Development of 5-FU-modified tumor suppressor microRNAs as a platform for novel microRNA-based cancer therapeutics.

MicroRNA (miRNAs) are pleiotropic post-transcriptional modulators of gene expression. Their inherently pleiotropic nature makes miRNAs strong candidates for the development of cancer therapeutics, yet despite their potential, there remains a challenge to deliver nucleic acid-based therapies into cancer cells. We developed a novel approach to modify miRNAs by replacing the uracil bases with 5-fluorouracil (5-FU) in the guide strand of tumor suppressor miRNAs, thereby combining the therapeutic effect of 5-FU with tumor-suppressive effect of miRNAs to create a potent, multi-targeted therapeutic molecule without altering its native RNAi function. To demonstrate the general applicability of this approach to other tumor-suppressive miRNAs, we screened a panel of 12 novel miRNA mimetics in several cancer types, including leukemia, breast, gastric, lung, and pancreatic cancer. Our results show that 5-FU-modified miRNA mimetics have increased potency (low nanomolar range) in inhibiting cancer cell proliferation and that these mimetics can be delivered into cancer cells without delivery vehicle both in vitro and in vivo, thus representing significant advancements in the development of therapeutic miRNAs for cancer. This work demonstrates the potential of fluoropyrimidine modifications that can be broadly applicable and may serve as a platform technology for future miRNA and nucleic acid-based therapeutics.

Efficacy of the triple-combination SM-1 in a 5-h phase advance transient insomnia model.

Thomas Dahl, PO Box 404, Guilford, CT 06437 USA. Email: tadahl@outlook.com. The objectives of the study were to demonstrate the efficacy and safety of SM-1 in a circadian challenge model of transient insomnia. Randomized, double-blind, placebo-controlled cross-over study utilizing a 5-h phase advance model of transient insomnia. Subjects were 85 healthy adults reporting a history of transient insomnia, with an average age of 38.9 years. Both SM-1 and placebo were administered to all subjects in a randomly assigned sequence, with at least 1 week between treatments. The primary endpoint was total sleep time determined by polysomnography. Secondary endpoints included wakefulness after sleep onset, latency to persistent sleep, number of awakenings, subjective total sleep time and subjective sleep onset latency, total sleep time by quarters of the night, subjective number of awakenings, and sleep quality. Safety endpoints included adverse events, Karolinska Sleepiness Scale, Digit Symbol Substitution Test, and predischarge evaluation (tandem gait and Romberg tests). SM-1 provided an increase of 94.4 min in total sleep time over placebo (p < 0.0001). Wakefulness after sleep onset, subjective total sleep time, subjective sleep onset latency, and total sleep time in the first quarter of the night also showed significant improvement. SM-1 was well-tolerated with both type and frequency of adverse events being comparable to placebo, and no residual sleepiness upon awakening (i.e., after 8 h). SM-1 provided a robust and statistically significant increase in total sleep time compared to placebo in a circadian model of transient insomnia, without evidence of next-day impairment.

Homogeneous antibody and CAR-T cells with improved effector functions targeting SSEA-4 glycan on pancreatic cancer.

Pancreatic cancer is usually asymptomatic in the early stages; the 5-y survival rate is around 9%; and there is a lack of effective treatment. Here we show that SSEA-4 is more expressed in all pancreatic cancer cell lines examined but not detectable in normal pancreatic cells; and high expression of SSEA-4 or the key enzymes B3GALT5 + ST3GAL2 associated with SSEA-4 biosynthesis significantly lowers the overall survival rate. To evaluate potential new treatments for pancreatic cancer, homogeneous antibodies with a well-defined Fc glycan for optimal effector functions and CAR-T cells with scFv construct designed to target SSEA-4 were shown highly effective against pancreatic cancer in vitro and in vivo. This was further supported by the finding that a subpopulation of natural killer (NK) cells isolated by the homogeneous antibody exhibited enhancement in cancer-cell killing activity compared to the unseparated NK cells. These results indicate that targeting SSEA-4 by homologous antibodies or CAR-T strategies can effectively inhibit cancer growth, suggesting SSEA-4 as a potential immunotherapy target for treating pancreatic disease.

Efficacy of SM-1 in a transient insomnia model.

