Bryostatin 1

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George R. Pettit - One of the best experts on this subject based on the ideXlab platform.

  • Neristatin 1 provides critical insight into Bryostatin 1 structure-function relationships.
    Journal of Natural Products, 2015
    Co-Authors: Noemi Kedei, Matthew B. Kraft, Gary E. Keck, Noeleen Melody, Cheery L Herald, George R. Pettit, Peter M. Blumberg
    Abstract:

    Bryostatin 1, a complex macrocyclic lactone isolated from Bugula neritina, has been the subject of multiple clinical trials for cancer. Although it functions as an activator of protein kinase C (PKC) in vitro, Bryostatin 1 paradoxically antagonizes most responses to the prototypical PKC activator, the phorbol esters. The bottom half of the Bryostatin 1 structure has been shown to be sufficient to confer binding to PKC. In contrast, we have previously shown that the top half of the Bryostatin 1 structure is necessary for its unique biological behavior to antagonize phorbol ester responses. Neristatin 1 comprises a top half similar to that of Bryostatin 1 together with a distinct bottom half that confers PKC binding. We report here that neristatin 1 is Bryostatin 1-like, not phorbol ester-like, in its biological activity on U937 promyelocytic leukemia cells. We conclude that the top half of the Bryostatin 1 structure is largely sufficient for Bryostatin 1-like activity, provided the molecule also possesses ...

  • Synergistic action of calcium ionophore A23187 and protein kinase C activator Bryostatin 1 on human B cell activation and proliferation.
    European Journal of Immunology, 2005
    Co-Authors: Hans G. Drexler, Suzanne M. Gignac, George R. Pettit, A. Victor Hoffbrand
    Abstract:

    In this study we have examined the immunostimulatory effects of the macrocyclic lactone Bryostatin 1 on various aspects of B cell activation and proliferation using human tonsillar B cells. Bryostatin 1 is an activator of protein kinase C (PKC) and its properties were compared to those of the classical PKC activator phorbol 12-myristate 13-acetate (PMA), a phorbol ester. Time-course kinetics and dose-response curves of RNA and DNA synthesis induced by Bryostatin 1 or PMA were comparable, albeit the phorbol ester was significantly more potent. The responses triggered by both Bryostatin 1 and PMA could be blocked by the PKC inhibitor H7. Bryostatin 1 and PMA mediated similar effects with regard to the activation parameters, increase in cell size, expression of activation-associated antigens and hyperexpression of major histocompatibility complex class II antigens. Addition of the calcium ionophore A23187 to Bryostatin 1-treated cultures resulted in synergistically enhanced activation and proliferation responses, and this potentiation by A23187 could be inhibited by cyclosporin A. Bryostatin 1 antagonized the effects of PMA-triggered stimulation in a time- and dose-dependent manner. The basis for this modulation of PMA-induced effects and the reason for the difference in the abilities of the two agents to stimulate B cells is unclear; possibly, Bryostatin 1 and PMA activate different isoforms of PKC and elicit different signals on intracellular biochemical pathways. Bryostatin 1 lacks the tumor-promoting activity of PMA and is a potent anti-neoplastic substance. These features together with its immunomodulatory properties qualify Bryostatin 1 as a candidate for in vivo use as a biological response modifier.

  • the transient increase of tight junction permeability induced by Bryostatin 1 correlates with rapid downregulation of protein kinase c α
    Experimental Cell Research, 2000
    Co-Authors: Hilary Clarke, George R. Pettit, Nicole Ginanni, K V Laughlin, Jeffrey B Smith, James M Mullin
    Abstract:

    Abstract The role of PKC-α in altered epithelial barrier permeability following the activation of PKC by TPA (12-O-tetradecanoyl phorbol 13-acetate) and Bryostatin 1 in LLC-PK1 cells was investigated in this study. Like TPA, Bryostatin 1 binds to and activates PKC but unlike TPA, it is not a tumor promoter. TPA at 10−7 M induced a sustained 95% decrease in transepithelial electrical resistance (Rt) across LLC-PK1 epithelial cell sheets, while 10−7 M Bryostatin 1 caused only a 30% decrease in Rt, which spontaneously reversed after 5 h. Simultaneous exposure of cell sheets to 10−7 M TPA and 10−7 M Bryostatin 1 blunted the increase in epithelial permeability observed with 10−7 M TPA alone. Co-incubation of cell sheets with Bryostatin 1 and MG-132, a proteasomal inhibitor, caused a further decrease in Rt at the 6-h time point and inhibited the recovery in Rt seen with Bryostatin 1 alone at this time point. TPA caused a rapid translocation of PKC-α from the cytosol to the membrane of the cell where it remained elevated. Bryostatin 1 treatment resulted in a slower translocation of PKC-α from the cytosol to the membrane and a much more rapid downregulation of PKC-α, with disappearance from this compartment after only 6 h. The classical PKC inhibitor Go6976 prevented the decrease in Rt seen with TPA. Treatment of cells with TPA and Bryostatin 1 resulted in a PKC-α translocation and downregulation profile which more closely resembled that seen with Bryostatin 1 alone. Co-incubation of cells with MG-132 and Bryostatin 1 caused a slower downregulation of PKC-α from the membrane fraction. Bryostatin 1 treatment of cells expressing a dominant/negative form of PKC-α resulted in a slower and less extensive decrease in Rt compared to the corresponding control cells. For both TPA and Bryostatin 1, the level of PKC-α in the membrane-associated fraction of the treated cells correlated closely with increased transepithelial permeability. Due to its transient effect on tight junction permeability, Bryostatin 1 offers a novel pharmacological tool to investigate junctional physiology.

