractory organics in the leachate (i.e., humic acid-like (HAL) and fulvic acid-like (FAL) substances), mainly through the free radical oxidation mechanism. The SO4•- degraded HAL substances efficiently, while •OH had a better degradation effect on FAL substances. Therefore, the MW/PS process had a stronger degradation effect on organics than the MW/H2O2 process due to the enhancement of ROS production. The final energy consumption analysis revealed that the energy conserved by the MW/PS process was about 614.2 kWh per unit volume of effluent compared to the MW/H2O2 process. This study provides a useful reference for the use of the MW/PS and MW/H2O2 processes to eliminate refractory organics in leachate effluent from an MBR.

A Reinforcement Learning Method Using Multifunctional Principal Component Analysis for Human-Like Grasping

Postural synergies allow a rich set of hand configurations to be represented in lower dimension space compared to the original joint space. In our previous works, we have shown that this can be extended to trajectories thanks to the multivariate functional principal component analysis, obtaining a set of basis functions able to represent grasping movements learned from human demonstration. In this article, we introduce a human cognition-inspired approach for generalizing and improving robot grasping skills in this motion synergies subspace. The use of a reinforcement learning (RL) algorithm allows the robot to explore the surrounding space and improve its capability to reach and grasp objects. The learning method is the policy improvement with path integrals, running in the policy space. Bootstrapped with synergy coefficients obtained from neural networks, the policy reward is based on a force closure grasp quality index computed at the end of the task, measuring how a firm is the grip. We finally show that combining neural networks and RL allows the robot manipulator to have a good initial estimate of the grasping configuration and faster convergence to an optimal grasp with respect to a database approach, the latter a less general solution in presence of new objects.

Targeting MDM2 and XIAP By Idasanutlin in Diffuse Large B-Cell Lymphoma

Purpose: The combination of rituximab and chemotherapy has improved overall survival of diffuse large B-cell lymphoma (DLBCL) patients in recent decades. Relapsed/refractory DLBCL patients previously treated with rituximab-containing regimen had significantly poorer response to salvage therapy. To study the mechanisms of this resistance, our laboratory developed several rituximab resistant cell lines. Previous study from our group found demonstrated an aberrant imbalance in the levels of pro- and anti-apoptotic proteins in these cell lines, including Bak/Bak, Mcl-1/BCLxL/Survivin and inhibitor of apoptosis (IAP) family proteins resulting in acquired chemotherapy resistance. MDM2 is an E3 ligase which regulates the degradation of multiple cellular protein targets such as pRB, HIF-1, p73, NF-kB, and E2F1 as well as FOXO3a. It is also the major negative regulator of p53. Recently, we found that MX69 (a MDM2 inhibitor), which blocks the MDM2 protein-XIAP RNA interaction, led to both MDM2 and XIAP degradation, and induced apoptosis-dependent cell killing activity. Moreover, MX69 re-sensitized resistant DLBCL cell lines to chemotherapy agents or small inhibitors (i.e. Venetoclax) in vitro. However, MX69 cannot be used clinically stressing the need to use a clinically available MDM2 inhibitor. To this end, we evaluated Idasanutlin, an investigational Nutlin family MDM2 antagonist, in B-cell lymphoma pre-clinical models. Methods: We used a panel of different DLBCL subtypes cell lines including activated B-cell lymphoma cell lines (TMD8, U2932); germinal center B-cell lymphoma (DOHH2, VAL, OCILY2, DH4, DH6, ROS50); rituximab-sensitive cell lines (Raji and RL); and rituximab/-resistant cell lines (Raji 4RH and RL 4RH). Cells were exposed to escalating doses of Idasanutlin (1nM-100µM) without or with other chemotherapeutic agents for 24, 48 and 72 hrs. Differences in cell viability were evaluated utilizing PrestoBlue. IC50 was calculated by GraphPad and Coefficient of synergy was calculated using CalcuSyn. Low mitochondrial potential was detected by staining cell with DiOC6 10ng/ml for 30 minutes and followed by flow cytometric analysis. Western blot was used to detect the changes of MDM2, XIAP and PUMA expression before and after exposure to Idasanutlin 1uM for 24h. Results:In vitro exposure Idasanutlin to DLBCL cell lines demonstrated a dose- and time-dependent cell death. IC50 dosage of the cells was ranged from 0.7uM to 63.07uM at 48h. Low dose of Idasanutlin (1uM) was able to disrupt mitochondria and induced low mitochondrial membrane potential in both ABC and GCB cell lines. At molecular level, Idasanutlin reduced expression level of MDM2 in TMD8, U2932, VAL, OCILy2 DHL4 and ROS50 cell lines. XIAP was reduced in Val and DHL4. Meanwhile, PUMA, the downstream of p53 activation, was increased after Idasanutlin exposure. Idasanutlin enhanced the anti-tumor activity of proteasome inhibitors (carfilzomib, ixazomib), Bcl-2 inhibitor venetoclax and Bryton kinase inhibitor ibrutinib. Conclusion: Idasanutlin showed potent anti-tumor activity as a single agent in DLBCL pre-clinical setting. Idasanutlin was able to decrease cellular mitochondrial membrane potential. At molecular level, Idasanutlin decreased MDM2 and XIAP protein level and induced PUMA expression. Moreover, Idasanutlin enhanced anti-tumor activities of other small inhibitors. Taking together, Idasanutlin could be used as a novel and promising drug in the clinical setting of DLBCL with relapsed/refractory disease. Disclosures No relevant conflicts of interest to declare.

Evolutionary Game Analysis on Knowledge-Sharing Behavior in the Construction Supply Chain

With the increasingly fierce global competition, supply chain members have to collaborate to respond to constant changes. Efficient knowledge sharing is the basis for the collaborative operation of the supply chain. Combined with evolutionary game theory, this paper studies the evolution path and stable strategies of knowledge-sharing behavior between construction supply chain enterprises, analyzing the factors that influence the establishment of a knowledge-sharing alliance. A numerical simulation is conducted to verify theoretical results and the effects of parameter adjustments on behavioral evolution. The results indicate that under different income relationships, knowledge-sharing behavior in construction supply chains presents different evolutionary trajectories. In addition, the probability of accepting the sharing strategy is positively correlated with the penalty coefficient, incentive coefficient, trust level, and synergy coefficient and negatively correlated with cost. This study provides a new perspective and theoretical guidance for establishing stable knowledge collaboration between enterprises and promoting the sustainable development of the construction supply chain.