OBJECTIVES: The objectives of this study were primarily to assess the efficacy and safety of SM-1 in a circadian challenge model of transient insomnia and secondarily, to assess the contribution of diphenhydramine to the combination. METHODS: Randomized, double-blind, placebo-controlled three-way cross-over study with a 5-hr phase advance. Subjects were 39 healthy adults reporting a history of transient insomnia. All treatments (SM-1, SM-1 without diphenhydramine, or placebo) were administered to all subjects in a randomly assigned sequence, with at least 1 week between treatments. The primary endpoint was total sleep time (TST) determined by polysomnography. Secondary endpoints included wakefulness after sleep onset (WASO), latency to persistent sleep, number of awakenings (NAW), subjective TST (sTST) and sleep latency (sSL), TST, and NAW by quarters of the night and sleep quality. Safety endpoints included adverse events, Karolinska Sleepiness Scale digit symbol substitution test, and subject-reported alertness level. RESULTS: SM-1 provided an increase of 126.7 min in TST over placebo (p < .001). WASO, sTST, sleep quality, and sSL also showed significant improvement. Diphenhydramine demonstrated a significant (p = .014) contribution of 43.7 min to TST. SM-1 was well-tolerated with type and frequency of adverse events comparable with placebo, and no residual sleepiness upon awakening after 8 hr. CONCLUSIONS: SM-1 provided a robust and statistically significant increase in TST compared with placebo in a circadian model of transient insomnia, without evidence of next-day impairment. Diphenhydramine contributed to the effect.

Pharmacodynamic and pharmacokinetic profile of SM-1, a triple-drug combination to increase total sleep time.

OBJECTIVE: The primary objective was to characterize the pharmacokinetics and pharmacodynamics of SM-1 after administration of a single oral dose to healthy volunteers in a placebo-controlled double-blind trial of daytime sedation. Secondary objectives were to determine the onset, duration, and offset of the sedative effects using subjective and objective measures of sedation. Safety and tolerability of SM-1 were also investigated. METHODS: Males and females 18-45 years of age received SM-1, a combination drug product comprised of diphenhydramine, zolpidem (delayed release), and lorazepam (delayed release). The pharmacokinetic profile of each drug was determined from blood samples. Sedative effects were assessed by visual analog scale, digit symbol substitution test, memory test, and quantitative electroencephalography. RESULTS: Similar number and severity of adverse events were observed following administration of SM-1 and placebo. Onset of sedation, as determined by subjective, performance, and electroencephalography measures, occurred 0.5-1 hr postdose, lasting about 7-7.5 hr. Plasma concentration curves for the two delayed-release components were altered compared with published data for unmodified drugs. Exposure values obtained with the combination product were in good agreement with published values of the drugs given individually. CONCLUSIONS: SM-1 was well tolerated and has pharmacologic activity starting within an hour of ingestion, lasting approximately 7-8 hr. Sedative activity was seen with subjective, psychomotor, and electroencephalography assays.

Analysis on the clinical features of 507 HDV-infected patients.

The objective of this study was to analyze the clinical feature of hepatitis delta virus (HDV)-infected patients and to discuss the pathological mechanism of hepatitis D. A total of 507 patients with hepatitis due to the infection of HDV were included. The incidence rates of various hepatitis subtypes, the sequelae, the clinical manifestation, the hepatic function, and the hepatic virus makers for all the 507 patients were analyzed statistically. A cohort of 213 patients with hepatitis B, who were also HDV free, served as the control. HDV infection significantly contributed to the increased incidence rate and mortality of severe hepatitis (SH) and cirrhosis (P < 0.01). HDV was also associated with higher incidence rates of hemorrhage in the gastrointestinal tract, abdominal ascites and hepatic encephalopathy, repetitive augmentation of alanine transaminase, and its enhanced magnitude (P < 0.01 or 0.05). The major liver function changes in patients with SH or chronic serious hepatitis was significant compared to the control (P < 0.01). Within the HDV(+) category, HBeAg(-) expression was significantly higher in the HBV DNA(-) group than the HBV DNA(+) group (P < 0.01). The expression of HDAg(+) HBeAg(-) in acute hepatitis, SH, and cirrhosis was significantly higher than that of HDAg(+) HBeAg(+) (P < 0.01 or P < 0.05). The HDV infection was closely associated with the development and prognosis of chronic serious hepatitis, SH, and cirrhosis. HDV infection could inhibit the HBV DNA replication or the HBcAg expression. The direct cytotoxicity of HDV might be the leading pathological factor in AH. HDV might play a major role in the deterioration and chronicization of HDV-co-infected hepatitis B and was responsible for the increased mortality of HBV/HDV patients.