  • The inhibitory effects of Bryostatin 1 administration on the growth of rabbit papillomas
    Cancer Letters, 1999
    Co-Authors: Jason M. Bodily, Cheery L Herald, David J Hoopes, Beverly L. Roeder, Sharon G Gilbert, Darin N Rollins, George R. Pettit, Richard A Robison
    Abstract:

    Abstract Bryostatin 1 is a protein kinase C modulator that shows antineoplastic activity in a variety of tumor systems. This study examined the effects of Bryostatin 1 administration on papilloma growth in rabbits. Investigations of optimal route, dose, and schedule were performed. Several groups of rabbits were inoculated with cottontail rabbit papillomavirus (CRPV) DNA. Bryostatin 1 was administered i.v., both daily and weekly, and intralesionally both weekly and bi-weekly. Intralesionally dosed papillomas were examined histologically for immune cell infiltration. In weekly and daily i.v. trials, 2.5 and 1.0 μg/kg, respectively, showed the greatest overall reduction in tumor size. Bryostatin 1 administered intralesionally also slowed papilloma growth. Treated lesions had significantly higher numbers of heterophils and eosinophils.

  • successful treatment of human chronic lymphocytic leukemia xenografts with combination biological agents auristatin pe and Bryostatin 1
    Clinical Cancer Research, 1998
    Co-Authors: Ramzi M. Mohammad, George R. Pettit, Mary L Varterasian, Victor P Almatchy, Ghadeer N Hannoudi, Ayad Alkatib
    Abstract:

    We tested the activity of dolastatin 10 (a natural product derived from the shell-less marine mollusk, Dolabella auricularia, a sea hare) and its structural modification, auristatin PE, alone and in combination with Bryostatin 1 (a protein kinase C activator derived from the marine bryozoan Bugula neritina) on a human B-cell chronic lymphocytic leukemia cell line (WSU-CLL) and in a severe combined immune deficient (SCID) mouse xenograft model bearing this cell line. WSU-CLL cells were cultured in RPMI 1640 at a concentration of 2 x 10(5)/ml using a 24-well plate. Agents were added to triplicate wells, and cell count, viability, mitosis, and apoptosis were assessed after 24 h of incubation at 37 degrees C. Results showed that dolastatin 10 had no apparent inhibition of cell growth at concentrations less than 500 pg/ml. Auristatin PE, on the other hand, showed significant growth inhibition at concentrations as low as 50 pg/ml. Auristatin PE-treated cultures, at this concentration, exhibited 27 and 4.5% mitosis and apoptosis, respectively. Dolastatin 10, at the same concentration, did not exert any effect and was comparable with that of control cultures. In the WSU-CLL-SCID mouse xenograft model, the efficacy of these agents alone and in combination with Bryostatin 1 was evaluated. Tumor growth inhibition (T/C), tumor growth delay (T-C), and log10 kill for dolastatin 10, auristatin PE, and Bryostatin 1 were 14%, 25 days, and 1.98; 2%, 25 days, and 1.98; 19%, 13 days, and 1.03, respectively. Auristatin-PE produced cure in three of five mice, whereas dolastatin 10 showed activity but no cures. When given in combination, auristatin PE + Bryostatin 1-treated animals were all free of tumors (five of five) for 150 days and were considered cured. Dolastatin 10 + Bryostatin 1-treated animals produced cure in only two of five mice. We conclude that: (a) auristatin-PE is more effective in this model than dolastatin 10; (b) auristatin PE can be administered at a concentration 10 times greater than dolastatin 10; (c) there is a synergetic effect between these agents and Bryostatin 1, which is more apparent in the Bryostatin 1 + auristatin PE combination. The use of these agents should be explored clinically in the treatment of CLL.

Peter M. Blumberg - One of the best experts on this subject based on the ideXlab platform.

  • Munc13 Is a Molecular Target of Bryostatin 1.
    Biochemistry, 2019
    Co-Authors: Francisco A. Blanco, Noemi Kedei, Agnes Czikora, Gary A. Mitchell, Satyabrata Pany, Anamitra Ghosh, Peter M. Blumberg
    Abstract:

    Bryostatin 1 is a natural macrolide shown to improve neuronal connections and enhance memory in mice. Its mechanism of action is largely attributed to the modulation of novel and conventional protein kinase Cs (PKCs) by binding to their regulatory C1 domains. Munc13-1 is a C1 domain-containing protein that shares common endogenous and exogenous activators with novel and conventional PKC subtypes. Given the essential role of Munc13-1 in the priming of synaptic vesicles and neuronal transmission overall, we explored the potential interaction between Bryostatin 1 and Munc13-1. Our results indicate that in vitro Bryostatin 1 binds to both the isolated C1 domain of Munc13-1 (Ki = 8.07 ± 0.90 nM) and the full-length Munc13-1 protein (Ki = 0.45 ± 0.04 nM). Furthermore, confocal microscopy and immunoblot analysis demonstrated that in intact HT22 cells Bryostatin 1 mimics the actions of phorbol esters, a previously established class of Munc13-1 activators, and induces plasma membrane translocation of Munc13-1, a h...

  • munc13 is a molecular target of Bryostatin 1
    Biochemistry, 2019
    Co-Authors: Francisco A. Blanco, Noemi Kedei, Agnes Czikora, Gary A. Mitchell, Satyabrata Pany, Anamitra Ghosh, Peter M. Blumberg
    Abstract:

    : Bryostatin 1 is a natural macrolide shown to improve neuronal connections and enhance memory in mice. Its mechanism of action is largely attributed to the modulation of novel and conventional protein kinase Cs (PKCs) by binding to their regulatory C1 domains. Munc13-1 is a C1 domain-containing protein that shares common endogenous and exogenous activators with novel and conventional PKC subtypes. Given the essential role of Munc13-1 in the priming of synaptic vesicles and neuronal transmission overall, we explored the potential interaction between Bryostatin 1 and Munc13-1. Our results indicate that in vitro Bryostatin 1 binds to both the isolated C1 domain of Munc13-1 ( Ki = 8.07 ± 0.90 nM) and the full-length Munc13-1 protein ( Ki = 0.45 ± 0.04 nM). Furthermore, confocal microscopy and immunoblot analysis demonstrated that in intact HT22 cells Bryostatin 1 mimics the actions of phorbol esters, a previously established class of Munc13-1 activators, and induces plasma membrane translocation of Munc13-1, a hallmark of its activation. Consistently, Bryostatin 1 had no effect on the Munc13-1H567K construct that is insensitive to phorbol esters. Effects of Bryostatin 1 on the other Munc13 family members, ubMunc13-2 and bMunc13-2, resembled those of Munc13-1 for translocation. Lastly, we observed an increased level of expression of Munc13-1 following a 24 h incubation with Bryostatin 1 in both HT22 and primary mouse hippocampal cells. This study characterizes Munc13-1 as a molecular target of Bryostatin 1. Considering the crucial role of Munc13-1 in neuronal function, these findings provide strong support for the potential role of Munc13s in the actions of Bryostatin 1.