Abstract 2501: Combining BH3-mimetics to target both BCL-2 and MCL1 has potent activity in diffuse large B-cell lymphoma (DLBCL) pre-clinical models

Abstract Background: Upregulation of the anti-apoptotic protein Mcl-1 has been implicated in chemotherapy resistance and poor clinical outcomes in activated B-cell like (ABC)-DLBCL. We hypothesized that singular inhibition of Mcl-1 by AMG-176 and/or dual inhibition of MCL-1 and Bcl-2 by combining AMG-176 with venetoclax could be an effective strategy for inducing cell death in DLBCL pre-clinical models. Materials and Methods: Using a panel of DLBCL cell lines, we conducted pre-clinical targeting of Mcl-1 with AMG-176. Cells were exposed to AMG-176 as a single agent (0-20uM) over 24, 48, 72 hrs and IC50 concentrations were calculated for each cell line. Subsequently, DOHH-2, OCI-LY1, DHL4, DHL6, TMD8, and VAL cells were exposed to AMG-176 (0-20 uM) in combination with Doxorubicin (0-1uM), Cisplatin (0-10uM), Cytarabine (0-10uM), Vincristine (0-10uM), Ixazomib (0-100nM), Carfilzomib (0-12.5nM), and Venetoclax (0-10uM) for 72 hours. Cell viability was determined by Cell Titerglo. Coefficient of synergy was calculated using CalcuSyn. In addition, induction of apoptosis was evaluated by flow cytometry (Annexin V/PI staining) as well as changes in cell cycle (PI staining). Bcl-2 family member protein expression was determined by Western blotting. Primary tumor cells isolated from a patient with a composite lymphoma: angioimmunoblastic T-cell lymphoma (AITL) and EBV-driven DLBCL were exposed ex vivo to AMG-176 +/- venetoclax. By immune-phenotype, the primary tumor cells isolated and expanded were ABC-DLBCL. Results: In vitro, AMG-176 single agent exposure induced cell death in a dose- and time-dependent manner in all DLBCL cell lines tested. AMG-176 IC50 values varied from 0.03 to 11.42 uM for OCI-LY1 and ROS-50 respectively. Western blotting studies confirmed changes in Bcl-2 family members, specifically in Bok, Bcl-XL, Mcl-1 and Bcl-2 following in vitro exposure to AMG-176. The combination of AMG-176 with Doxorubicin, Cisplatin, Cytarabine, Vincristine, Ixazomib, Carfilzomib, or Venetoclax resulted in significant synergistic activity for each combination. The strongest coefficient of synergy was observed when AMG-176 was combined with venetoclax. Similar synergy results were noted in primary tumor cells isolated from the lymphoma patient. Annexin/PI staining with AMG-176 demonstrated caspase dependent apoptosis. No significant changes in cell cycle were observed with AMG-176 exposure. Conclusion: Our data suggests that AMG-176 exhibits strong synergistic activity with various chemotherapy agents and small molecule inhibitors especially venetoclax. In vivo experiments are planned. Combination therapy targeting Mcl-1 and Bcl-2 may provide an alternative therapeutic approach in DLBCL. Citation Format: Cory Mavis, Juan Gu, Matthew Barth, Sumera Khan, Pallawi Torka, Francisco Hernandez-Ilizaliturri. Combining BH3-mimetics to target both BCL-2 and MCL1 has potent activity in diffuse large B-cell lymphoma (DLBCL) pre-clinical models [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 2501.

Abstract 723: MX69 induces apoptosis by inhibiting XIAP in both rituximab sensitive and resistant lymphomas

Abstract Background: X-linked inhibitor of apoptosis protein (XIAP) has been implicated in development of resistance to chemotherapy via its direct inhibition of caspases 3,7 and 9. It is overexpressed in several lymphoma cell lines including rituximab-resistant cell lines (RRCL), Raji4RH and RL4RH, produced by our group. We have previously demonstrated that XIAP is critical for chemotherapy-resistance and survival in RRCL by knocking down XIAP using siRNA interference which demonstrated that XIAP is critical for chemotherapy-sensitivity and survival in RRCL. MDM2 inhibition activates p53 transcription and inhibits translation of XIAP, thus promoting apoptosis of cancer cells. Here, we targeted XIAP at a translational level by inhibiting it with MX69, a dual inhibitor of MDM2 and XIAP. Materials and Methods: Cell lines representing Burkitt’s lymphoma (BL), activated B-cell like diffuse large B-cell lymphoma (ABC-DLBCL, germinal center B-cell like DLBCL (GCB-DLBCL), mantle cell lymphoma (MCL) and B-cell lymphoblastic leukemia were exposed to MX69 as a single agent (0-80 uM) over 24, 48 and 72 hrs and IC50 concentrations were calculated. Subsequently, Raji, Raji4RH, RL, RL4RH, HBL-2, TMD-8, Daudi and Reh cell lines were exposed to MX69 (0-80 uM), in combination with doxorubicin (0-1 uM), cytarabine (0-50 uM), vincristine (0-10 nM), etoposide (0-50 uM), carboplatin (0-10 uM), ixazomib (0-1.5 uM), ibrutinib (0-10 uM) and venetoclax (0-10 uM) for 48 hours. Cell viability was determined by Cell Titer-Glo. Coefficient of synergy was calculated using CalcuSyn. Induction of apoptosis was evaluated by flow cytometry (Annexin V/PI stain). MDM2, p53, XIAP and PARP protein expression was determined by Western blotting. Results: MX69 induced cell death in a dose- and time-dependent manner in all cell lines. Annexin/PI staining showed caspase-dependent apoptosis with MX69 in all cell lines. Western blotting confirmed significant inhibition of MDM2, XIAP and changes in p53 and PARP following exposure to MX69. MX69 demonstrated significant synergistic activity when combined with doxorubicin, ixazomib, ibrutinib or venetoclax; synergy was strongest for the MX69- venetoclax combination. Conclusion: Our data suggests that in vitro exposure to MX69 resulted in anti-tumor activity in a wide variety of B-cell lymphoma cells lines (including BL, DLBCL, MCL). Perhaps related to its anti-tumor effects, MX69 inhibited XIAP levels. These findings are similar to prior siRNA XIAP knockdown experiments. Strong synergistic activity was observed when XIAP was combined with various chemotherapy agents and small molecules inhibitors (such as venetoclax, ixazomib or ibrutinib). Ex vivo experiments using primary tumor cells isolated from lymphoma patients and lymphoma mouse models have been planned. Targeting MDM2 and XIAP can be an attractive therapeutic strategy in patients with rituximab-sensitive or -resistant B-cell lymphoma. Citation Format: Sumera Khan, Cory Mavis, Ahmad Hanif, Juan Gu, Pallawi Torka, Francisco Hernandez-Ilizaliturri. MX69 induces apoptosis by inhibiting XIAP in both rituximab sensitive and resistant lymphomas [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 723.

