6 research outputs found
Oxygen Sensing Difluoroboron Dinaphthoylmethane Polylactide
Dual
emissive properties of solid-state difluoroboron β-diketonate-polyÂ(lactic
acid) (BF<sub>2</sub>bdk-PLA) materials have been utilized as biological
oxygen sensors. Dyes with red-shifted absorption and emission are
important for multiplexing and <i>in vivo</i> imaging, thus
hydroxyl-functionalized dinaphthoylmethane initiators and dye-PLA
conjugates BF<sub>2</sub>dnmÂ(X)ÂPLA (X = H, Br, I) with extended conjugation
were synthesized. The luminescent materials show red-shifted absorbance
(∼435 nm) and fluorescence tunability by molecular weight.
Fluorescence colors range from yellow (∼530 nm) in 10–12
kDa polymers to green (∼490 nm) in 20–30 kDa polymers.
Room-temperature phosphorescence (RTP) and thermally activated delayed
fluorescence (TADF) are present under a nitrogen atmosphere. For the
iodine-substituted derivative, BF<sub>2</sub>dnmÂ(I)ÂPLA, clearly distinguishable
fluorescence (green) and phosphorescence (orange) peaks are present,
making it ideal for ratiometric oxygen-sensing and imaging. Bromide
and hydrogen analogues with weaker relative phosphorescence intensities
and longer phosphorescence lifetimes can be used as highly sensitive,
concentration independent, lifetime-based oxygen sensors or for gated
emission detection. BF<sub>2</sub>dnmÂ(I)ÂPLA nanoparticles were taken
up by T41 mouse mammary cells and successfully detected differences in oxygen levels during <i>in vitro</i> ratiometric imaging
Discovery of Manassantin A Protein Targets Using Large-Scale Protein Folding and Stability Measurements
Manassantin
A is a natural product that has been shown to have
anticancer activity in cell-based assays, but has a largely unknown
mode-of-action. Described here is the use of two different energetics-based
approaches to identify protein targets of manassantin A. Using the
stability of proteins from rates of oxidation technique with an isobaric
mass tagging strategy (iTRAQ-SPROX) and the pulse proteolysis technique
with a stable isotope labeling with amino acids in cell culture strategy
(SILAC-PP), over 1000 proteins in a MDA-MB-231 cell lysate grown under
hypoxic conditions were assayed for manassantin A interactions (both
direct and indirect). A total of 28 protein hits were identified with
manassantin A-induced thermodynamic stability changes. Two of the
protein hits (filamin A and elongation factor 1α) were identified
using both experimental approaches. The remaining 26 hit proteins
were only assayed in either the iTRAQ-SPROX or the SILAC-PP experiment.
The 28 potential protein targets of manassantin A identified here
provide new experimental avenues along which to explore the molecular
basis of manassantin A’s mode of action. The current work also
represents the first application iTRAQ-SPROX and SILAC-PP to the large-scale
analysis of protein–ligand binding interactions involving a
potential anticancer drug with an unknown mode-of-action
Synthesis and Biological Evaluation of Manassantin Analogues for Hypoxia-Inducible Factor 1α Inhibition
To cope with hypoxia, tumor cells
have developed a number of adaptive
mechanisms mediated by hypoxia-inducible factor 1 (HIF-1) to promote
angiogenesis and cell survival. Due to significant roles of HIF-1
in the initiation, progression, metastasis, and resistance to treatment
of most solid tumors, a considerable amount of effort has been made
to identify HIF-1 inhibitors for treatment of cancer. Isolated from <i>Saururus cernuus</i>, manassantins A (<b>1</b>) and B
(<b>2</b>) are potent inhibitors of HIF-1 activity. To define
the structural requirements of manassantins for HIF-1 inhibition,
we prepared and evaluated a series of manassantin analogues. Our SAR
studies examined key regions of manassantin’s structure in
order to understand the impact of these regions on biological activity
and to define modifications that can lead to improved performance
and drug-like properties. Our efforts identified several manassantin
analogues with reduced structural complexity as potential lead compounds
for further development. Analogues <b>MA04</b>, <b>MA07</b>, and <b>MA11</b> down-regulated hypoxia-induced expression
of the HIF-1α protein and reduced the levels of HIF-1 target
genes, including cyclin-dependent kinase 6 (Cdk6) and vascular endothelial
growth factor (VEGF). These findings provide an important framework
to design potent and selective HIF-1α inhibitors, which is necessary