  • Neristatin 1 provides critical insight into Bryostatin 1 structure-function relationships.
    Journal of Natural Products, 2015
    Co-Authors: Noemi Kedei, Matthew B. Kraft, Gary E. Keck, Noeleen Melody, Cheery L Herald, George R. Pettit, Peter M. Blumberg
    Abstract:

    Bryostatin 1, a complex macrocyclic lactone isolated from Bugula neritina, has been the subject of multiple clinical trials for cancer. Although it functions as an activator of protein kinase C (PKC) in vitro, Bryostatin 1 paradoxically antagonizes most responses to the prototypical PKC activator, the phorbol esters. The bottom half of the Bryostatin 1 structure has been shown to be sufficient to confer binding to PKC. In contrast, we have previously shown that the top half of the Bryostatin 1 structure is necessary for its unique biological behavior to antagonize phorbol ester responses. Neristatin 1 comprises a top half similar to that of Bryostatin 1 together with a distinct bottom half that confers PKC binding. We report here that neristatin 1 is Bryostatin 1-like, not phorbol ester-like, in its biological activity on U937 promyelocytic leukemia cells. We conclude that the top half of the Bryostatin 1 structure is largely sufficient for Bryostatin 1-like activity, provided the molecule also possesses ...

  • abstract 1642 Bryostatin 1 and the simplified analog merle 23 have similar and opposing properties on mouse epidermal cells and mouse skin
    Cancer Research, 2014
    Co-Authors: Jessica S. Kelsey, Noemi Kedei, Gary E. Keck, Christophe Cataisson, Stuart H. Yuspa, Mark E. Petersen, Peter M. Blumberg
    Abstract:

    Protein kinase C (PKC) is differentially regulated in a range of cancers and has become an attractive therapeutic target. A number of natural compounds exist that have been found to regulate PKCs, and the biological responses of each compound can vary. The PKC activator PMA is an established tumor promoter, while others, such as the Bryostatins, are non-tumor promoting and have inhibitory effects on the PKC pathway. Because of its effects on the PKC pathway, Bryostatin 1 is currently in clinical trials as an anti-cancer agent. Limited availability and the difficulty involved in synthesizing Bryostatin 1 make its use as an anti-cancer agent less attractive. Merle 23, a simplified synthetic analog of Bryostatin 1, could be an alternative to Bryostatin 1 if it acted similarly. Initial investigations of Merle 23 have revealed biological responses both similar to Bryostatin 1 and similar to PMA, depending on the system. In U937 leukemia cells Merle 23 acts similar to PMA, and in the LNCaP prostate cancer cell line it behaves more like Bryostatin 1. Its behavior in mouse skin is of particular interest, since this is the system in which the tumor promoting activity of PMA and the anti-tumor promoting activity of Bryostatin 1 have been characterized. The activity of Merle 23 was compared to PMA and Bryostatin 1 in mouse epidermal cells and mouse skin. In mouse primary keratinocytes PMA produces a prolonged morphological response, while the morphological response to Bryostatin 1 is transient. The response to Merle 23 is intermediate, but more similar to Bryostatin 1. When applied simultaneously, Merle 23, like Bryostatin 1, is able to protect the cells from the morphological response induced by PMA. In the PKC pathway, all three compounds promote similar levels of initial PKCδ activation and ERK1/2 phosphorylation. By 24 hrs, however, the responses differ. At high concentrations, Bryostatin 1 protects PKCδ from down regulation whereas Merle 23 does not. Bryostatin 1 is also much more potent for down regulation of PKCα, whereas Merle 23 and PMA are similar. The inflammatory response was measured by monitoring the transcript levels of various genes involved in inflammation. PMA induced the largest increase in transcript levels, while Merle 23 was more similar to Bryostatin 1. Short-term in vivo analysis reveals no epidermal thickening in response to low doses of Merle 23 whereas the corresponding dose of PMA induced significant hyperplasia. The in vivo analysis of Merle 23 is ongoing. The results presented suggest that Merle 23 behaves more similarly to Bryostatin 1 in the mouse epidermal cells at the level of biological response, raising the possibility that it may likewise be non-promoting. The results emphasize that the structure activity relations responsible for the distinct behavior between PMA and Bryostatin 1 are distinct for different cellular systems and different biological endpoints. Citation Format: Jessica Kelsey, Noemi Kedei, Christophe Cataisson, Mark Petersen, Stuart Yuspa, Gary Keck, Peter Blumberg. Bryostatin 1 and the simplified analog Merle 23 have similar and opposing properties on mouse epidermal cells and mouse skin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1642. doi:10.1158/1538-7445.AM2014-1642

  • Abstract 2454: Comparison of the activity of the Bryostatin derivative Merle 23 with that of Bryostatin 1 and phorbol ester in mouse epidermal cells.
    Cancer Research, 2013
    Co-Authors: Jessica S. Kelsey, Noemi Kedei, Gary E. Keck, Christophe Cataisson, Stuart H. Yuspa, Mark E. Petersen, Peter M. Blumberg
    Abstract:

