Phase II trial of trimelamol in refractory ovarian cancer

Trimelamol is an analogue of hexamethymelamine which exhibited activity against refractory ovarian cancer in phase I clinical trial. The dose limiting toxicity was leukopenia. In a phase II study, 42 patients with recurrent, or platinum-complex resistant, advanced ovarian cancer were treated using the dose schedule 800 mg m-2 i.v. daily for 3 days. There were one complete, three partial and five minor responses, objective response rate: 9.5%. The main toxicity observed was nausea and vomiting, myelosuppression was minor. The role of Trimelamol in the treatment of ovarian cancer remains to be defined, but its activity is limited in refractory disease

Pharmacokinetic evaluation of a novel transdermal ketoprofen formulation in healthy dogs

Dogs undergo various surgical procedures such as castration, ovariohysterectomy, and other orthopedic procedures, which are known to cause inflammation and pain. Non-steroidal anti-inflammatory drugs (NSAIDs) are very effective analgesics for alleviating postoperative pain in veterinary medicine. Ketoprofen is currently approved in Australia and the United States for treating different painful conditions in dogs. This study evaluated the pharmacokinetic parameters of ketoprofen after intravenous (IV) and transdermal (TD) administration in healthy dogs. A novel transdermal ketoprofen (TDK) formulation containing 20% ketoprofen, dissolved in a combination of 45:45% isopropanol and Transcutol, along with 10% eucalyptus oil, was developed and evaluated for in vitro dermal permeation using Franz diffusion cells. A crossover study was then conducted to determine the pharmacokinetic parameters of the formulation in six dogs following IV ketoprofen (1 mg/kg) and TDK (10 mg/kg) administration. A liquid chromatography–mass spectrometry (LC-M/MS) method was used to measure plasma concentrations of ketoprofen over time, and a non-compartmental analysis determined the pharmacokinetic parameters. The mean terminal elimination half-life (T½ h), AUC0-t (µg·h/mL), and mean residence time (MRT, h) between IV and TDK groups were 4.69 ± 1.33 and 25.77 ± 22.15 h, 15.75 ± 7.72 and 8.13 ± 4.28 µg·h/mL, and 4.86 ± 1.81 and 41.63 ± 32.33 h, respectively. The calculated bioavailability (F%) was ~7%, with a lag time of 30 min to achieve effective plasma concentrations after the application of TDK

RNA-binding proteins ZFP36L1 and ZFP36L2 promote cell quiescence.

Progression through the stages of lymphocyte development requires coordination of the cell cycle. Such coordination ensures genomic integrity while cells somatically rearrange their antigen receptor genes [in a process called variable-diversity-joining (VDJ) recombination] and, upon successful rearrangement, expands the pools of progenitor lymphocytes. Here we show that in developing B lymphocytes, the RNA-binding proteins (RBPs) ZFP36L1 and ZFP36L2 are critical for maintaining quiescence before precursor B cell receptor (pre-BCR) expression and for reestablishing quiescence after pre-BCR-induced expansion. These RBPs suppress an evolutionarily conserved posttranscriptional regulon consisting of messenger RNAs whose protein products cooperatively promote transition into the S phase of the cell cycle. This mechanism promotes VDJ recombination and effective selection of cells expressing immunoglobulin-μ at the pre-BCR checkpoint.This work was funded by the Biotechnology and Biological Sciences Research Council, a Medical Research Council CASE studentship with GSK, an MRC centenary award (A.G) and project grants from Bloodwise. DJH was supported by a Medical Research Council Clinician Scientist FellowshipThis is the author accepted manuscript. The final version is available from the American Association for the Advancement of Science via http://dx.doi.org/10.1126/science.aad597

Resistance of stem-like cells from neuroblastoma cell lines to commonly used chemotherapeutic agents

