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Studies have shown a high prevalence (40–83%) of complementary and alternative medicine (CAM) use among cancer patients in the U.S.A cross-sectional, mixed-methods pilot study was completed. This paper focuses on the quantitative analysis conducted on demographic predictors of complementary medicine (CM) use, reasons to use CM, and disclosure to healthcare provider data. Surveys were interview-administered at the Loma Linda University Medical Center Cancer Center. Participants, 18 years or older, were selected from a convenient sample. Eighty-seven percent (87.9%) of participants reported to have used CM as a cancer treatment and most reported to have used it “to help fight the cancer.” Women were eight-times more likely to use prayer. All non-Caucasian and Hispanic participants reported to use CM as a cancer therapy and none reported to use a CM provider. More women (72%) disclosed their CM use than men (53.3%). Different prevalences and predictors exist when differentiating CM modalities, reasons to use CM vary by gender, and disclosure proportions vary by gender.

There is a need for efficacious therapies for metastatic castration-resistant prostate cancer (mCRPC) after disease progression on docetaxel. The SRC tyrosine kinase and its related family members may be important drivers of prostate cancer and can be inhibited by dasatinib. mCRPC patients, after one previous chemotherapy, started dasatinib at 70 mg twice daily, amended to 100 mg daily. The primary endpoint was the disease control (DC) rate, defined as complete response (CR), partial response (PR), or stable disease (SD) in prostate specific antigen (PSA), RECIST, bone scan, and FACT-P score. Up to 41 patients were to be accrued (two-stage design, 21+20) to rule out a null-hypothesized effect of 5 versus 20% (α=0.05, β=0.1). Secondary endpoints included progression-free survival, toxicity, and pharmacokinetic and pharmacodynamic correlatives. Of 38 patients, 27 were evaluable for response or toxicity. The median duration of therapy was 55 days (6-284). Five patients showed DC after 8 weeks of therapy (18.5% DC, 95% CI: 6.3-38.1%). One PR (3.7% response rate, 95% CI: 0.1-19.0%) was observed in a patient treated for 284 days. Twelve patients (43%) discontinued treatment for toxicity. Dasatinib induced a decrease in phytohemagglutinin-stimulated CSF2, CD40L, GZMB, and IL-2 mRNAs in blood cells, indicating target engagement. Decreases in plasma IL-6 and bone alkaline phosphatase, and in urinary N-telopeptide, were associated with DC. Dasatinib has definite but limited activity in advanced mCRPC, and was poorly tolerated. The observation of a patient with prolonged, objective, clinically significant benefit warrants molecular profiling to select the appropriate patient population.

Survivin is a member of the inhibitor of apoptosis (IAP) family and has multifunctional properties that include aspects of proliferation, invasion and cell survival control. Survivin is a promising candidate for targeted cancer therapy as its expression is associated with poor clinical outcome, more aggressive clinico-pathologic features, and resistance to radiation and chemotherapy. In the present review the different properties of the Survivin splice variants are discussed and their activities correlated with different aspects of cancer cell biology, to include subcellular location. Special emphasis is placed on our current understanding of these Survivin splice variants influence on each other and on the phenotypic responses to therapy that they may control.

Summary Objectives Novel targeted therapeutic strategies to overcome radio-resistance of cancer cells traditionally treated with radiation may improve patient survival with the added benefit of reduced systemic toxicity. Herein, we tested the feasibility of Linifanib (ABT-869), a multi-receptor tyrosine kinase inhibitor of members of vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF) receptor families, on radio-sensitization of Head and Neck Squamous Cell Carcinoma (HNSCC). Materials and methods UMSCC-22A and UMSCC-22B cells were treated with Linifanib and γ-radiation response was determined. Cell viability, cytotoxicity, apoptosis induction and cell cycle distribution were examined by MTT assay, colony formation assay and flow cytometry. In addition, expression of STAT3 and downstream signaling proteins were assessed using western immunoblotting. Results Treatment with Linifanib resulted in cell growth inhibition, G2/M cell cycle arrest, induction of cell death via apoptosis, reduced phosphorylation of STAT3, which has been linked to radio-resistance, lower expression of cyclin D1, survivin and increased PARP cleavage. In addition, Linifanib overcame the radio-resistance of the cell lines and significantly enhanced radiation-induced cytotoxicity (p Conclusion These data suggest the possibility of combining targeted therapeutic such as Linifanib with radiation to enhance inhibition of cell growth and apoptosis in HNSCC cells. Thus, it may provide a novel therapeutic strategy and improve efficacy of radiation against HNSCC in the future.