Research on Regional Sustainable Development Based on Evaluation of Tourism Competitiveness of 21 Cities (States) in Sichuan Province

In accordance with the principles of systemic, hierarchical, and operability, this paper divides into tourism status competitiveness, tourism environmental competitiveness and tourism potential competitiveness to construct an evaluation index system for urban tourism competitiveness. Taking 21 cities in Sichuan Province as the research objects, the tourism competitiveness of each city was evaluated by factor analysis and grey correlation analysis. The evaluation results of these two methods were further evaluated by Kendall synergy coefficient test method. The consistency test and the combination evaluation were conducted, and the tourism competitiveness rankings of each city in Sichuan Province were given. Then, the system cluster analysis classified 21 cities into five levels, according to the evaluation results. Finally, some suggestions on the regional sustainable development in Sichuan Province were listed.

GSK458 Is a Novel Dual PI3K/mTOR Inhibitor with Preclinical Antitumor Activity in T Cell Lymphomas As a Single Agent and in Combination Therapy

Introduction: T-cell lymphoma (TCL) accounts approximately 15% of all Non-Hodgkin's lymphoma cases. TCL are often divided into either systemic or cutaneous TCL (CTCL). The management of TCL systemic or cutaneous is challenging because the emergence of chemotherapy resistance that lead to early death (systemic T-cell lymphomas) or chronic debilitating clinical course (CTCL). To improve the clinical outcomes and reduce treatment-related toxicity, research need to be done focusing on understanding and targeting the molecular mechanism driving TCL subtypes. Recently, studies showed that the PI3K AKT/mTOR pathway was activated in TCL. GSK458 is a potent oral dual inhibitor of pan PI3K (α, β, γ and δ) and mTOR (mTOR1 and mTOR2). Preclinical studies in B-cell lymphomas showed GSK458 had broad antitumor activity in vitro and in vivo. In 2016, Phase I clinical trial of GSK458 was competed at the maximal dosage of 2.5mg twice daily. However, the effects of GSK458 on T-cell malignancies remain utterly unknown. Here, we evaluated the activity of GSK458 in preclinical T cell lymphoma models. Methods: We used a panel of T-cell lymphoma cell lines representing PTCL (J45), T-cell lymphoblastic lymphoma (SupT-1), and Mycosis Fungoides (MF)(MJ, HH and H9). TCL cell lines were exposed to escalating doses of GSK458 (1nM-100µM) without or with chemotherapeutic agents (doxorubicin, cisplatin, carboplatin, and dexamethasone); Bcl-2 inhibitor (Venetoclax); proteasome inhibitors ( bortezomib, carfilzomib, Ixazomib); or HDAC inhibitors (SAHA) for 48 and 72 hrs. Differences in cell viability, ATP levels, low mitochondria potential, glucose update, apoptosis and cell cycle distribution were evaluated utilizing PrestoBlue, Cell-Titer Glo assays, DiOC6, 2-NDG, Annexin V and propidium iodide staining followed by flow cytometric analysis, respectively. IC50 was calculated by GraphPad. PI3K and mTOR downstream pathway phosphorylation status, such as p-AKT Ser473, p-AKT Thr308, p-mTOR and p-GSK3β were detected by internally staining of FITC conjugated-antibodies followed by flow cytometry. Apoptosis proteins (MCL-1, PARP, p53, XIAP etc.) were detected by western blot. The additive/synergistic activity of GSK458 was detected by presto blue assay and Coefficient of synergy was calculated using CalcuSyn. Results:In vitro exposure of TCL cell lines to GSK458 demonstrated a dose- and time-dependent cell death. The IC50 of the cells were ranged from 3nM to 1.05uM at 72 hours. At 72h, GSK458 10nM lowered cellular mitochondrial potential, ATP levels and glucose uptake. GSK458 induced apoptosis and arrested the cell cycle at G1. At molecular level, GSK458 reduced phosphorylation status of AKT ser473 and Thr308, mTOR and GSK3β. Interestingly, GSK458 inhibited Mcl-1 expression level. GSK458 exhibited synergistic activity when combined with doxorubicin and dexamethasone. To a lesser degree, GSK458 enhanced the anti-tumor activity of Venetoclax, proteasome and HDAc inhibitors. Conclusion: GSK458 is active as a single agent or in combination with chemotherapy agents or small molecule inhibitors in a variety of T-cell pre-clinical models representing forms of systemic or cutaneous T-cell lymphoma. GSK458 was able to inhibit phosphorylation of AKT, mTOR and GSK3β, which may be the mechanism to reduce ATP production and glucose uptake in the cancer cells. Moreover, GSK458 arrested cell cycle at G1 arrest. Our data supports the clinical evaluation of GSK458 in relapsed/refractory T-cell lymphoma patients. (Supported by Roswell Park Cancer Institute Alliance Foundation Grant) DisclosuresNo relevant conflicts of interest to declare.