    Proceedings: AACR 104th Annual Meeting 2013; Apr 6-10, 2013; Washington, DC Tumor promoting phorbol esters, such as PMA, are potent activators of the PKC pathway. The Bryostatins, marine natural products, activate PKCs similarly to phorbol esters but paradoxically inhibit most phorbol ester responses, including skin tumor promotion. Because of this inhibitory activity on the PKC pathway, Bryostatin 1 has been the subject of extensive clinical trials. Serious impediments to clinical development, however, have been the scarcity of natural supplies coupled with great structural complexity, making synthesis problematic. Recent synthetic advances are now yielding simplified structural derivatives. Here, we compare the behavior of the Bryostatin derivative Merle 23 with that of Bryostatin 1 and phorbol 12-myristate 13-acetate (PMA) in mouse epidermal cells. While having similar binding activity on PKC as does Bryostatin 1, Merle 23 behaves like PMA rather than like Bryostatin 1 in U937 human leukemia cells but conversely behaves more like Bryostatin 1 in the human prostate cancer cell line LNCaP. As a first step in assessing whether Merle 23 possesses or lacks skin tumor promoting activity, its activity in mouse primary keratinocyte cultures was assessed. PMA induces a prominent, prolonged morphological response in keratinocytes. Bryostatin 1 induces a similar initial response, which is however very transient. The morphological response to Merle 23 is more prolonged than for Bryostatin 1 but much shorter than for PMA. For initial activation of PKCδ or induction of ERK1/2 phosphorylation, all three ligands show similar activity. By 24 hrs, however, Bryostatin 1 is much more potent for down regulation of PKCα, whereas Merle 23 and PMA are similar. For PKCδ, Bryostatin 1 at higher concentrations protects PKCδ from down regulation whereas Merle 23 does not. Transcriptional responses of various inflammation genes were monitored by qPCR. PMA causes the greatest increase in transcript levels for all genes investigated. In many cases Bryostatin 1 induces a smaller response and the response to Merle 23 generally resembles that of Bryostatin 1. While these studies are ongoing, they suggest that Merle 23 largely behaves like Bryostatin 1 in the mouse epidermal cells. Our findings emphasize the important role of cellular context in determining the pattern of behavior of Merle 23. An important implication is that different cellular environments reveal different structure activity relations for Bryostatin 1-like behavior. Finally, our studies raise the possibility that Merle 23, which is significantly simplified relative to Bryostatin 1 in its upper two rings, may be non-promoting, at least in the mouse skin tumor promotion system. Citation Format: Jessica Kelsey, Noemi Kedei, Christophe Cataisson, Mark Petersen, Stuart Yuspa, Gary Keck, Peter Blumberg. Comparison of the activity of the Bryostatin derivative Merle 23 with that of Bryostatin 1 and phorbol ester in mouse epidermal cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2454. doi:10.1158/1538-7445.AM2013-2454

Andrew S Kraft - One of the best experts on this subject based on the ideXlab platform.

  • Treatment of patients with metastatic melanoma with Bryostatin-1--a phase II study.
    Melanoma Research, 1999
    Co-Authors: Rene Gonzalez, Scot Ebbinghaus, Thomas K. Henthorn, D. Miller, Andrew S Kraft
    Abstract:

    : Bryostatin-1 is a protein kinase C regulator which has shown antitumour activity against B16 melanoma in animal models. Safety trials revealed this agent to be minimally toxic, thus a phase II trial of Bryostatin-1 was conducted to determine its efficacy In patients with melanoma. Eighteen patients with metastatic melanoma, seven of whom had been previously treated, were enrolled in the study. Patients received Bryostatin-1 25 microg/m2 intravenously weekly over 1 h for 3 out of 4 weeks. No objective responses were observed. One patient who had not previously received chemotherapy had stable disease for 4 months, and two patients (one previously treated) had a marked decrease in the skin component of their disease. The major toxicity was myalgia (one patient with grade III, two patients with grade II and five patients with grade I), with no grade IV toxicities reported. To Indirectly evaluate the stimulation of protein kinase C, a sensitive assay that measures the upregulation of the activated form of CD62 (glycoprotein IIb/IIIa) on platelets was performed. There was a statistically significant upregulation of this antigen 1 h after Bryostatin-1 therapy. A bioassay based on the ability of Bryostatin-1 to bind protein kinase C was used to measure Bryostatin-1 levels in serum. This assay showed that Bryostatin-1 has a volume of distribution of 2.1 l/m2, an elimination clearance of 32.9 ml/min per m2 and a half-life of 43.9 min. In conclusion, this phase II trial demonstrates that, although it is relatively non-toxic, Bryostatin-1 therapy had minimal activity in metastatic melanoma.

  • a phase i trial of Bryostatin 1 in children with refractory solid tumors a pediatric oncology group study
    Clinical Cancer Research, 1999
    Co-Authors: Steven D Weitman, Andrew S Kraft, Roger L. Berkow, Annemarie R Langevin, Paul J Thomas, Craig A Hurwitz, Ronald L Dubowy, Debra L Smith, Mark L Bernstein
    Abstract:

    Bryostatin-1, a macrocyclic lactone, appears to elicit a wide range of biological responses including modulation of protein kinase C (PKC). PKC, one of the major elements in the signal transduction pathway, is involved in the regulation of cell growth, differentiation, gene expression, and tumor promotion. Because of the potential for a unique mechanism of interaction with tumorgenesis, a Phase I trial of Bryostatin-1 was performed in children with solid tumors to: ( a ) establish the dose-limiting toxicity (DLT) and maximum-tolerated dose (MTD); ( b ) establish the pharmacokinetic profile in children; and ( c ) document any evidence of antitumor activity. A 1-h infusion of Bryostatin-1 in a PET formulation (60% polyethylene glycol 400, 30% ethanol, and 10% Tween 80) was administered weekly for 3 weeks to 22 children (age range, 2–21 years) with malignant solid tumors refractory to conventional therapy. Doses ranged from 20 to 57 μg/m 2 /dose. Pharmacokinetics were performed in at least three patients per dose level. The first course was used to determine the DLT and MTD. Twenty-two patients on five dose levels were evaluable for toxicities. At the 57 μg/m 2 /dose level dose-limiting myalgia (grade 3) was observed in three patients; two of those patients also experienced photophobia or eye pain, and one experienced headache. Symptoms occurred in all patients within 24–72 h after the second dose of Bryostatin-1 with resolution within 1 week of onset. Other observed toxicities (grades 1 and 2) included elevation in liver transaminases, thrombocytopenia, fever, and flu-like symptoms. The Bryostatin-1 infusion was typically well tolerated. Although stable disease was noted in several patients, no complete or partial responses were observed. The recommended Phase II dose of Bryostatin-1 administered as a 1-h infusion weekly for 3 of every 4 weeks to children with solid tumors is 44 μg/m 2 /dose. Myalgia, photophobia, or eye pain, as well as headache, were found to be dose limiting.