[Differential expression of protease-activated receptor 2 in a bile duct ligation-induced rat model of liver fibrosis].

OBJECTIVE: To observe the dynamic change in expression of protease-activated receptor 2 (PAR2) during onset and progression of liver fibrosis by using a rat model. METHODS: A cholestatic liver fibrosis model was established in Sprague-Dawley rats (aged 8-9 weeks, body weight 350 - 400 g) by bile duct ligation surgery. Rats receiving a sham operation and unoperated rats served as the negative and normal control groups, respectively. At baseline (pre-surgery) and post-surgery weeks 2, 4, 6, and 8, five rats from each group were sacrificed for whole liver resection. The protein and mRNA expressions of PAR2 and collagen I/III were detected by western blotting and RT-PCR, respectively. Between-group differences were assessed by analysis of variance testing. RESULTS: At post-surgery week 2, the liver fibrosis group showed higher expression of PAR2 mRNA and protein than either control group. The expression levels of PAR2 continued to rise over time in the liver fibrosis group (peaking at week 8), and were significantly higher than those detected in the control groups (weeks 4-6: P less than 0.05; week 8, P less than 0.05). A similar trend was observed for the expression of collagen I/III. CONCLUSION: Dynamic expression of PAR2 observed in a cholestatic liver fibrosis rat model may indicate a role for this factor in the formation of liver fibrosis.

The antidepressant sertraline downregulates Akt and has activity against melanoma cells.

Melanoma is a common malignancy which is poorly responsive to chemotherapy and radiation. One of the major reasons melanoma responds poorly to these modalities is constitutive expression of Akt, which protects against apoptosis. The antidepressant sertraline was found to be a potent cytotoxic agent against A375 human melanoma. To determine the mechanism by which sertraline kills melanoma cells, Western blot analysis of signaling molecules, including phosphorylated Akt, caspase 9 and phospho-p70 S6 kinase was performed. Finally, the effects of sertraline on A375 xenografts in mice were assessed. Sertaline potently inhibited the phosphorylation of Akt, and caused cell death through induction of endoplasmic reticulum in vitro. Sertraline monotherapy demonstrated activity against A375 xenografts in vivo. Akt is a major cause of resistance of melanoma to current therapy. Antidepressants are commonly used to prevent interferon-induced depression. Use of antidepressants that decrease Akt may improve the efficacy of interferon and other therapies against melanoma. Further studies are needed to elucidate whether sertraline acts as an Akt inhibitor in melanoma.

Phosphatidylinositol-3-kinase signaling mediates vascular smooth muscle cell expression of periostin in vivo and in vitro.

OBJECTIVE: Periostin is dramatically upregulated in rat carotid arteries after balloon injury. The objective of the present study was to understand mechanisms underlying periostin upregulation in balloon-injured rat carotid arteries and in cultured vascular smooth muscle cells (VSMCs). METHODS AND RESULTS: Periostin protein was strongly expressed at 3 days (in the medial SMCs) and 7 days (in the neointima) after injury. It was also abundantly expressed in the neointima in the late phase (at 14 and 28 days) after injury. Periostin upregulation was mediated through PI-3-kinase-dependent signaling pathway. In vivo, wortmannin, a PI-3-kinase inhibitor, inhibited balloon injury-induced Akt phosphorylation and periostin mRNA expression. In vitro, periostin mRNA expression in cultured VSMCs was stimulated by growth factors (transforming growth factor-beta1 (TGF-beta1), fibroblast growth factors (FGFs), PDGF-BB, and angiotensin II). This stimulatory effect was inhibited by the PI-3-kinase inhibitor LY294002. Further, periostin protein was mostly located in the cytoplasma of VSMCs in culture and abundantly secreted into the culture medium (CM) after stimulation with FGF-2, which significantly promoted VSMC migration in vitro. Immunodepletion of periostin from the VSMC-CM or blockade of periostin function with an anti-periostin antibody significantly reduced VSMC migration. CONCLUSIONS: Upregulation of periostin expression in rat carotid arteries following balloon injury and in cultured VSMCs after stimulation by growth factors is mediated through PI-3-kinase-dependent signaling pathway. Periostin protein secreted by VSMCs plays a significant role in regulating VSMC migration in vitro.

Genome-wide expression analysis of therapy-resistant tumors reveals SPARC as a novel target for cancer therapy.

