A 14-Protein Signature for Rapid Identification of Poor Prognosis Stage III Metastatic Melanoma.

Purpose: To validate differences in protein levels between good and poor prognosis American Joint Committee on Cancer (AJCC) stage III melanoma patients and compile a protein panel to stratify patient risk.

Experimental Design: Protein extracts from melanoma metastases within lymph nodes in patients with stage III disease with good (n = 16, >4 years survival) and poor survival (n = 14, <2 years survival) were analyzed by selected reaction monitoring (SRM). Diagonal Linear Discriminant Analysis (DLDA) was performed to generate a protein biomarker panel.

Melanoma and the Unfolded Protein Response.

The UPR (unfolded protein response) has been identified as a key factor in the progression and metastasis of cancers, notably melanoma. Several mediators of the UPR are upregulated in cancers, e.g.

Surface Antigen Profiling of Surgical Melanoma Specimens.

An antibody microarray (DotScan) has been developed for profiling clinical melanoma specimens. Immobilized antibodies capture live cells expressing corresponding antigens to produce a dot pattern that represents the surface profile or immunophenotype. The unique signatures obtained may correlate with disease subtype, tumor progression, and clinical outcome.

The Hsp90 inhibitor SNX-7081 is synergistic with fludarabine nucleoside via DNA damage and repair mechanisms in human, p53-negative chronic lymphocytic leukemia.

Clinical trials of heat shock protein 90 (Hsp90) inhibitors have been limited by high toxicity. We previously showed that the Hsp90 inhibitor, SNX-7081, synergizes with and restores sensitivity to fludarabine nucleoside (2-FaraA) in human chronic lymphocytic leukemia (CLL) cells with lesions in the p53 pathway (Best OG, et al., Leukemia Lymphoma 53:1367-75, 2012).

Protein profiles distinguish stable and progressive chronic lymphocytic leukemia.

Patients with a stable chronic lymphocytic leukemia (CLL) double their blood lymphocyte count in >5 years, but may develop progressive disease with lymphocytes doubling in <12 months. To identify a protein signature for progressive CLL, whole cell extracts of peripheral blood mononuclear cells from patients with CLL (n=27) were screened using iTRAQ (isobaric tags for relative and absolute quantification) analysis. A total of 84 differentially abundant proteins were identified from patients with stable and progressive CLL.

Protein signatures correspond to survival outcomes of AJCC stage III melanoma patients.

Outcomes for melanoma patients with stage III disease differ widely even within the same subcategory. Molecular signatures that more accurately predict prognosis are needed to stratify patients according to risk. Proteomic analyses were used to identify differentially abundant proteins in extracts of surgically excised samples from patients with stage IIIc melanoma lymph node metastases.

Mechanisms of action of fludarabine nucleoside against human Raji lymphoma cells.

Fludarabine (2-FaraAMP) is a purine analog that is effective against chronic lymphocytic leukemia (CLL) and non-Hodgkins lymphoma (NHL). For some cases of CLL, 2-FaraAMP as a single agent can clear the blood of leukemia cells, but leukemia stem cells usually remain protected in sanctuary sites. It is clear that 2-FaraAMP has multiple mechanisms of action that may collectively result in strand breaks in DNA, accumulation of phosphorylated p53 and apoptosis.

Surface antigen profiles of leukocytes and melanoma cells in lymph node metastases are associated with survival in AJCC stage III melanoma patients.

There is an urgent need to identify more accurate prognostic biomarkers in melanoma patients, particularly in those with metastatic disease. This study aimed to identify melanoma and leukocyte surface antigens predictive of survival in a prospective series of AJCC stage IIIb/c melanoma patients (n = 29). Live cell suspensions were prepared from melanoma metastases within lymph nodes (LN).

Hsp90 Inhibitor SNX-7081 dysregulates proteins involved with DNA repair and replication and the cell cycle in human chronic lymphocytic leukemia (CLL) cells.

The proteomic effects of the Hsp90 inhibitor, SNX-7081, have been determined on the p53-mutated B-cell chronic lymphocytic leukemia (CLL) cell line, MEC1. Following SNX-7081 treatment (500 nM, 24 h), 51 proteins changed abundance by more than 2-fold (p < 0.05); 7 proteins increased while 44 proteins decreased.

