The ansamycin antibiotic, rifamycin SV, inhibits BCL6 transcriptional repression and forms a complex with the BCL6-BTB/POZ domain

BCL6 is a transcriptional repressor that is over-expressed due to chromosomal translocations, or other abnormalities, in ~40% of diffuse large B-cell lymphoma. BCL6 interacts with co-repressor, SMRT, and this is essential for its role in lymphomas. Peptide or small molecule inhibitors, which prevent the association of SMRT with BCL6, inhibit transcriptional repression and cause apoptosis of lymphoma cells in vitro and in vivo. In order to discover compounds, which have the potential to be developed into BCL6 inhibitors, we screened a natural product library. The ansamycin antibiotic, rifamycin SV, inhibited BCL6 transcriptional repression and NMR spectroscopy confirmed a direct interaction between rifamycin SV and BCL6. To further determine the characteristics of compounds binding to BCL6-POZ we analyzed four other members of this family and showed that rifabutin, bound most strongly. An X-ray crystal structure of the rifabutin-BCL6 complex revealed that rifabutin occupies a partly non-polar pocket making interactions with tyrosine58, asparagine21 and arginine24 of the BCL6-POZ domain. Importantly these residues are also important for the interaction of BLC6 with SMRT. This work demonstrates a unique approach to developing a structure activity relationship for a compound that will form the basis of a therapeutically useful BCL6 inhibitor

Aberration in DNA Methylation in B-Cell Lymphomas Has a Complex Origin and Increases with Disease Severity

Despite mounting evidence that epigenetic abnormalities play a key role in cancer biology, their contributions to the malignant phenotype remain poorly understood. Here we studied genome-wide DNA methylation in normal B-cell populations and subtypes of B-cell non-Hodgkin lymphoma: follicular lymphoma and diffuse large B-cell lymphomas. These lymphomas display striking and progressive intra-tumor heterogeneity and also inter-patient heterogeneity in their cytosine methylation patterns. Epigenetic heterogeneity is initiated in normal germinal center B-cells, increases markedly with disease aggressiveness, and is associated with unfavorable clinical outcome. Moreover, patterns of abnormal methylation vary depending upon chromosomal regions, gene density and the status of neighboring genes. DNA methylation abnormalities arise via two distinct processes: i) lymphomagenic transcriptional regulators perturb promoter DNA methylation in a target gene-specific manner, and ii) aberrant epigenetic states tend to spread to neighboring promoters in the absence of CTCF insulator binding sites

A repurposing strategy for Hsp90 inhibitors demonstrates their potency against filarial nematodes

Novel drugs are required for the elimination of infections caused by filarial worms, as most commonly used drugs largely target the microfilariae or first stage larvae of these infections. Previous studies, conducted in vitro, have shown that inhibition of Hsp90 kills adult Brugia pahangi. As numerous small molecule inhibitors of Hsp90 have been developed for use in cancer chemotherapy, we tested the activity of several novel Hsp90 inhibitors in a fluorescence polarization assay and against microfilariae and adult worms of Brugia in vitro. The results from all three assays correlated reasonably well and one particular compound, NVP-AUY922, was shown to be particularly active, inhibiting Mf output from female worms at concentrations as low as 5.0 nanomolar after 6 days exposure to drug. NVP-AUY922 was also active on adult worms after a short 24 h exposure to drug. Based on these in vitro data, NVP-AUY922 was tested in vivo in a mouse model and was shown to significantly reduce the recovery of both adult worms and microfilariae. These studies provide proof of principle that the repurposing of currently available Hsp90 inhibitors may have potential for the development of novel agents with macrofilaricidal properties

The anti-myeloma activity of a novel purine scaffold HSP90 inhibitor PU-H71 is via inhibition of both HSP90A and HSP90B1

<p>Abstract</p> <p>Background</p> <p>Heat shock protein 90 (HSP90) inhibitors have emerged as a promising class of anti-cancer drugs in both solid and hematologic malignancies. The HSP90 family includes the cytosolic HSP90 (HSP90AA1), the ER paralogue gp96 (HSP90B1) and the mitochondrial member TRAP1 (HSP90L). We evaluated the <it>in vitro </it>anti-tumor activity and mechanism of action of PU-H71, a novel purine scaffold HSP90 inhibitor in human multiple myeloma cell lines.</p> <p>Methods</p> <p>Multiple human myeloma cell lines including cells that are resistant to corticosteroids and bortezimab were treated with PU-H71, followed by analysis of cell viability, cell cycle progression and apoptosis, by flow cytometry and caspase 3 immunoblot. Induction of unfolded protein response was studied by XBP-1 s immunoblot. The role of gp96 was further assessed by small hairpin RNA knockdown of gp96 before treatment with PU-H71.</p> <p>Results</p> <p>PU-H71 has potent <it>in vitro </it>anti-myeloma activity in both drug-sensitive and drug-resistant cell lines. PU-H71 activates the unfolded protein response and induces caspase-dependent apoptosis. The stable gp96 knockdown human myeloma cell line was found to be more resistant to PU-H71 and other HSP90 inhibitors including 17-AAG and 17-DMAG, even though these cells are more sensitive to conventional anti-myeloma drugs.</p> <p>Conclusion</p> <p>We conclude that PU-H71 is a promising drug for the treatment of myeloma. Our finding further suggests that PU-H71 and the geldanamycin analogues work in part by inhibiting the endoplasmic reticulum gp96 along with the cytosolic HSP90.</p