7030Background: B cell acute lymphoblastic leukemia with overexpression of CRLF2 (CRLF2 B-ALL) is a Ph-like leukemia with poor outcome in children, as well as in adults. Activation of the CRLF2 rec...

The influence of adult stem cells on tumor growth is paradoxical. On one hand, angiogenic factors secreted by stem cells are known to be essential for tumor vascularization. On the other hand, stem cell-derived factors can reportedly induce tumor differentiation or direct death of tumor cells. Both the placenta and umbilical cord are rich sources of stem cells with immune modulatory and tissue-healing properties; however, the effects of placental components on cancer cells have not been fully defined. Here we demonstrate that extracts of placental lysates reduce the malignancy of a variety of human tumor cell lines in a species-unrestricted manner. Using a standard model of leukemia cell differentiation, we demonstrated that addition of placental extracts to tumor cells, or co-culture of tumor cells with the CD34(+) cells from umbilical cord blood, induced tumor cell differentiation. Inhibition of tumor growth and metastasis in vivo was also observed following administration of placental extracts. These data support the concept of non-toxic biological therapy of cancer using stem cell derivatives, possibly through the induction of tumor cell differentiation.

Introduction. The high prevalence of complementary and alternative medicine (CAM) use among cancer patients (40%-83%) receiving conventional treatment and the complex relationship between the psychosocial factors that may contribute to or result from CAM use requires further understanding. The authors conducted a descriptive mixed-methods pilot study to understand CAM practices, attitudes, and beliefs among cancer patients at the Loma Linda University Medical Center. Methods. This was the qualitative phase of the study, and no hypotheses were set. A total of 23 face-to-face interviews were conducted, and thematic coding was used to analyze 22 interview transcriptions. There were 14 CAM users (64%) and 8 nonusers (36%). Findings. The themes present among those who used CAM were the following: physicians viewed as one aspect of health care options, a holistic view on well-being, satisfaction with CAM use, and 3 key coping methods (confrontive, supportive, and optimistic) to confront cancer. Themes were not independent of each other. Two themes were present among nonusers; nonusers trusted their physician and were more likely to express evasive coping methods. Discussion. Perceptions and behavioral patterns are complex predictors of CAM use. A better understanding of CAM, medical pluralism, and the perceptions of patients would help health care providers deliver a better quality of care. The promotion of integrative care may help health care providers better identify medical pluralism and would shift focus to patient-centered care.

Cytogenetic studies have previously identified abnormalities of chromosome band 11q23 in many cases of infant acute leukemia. Recent studies by ourselves and others have demonstrated breakpoint clustering in acute leukemias bearing translocations involving 11q23, and a Drosophila trithorax gene homologue (called MLL, HRX, or ALL-1) has been shown to span the 11q23 breakpoints of these translocations. To determine if this gene is affected in infant acute myeloid leukemia (AML), we have analyzed 26 infant AML cases for molecular alterations of this 11q23 gene. 15 out of 26 cases studied (58%) showed rearrangement of the MLL gene at the molecular level, and these rearrangements were clustered within an approximately 11-kb region containing nine exons of this gene. Moreover, 14 of the 15 cases with 11q23 rearrangements (93%) had myelomonocytic or monocytic phenotypes (M4 or M5 FAB subtypes, respectively), both of which are associated with a poor prognosis in childhood AML. In contrast, only 1 of 11 nonrearranged cases had an M4 or M5 phenotype (P = 0.00002). Rearrangement also correlated significantly with hyperleukocytosis (P = 0.02), another clinical parameter associated with poor outcome in this disease. Our results demonstrate that molecular rearrangements of MLL are common in M4 or M5 infant AML, and suggest that alteration of this gene may result in abnormal control of proliferation and differentiation in monocytic progenitor cells.