  • preclinical pharmacology of the natural product anticancer agent Bryostatin 1 an activator of protein kinase c
    Cancer Research, 1996
    Co-Authors: George R. Pettit, Xueshu Zhang, Ruiwen Zhang, Hui Zhao, Kimberly A Gush, Russell G Kerr, Andrew S Kraft
    Abstract:

    Abstract Bryostatin 1, a natural product anticancer agent isolated from a marine bryozoan, has been shown in tissue culture to activate protein kinase C. This agent has recently undergone Phase I testing in humans given either as a bolus i.v. injection or a continuous infusion. To understand how Bryostatin 1 might be used best as an anticancer agent, a study of the pharmacokinetics, tissue distribution, metabolism, and elimination of Bryostatin 1 in mice was undertaken, using [C26- 3 H]-labeled Bryostatin 1. Following i.v. administration, the plasma disappearance curve for Bryostatin 1 could be described by a two-compartment model, with half-lives of 1.05 and 22.97 h, respectively. In contrast, the plasma disappearance curve for Bryostatin 1 administered i.p. was better described by a first order absorption one-compartment model, with an absorption half-life of 0.81 h and an elimination half-life of 28.76 h, respectively. The majority of radioactivity in plasma was associated with the intact drug for up to 24 h after dosing. In the first 12 h after i.v administration, urinary excretion represented the major pathway of elimination, with 23.0 ± 1.9% (mean ± SD) of the administered dose excreted. Within 72 h after i.v. administration, approximately equal amounts of radioactivity (40%) were excreted in feces compared to urine. Bryostatin 1 was widely distributed in many organs but concentrated in the lung, liver, gastrointestinal tract, and fatty tissue. The concentration in the gastrointestinal tract, along with the fecal excretion, suggests the possibility of enterohepatic circulation of this drug. In summary, this study demonstrates that Bryostatin 1 is relatively stable in vivo , widely distributed but concentrated in some major tissues, and rapidly excreted first through urine and at later times through the feces. The data from this animal study should be useful in the design of future human trials with this anticancer drug.

  • inhibition of leukemic cell growth by the protein kinase c activator Bryostatin 1 correlates with the dephosphorylation of cyclin dependent kinase 2
    Cancer Research, 1995
    Co-Authors: Clement Asiedu, Andrew S Kraft, Michael B. Lilly, Joseph R Biggs
    Abstract:

    Bryostatin 1 is a natural antineoplastic agent that activates protein kinase C. Treatment of U937 human leukemic cells with Bryostatin 1 caused a 60% reduction in cell growth, whereas another protein kinase C activator, phorbol myristate acetate (PMA), completely inhibited U937 cell growth. Both Bryostatin 1 and PMA induced inhibition of cyclindependent kinase 2 (cdk2) activity. The first phase of cdk2 inhibition correlated with the transient induction of p21, a known inhibitor of cdk2. In contrast, the second phase of cdk2 inhibition correlated with the dephosphorylation of cdk2 on threonine-160, which must be phosphorylated for cdk2 activity. The level of growth inhibition induced by these two compounds correlated with the degree of cdk2 dephosphorylation as follows: Bryostatin 1, 60%; PMA, 100%.

  • In Vivo Administration of the Anticancer Agent Bryostatin 1 Activates Platelets and Neutrophils and Modulates Protein Kinase C Activity
    Cancer Research, 1993
    Co-Authors: Roger L. Berkow, George R. Pettit, Larry Schlabach, Robert W. Dodson, William H. Benjamin, Pradip K. Rustagi, Andrew S Kraft
    Abstract:

    Bryostatin 1 is a naturally occurring macrocyclic lactone which when applied to cells in culture activates protein kinase C (PKC). In vivo Bryostatin 1 functions as an anticancer agent with activity against murine lymphomas, leukemias, and melanoma. Because all organs and tissues contain PKC, normal cells would also be a likely target for this agent. Here we demonstrate that in vivo administration of Bryostatin 1 activates platelets over a dose range of 0.4 to 40 µg/kg with half-maximal activation occurring at 3 µg/kg and stimulation of neutrophils over a similar dose range. This in vivo activation of neutrophils is associated with a rapid decrease in measurable cytosolic PKC, a finding consistent with translocation of the enzyme to the membrane. In contrast, no statistically significant change in PKC location was found in liver, spleen, brain, or L10A B-cell lymphoma. However, in culture the L10A lymphoma did respond to Bryostatin 1 with translocation of PKC. To evaluate whether the lack of effect of Bryostatin 1 on PKC in organs was secondary to rapid degradation, we developed a bioassay to measure the levels of Bryostatin 1 in the blood. To measure the presence of Bryostatin 1, human neutrophils were incubated with plasma from mice given injections of different concentrations of Bryostatin 1. Using this assay, Bryostatin 1 at levels as low as 60 nm could be measured in the plasma. A time course with this bloassay demonstrated that less than 10% of the Bryostatin 1 injected was detectable after 2.5 min. These results demonstrate that Bryostatin 1 is capable of activating platelets and neutrophils and modulating PKC in vivo . The lack of effect of Bryostatin 1 on specific organs may be secondary to the rapid clearance/degradation of this compound from the blood.

Steven Grant - One of the best experts on this subject based on the ideXlab platform.