Targeting XIAP Via Degradation of MDM-2 By MX69 in Aggressive Lymphomas

Background: The addition of Rituximab to front-line therapy has improved clinical outcomes in diffuse large B-cell lymphoma (DLBCL), but it has also altered the biology of relapsed/refractory disease. To better understand the mechanisms responsible for Rituximab associated chemotherapy cross-resistance our group developed and characterized several Rituximab resistance cell lines (RRCL). We previously demonstrated using SiRNA interference, that X-linked inhibitor of apoptosis (XIAP) is critical for chemotherapy sensitivity and survival in RRCL. MX69, a dual inhibitor of Mdm2 and XIAP that indirectly downregulates XIAP, is undergoing pre-clinical testing. MX69 affects XIAP levels by its effects on the ubiquitination and degradation of endogenous MDM-2, resulting in decrease XIAP translation and activation of caspase 3, 7 and 9 as well as PARP cleavage leading to apoptosis of cancer cells. In our current work, we pharmacologically inhibited XIAP in lymphoma pre-clinical models using MX69.Materials and Methods: A panel of Burkitt's Lymphoma (BL, including RRCL), germinal center B-cell (GCB)-DLBCL (including RRCL), activated B-cell (ABC)-DLBCL, Mantle cell Lymphoma (MCL) and Pre-B cell Leukemia cell lines were exposed to MX69 as a single agent (0-80uM) over 24, 48, 72 hrs and IC50 concentrations were calculated for each cell line. Changes in Mdm2, p53, XIAP and PARP expressions were determined following MX69 exposure (at IC50 doses) for 24 hrs. Induction of apoptosis was evaluated by Annexin V/propidium iodine staining. Subsequently, cell lines were exposed to MX69 (0-80 uM), in combination with Doxorubicin (0-1uM), Cytarabine(0-50uM), Vincristine (0-10nM), Etoposide(0-50uM), Carboplatin (0-20uM), Ixazomib (0-1.5uM), Ibrutinib (0-20uM) and Venetoclax (0-10uM) for 48 hours. Cell viability was determined by Cell Titerglo. Coefficient of synergy was calculated using CalcuSyn.Results: In vitro, MX69 single agent exposure induced cell death in a dose and time-dependent manner in all cell lines tested. Western blotting studies confirmed downregulation of Mdm2, XIAP and changes in P53 and PARP, following in vitro exposure to MX69. Induction of apoptosis was observed by flow cytometry in all cell lines tested. The combination of MX69 with Doxorubicin, Cytarabine, Vincristine, Ixazomib, Carboplatin, Etoposide, Ibrutinib, and Venetoclax resulted in significant synergistic activity. The strongest CI of synergy was observed when cell lines were exposed to MX69 and Venetoclax, Ixazomib, Etoposide or Ibrutinib.Conclusion: Our data suggests that in vitro exposure of a wide variety of B-cell lymphoma cell lines (including BL, DLBCL, MCL or RRCL) to MX69 resulted in anti-tumor activity. Perhaps related to its anti-tumor effects, MX69 inhibited XIAP levels. These findings are similar to prior SiRNA XIAP knockdown experiments. Strong synergistic activity was observed when XIAP was combined with various chemotherapy agents and small molecules inhibitors (such as Venetoclax, ixazomib or ibrutinib). Ex vivo experiments using primary tumor cells isolated from lymphoma patients and lymphoma mouse models are been planned. Targeting Mdm2 and XIAP can be an attractive therapeutic strategy in patients with Rituximab-sensitive or -resistant B-cell lymphoma. DisclosuresNo relevant conflicts of interest to declare.

Selectively Targeting Checkpoint Kinase By Prexasertib Had Potent Anti-Tumor Activity As Single Agent or Combined with Other Chemotherapeutic Drugs in Diffuse Large B-Cell Lymphoma, Especially Double Hit Lymphoma

Background: Relapsed/Refractory diffuse large B-cell lymphoma (DLBCL) patients had a poor prognosis, especially those relapsing within 12 months of completion therapy. Clinically, there is a need to identify and target pathways associated with acquired treatment resistance. Using rituximab-chemotherapy resistant DLBCL cell lines generated in our laboratory, we demonstrated the acquirement of rituximab resistance is associated with changes in the cell cycle distribution (S phase arrest) and expression of cell cycle regulatory proteins (cyclin B, CDC2, CDK7, Wee1, checkpoint kinase 1 [CHK1] and checkpoint kinase 2 [CHK2]). CHK1 and 2 are serine/threonine kinases that control: 1) G2/M phase transition in response of DNA damage, 2) stabilization of DNA replication fork and 3) coordinate the mitosis. Inhibition of CHKs is an attractive strategy in cancer medicine. CHK1 inhibitors are been studied in phase I clinical trials in solid tumor malignancies (i.e. breast, pancreas or lung cancer). We hypothesized that CHK inhibition by Prexasertib has effective anti-tumor activity in DLBCL pre-clinical models. Methods: We used a panel of DLBCL cell lines including rituximab sensitive (Raji, RL, DHL4, DHL6, TMD8, RIVA, and U2932), rituximab resistant (Raji 4RH and RL 4RH), and double hit DLBCL (DOH2, ROS50 and VAL). In addition, experiments were conducted in primary tumor cells isolated from lymphoma patients. Cells were exposed to prexasertib alone or in combination with chemotherapy agents. Cell viability were determined by Presto Blue assay. IC50 was calculated by Graphpad software and coefficient of synergy was calculated using the CalcuSyn software. Subsequently, cells were expose to prexasertib and changes in cell cycle distribution and apoptosis were determined by propidium iodide staining (Pi), annexing V/PI staining and by Caspase3/7 activity detection assay. Changes in mitochondrial potential were determined by DiOC6 staining. Induction of DNA double strain break was investigated by western blot probed with phosphory-H2A.X antibody. At molecular basis, downstream pathway activities ( Mcl-1, phosphor-GSK3β, phosphory-H3, and wee1) were determined by western blot. Results: Prexasertib exhibited a dose dependent anti-lymphoma activity in all all cell lines tested including DHL cell lines. The IC50 of the cells ranged from 11nM to 2.8uM at 72 hours. Additive/synergistic effects were observed by combining prexasertib and doxorubicin, etoposide, vincristine, gemcitabine and proteasome inhibitors. At the dosage of 10nM and 25nM, prexasertib induced cell cycle arrest at G2/M phase. It also induced low membrane potential, caspse3/7 activation and apoptosis in all DLBCL cell lines. Interestingly, western blot also showed CHK inhibitor at 10nM was able to reduce Mcl-1 and p-GSK 3beta. Conclusion: Our data suggests that targeting checkpoint kinase is a promising therapeutic strategy in the lymphoma, including in double hit lymphoma pre-clinical models. In vivo studies are planned. Our data supports the clinical development of prexasertib in DLBCL. (Supported by Roswell Park Cancer Institute Alliance Foundation Grant) DisclosuresNo relevant conflicts of interest to declare.