Fludarabine and cladribine induce changes in surface proteins on human B-lymphoid cell lines involved with apoptosis, cell survival, and antitumor immunity.

Fludarabine and cladribine are purine analogues used to treat hematological malignancies. Alone or in combination with therapeutic antibodies, they are effective in treating patients with chronic lymphocytic leukemia and non-Hodgkin's lymphoma. However, the mechanisms of action of these drugs are not well understood.

Fludarabine nucleoside induces accumulations of p53, p63 and p73 in the nuclei of human B-lymphoid cell lines, with cytosolic and mitochondrial increases in p53.

Purpose: Human Raji cells treated with fludarabine nucleoside (2-FaraA, 3 μM) undergo apoptosis with accumulation of p53 in the nuclei as multiple phosphorylated isoforms and C-terminal truncated derivatives. Changes induced by 2-FaraA in the levels of p53, p63 and p73 in the nuclear, cytosolic and mitochondrial fractions have been determined in four human B-lymphoid cell lines that are TP53-functional (Raji and IM9) and TP53-mutated (MEC1 and U266).

Experimental Design: The B-lymphoid cell lines were treated with 2-FaraA (3 μM, 24 h, 48 h) and viability determined.

The phosphoinositide 3-kinase inhibitor LY294002, decreases aminoacyl-tRNA synthetases, chaperones and glycolytic enzymes in human HT-29 colorectal cancer cells.

The proposed anticancer drug LY294002, inhibits phosphoinositide-3 kinase (PI3K) that initiates a signalling pathway often activated in colorectal cancer (CRC). The effects of LY294002 (10 μM, 48 h) on the cytosolic, mitochondrial and nuclear proteomes of human HT-29 CRC cells have been determined using iTRAQ (isobaric tag for relative and absolute quantitation) and tandem mass spectrometry (MS/MS). Analysis of cells treated with LY294002 identified 26 differentially abundant proteins that indicate several mechanisms of action.

Fludarabine nucleoside modulates nuclear "survival and death" proteins in resistant chronic lymphocytic leukemia cells.

The nuclear mechanisms by which fludarabine nucleoside (F-ara-A) induces apoptosis have been investigated in human MEC1 cells derived from B-cell chronic lymphocytic leukemia. Upon treatment of cells with F-ara-A (100 μM, 72 hours), 15 nuclear proteins changed in abundance by more than 2-fold. Nuclear proteins up-regulated included calmodulin (4.

Comprehensive proteomic analysis of the effects of purine analogs on human Raji B-cell lymphoma.

Cladribine (CdA) and fludarabine (FdAMP) are purine analogs that induce apoptosis in chronic lymphocytic leukemia and non-Hodgkin's lymphoma, but the mechanisms are undefined. The effects of CdA and fludarabine nucleoside (FdA) on the cytosolic, mitochondrial, and nuclear proteomes in human Raji lymphoma cells have been determined using two-dimensional fluorescence difference gel electrophoresis (DIGE) and mass spectrometry. Differentially abundant proteins have provided new insights into CdA- and FdA-induced apoptosis.

An extended antibody microarray for surface profiling metastatic melanoma.

An antibody microarray was developed for profiling the surface proteome of melanoma cells, which may facilitate melanoma sub-classification and provide important prognostic information useful in predicting the clinical behavior of the melanoma (e.g., likely sites of metastatic spread), patient outcome and treatment response.

Cladribine and Fludarabine Nucleoside Change the Levels of CD Antigens on B-Lymphoproliferative Disorders.

The purine analogs, fludarabine nucleoside (FdA), and cladribine (CdA) (1 μM, 24 hours), significantly changed the levels of some surface antigens on the human B-cell lines MEC2 and Raji. Changes in the surface proteins were identified using a Cluster of Differentiation (CD) antibody microarray that captures live cells and confirmed by flow cytometry. For Raji cells, CdA up-regulated CD10, CD54, CD80, and CD86, with repression of CD22, while FdA up-regulated CD20, CD54, CD80, CD86 and CD95.