  • Induction of tumor necrosis factor by Bryostatin 1 is involved in synergistic interactions with paclitaxel in human myeloid leukemia cells.
    Blood, 2003
    Co-Authors: Shujie Wang, Paul Dent, Zhiliang Wang, Steven Grant
    Abstract:

    Interactions between the protein kinase C (PKC) activator/down-regulator Bryostatin 1 and paclitaxel have been examined in human myeloid leukemia cells (U937) and in highly paclitaxel-resistant cells ectopically expressing a Bcl-2 phosphorylation loop–deleted protein (Bcl-2). Treatment (24 hours) of wild-type cells with paclitaxel (eg, 5 to 20 nM) in combination with 10 nM Bryostatin 1 induced a marked increase in mitochondrial damage (eg, cytochrome c and Smac/DIABLO [second mitochondria-derived activator of caspases/direct IAP binding protein with low pI] release), caspase activation, Bid cleavage, and apoptosis; moreover, Bryostatin 1 circumvented the block to paclitaxel-induced mitochondrial injury and apoptosis conferred by ectopic expression of the loop-deleted protein. Coadministration of tumor necrosis factor (TNF) soluble receptors, or ectopic expression of CrmA or dominant-negative caspase-8, abrogated potentiation of paclitaxel-induced mitochondrial injury and apoptosis by Bryostatin 1, implicating the extrinsic apoptotic pathway in this process. Similar events occurred in HL-60 leukemia cells. Potentiation of paclitaxelinduced apoptosis in wild-type and mutant cells by Bryostatin 1 was associated with increases in TNF- mRNA and protein and was mimicked by exogenous TNF-. Coadministration of the selective PKC inhibitor GFX (1 M) blocked the increase in TNF- mRNA levels and apoptosis in Bryostatin 1/paclitaxel–treated cells. Lastly, synchronization of cells in G2M increased their sensitivity to TNF-– associated lethality. Collectively, these findings indicate that in U937 cells, Bryostatin 1 promotes paclitaxel-mediated mitochondrial injury and apoptosis, and circumvents resistance to cell death conferred by loss of the Bcl-2 phosphorylation domain, through the PKC-dependent induction of TNF-. They further suggest that this process is amplified by paclitaxel-mediated arrest of cells in G2M, where they are more susceptible to TNF–induced lethality. (Blood. 2003;101: 3648-3657)

  • Bryostatin 1 and UCN-01 Potentiate 1-β-d-Arabinofuranosylcytosine-Induced Apoptosis in Human Myeloid Leukemia Cells through Disparate Mechanisms
    Molecular Pharmacology, 2003
    Co-Authors: Shujie Wang, Zhiliang Wang, Steven Grant
    Abstract:

    The effects of the PKC activator and down-regulator Bryostatin 1 and the PKC and Chk1 inhibitor 7-hydroxystaurosporine (UCN-01) were compared with respect to potentiation of 1-β-d-arabinofuranosylcytosine (ara-C)-induced apoptosis in human myelomonocytic leukemia cells (U937). Whereas Bryostatin 1 and UCN-01 both markedly enhanced ara-C–induced mitochondrial injury (e.g., cytochrome c and Smac/DIABLO release, loss of mitochondrial membrane potential), caspase activation, and apoptosis, ectopic expression of an N-terminal loop-deleted Bcl-2 mutant protein protected cells from ara-C/UCN-01– but not ara-C/Bryostatin 1-mediated lethality. Conversely, ectopic expression of CrmA or dominant-negative caspase-8 abrogated potentiation of ara-C–mediated apoptosis by Bryostatin 1 but not by UCN-01. Exposure of cells to ara-C and Bryostatin 1 (but not UCN-01) resulted in sustained release of tumor necrosis factor (TNF) α; moreover, potentiation of ara-C lethality by Bryostatin 1 (but not by UCN-01) was reversed by coadministration of TNF soluble receptors or the selective PKC inhibitor bisindolylmaleimide (1 μM). Finally, similar events were observed in the human promyelocytic leukemia cell line HL-60. Together, these findings suggest that potentiation of ara-C lethality in human myeloid leukemia cells by Bryostatin 1 but not UCN-01 involves activation of the extrinsic, receptor-mediated apoptotic pathway, and represents a consequence of Bryostatin 1-mediated release of TNF-α. They also argue that the mechanism by which Bryostatin 1 promotes ara-C–induced mitochondrial injury, caspase activation, and apoptosis involves factors other than or in addition to PKC down-regulation or modulation of Bcl-2 phosphorylation status.

  • Bryostatin 1 enhances paclitaxel induced mitochondrial dysfunction and apoptosis in human leukemia cells u937 ectopically expressing bcl xl
    Leukemia, 1999
    Co-Authors: Shujie Wang, Paul Dent, Zhiliang Wang, Lawrence H Boise, Steven Grant
    Abstract:

    The effects of the protein kinase C (PKC) activator and down-regulator Bryostatin 1 were examined with respect to paclitaxel-induced apoptosis and antiproliferative activity in human myeloid leukemia cells (U937) displaying enforced expression of the anti-apoptotic protein Bcl-xL. Overexpression of Bcl-xL blocked various aspects of paclitaxel-mediated apoptosis, including caspase-3 activation, degradation of poly(ADP-ribose) polymerase (PARP), loss of mitochondrial membrane potential (Δ Ψm), and release of cytochrome c. However, subsequent (but not prior) exposure of paclitaxel-treated U937/Bcl-xL cells (500 nM; 6 h) to Bryostatin 1 (10 nM; 15 h) restored the extent of apoptosis, caspase activation, and mitochondrial damage to levels approximating those in paclitaxel-treated empty-vector control cells (U937/Neo). Potentiation of paclitaxel-induced apoptosis by Bryostatin 1 in U937/Bcl-xL cells occurred primarily in the G2M cell population, and was associated with alterations in Bcl-xL gel mobility and a reduction in paclitaxel-mediated stimulation of CDK1 activity. Enhancement of paclitaxel-induced apoptosis by Bryostatin 1 in Bcl-xL overexpressors was accompanied by a corresponding reduction in clonogenic potential. In contrast to its effects on apoptosis, Bryostatin 1 failed to restore paclitaxel-mediated increases in free Bax levels in U937/Bcl-xL cells. Lastly, the actions of Bryostatin 1 were mimicked by a pharmacologic inhibitor of the MEK1/MAP kinase pathway (PD98059), but not by SB203580, an inhibitor of p 38 MAP kinase. Moreover, sequential exposure of both U937/Neo or/Bcl-xL cells to paclitaxel followed by Bryostatin 1 or PD98059 was associated with a net reduction in MAP kinase activity. Collectively, these findings indicate that protection against paclitaxel-mediated mitochondrial dysfunction and apoptosis in human U937 leukemia cells conferred by Bcl-xL overexpression can be substantially overcome by Bryostatin 1 and possibly other agents that interrupt the MAP kinase signal transduction pathway.