Effects of Proteasome Inhibition in the Anti-Tumor Activity of Venetoclax in Mantle Cell Lymphoma Pre-Clinical Models

Background: At the molecular level, mantle cell lymphoma (MCL) is characterized by the deregulation of Bcl-2 family members (Mcl-1, BIM) and cell cycle (cyclin D1) regulatory proteins. Perhaps related to this, the clinical outcome of MCL continues to be poor specially for those patients with disease progression after high dose chemotherapy and autologous stem cell rescue and/or BTK inhibitors, stressing the need to develop novel therapeutic strategies or optimize current available options. Venetoclax (V), a highly selective Bcl-2 inhibitor, has shown modest activity against relapsed/refractory MCL. Over-expression of Mcl-1 has been postulated to be a mechanism of resistance to V limiting its anti-tumor activity in subtypes of lymphoma including MCL. The lethality by proteasome inhibitors (PIs) has been associated with changes in the Bcl-2 family members (Bax, Noxa, Mcl-1 and Bcl-XL) in lymphoma pre-clinical models, making them ideal agents to combine with V. To this end, we studied the anti-tumor activity of combining PIs with V in MCL pre-clinical models. Materials and Methods: A panel cytarabine sensitive (Rec-1, Jeko, Granta, HBL-2, Z-138 and Mino) and resistant (araC) cell lines (Jeko araC, HBL-2 araC, and Mino araC) were exposed to V, Bortezomib (BTZ), carfilzomib (CFZ), or ixazomib (IXZ) for 24, 48 and 72 hours. Cell viability was calculated measuring the ATP content. IC50 drug concentrations were calculated for each agent. Subsequently, MCL cell lines were exposed to escalating doses of V (0.001uM-5uM) and CFZ (1.5625nM-50nM), BTZ (3.125nM-100nM) or IXZ (3.125nM-100nM). In addition, primary tumor cells isolated from B-cell lymphoma patients (N=21) including MCL patients were exposed to V +/- BTZ or CFZ for 48 hrs. Cell viability was determined by Cell Titerglo. Coefficient of synergy were calculated using CalcuSyn software program. Induction of apoptosis was detected by Annexin V/Propidium iodine staining and PARP cleavage. Changes in Bcl-2 and cell cycle regulatory proteins were evaluated by Western blotting in HBL-2 cells. For in vivo experiments, 6-8 weeks old severe combined immunodeficiency (SCID) mice were inoculated with 10x106 HBL-2 cells via tail vein injection (IV). Subsequently, SCID mice were treated with V (100mg/kg/dose via gastric lavage on days 3-7, 10-14 and 17-21) or IXZ (6mg/kg/dose IV days 3, 6, 10, 13, 17 and 20) or combination of both agents. A group of untreated animals was used as a control. Differences in survival were evaluated between treatment groups. Results: In vitro exposure of MCL cell lines to either V, BTZ, CFZ, and IXZ induced cell death in a dose- and time-dependent manner. Significant synergistic activity was observed by combining both V with CFZ or IXZ at known sub-therapeutic and therapeutic doses of individual agents measured by ATP content and apoptosis potential. Anti-tumor activity was observed in cytarabine sensitive and resistant cell lines. Similar findings were observed in primary tumor cells isolated from B-cell lymphoma patients. In vitro exposure of MCL cell lines with the lowest IC50 (HBL-2) to V and PIs (BTZ, CFZ, or IXZ) resulted in the upregulation of Noxa, BIM, Mcl-1 cleavage form (pro-apoptotic) and downregulation of Bcl-XL leading to PARP cleavage and apoptosis. In vivo treatment of MCL bearing SCID mice with V resulted in significant anti-tumor activity when compared to single agent IXZ treated or control animals. Of interest, MCL bearing SCID animals treated with V and IXZ exhibited a better disease control and the survival was longer than SCID animals treated with V or IXZ single agent (P<0.05). Conclusion: Our data suggests that V exhibits strong synergistic activity with PIs, especially with CFZ (in vitro) or IXZ (in vitro and in vivo). Together, our data supports the evaluation of V in combination with readily available novel PIs (IXZ or CFZ) in relapsed/refractory MCL. (Supported by LRF grant 555463, an NIH grant R01 CA136907-01A1 and a grant from The Roswell Park Cancer Institute Alliance Foundation) DisclosuresNo relevant conflicts of interest to declare.