to aid translation of manassantin-derived natural products to the
clinic as novel therapeutics for cancers
Synthesis and Biological Evaluation of Manassantin Analogues for Hypoxia-Inducible Factor 1α Inhibition
To cope with hypoxia, tumor cells
have developed a number of adaptive
mechanisms mediated by hypoxia-inducible factor 1 (HIF-1) to promote
angiogenesis and cell survival. Due to significant roles of HIF-1
in the initiation, progression, metastasis, and resistance to treatment
of most solid tumors, a considerable amount of effort has been made
to identify HIF-1 inhibitors for treatment of cancer. Isolated from <i>Saururus cernuus</i>, manassantins A (<b>1</b>) and B
(<b>2</b>) are potent inhibitors of HIF-1 activity. To define
the structural requirements of manassantins for HIF-1 inhibition,
we prepared and evaluated a series of manassantin analogues. Our SAR
studies examined key regions of manassantin’s structure in
order to understand the impact of these regions on biological activity
and to define modifications that can lead to improved performance
and drug-like properties. Our efforts identified several manassantin
analogues with reduced structural complexity as potential lead compounds
for further development. Analogues <b>MA04</b>, <b>MA07</b>, and <b>MA11</b> down-regulated hypoxia-induced expression
of the HIF-1α protein and reduced the levels of HIF-1 target
genes, including cyclin-dependent kinase 6 (Cdk6) and vascular endothelial
growth factor (VEGF). These findings provide an important framework
to design potent and selective HIF-1α inhibitors, which is necessary
to aid translation of manassantin-derived natural products to the
clinic as novel therapeutics for cancers
Fluorescence Linked Enzyme Chemoproteomic Strategy for Discovery of a Potent and Selective DAPK1 and ZIPK Inhibitor
DAPK1 and ZIPK (also called DAPK3)
are closely related serine/threonine
protein kinases that regulate programmed cell death and phosphorylation
of non-muscle and smooth muscle myosin. We have developed a fluorescence
linked enzyme chemoproteomic strategy (FLECS) for the rapid identification
of inhibitors for any element of the purinome and identified a selective
pyrazoloÂ[3,4-<i>d</i>]Âpyrimidinone (HS38) that inhibits
DAPK1 and ZIPK in an ATP-competitive manner at nanomolar concentrations.
In cellular studies, HS38 decreased RLC20 phosphorylation. In <i>ex vivo</i> studies, HS38 decreased contractile force generated
in mouse aorta, rabbit ileum, and calyculin A stimulated arterial
muscle by decreasing RLC20 and MYPT1 phosphorylation. The inhibitor
also promoted relaxation in Ca<sup>2+</sup>-sensitized vessels. A
close structural analogue (HS43) with 5-fold lower affinity for ZIPK
produced no effect on cells or tissues. These findings are consistent
with a mechanism of action wherein HS38 specifically targets ZIPK
in smooth muscle. The discovery of HS38 provides a lead scaffold for
the development of therapeutic agents for smooth muscle related disorders
and a chemical means to probe the function of DAPK1 and ZIPK across
species
Fluorescence Linked Enzyme Chemoproteomic Strategy for Discovery of a Potent and Selective DAPK1 and ZIPK Inhibitor
DAPK1 and ZIPK (also called DAPK3)
are closely related serine/threonine
protein kinases that regulate programmed cell death and phosphorylation
of non-muscle and smooth muscle myosin. We have developed a fluorescence
linked enzyme chemoproteomic strategy (FLECS) for the rapid identification
of inhibitors for any element of the purinome and identified a selective
pyrazoloÂ[3,4-<i>d</i>]Âpyrimidinone (HS38) that inhibits
DAPK1 and ZIPK in an ATP-competitive manner at nanomolar concentrations.
In cellular studies, HS38 decreased RLC20 phosphorylation. In <i>ex vivo</i> studies, HS38 decreased contractile force generated
in mouse aorta, rabbit ileum, and calyculin A stimulated arterial
muscle by decreasing RLC20 and MYPT1 phosphorylation. The inhibitor
also promoted relaxation in Ca<sup>2+</sup>-sensitized vessels. A
close structural analogue (HS43) with 5-fold lower affinity for ZIPK
produced no effect on cells or tissues. These findings are consistent
with a mechanism of action wherein HS38 specifically targets ZIPK
in smooth muscle. The discovery of HS38 provides a lead scaffold for
the development of therapeutic agents for smooth muscle related disorders
and a chemical means to probe the function of DAPK1 and ZIPK across
species