Overcoming resistance to chemotherapy and radiation therapy has been a difficult but important goal in the effort to cure cancer. We used gene-expression microarrays to identify differentially expressed genes involved in colorectal cancer resistance to chemotherapy and identified secreted protein, acidic and rich in cysteine (osteonectin) (SPARC) as a putative resistance-reversal gene by demonstrating low SPARC expression in refractory human MIP101 colon cancer cells. We were able to achieve restoration of their radiosensitivity and sensitivity to 5-fluorouracil and irinotecan by reexpression of SPARC in tumor xenografts. Moreover, treatment of mice with SPARC conferred increased sensitivity to chemotherapy and led to significant regression of xenografted tumors. The results show that modulation of SPARC expression affects colorectal cancer sensitivity to radiation and chemotherapy. SPARC-based gene or protein therapy may ameliorate the emergence of resistant clones and eradicate existing refractory clones and offers a novel approach to treating cancer.

Periostin induction in tumor cell line explants and inhibition of in vitro cell growth by anti-periostin antibodies.

Several factors have been shown to promote the growth of colorectal cancers. Here, we provide evidence that periostin, a protein with structural and sequence homology with a TGF-beta-inducible gene, beta ig-h3, is upregulated in colorectal cancers and their liver metastasis, and it may play a role in promoting growth in these tumors. In vitro studies reveal that periostin promotes growth and cell proliferation in colorectal cancers and that this effect can be abrogated with antibodies to periostin. Furthermore, exposure of colorectal cancer cells to anti-periostin antibodies activates apoptosis and potentiates the effects of 5-fluorouracil chemotherapy. The results demonstrate the growth-promoting properties of periostin, and a possible role of targeting this protein as a therapeutic option in colorectal cancers.

A rare-cell detector for cancer.

Although a reliable method for detection of cancer cells in blood would be an important tool for diagnosis and monitoring of solid tumors in early stages, current technologies cannot reliably detect the extremely low concentrations of these rare cells. The preferred method of detection, automated digital microscopy (ADM), is too slow to scan the large substrate areas. Here we report an approach that uses fiber-optic array scanning technology (FAST), which applies laser-printing techniques to the rare-cell detection problem. With FAST cytometry, laser-printing optics are used to excite 300,000 cells per sec, and emission is collected in an extremely wide field of view, enabling a 500-fold speed-up over ADM with comparable sensitivity and superior specificity. The combination of FAST enrichment and ADM imaging has the performance required for reliable detection of early-stage cancer in blood.

Reliable and sensitive identification of occult tumor cells using the improved rare event imaging system.

PURPOSE: The purpose of this study was to assess the feasibility of using rare event imaging system (REIS)-assisted analysis to detect occult tumor cells (OTCs) in peripheral blood (PB). The study also sought to determine whether REIS-assisted OTC detection presents a clinically viable alternative to manual microscopic detection to establish the true significance of OTC from solid epithelial tumors. EXPERIMENTAL DESIGN: We recently demonstrated proof of concept using a fluorescence-based automated microscope system, REIS, for OTC detection from the PB. For this study, the prototype of the system was adopted for high-throughput and high-content cellular analysis. RESULTS: The performance of the improved REIS was examined using normal blood (n = 10), normal blood added to cancer cells (n = 20), and blood samples obtained from cancer patients (n = 80). Data from the screening of 80 clinical slides from breast and lung cancer patients, by manual microscopy and by the REIS, revealed that as many as 14 of 35 positive slides (40%) were missed by manual screening but positively identified by REIS. In addition, REIS-assisted scanning reliably and reproducibly quantified the total number of cells analyzed in the assay and categorized positive cells based on their marker expression profile. CONCLUSIONS: REIS-assisted analysis provides excellent sensitivity and reproducibility for OTC detection. This approach may enable an improved method for screening of PB samples and for obtaining novel information about disease staging and about risk evaluation in cancer patients.

Automated detection of immunofluorescently labeled cytomegalovirus-infected cells in isolated peripheral blood leukocytes using decision tree analysis.