Criteria evaluation for pricing decisions in strategic marketing management using an intuitionistic cognitive map approach

Marketing decisions are challenging in profit-oriented companies because of their complex nature. Many factors influence the marketing strategy in the new product development (NPD) process. With this aspect, strategic marketing decisions for launching a product to the market can be observed as multiple criteria decision-making (MCDM) problems. This study proposes a novel approach called intuitionistic cognitive map (ICM), for assessing the criteria that influence the pricing strategy of a company in earlier stages of the product’s life cycle in the market. A framework is formed based on a profound analysis of literature and experts’ opinions, in terms of criteria affecting pricing strategy and the causal relationships between them. Intuitionistic fuzzy sets and cognitive mapping are used together to capture the hesitation of the decision makers caused by lack of information and to define cause-and-effect relations between the criteria to represent the complexity of strategic marketing decisions. Contrary to conventional MCDM methods that require complete data, ICM method is able to deal with lack of information and hesitancy of the decision makers. In addition, ICM method has a new feature called “the coefficient of synergy” incorporating the synergy effect of the application field in the model. The case study is conducted in a technological-device-manufacturing company. Seventeen factors that influence the company’s pricing decisions are determined and evaluated with three marketing experts, and in the numerical application, brand image, market share, consumer fidelity, market/segment size, and new product capability criteria had the maximum positive influence on pricing decisions.

Neural synergies for controlling reach and grasp movement in macaques

It has been suggested that the brain adopts a simplified strategy to coordinate a large number of degrees of freedom in motor control. Synergies have been proposed as a strategy to produce movements by recruitment of a small number of fixed modular patterns. However, there is no direct support for a synergistic organization of the brain itself. In this study, we recorded neural activities from the dorsal premotor cortex (PMd) of monkeys trained to reach and grasp differently shaped objects (grasping task) or the same object in different positions (reaching task). Non-negative matrix factorization (NNMF) was applied to the neural data to extract neural synergies, whose functional roles were verified in several ways. We found that motor cortex used similar neural synergies for grasping different objects; combining only a few of the synergies accounted for most of the variance in the original data. When used for single-trial task decoding, the synergy coefficients performed as well and robustly as the original data in both tasks. The synergy amplitudes for each unit were significantly correlated with the corresponding neuron’s firing rate. In addition, we also observed synergies shared between tasks and task-specific synergies, as shown before for muscle synergies. Altogether, we demonstrated that neural synergies are effective in describing neural population activity during reach to grasp movements and provide a new tool for interpreting neural data for movement control.

Children With and Without Dystonia Share Common Muscle Synergies While Performing Writing Tasks.

Childhood dystonia is a movement disorder characterized by muscle overflow and variability. This is the first study that investigates upper limb muscle synergies in childhood dystonia with the twofold aim of deepening the understanding of neuromotor dysfunctions and paving the way to possible synergy-based myocontrol interfaces suitable for this neurological population. Nonnegative matrix factorization was applied to the activity of upper-limb muscles recorded during the execution of writing tasks in children with dystonia and age-matched controls. Despite children with dystonia presented compromised kinematics of the writing outcome, a strikingly similarity emerged in the number and structure of the synergy vectors extracted from children in the two groups. The analysis also revealed that the timing of activation of the synergy coefficients did not significantly differ, while the amplitude of the peaks presented a slight reduction. These results suggest that the synergy analysis has the ability of capturing the uncorrupted part of the electromyographic signal in dystonia. Such an ability supports a possible future use of muscle synergies in the design of myocontrol interfaces for children with dystonia.

Itraconazole, an Oral Antifungal Drug, Is Active in Chemotherapy Resistant B-Cell Non-Hodgkin Lymphoma and Enhances the Anti-Tumor Activity of Chemotherapy Agents

Background: Relapsed/refractory diffuse large B-cell lymphoma (DLBCL) patients previously treated with rituximab-based therapy have poor clinical outcome, according to the results from collaborative trial in relapsed aggressive lymphoma (CORAL) study. It stresses the need to identify and/or optimize novel targeted agents. To better understand the molecular mechanisms underlining the acquired resistance to rituximab, we generated and characterized several rituximab-resistant DLBCL cell lines (RRCLs). Itraconazole, an oral antifungal agent, was reported had novel anticancer activity in basal cell carcinoma, non-small cell lung cancer and prostate cancer. In our current work, we define and characterize the anticancer activity of itraconazole in preclinical rituximab-sensitive or -resistant lymphoma models.Methods: A panel of rituximab-sensitive (RSCL) and rituximab-resistant (RRCL) cell lines were exposed to escalating doses of itraconazole (0-20μM) for 24, 48 and 72h. Changes in cell viability and cell cycle distribution were evaluated using the Presto Blue assay and flow cytometry respectively. IC50 was calculated by Graphpad Prism6 software. Loss of mitochondrial membrane potential (Δψm) following itraconazole exposure was assessed by DiOC6 and flow cytometry. Subsequently lymphoma cells were exposed to itraconazole or vehicle and various chemotherapy agents such as doxorubicin (1µM), dexamethasone (1µM), cDDP (20μg/ml), bortezomib (20nM), carfilzomib (20nM) or MLN2238 (20nM) for 48 hours. Coefficient of synergy was calculated using the CalcuSyn software. Changes in hexokinase II (HKII), Voltage dependent anion channel protein (VDAC), LC3 and BCL-xL expression levels were determined by western blotting after exposure cells to itraconazole. VDAC-HKII interactions following in vitro exposure to itraconazole were determined by immunoprecipitation of VDAC and probing for HKII in RSCL and RRCLs.Result:Itraconazole consistently showed potent, specific, dose-and time- dependent inhibition of all our sensitive and resistant lymphoma cell lines. In vitro exposure cells to itraconazole resulted in a loss of mitochondrial membrane potential and caused G2 cell cycle arrest. Itraconazole significantly had a synergistic anti-tumor effect combined with various chemotherapeutic agents, including doxorubicin, dexamethasone, cisplatin and different generations of proteasome inhibitors (bortezomib, carfilzomib or ixazomib) in both RSCL and RRCL. Western blot and immunoprecipitation studies demonstrated that following exposure to itraconazole, HKII bound less to mitochondrial specific protein VDAC. Complete silencing of HKII (using HKII siRNA interference) resulted in a rescue of loss in the mitochondrial membrane potential induced by intraconazole.Conclusion: Taking together, our data suggest that itraconazole had a potent anti-tumor activity against rituximab-sensitive or resistant pre-clinical models. The disruption of HKII from mitochondria following itraconazole exposure may contribute to lower the mitochondrial membrane potential and enhance the chemotherapeutic efficacy. Our finding highlights itraconazole as a potential therapeutic agent in the treatment of B-cell malignancies, and strongly supports clinical translation of its use. DisclosuresNo relevant conflicts of interest to declare.

