PubMed Health⌕ Search

Biomedical subjects

Michael K Manion

Publications and source records attributed to Michael K Manion.

6 recordsLinked to original sources
PubMed2006

Small-molecule inhibitors of Bcl-2.

Cancer cells with elevated levels of Bcl-2 and the related anti-apoptotic proteins Bcl-x(L), Mcl-1 and Bcl-W are broadly resistant to standard anticancer drugs and other therapeutic modalities. Antisense oligodeoxynucleotides and, more recently, small-molecule ligands for Bcl-2 and Bcl-x(L), sensitize cancer cells to cytotoxic therapies. In some cases, Bcl-2-targeted therapies can function as single therapeutic agents to kill tumor cells, suggesting that Bcl-2 has an important role in the critical functions of cancer cells. The molecular mechanisms of Bcl-2 are not completely understood, therefore, the validation of cytotoxic mechanisms related to Bcl-2 as well as the identification of surrogate markers for Bcl-2 function are significant obstacles for drug development. Despite these problems, two Bcl-2 small-molecule inhibitors are currently undergoing phase I/II clinical trials and several other compounds are in preclinical development. Ongoing studies with these investigational drugs should provide new insights into optimal strategies to disrupt Bcl-2 survival functions to selectively kill cancer cells.

Animals↗
PubMed2005

BCL-XL dimerization by three-dimensional domain swapping.

Dimeric interactions among anti- and pro-apoptotic members of the BCL-2 protein family are dynamically regulated and intimately involved in survival and death functions. We report the structure of a BCL-X(L) homodimers a 3D-domain swapped dimer (3DDS). The X-ray crystal structure demonstrates the mutual exchange of carboxy-terminal regions including BH2 (Bcl-2 homology 2) between monomer subunits, with the hinge region occurring at the hairpin turn between the fifth and sixth alpha helices. Both BH3 peptide-binding hydrophobic grooves are unoccupied in the 3DDS dimer and available for BH3 peptide binding, as confirmed by sedimentation velocity analysis. BCL-X(L) 3DDS dimers have increased pore-forming activity compared to monomers, suggesting that 3DDS dimers may act as intermediates in membrane pore formation. Chemical crosslinking studies of Cys-substituted BCL-X(L) proteins demonstrate that 3DDS dimers form in synthetic lipid vesicles.

Cross-Linking Reagents↗
PubMed2004

A novel approach for monitoring extracellular acidification rates: based on bead injection spectrophotometry and the lab-on-valve system.

Monitoring extracellular acidification rates (ECARs) is important for the study of cellular activities, since it allows for the evaluation of factors that alter metabolic function, such as stimulants, inhibitors, toxins as well as receptor and non-receptor mediated events. While the light addressable potentiometric sensor (Cytosensor Microphysiometer) has been the principal tool for ECARs measurement in the past, this work introduces a novel method that exploits an immobilized pH indicator on the surface of microcarrier beads (Sephadex) and is probed with a fiber optic coupled spectrophotometer. Likewise, live cells under investigation were also immobilized on microcarrier beads (Cytopore). These beads are metered, transported and monitored within a microfluidic system, termed as the Lab-on-Valve (LOV). Use of carrier beads in conjunction with Bead Injection Spectrophotometry and a Lab-on-Valve module (BIS-LOV), makes ECAR measurements reliable and automated. The feasibility of the BIS-LOV approach is demonstrated measuring ECARs of the mouse hepatocyte cell line, TABX.2S, grown on Cytopore beads packed within the central channel of the LOV system. These immobilized cells were perfused in a phosphate buffer carrier solution (capacity: 1 mmol L(-1), pH 7.4). Protons extruded from 10(5) to 10(6) cells were accumulated during a stopped flow period of 220 s followed by a pH measurement, detected by changes in absorbance of the pH indicator bonded to the microcarrier beads. Addition of metabolic inhibitors (sodium azide, oxamic acid) to the carrier buffer solution can induced an increase or decrease of the basal proton extrusion rate in a very reproducible manner. Comparison of the BIS-LOV technique to the Cytosensor microphysiometer and literature confirms the validity of this novel approach, highlighting its advantages and suggesting future improvements that will make the BIS-LOV a practical tool for routine ECARs measurement.

Animals↗
PubMed2003

Bcl-XL mutations suppress cellular sensitivity to antimycin A.

Cells expressing high levels of the BCL-X(L) anti-apoptotic protein are preferentially killed by the mitochondrial inhibitor antimycin A (AA). Computational modeling predicts a binding site for AA in the extended hydrophobic groove on BCL-X(L), previously identified as an interface for dimerization to BAX and related proapoptotic proteins. Here, we identify BCL-X(L) hydrophobic groove mutants with normal cellular anti-apoptotic function but suppressed sensitivity to AA. The LD(50) of AA for cells expressing BCL-X(L) mutants directly correlates with the measured in vitro dissociation constants for AA binding. These results indicate that BCL-X(L) is a principal target mediating AA cytotoxicity.

Amino Acid Sequence↗
PubMedDate not supplied

Targeting BCL-2-related proteins in cancer therapy.

The BCL-2 family proteins are attractive targets for drug design. As pivotal regulators of apoptotic cell death, the logic of manipulating BCL-2 functions for anti-tumor effects is perhaps the strongest for any of the molecular targets proposed for cancer therapeutics. Moreover, elevated levels of anti-apoptotic proteins have been demonstrated in virtually every type of human cancer. BCL2-specific antisense oligonucleotides have shown broad anti-cancer activities in pre-clinical models and are currently in several phase III trials. Rational drug design to manipulate the functions of these proteins has been hampered by the lack of a clear understanding of biochemical or molecular functions. Initial efforts have been centered on disrupting protein-protein interactions within the BCL-2 homology (BH) family. Substantial progress in this task has been made using molecular modeling and drug leads.

Apoptosis↗