BACKGROUND: Cytomegalovirus (CMV) infection continues to be a major problem for immunocompromised patients. Detection of viral antigens in leukocytes (antigenemia assay) is widely used for the diagnosis of CMV infection and for guiding antiviral therapy. The antigenemia technique, contingent upon the manual microscopic analysis of rare cells, is a laborious task that is subject to human error. In this study, we combine automated microscopy with artificial intelligence for reliable detection of fluorescently labeled CMV-infected cells. METHODS: Cytospin preparations of peripheral blood leukocytes were immunofluorescently labeled for the CMV lower matrix phosphoprotein (pp65) and scanned in the Rare Event Imaging System (REIS), a fully automated image cytometer. The REIS detected potential positive objects and digitally recorded 49 measured cellular features for each identified case. The measurement data of these objects were analyzed by the See5 decision tree (DT) algorithm to ascertain whether they were true-positive detections. RESULTS: The DT was built from the measurement data of 2,047 true- and 2,028 false-positive detections, collected from 32 patient samples. By designating misclassifications of false-negatives three times more costly, the 10-fold cross-validation sensitivity, specificity, and misclassification error of the assay was 94.3%, 56.2%, and 25%, respectively. The method was also validated using an independent test set of 21 patient samples, in which similar results were obtained. CONCLUSIONS: To our knowledge, this study represents the first attempt to improve the accuracy of rare event image cytometry through the implementation of artificial intelligence methodology. Results suggest that cost-sensitive decision tree analysis of digitally measured cellular features vastly improves the performance of rare event image cytometry.

Effects of pressure overload on extracellular matrix expression in the heart of the atrial natriuretic peptide-null mouse.

This study tested the hypothesis that atrial natriuretic peptide has direct antihypertrophic actions on the heart by modulating expression of genes involved in cardiac hypertrophy and extracellular matrix production. Hearts of male, atrial natriuretic peptide-null and control wild-type mice that had been subjected to pressure overload after transverse aortic constriction and control unoperated hearts were weighed and subjected to microarray, Northern blot, and immunohistochemical analyses. Microarray and Northern blot analyses were used to identify genes that are regulated differentially in response to stress in the presence and absence of atrial natriuretic peptide. Immunohistochemical analysis was used to identify and localize expression of the protein products of these genes. Atrial natriuretic peptide-null mice demonstrated cardiac hypertrophy at baseline and an exaggerated hypertrophic response to transverse aortic constriction associated with increased expression of the extracellular matrix molecules periostin, osteopontin, collagen I and III, and thrombospondin, as well as the extracellular matrix regulatory proteins, matrix metalloproteinase-2 and tissue inhibitor of metalloproteinase-3, and the novel growth factor pleiotrophin compared with wild-type controls. These results support the hypothesis that atrial natriuretic peptide protects against pressure overload-induced cardiac hypertrophy and remodeling by negative modulation of genes involved in extracellular matrix deposition.

Circulating tumor cells and serum tumor biomarkers in small cell lung cancer.

Lung cancer accounts for approximately 30% of all cancer mortalities in the United States. Small cell lung cancer (SCLC), which is an aggressive malignancy with frequent and early metastases, accounts for about 15% of all of the lung cancer cases with a dismal 5-year survival rate of < 5% with current standard therapies. Early detection of SCLC is challenging, in part due to the lack of adequate serum tumor markers. The goal of this review is to summarize the current knowledge of circulating tumor cells and serum biomarkers in small cell lung cancer. The role of circulating tumor cells in prognostication is controversial, but may be better defined with advancing technologies of detection of such cells with higher precision, and improved clinico-pathological correlations. The current knowledge on the known serum cytokines and tumor biomarkers of SCLC, such as CEA, chromogranin-A and neuron-specific enolase will be presented. Serum cytokines, such as vascular endothelial growth factor (VEGF), stem cell factor (SCF) and hepatocyte growth factor/scatter factor (HGF/SF) are also discussed. New findings in the search for novel diagnostic and therapeutic molecular markers using the emerging genomics and proteomics technologies are emphasized. It is our hope that validation of these new research platforms and technologies will result in improved early detection, prognostication and finally treatment of SCLC with potential novel molecularly-targeted therapeutics.

2-methoxyestradiol alters cell motility, migration, and adhesion.