Integration of anodic and cathodic processes for the synergistic electrochemical production of peracetic acid

Efficient in-situ production of peracetic acid is an unreached milestone of electrochemical engineering. Previous attempts in the production of peracetic acid were focused either on the cathodic production of hydrogen peroxide and its further addition to acetic acid solutions or on the oxidation of a suitable raw material (v. g. acetic acid, acetaldehyde, ethanol). In the present work, the oxidation of acetic acid by a boron doped diamond (BDD) anode was integrated with the cathodic production of hydrogen peroxide using a carbon felt gas diffusion electrode. A marked synergistic effect (synergy coefficient of 192.0±13.1%) is observed when the oxidation of acetic acid by hydroxyl radicals is performed together with the cathodic production of hydrogen peroxide. A maximum PAA production efficiency of 19.87% was obtained, a value much higher than previous works based on the oxidation of acetic acid by BDD anodes and approximately double the optimal value reported in studies based on the production of hydrogen peroxide.

Synergistic integration of sonochemical and electrochemical disinfection with DSA anodes

This work focuses on the disinfection actual urban wastewater by the combination of ultrasound (US) irradiation and electrodisinfection with Dimensionally Stable Anodes (DSA). First, the inactivation of Escherichia coli (E. coli) during the sonochemical disinfection was studied at increasing ultrasound power. Results showed that it was not possible to achieve a complete disinfection, even at the highest US power (200 W) dosed by the experimental device used. Next, the electrodisinfection with DSA anodes at different current densities was studied, finding that it was necessary a minimum current density of 11.46 A m−2 to reach the complete disinfection. Finally, an integrated sonoelectrodisinfection process was studied. Results showed a synergistic effect when coupling US irradiation with DSA electrodisinfection, with a synergy coefficient higher than 200% of the disinfection rate attained for the highest US power applied. In this process, hypochlorite and chloramines were identified as the main reagents for the disinfection process (neither chlorate nor perchlorate were detected), and the presence of trihalomethanes was far below acceptable values. Confirming this synergistic effect with DSA anodes opens the door to novel efficient disinfection processes, limiting the occurrence of hazardous disinfection by-products.

Abstract 3038: Investigating novel targeted therapies for double hit diffuse large B-cell lymphoma (DH-DLBCL)

Abstract Background: Molecular studies divide DLBCL into three subtypes with distinct pathogenesis and clinical outcomes: activated B-cell (ABC), germinal center B-cell (GCB) and primary mediastinal lymphoma (PML). Florescence in situ hybridization (FISH) studies identified another subgroup of DLBCL, classified as DH-DLBCL, with a poor clinical outcome harboring concurrent gene rearrangements of the c-MYC, BCL2 and/or BCL6 proto-oncogenes, resulting in the over-expression of c-Myc, Bcl2 and Bcl6 proteins. Previously, our retrospective review from single institution series revealed that 30 out of 611 DLBCL patients had aberrations in c-MYC and BCL2 or BCL6 by FISH. These patients exhibited inferior response rates (RR) to rituximab-based chemotherapy, and a shorter progression-free survival (PFS)/overall survival (OS), suggesting that newer therapies are in dire need. DH-DLBCL is characterized by de-regulation of apoptosis and cell cycle progression, resulting in rapid cellular proliferation and resistance to apoptotic stimuli. In ABC-DLBCL, anti-apoptotic factor MCL-1 is implicated in poor prognosis leading to resistance to standard chemotherapy. C-MYC transcriptionally upregulates Mcl1. Translocation of c-MYC in DH-DLBCL may contribute to the aggressive phenotype and chemotherapy resistance via the MCL-1 pathway. We hypothesize that dual inhibition of both anti-apoptotic proteins BCL2 and MCL1 is an effective strategy in inducing lymphoma cell death in DH-DLBCL. Materials &amp; Methods: At the pre-clinical level, we studied 3 novel therapeutic agents targeting BCL2 (ABT-199), c-MYC (JQ-1), and various cell cycle regulatory proteins (p21) and other BCL2 family members affecting ABT-199 activity (irreversible proteasome inhibitor carfilzomib(CFZ)) using DH lymphoma (DHL) cell lines (Val, DOHH-2, ROS-50). DHL cell lines were exposed to ABT-199 (0-10 uM), JQ-1 (0-100 uM) and carfilzomib (CFZ) (0-50 nM) at 24, 48 and 72 hours. Changes in cell viability were evaluated using Presto Blue assay. Subsequently, DHL cells were exposed to doublet combinations of ABT-199, JQ-1 and CFZ for 48 hours. Coefficient of synergy was calculated using CalcuSyn. Results: In vitro, ABT199, JQ-1, and CFZ induced cell death in a dose- and time-dependent manner. Significant synergistic activity was observed by combining ABT199 with CFZ and to a lesser degree with JQ-1. Conclusion: ABT199 exhibited strong synergistic activity with CFZ. Dual targeting of BCL2 and c-MYC pathways results in synergistic activity in DHL cell lines. Of interest, this pharmacological interaction could be related to the effects of proteasome inhibition on MCL1 and p21 levels in lymphoma cells, further enhancing the activity of ABT199. Using combination therapy to inhibit c-MYC and the proteasome and in turn decreasing MCL1 will render ABT-199 more effective and be a more potent combination in causing apoptosis and lymphoma cell death. Further pre-clinical work is ongoing. Citation Format: Priyank P. Patel, Alison Zeccola, Juan Gu, Cory Mavis, Sheila N. J. Sait, Vishala Neppalli, Francisco J. Hernandez-Ilizaliturri. Investigating novel targeted therapies for double hit diffuse large B-cell lymphoma (DH-DLBCL). [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 3038.