Inhibition of Anaplastic Lymphoma Kinase (ALK) Activity Provides a Therapeutic Approach for CLTC-ALK-Positive Human Diffuse Large B Cell Lymphomas

ALK positive diffuse large B-cell lymphomas (DLBCL) are a distinct lymphoma subtype associated with a poor outcome. Most of them feature a t(2;17) encoding a clathrin (CLTC)-ALK fusion protein. The contribution of deregulated ALK-activity in the pathogenesis and maintenance of these DLBCLs is not yet known. We established and characterized the first CLTC-ALK positive DLBCL cell line (LM1). LM1 formed tumors in NOD-SCID mice. The selective ALK inhibitor NVP-TAE684 inhibited growth of LM1 cells in vitro at nanomolar concentrations. NVP-TAE684 repressed ALK-activated signalling pathways and induced apoptosis of LM1 DLBCL cells. Inhibition of ALK-activity resulted in sustained tumor regression in the xenotransplant tumor model. These data indicate a role of CLTC-ALK in the maintenance of the malignant phenotype thereby providing a rationale therapeutic target for these otherwise refractory tumors

Functional and molecular interactions between ERK and CHK2 in diffuse large B-cell lymphoma

Distinct oncogenic signalling cascades have been associated with non-Hodgkin lymphoma. ERK1/2 signalling elicits both transcriptional and post-transcriptional effects through phosphorylation of numerous substrates. Here we report a novel molecular relationship between ERK1/2 and CHK2, a protein kinase that is a key mediator of the DNA damage checkpoint that responds to DNA double-strand breaks. Our studies are the first to demonstrate the co-localization and overexpression of ERK1/2 and CHK2 in diffuse large B-cell lymphoma (DLBCL). The physical interaction between ERK and CHK2 was highly dependent on phosphorylated Thr 68 of CHK2. Concurrent administration of an ERK inhibitor enhances the antitumour activity of CHK2 inhibition in both a human DLBCL xenograft model as well as primary human DLBCL cells. Our data suggest a functional interaction between ERK and CHK2 and support the potential combined therapeutic targeting of ERK and CHK2 in human DLBCL

The association of hydration status with physical signs, symptoms and survival in advanced cancer-The use of Bioelectrical Impedance Vector Analysis (BIVA) technology to evaluate fluid volume in palliative care: An observational study

Background Hydration in advanced cancer is a controversial area; however, current hydration assessments methods are poorly developed. Bioelectrical impedance vector analysis (BIVA) is an accurate hydration tool; however its application in advanced cancer has not been explored. This study used BIVA to evaluate hydration status in advanced cancer to examine the association of fluid status with symptoms, physical signs, renal biochemical measures and survival. Materials and methods An observational study of 90 adults with advanced cancer receiving care in a UK specialist palliative care inpatient unit was conducted. Hydration status was assessed using BIVA in addition to assessments of symptoms, physical signs, performance status, renal biochemical measures, oral fluid intake and medications. The association of clinical variables with hydration was evaluated using regression analysis. A survival analysis was conducted to examine the influence of hydration status and renal failure. Results The hydration status of participants was normal in 43 (47.8%), 'more hydrated' in 37 (41.1%) and 'less hydrated' in 10 (11.1%). Lower hydration was associated with increased symptom intensity (Beta = -0.29, p = 0.04) and higher scores for physical signs associated with dehydration (Beta = 10.94, p = 0.02). Higher hydration was associated with oedema (Beta = 2.55, p<0.001). Median survival was statistically significantly shorter in 'less hydrated' patients (44 vs. 68 days; p = 0.049) and in pre-renal failure (44 vs. 100 days; p = 0.003). Conclusions In advanced cancer, hydration status was associated with clinical signs and symptoms. Hydration status and pre-renal failure were independent predictors of survival. Further studies can establish the utility of BIVA as a standardised hydration assessment tool and explore its potential research application, in order to inform the clinical management of fluid balance in patients with advanced cancer