Attenuation Tomography Using Low-Frequency Ultrasound for Thorax Imaging: Feasibility Study.

Low-frequency ultrasound can permeate human thorax and can be applied in functional imaging of the respiratory system. In this study, we investigate the transmission of low-frequency ultrasound through the human thorax and propose a waveform matching method to track the changes in the transmission signal during subject's respiration. The method's effectiveness is validated through experiments involving ten human subjects. Furthermore, the experimental findings indicate that the traveltime of the first-arrival signal remains consistent throughout the respiratory cycle. Leveraging this observation, we introduce an algorithm for ultrasound thorax attenuation factor differential imaging. By computing the paths and energy variation of the first-arrival signal from the received waveform, the algorithm reconstructs the distribution of attenuation factor differences between two different thorax states, providing insights into the functional status of the respiratory system. Numerical experiments, using both normal thorax and defective thorax models, confirm the algorithm's feasibility and its robustness against noise, variations in transducer position and orientation. These results highlight the potential of low-frequency ultrasound for bedside, continuous monitoring of human respiratory system through functional imaging.

A small molecule compound inhibits AKT pathway in ovarian cancer cell lines.

BACKGROUND AND OBJECTIVE: Overactivation of AKT1 and gene amplification of AKT2 are frequently detected in ovarian cancer. Activated AKT kinases provide a cell survival signal that may confer resistance to apoptosis induced by conventional therapies in cancer cells. Therefore, development of potent inhibitors that block AKT pathway is an attractive therapeutic strategy for treating ovarian carcinoma. METHODS: Ovarian cancer cell lines, A2780, MDAH2774, OVCAR-8, Caov-3, and normal murine fibroblasts (NIH3T3) were used. Cells were treated with different doses of a non-peptide small molecule compound, 9-methoxy-2-methylellipticinium acetate (termed API-59-OME) that potentially inhibit AKT pathway. Kinase assays and the phosphorylation of AKT, GSK-3alpha/beta, PDK1, ERK1/2, SGK, p38, FAK, EGFR, JAK2, PKC isoforms, and the cleavage of poly (ADP-ribose) polymerase (PARP) were examined in treated and untreated cell lines. Further, cells treated with API-59-OME were analyzed for induction of apoptosis using sub-G1 profile with propidium iodide staining. RESULTS: API-59-OME inhibited AKT kinase activity but did not inhibit ERK or JNK kinase activities in A2780, MDAH2774, and OVCAR-8 cell lines. API-59-OME did not reduce phosphorylation of other protein kinases in these cell lines. API-59-OME induced apoptosis and the cleavage of PARP in A2780, MDAH2774, and OVCAR-8 ovarian cancer cell lines that express elevated levels of phosphorylated AKT. In contrast, in Caov-3 and NIH3T3 cell lines, which lack constitutive AKT activity, API-59-OME only had minimal effect to induce apoptosis. CONCLUSION: These data suggest that API-59-OME may be a potent agent to target constitutively activated AKT pathway in ovarian cancer cells.

Role of phorbol ester localization in determining protein kinase C or RasGRP3 translocation: real-time analysis using fluorescent ligands and proteins.

The diacylglycerol signaling pathway, involving protein kinase C (PKC) and RasGRP, is a promising therapeutic target for both cancer and other indications. The phorbol esters, ultrapotent diacylglycerol analogues, bind to and activate PKC and RasGRP. Here, using fluorescent phorbol esters and complementary fluorescent PKC and RasGRP constructs, we determined the localization of the phorbol ester as a function of time after addition and how the resultant PKC or RasGRP3 translocation related to ligand localization. For these studies, we prepared fluorescently labeled phorbol esters of varying lipophilicities based on the BODIPY FL (green) or BODIPY 581/591 (red) fluorophores, and by using fusion constructs of green fluorescent protein or DsRed with PKC isoforms or RasGRP3 expressed in Chinese hamster ovary cells, we simultaneously compared the kinetics and pattern of localization of PKC or RasGRP3 with that of the fluorescent red or green phorbol esters. Binding assays showed that the fluorescent derivatives were potent ligands. Uptake followed a one-compartment pharmacokinetic model with a half-time of minutes to hours, depending on the ligand, and all of the fluorescent phorbol esters localized primarily to intracellular membranes, with little plasma membrane localization. The fluorescent phorbol esters induced translocation of and generally colocalized with PKCdelta or RasGRP3. However, PKCalpha and, initially, PKCdelta, translocated to the plasma membrane, in which little phorbol ester accumulated. The findings argue that the rate of uptake of phorbol esters influences the subsequent pattern of PKCdelta translocation, and that the specificity for PKCalpha translocation is dominated by factors other than the localization of the ligand.

Breast cancer cells can evade apoptosis-mediated selective killing by a novel small molecule inhibitor of Bcl-2.

Proteins of the Bcl-2 family are key regulators of caspase activation and apoptosis. Some members of this family, notably Bcl-2 and Bcl-x(L), are overexpressed in cancer cells, which have been associated with chemoresistance. We have designed and synthesized a small molecule inhibitor of Bcl-2, named YC137, and studied its role in cancer cells. In vitro studies showed that YC137 inhibits the binding of the Bid BH3 peptide to Bcl-2, thus disrupting an interaction essential for the antiapoptotic activity of Bcl-2. This inhibitor induces apoptosis of hematopoietic progenitors overexpressing Bcl-2 but not Bcl-x(L) and breast cancer cells that express high levels of Bcl-2. On the contrary, a variety of normal primary cells, including CD34(+) progenitors, myoblasts, and peripheral blood mononuclear cells, do not respond to the inhibitor. A breast cancer cell line resistant to YC137 was generated. Analysis of resistant cells revealed a reduced expression of Bcl-2, which correlated with low activation of signal transducer and activator of transcription-3 (Stat3) and reduced expression of the human epidermal growth factor receptor-2 (HER2). Of note, YC137-resistant cells were more sensitive to apoptosis induced by chemotherapy. Because HER2 has not been linked previously to the Stat3-Bcl-2 transcriptional pathway, we additionally confirmed that specific blockade of HER2 in breast cancer cells resulted in down-regulation of Stat3 activity and reduced levels of Bcl-2. Consistently, HER2 blockade led to YC137 resistance. These data provide evidence for the selective killing of tumor cells by YC137 and represent the first example of in vitro selection of cancer cells refractory to a Bcl-2 inhibitor.

Iron-mediated preparation of vinylcyclopropanecarboxylates: scope, mechanism, and applications.

The addition of stabilized carbon nucleophiles to tricarbonyl(1-methoxycarbonylpentadienyl)iron(1+) cation (1a) proceeds via attack at C2 on the face of the ligand opposite the Fe(CO)(3) group to generate tricarbonyl(pentenediyl)iron complexes 2. Oxidation of complexes 2 affords vinylcyclopropanecarboxylates in good yield. In general, the relative stereochemistry about the cyclopropane ring reflects reductive elimination with retention of configuration. In cases where the C2 substituent is bulky (i.e., 2b) the major cyclopropane product 9b represents ring closure with inversion at C3. A mechanism involving pi-sigma-pi rearrangement of the initially oxidized (pentenediyl)iron species is proposed to account for these results. Experiments which probe the stereochemistry of deuterium labeling in the vinyl group of the vinylcyclopropanecarboxylate products were carried out, and these results are consistent with the proposed mechanism. This methodology for the preparation of vinylcyclopropanecarboxylates was applied to the synthesis of 2-(2'-carboxycyclopropyl)glycines (+)-22 and (-)-23 and the cyclopropane triester (-)-26.