Palbociclib (PD-0332991), a Potent Selective CDK4/6 Inhibitor, Overcomes Therapy-Resistant and Sensitizes Diffuse Large B-Cell Lymphoma Pre-Clinical Models to Chemo-Reagents

Background: Results from the Collaborative trial in relapsed aggressive lymphoma (CORAL) study highlighted the poor clinical outcome of relapsed/refractory diffuse large B-cell lymphoma (DLBCL) patients previously treated with rituximab-based therapy. To better understand the molecular mechanisms underlining the acquired resistance to rituximab, we generated and characterized several rituximab-resistant DLBCL cell lines (RRCLs). We found that RRCL exhibited a unique cell cycle distribution (accumulation in S-phase) and an increase in cyclin D Kinase-4 (CDk4) mRNA/protein levels. In our current work, we evaluated Palbociclib, a novel orally bioavailable CDK4/6 inhibitor recently approved by the Food and Drug Administration for the treatment of breast cancer in rituximab-sensitive and -resistant lymphoma pre-clinical models.Methods: A panel of rituximab-sensitive cell lines (RSCL) or RRCL were exposed to escalating dosages of PD0332991 (0-80µM) for 24-72 hrs. Changes in cell viability and cell cycle distribution were evaluated using the Presto Blue assay and flow cytometry respectively. IC50 values were calculated using the GraphPad Prism6 software. Subsequently RSCL or RRCL were exposed to PD0332991 or vehicle control and doxorubicin (0.5, 1, 2µM), dexamethasone (1µM), ibrutinib (1µM), bortezomib (10-20nM) or carfilzomib (10nM) for 48 hours. Changes in cell viability were determined by the Presto Blue assay. Coefficient of synergy was calculated using the CalcuSyn software.Results: Palbociclib induced dose- and time-dependent decrease in cell viability in both RSCL and RRCL. Of interest, palbociclib was more potent in RRCL than RSCL. The IC50 values were lower in RRCL Raji-4RH and RL-4RH (12.12µM and 13.43µM) than in RSCL Raji and RL cells (14.7µM and 19.99µM). In vitro exposure of lymphoma cells to palbociclib resulted in G1 cell cycle arrested and it was more evident in RRCL than in RSCL. Synergistic activity was observed by combining palbociclib with doxorubicin, dexamethasone and proteasome inhibitors (bortezomib and carfilzomib) in vitro. Our data suggests that, palbociclib is active against in rituximab-sensitive or resistant pre-clinical models. In addition, CDK4/6 inhibition resulted in synergistic effects with several chemotherapeutic agents. A better understanding in the molecular events triggered by palbociclib is necessary to better develop optimal combination strategies using this novel agent in future clinical trials. (Research, in part, supported by a NIH grant R01 CA136907-01A1 awarded to Roswell Park Cancer Institute and The Eugene and Connie Corasanti Lymphoma Research Fund). DisclosuresCzuczman:Celgene: Employment; MorphoSys: Consultancy; Immunogen: Other: Advisory board; Boehringer-Ingelheim: Other: Advisory Board.

JQ1, a Potent c-MYC Inhibitor Overcomes Rituximab-Chemotherapy Resistance in Lymphoma Pre-Clinical Models

Background: The poor clinical outcomes of patients with aggressive B-cell lymphoma in the post-rituximab era, stress the need to identify and/or optimize novel targeted agents. Several retrospective and prospective clinical studies had demonstrated that C-myc expression correlates with a poor clinical outcome in patients with newly diagnosed or relapsed/refractory diffuse large B-cell lymphoma (DLBCL). To this end, we evaluated the therapeutic effects of targeting C-myc using JQ1, a novel bromodomain inhibitor in rituximab-sensitive or -resistant models.Methods: A panel of rituximab-sensitive (RSCL) or rituximab-resistant (RSCL) cell lines was exposed to JQ1 (0-100 µM) for 24-72 hrs. Changes in cell viability and cell cycle distribution were evaluated using the Presto Blue assay and flow cytometry respectively. IC50 values were calculated using the GraphPad Prism6 software. Subsequently lymphoma cells were exposed to JQ1 or vehicle and various chemotherapy agents such as doxorubicin (0.5, 1, 2µM), dexamethasone (1µM), Ibrutinib (1µM), bortezomib (10-20nM) or carfilzomib (10nM) for 48 hours. Coefficient of synergy was calculated using the CalcuSyn software.Results:In vitro exposure of RRCL and to a lesser degree RSCL to JQ1 resulted in a dose- and time-dependent cell death. Strong synergistic activity was observed when JQ1 was combined with doxorubicin, dexamethasone bortezomib or carfilzomib in vitro. Cell cycle analysis demonstrated that in vitro of RSCL or RRCL to JQ1 resulted in G1 cell cycle arrest.Conclusions: In summary, our data suggests that targeting C-myc expression using JQ1 results in anti-tumor activity against RSCL and RRCL. In addition, JQ1 exhibited synergistic activity when combined with chemotherapy agents (doxorubicin or dexamethasone) or targeted agents (bortezomib or carfilzomib). On going studies are aimed to study the mechanisms by which c-myc inhibition results in cell death in RSCL and RRCL. JQ1 is a distinct targeted agent undergoing clinical evaluation in patients with relapsed/refractory lymphomas. Molecular studies dissecting the cellular pathways affected by JQ1 are important in order to further advance the clinical development of c-myc inhibitors in lymphoid malignancies. (Research, in part, supported by a NIH grant R01 CA136907-01A1 awarded to Roswell Park Cancer Institute and The Eugene and Connie Corasanti Lymphoma Research Fund) DisclosuresCzuczman:Boehringer-Ingelheim: Other: Advisory Board; Immunogen: Other: Advisory board; MorphoSys: Consultancy; Celgene: Employment.