  • effect of Bryostatin 1 on taxol induced apoptosis and cytotoxicity in human leukemia cells u937
    Biochemical Pharmacology, 1998
    Co-Authors: Shujie Wang, Anthony Castillo, Paul Dent, Steven Grant
    Abstract:

    Abstract We have examined the effects of the macrocyclic lactone protein kinase C (PKC) activator Bryostatin 1 on taxol-induced apoptosis and inhibition of clonogenicity in the human monocytic leukemia cell line U937. Exposure of cells to Bryostatin 1 (10 nM; 15 hr) after (but not before) a 6-hr incubation with 0.5 μM taxol significantly increased apoptosis and resulted in an approximately 3 log reduction in clonogenicity. Cell cycle analysis revealed that the increase in apoptotic cells following Bryostatin 1 treatment occurred primarily in the population undergoing taxol-mediated G2M arrest. The actions of Bryostatin 1 were not attributable to potentiation of taxol-induced tubulin stabilization or to a reduction in the intracellular retention of taxol. Following exposure of cells to taxol, the Bcl-2 protein displayed an alteration in mobility that was not modified appreciably by Bryostatin 1 treatment. The mobility shift in Bcl-2 protein from cells exposed to taxol followed by Bryostatin 1 was eliminated by treatment of lysates with the protein phosphatase 2A (PP2A); the latter effect was blocked by okadaic acid. Treatment of cells with taxol followed by Bryostatin 1 did not increase the amount of total Bax (compared with treatment with taxol alone), but did increase the amount of free Bax in the supernatant fraction. Finally, the ability of Bryostatin 1 to potentiate taxol-induced apoptosis in U937 cells was mimicked closely by 2′-amino-3′-methoxyflavone (PD98059), a specific inhibitor of the mitogen-activated protein kinase (MAPK) kinase (MEK). Collectively, these findings indicate that Bryostatin 1 increases the susceptibility of U937 cells to taxol-induced apoptosis and inhibition of clonogenicity. They also raise the possibility that this phenomenon may involve functional alterations in Bcl-2 and/or other proteins involved in regulation of the cell death pathway.

  • divergent effects of Bryostatin 1 and phorbol myristate acetate on cell cycle arrest and maturation in human myelomonocytic leukemia cells u937
    Differentiation, 1998
    Co-Authors: Julie A Vrana, Aida M Saunders, Srikumar Chellappan, Steven Grant
    Abstract:

    Bryostatin 1 and the phorbol ester, phorbol myristate acetate (PMA), both bind to and activate protein kinase C (PKC) but exhibit divergent biological actions. Bryostatin 1 exerts variable effects on leukemic cell differentiation, and has been reported by some investigators to inhibit the proliferation of the monocytic leukemic cell line U937. In this study, we have compared the efficacy of Bryostatin 1 and PMA with respect to U937 cell maturation, with a major emphasis on differential actions on the cell cycle arrest machinery. At equimolar concentrations (10 nM), PMA, in contrast to Bryostatin 1, induced cellular differentiation of U937 cells, reflected by growth inhibition, increased plastic adhesion, and expression of the monocytic differentiation marker, CD11b. Consistent with these results, Bryostatin 1 was less effective in inducing G0/G1 arrest and inhibiting cyclin-dependent kinase 2 (CDK2) activity. Bryostatin 1, unlike PMA, failed to induce expression of the cyclin-dependent kinase inhibitor (CDKI), p21CIP1/WAF1, and blocked the ability of PMA to induce this protein. Bryostatin 1 exposure resulted in increased expression of the CDKI p27KIP1 in these cells, although the kinetics differed from PMA. In addition, Bryostatin 1 was less effective than PMA in dephosphorylating pRb, modifying E2F complexes, and downregulating c-Myc. Co-administration of Bryostatin 1 with PMA antagonized the latter's differentiation-inducing capacity and anti-proliferative effects, actions that were accompanied by a reduction in PMA-mediated p21CIP1/WAF1 induction, CDK2 inhibition, pRb dephosphorylation, and c-Myc downregulation. Antagonistic effects of Bryostatin 1 on PMA-related cell cycle events were mimicked by the specific PKC inhibitor GF109203X. Together, these studies indicate that Bryostatin 1 is a considerably weaker stimulus than PMA for U937 cell differentiation, and raise the possibility that this deficiency arises from its failure to induce p21CIP1/WAF1 and trigger cell cycle arrest.

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  • phase ii study of Bryostatin 1 in patients with relapsed multiple myeloma
    Investigational New Drugs, 2001
    Co-Authors: Mary L Varterasian, Pamela Pemberton, Kim Hulburd, Dorothy H Rodriguez, Anthony J Murgo, Ayad Alkatib
    Abstract:

    Bryostatin 1, a macrocyclic lactoneisolated from the marine bryozoan Bugula neritina,is a protein kinase C (PKC) modulator which has shown bothpreclinical and clinical activity inlymphoid malignancies. We conducted aphase II trial of Bryostatin 1 administeredat a dose of 120 μg/m2 by 72-hcontinuous infusion every 2 weeks inpatients with relapsed multiple myeloma. Treatment was well tolerated with myalgiasconstituting the primaray toxicity. Therewere no responses in nine evaluablepatients. The preclinical anti-lymphoidactivity is strong enough to supportfurther exploration of Bryostatin 1 indifferent schedules and in combinationtherapy for multiple myeloma.