Background Cancer stem cell theory suggests that the presence of tumor initiating stem-like cells in cancers may be responsible for cancer progression and relapse. CD133 cell surface maker expression has been used to identify stem-like cells in cancer cell lines. Our goal was to identify such cells in neuroblastoma cell lines and to study the cytotoxicity of common anticancer drugs for those cells. Materials and Methods CD133+ cells from SK-N-SH and SK-N-BE cell lines were isolated using magnetic microbeads. Cytotoxicity of four anticancer drugs was studied on CD133+ and CD133− populations. The percentage of live, apoptotic, and dead cells in each population after drug treatment was estimated by MTT and PI/Annexin-binding assays. Western blot analyses were used to identify differences in the expression of kinases. Results Eight to 10% of SK-N-SH and 3–5% of SK-N-BE cells were CD133+. These cells were more resistant than CD133− cells to all four chemotherapeutic agents tested in the MTT assay. Decreased apoptosis was observed in CD133+ cells compared to CD133− cells by PI/Annexin V-binding assay. Western blot analysis showed that CD133+ cells expressed less MKP-1. Phosphorylated forms of both ERK and P-38 kinases were expressed at higher levels in CD133+ cells than in CD133− cells. Conclusions This study suggests that CD133+ cells are more resistant to anticancer drugs than CD133− cells. Differences in the expression and phosphorylation of kinases could be partially responsible for this difference. Targeting CD133-expressing cells could be a strategy to develop more effective treatments for neuroblastoma. Pediatr Blood Cancer 2010;54:361–368. © 2009 Wiley-Liss, Inc.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/64907/1/22351_ftp.pd

CCN3: a key growth regulator in Chronic Myeloid Leukaemia

Chronic Myeloid Leukaemia (CML) is characterized by expression of the constitutively active Bcr-Abl tyrosine kinase. We have shown previously that the negative growth regulator, CCN3, is down-regulated as a result of Bcr-Abl kinase activity and that CCN3 has a reciprocal relationship of expression with BCR-ABL. We now show that CCN3 confers growth regulation in CML cells by causing growth inhibition and regaining sensitivity to the induction of apoptosis. The mode of CCN3 induced growth regulation was investigated in K562 CML cells using gene transfection and treatment with recombinant CCN3. Both strategies showed CCN3 regulated CML cell growth by reducing colony formation capacity, increasing apoptosis and reducing ERK phosphorylation. K562 cells stably transfected to express CCN3 showed enhanced apoptosis in response to treatment with the tyrosine kinase inhibitor, imatinib. Whilst CCN3 expression was low or undetectable in CML stem cells, primary CD34+ CML progenitors were responsive to treatment with recombinant CCN3. This study shows that CCN3 is an important growth regulator in haematopoiesis, abrogation of CCN3 expression enhances BCR-ABL dependent leukaemogenesis. CCN3 restores growth regulation, regains sensitivity to the induction of apoptosis and enhances imatinib cell kill in CML cells. CCN3 may provide an additional therapeutic strategy in the management of CML

ERK1/2 signalling protects against apoptosis following endoplasmic reticulum stress but cannot provide long-term protection against BAX/BAK-independent cell death