The effect of 2-methoxyestradiol, 2ME2, an endogenous metabolite of 17beta-estradiol (E2), on cell growth and cytoskeletal functions in a BCR-ABL-transformed cell line model was investigated. We determined the interaction of 2ME2 with STI571 (Gleevec, imatinib mesylate) in STI571 drug-sensitive and -resistant cell lines. In cells expressing BCR-ABL, STI571 cooperated with 2ME2 in reducing cell growth, and STI571-resistant cells were sensitive to 2ME2 treatment. 2ME2 also inhibited growth of several cancer cell lines by a mechanism independent of BCR-ABL. BCR-ABL transformation leads to altered motility, increased adhesion, and spontaneous migration in different in vitro model systems. 2ME2 was found to specifically inhibit the spontaneous motility of BCRABL-transformed Ba/F3 cells and to change the morphology and volume of treated cells. Cells attached to fibronectin-coated surfaces showed a reduced number of filipodia and lamellipodia. In addition, 2ME2 significantly reduced BCRABL-mediated adhesion to fibronectin. The spontaneous migration of BCR-ABL-transformed cells through a transwell membrane also was found to be significantly decreased by 2ME2. Cytoskeletal changes were accompanied by alteration of tubulin formation, distinct from paclitaxel treatment. These results demonstrate that 2ME2 treatment of transformed cells strongly reduces cytoskeletal functions and may also be useful for the treatment of cancers with high metastatic potential. Combination of 2ME2 with other anticancer drugs may be beneficial to treatment of drug-resistant cancers.

Elevated serum periostin levels in patients with bone metastases from breast but not lung cancer.

Periostin is a recently identified gene that is preferentially expressed in periosteum, indicating a potential role in bone formation and maintenance of structure. We independently identified and isolated periostin from cancer tissue, using the palindromic PCR-driven cDNA Differential Display technique. For the present work, we developed a novel sandwich chemiluminescence assay to detect serum periostin level using newly developed monoclonal and polyclonal antibodies. We investigated serum periostin levels in breast cancer and small cell lung cancer patients, especially in patients with bone metastasis. The study included 58 breast cancer and 44 small cell lung cancer patients. Serum periostin levels were elevated in breast cancer patients presenting with bone metastases (92.0 +/- 28.6 ng/ml) compared to similar breast cancer patients without evidence of bone metastasis (55.0 +/- 16.6 ng/ml, p = 0.04). No correlation was found between the serum periostin level and any other prognostic factors, such as clinical stage and lymph node metastasis in breast cancer. Serum periostin levels thus appear to serve as a marker of bone metastasis from breast cancer. In contrast, serum periostin levels were similar in samples from patients with small cell lung cancer who did or did not have bone metastasis. However, increasing T-stage and N-stage of patients with small cell lung cancer were correlated with higher periostin levels (T4, 126.5 +/- 29.7 ng/ml v.s. T2, 64.9 +/- 16.1 ng/ml, p = 0.03; and T4 v.s. T1, 36.3+/- 7.5 ng/ml, p = 0.01; N3, 108.7 +/- 17.3 ng/ml v.s. N2, 49.7+/- 10.9 ng/ml, p = 0.01). Periostin has a substantial homology with the insect cell adhesion molecule, fasciclin I. Thus, expression of periostin may facilitate tumor cell adhesion to the bone surface. In fact, we found by in situ RNA hybridization, that the periostin gene was highly expressed in the stromal cells immediately surrounding the tumor, but not within the breast cancer cells themselves.

Identification of genes regulated by 2-methoxyestradiol (2ME2) in multiple myeloma cells using oligonucleotide arrays.

Our previous study demonstrated that 2-methoxyestradiol (2ME2), an estrogen derivative, induces apoptosis in multiple myeloma (MM) cells; however, the related transcriptional events are unclear. In the present study, we used oligonucleotide microarrays to identify genes altered during 2ME2-induced apoptosis in MM cells. 2ME2 triggers an early transient induction of genes known to trigger cell death and repression of growth/survival-related genes. Many genes regulating cell defense/repair machinery also were transiently induced. Since 2ME2 also induces apoptosis in MM cells resistant to conventional therapies such as dexamethasone (Dex), we compared the gene profiles of 2ME2-treated and Dex-resistant MM cells. Our results suggest that 2ME2 overcomes Dex resistance by modulating genes that confer chemoresistance in MM cells. Microarray results were confirmed by Northern and Western blot analyses. A comparative analysis of selected genes from freshly isolated MM patient cells and 2ME2-treated MM.1S MM cells further provides an in vivo relevance of our in vitro studies. Collectively, these findings suggest genetic events mediating anti-MM activity of 2ME2, as well as mechanisms whereby 2ME2 overcomes Dex resistance in MM cells. These studies may therefore allow improved therapeutic use of 2ME2, based upon targeting genes that regulate MM cell growth and survival.