  • Phase II Trial of Bryostatin 1 in Patients with Relapsed Low-Grade Non-Hodgkin’s Lymphoma and Chronic Lymphocytic Leukemia
    Clinical Cancer Research, 2000
    Co-Authors: Mary L Varterasian, Ramzi M. Mohammad, Kim Hulburd, Dorothy H Rodriguez, Anthony J Murgo, Muhammad Shurafa, Pamela A. Pemberton, Virginia Spadoni, David Eilender, Nathan R. Wall
    Abstract:

    Bryostatin 1 is a natural product isolated from the marine bryozoan Bugula neritina in 1982 and is currently undergoing evaluation in a number of malignancies. Twenty-five patients with relapsed, low-grade non-Hodgkin’s lymphoma or chronic lyphocytic leukemia (CLL) received Bryostatin 1 by 72-h continuous infusion every 2 weeks at a dose of 120μ g/m 2 per course. Patients who progressed while receiving Bryostatin 1 alone could participate in a feasibility study by receiving vincristine administered by bolus i.v. injection immediately after the completion of the Bryostatin 1 infusion. The dose of vincristine was escalated in groups of three patients as follows: level 1, 0.5 mg/m 2 ; level 2, 1.0 mg/m 2 ; and level 3, 1.4 mg/m 2 with vincristine doses capped at 2.0 mg for all patients. Bryostatin 1 alone resulted in one complete remission and two partial remissions. Nine patients received sequential treatment with Bryostatin 1 and vincristine. The addition of vincristine at a dose of 2 mg was feasible and caused the expected dose-related sensory neuropathy. Phenotypic analysis by flow cytometric analysis on pre- and post-Bryostatin 1-treated peripheral blood lymphocytes revealed up-regulation in the coexpression of CD11c/CD22 on CD20 + B cells in two of four CLL patients studied, which is consistent with in vitro findings of differentiation of CLL cells to a hairy cell phenotype.

  • phase ii trial of Bryostatin 1 in patients with relapsed low grade non hodgkin s lymphoma and chronic lymphocytic leukemia
    Clinical Cancer Research, 2000
    Co-Authors: Mary L Varterasian, Ramzi M. Mohammad, Pamela Pemberton, Kim Hulburd, Dorothy H Rodriguez, Anthony J Murgo, Muhammad Shurafa, Virginia Spadoni, David Eilender, Nathan R. Wall
    Abstract:

    Bryostatin 1 is a natural product isolated from the marine bryozoan Bugula neritina in 1982 and is currently undergoing evaluation in a number of malignancies. Twenty-five patients with relapsed, low-grade non-Hodgkin’s lymphoma or chronic lyphocytic leukemia (CLL) received Bryostatin 1 by 72-h continuous infusion every 2 weeks at a dose of 120μ g/m 2 per course. Patients who progressed while receiving Bryostatin 1 alone could participate in a feasibility study by receiving vincristine administered by bolus i.v. injection immediately after the completion of the Bryostatin 1 infusion. The dose of vincristine was escalated in groups of three patients as follows: level 1, 0.5 mg/m 2 ; level 2, 1.0 mg/m 2 ; and level 3, 1.4 mg/m 2 with vincristine doses capped at 2.0 mg for all patients. Bryostatin 1 alone resulted in one complete remission and two partial remissions. Nine patients received sequential treatment with Bryostatin 1 and vincristine. The addition of vincristine at a dose of 2 mg was feasible and caused the expected dose-related sensory neuropathy. Phenotypic analysis by flow cytometric analysis on pre- and post-Bryostatin 1-treated peripheral blood lymphocytes revealed up-regulation in the coexpression of CD11c/CD22 on CD20 + B cells in two of four CLL patients studied, which is consistent with in vitro findings of differentiation of CLL cells to a hairy cell phenotype.

  • successful treatment of human chronic lymphocytic leukemia xenografts with combination biological agents auristatin pe and Bryostatin 1
    Clinical Cancer Research, 1998
    Co-Authors: Ramzi M. Mohammad, George R. Pettit, Mary L Varterasian, Victor P Almatchy, Ghadeer N Hannoudi, Ayad Alkatib
    Abstract:

    We tested the activity of dolastatin 10 (a natural product derived from the shell-less marine mollusk, Dolabella auricularia, a sea hare) and its structural modification, auristatin PE, alone and in combination with Bryostatin 1 (a protein kinase C activator derived from the marine bryozoan Bugula neritina) on a human B-cell chronic lymphocytic leukemia cell line (WSU-CLL) and in a severe combined immune deficient (SCID) mouse xenograft model bearing this cell line. WSU-CLL cells were cultured in RPMI 1640 at a concentration of 2 x 10(5)/ml using a 24-well plate. Agents were added to triplicate wells, and cell count, viability, mitosis, and apoptosis were assessed after 24 h of incubation at 37 degrees C. Results showed that dolastatin 10 had no apparent inhibition of cell growth at concentrations less than 500 pg/ml. Auristatin PE, on the other hand, showed significant growth inhibition at concentrations as low as 50 pg/ml. Auristatin PE-treated cultures, at this concentration, exhibited 27 and 4.5% mitosis and apoptosis, respectively. Dolastatin 10, at the same concentration, did not exert any effect and was comparable with that of control cultures. In the WSU-CLL-SCID mouse xenograft model, the efficacy of these agents alone and in combination with Bryostatin 1 was evaluated. Tumor growth inhibition (T/C), tumor growth delay (T-C), and log10 kill for dolastatin 10, auristatin PE, and Bryostatin 1 were 14%, 25 days, and 1.98; 2%, 25 days, and 1.98; 19%, 13 days, and 1.03, respectively. Auristatin-PE produced cure in three of five mice, whereas dolastatin 10 showed activity but no cures. When given in combination, auristatin PE + Bryostatin 1-treated animals were all free of tumors (five of five) for 150 days and were considered cured. Dolastatin 10 + Bryostatin 1-treated animals produced cure in only two of five mice. We conclude that: (a) auristatin-PE is more effective in this model than dolastatin 10; (b) auristatin PE can be administered at a concentration 10 times greater than dolastatin 10; (c) there is a synergetic effect between these agents and Bryostatin 1, which is more apparent in the Bryostatin 1 + auristatin PE combination. The use of these agents should be explored clinically in the treatment of CLL.

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