Disruption of protein folding in the endoplasmic reticulum (ER) causes ER stress. Activation of the unfolded protein response (UPR) acts to restore protein homeostasis or, if ER stress is severe or persistent, drive apoptosis, which is thought to proceed through the cell intrinsic, mitochondrial pathway. Indeed, cells that lack the key executioner proteins BAX and BAK are protected from ER stress-induced apoptosis. Here we show that chronic ER stress causes the progressive inhibition of the extracellular signal-regulated kinase (ERK1/2) signalling pathway. This is causally related to ER stress since reactivation of ERK1/2 can protect cells from ER stress-induced apoptosis whilst ERK1/2 pathway inhibition sensitises cells to ER stress. Furthermore, cancer cell lines harbouring constitutively active BRAFV600E are addicted to ERK1/2 signalling for protection against ER stress-induced cell death. ERK1/2 signalling normally represses the pro-death proteins BIM, BMF and PUMA and it has been proposed that ER stress induces BIM-dependent cell death. We found no evidence that ER stress increased the expression of these proteins; furthermore, BIM was not required for ER stress-induced death. Rather, ER stress caused the PERK-dependent inhibition of cap-dependent mRNA translation and the progressive loss of pro-survival proteins including BCL2, BCLXL and MCL1. Despite these observations, neither ERK1/2 activation nor loss of BAX/BAK could confer long-term clonogenic survival to cells exposed to ER stress. Thus, ER stress induces cell death by at least two biochemically and genetically distinct pathways: a classical BAX/BAK-dependent apoptotic response that can be inhibited by ERK1/2 signalling and an alternative ERK1/2- and BAX/BAK-independent cell death pathway

Erk1/2 activation and modulation of STAT3 signaling in oral cancer

Constitutive activation of the signal transducer and activator of transcription 3 (STAT3) signaling pathway possesses confirmed oncogenic potential in oral squamous cell carcinoma (OSCC). Crosstalk with other molecular pathways contributes to STAT3 regulation in cancer. The effects of mitogen-activated protein kinases (MAPKs) and particularly extracellular signal-regulated kinase 1/2 (Erk1/2) on STAT3 signaling in OSCC have not been thoroughly investigated. The present study examined the effects of Erk1/2 modulation on STAT3 signaling and cell growth in OSCC cells. Constitutive expression levels of phosphorylated (tyrosine and serine) and total STAT3, Erk1/2 and cyclin D1 were assessed in OSCC cell lines. Erk1/2 modulation was achieved by pharmacological agents; siRNA silencing against Erk1/2 was also performed. Cell proliferation and viability were assessed. Erk1/2 inhibition with either U0126 treatment or specific siRNA silencing resulted in decreases in p-ser STAT3 and cyclin D1 levels and increases in p-tyr STAT3 in OSCC cells. Moreover, Erk1/2 inhibition resulted in a dose-dependent reduction in OSCC cell growth and viability. Erk1/2 induction had the opposite effects. Taken together, these results are supportive of an active crosstalk between the oncogenic Erk1/2 and STAT3 pathways in OSCC, the significance of which requires further investigation

ERK1/2 MAP kinases promote cell cycle entry by rapid, kinase-independent disruption of retinoblastoma–lamin A complexes

When in the nucleus, ERK1/2 dislodges the retinoblastoma protein from lamin A, facilitating its rapid phosphorylation

BAY61-3606 Affects the Viability of Colon Cancer Cells in a Genotype-Directed Manner

Background: K-RAS mutation poses a particularly difficult problem for cancer therapy. Activating mutations in K-RAS are common in cancers of the lung, pancreas, and colon and are associated with poor response to therapy. As such, targeted therapies that abrogate K-RAS-induced oncogenicity would be of tremendous value. Methods: We searched for small molecule kinase inhibitors that preferentially affect the growth of colorectal cancer cells expressing mutant K-RAS. The mechanism of action of one inhibitor was explored using chemical and genetic approaches. Results: We identified BAY61-3606 as an inhibitor of proliferation in colorectal cancer cells expressing mutant forms of K-RAS, but not in isogenic cells expressing wild-type K-RAS. In addition to its anti-proliferative effects in mutant cells, BAY61-3606 exhibited a distinct biological property in wild-type cells in that it conferred sensitivity to inhibition of RAF. In this context, BAY61-3606 acted by inhibiting MAP4K2 (GCK), which normally activates NFκβ signaling in wild-type cells in response to inhibition of RAF. As a result of MAP4K2 inhibition, wild-type cells became sensitive to AZ-628, a RAF inhibitor, when also treated with BAY61-3606. Conclusions: These studies indicate that BAY61-3606 exerts distinct biological activities in different genetic contexts