BORIS, or CTCFL, the so called Brother of the Regulator of Imprinted Sites because of the extensive homology in the central DNA binding region of the protein to the related regulator, CTCF, is expressed in early gametogenesis and in multiple cancers but not in differentiated somatic cells. Thus it is a member of the cancer testes antigen group (CTAs). Since BORIS and CTCF target common DNA binding sites, these proteins function on two levels, the first level is their regulation via the methylation context of the DNA target site and the second level is their distinct and different epigenetic associations due to differences in the non-homologous termini of the proteins. The regulation on both of these levels is extensive and complex and the sphere of influence of each of these proteins is associated with vastly different cellular signaling processes. On the level of gene expression, BORIS has three known promoters and multiple spliced mRNAs which adds another level of complexity to this intriguing regulator. BORIS expression is observed in the majority of cancer tissues and cell lines analyzed up to today. The expression profile and essential role of BORIS in cancer make this molecule very attractive target for cancer immunotherapy. This review summarizes what is known about BORIS regarding its expression, structure, and function and then presents some theoretical considerations with respect to its genome wide influence and its potential for use as a vaccine for cancer immunotherapy.

The slow-releasing hydrogen sulfide (H₂S) donor, GYY4137, caused concentration-dependent killing of seven different human cancer cell lines (HeLa, HCT-116, Hep G2, HL-60, MCF-7, MV4-11 and U2OS) but did not affect survival of normal human lung fibroblasts (IMR90, WI-38) as determined by trypan blue exclusion. Sodium hydrosulfide (NaHS) was less potent and not active in all cell lines. A structural analogue of GYY4137 (ZYJ1122) lacking sulfur and thence not able to release H₂S was inactive. Similar results were obtained using a clonogenic assay. Incubation of GYY4137 (400 µM) in culture medium led to the generation of low (

A single arm phase II trial of single-agent bortezomib (BZM) alone or combined with hormone blockade was conducted in patients with early PSA recurrence after definitive local therapy. The primary endpoint of this study was to determine the time to PSA relapse after BZM therapy alone or when BZM was combined with hormone blockade. The secondary endpoint was to determine the safety of combination therapy. Part A of the treatment schedule consisted of three cycles of BZM 1.3 mg/m2 IV given on days 1,4,8,11. If patients progressed on Part A, they were entered on Part B which consisted of a single dose of LH-RH antagonist, daily oral antiandrogen, and weekly BZM 1.3 mg/m2 for three out of four weeks for a total of three months. BZM treatment significantly decreased the slope of the log PSA (p=0.024) demonstrating that this agent alone was capable of slowing the rise of the PSA. Of eight patients treated with BZM alone five had stable disease, two progressed and 1 went off study secondary to toxicity. The major toxicity was neurotoxicity requiring discontinuation of therapy in three patients and treatment interruption in nine patients. Of those receiving Parts A and B or B only, there were 11 of 15 CRs with the average time to progression of 5.5 months. BZM treatment can change the slope of PSA rise and can be combined with hormone deprivation therapy without significant additional side effects; these agents are associated with a median time to CR of 42 days.

The history of ascorbic acid (AA) and cancer has been marked with controversy. Clinical studies evaluating AA in cancer outcome continue to the present day. However, the wealth of data suggesting that AA may be highly beneficial in addressing cancer-associated inflammation, particularly progression to systemic inflammatory response syndrome (SIRS) and multi organ failure (MOF), has been largely overlooked. Patients with advanced cancer are generally deficient in AA. Once these patients develop septic symptoms, a further decrease in ascorbic acid levels occurs. Given the known role of ascorbate in: a) maintaining endothelial and suppression of inflammatory markers; b) protection from sepsis in animal models; and c) direct antineoplastic effects, we propose the use of ascorbate as an adjuvant to existing modalities in the treatment and prevention of cancer-associated sepsis.

The 3-Deazaneplanocin A (DZNep), one of S-adenosylhomocysteine (AdoHcy) hydrolase inhibitors, has shown antitumor activities in a broad range of solid tumors and acute myeloid leukemia. Here, we examined its effects on multiple myeloma (MM) cells and found that, at 500 nM, it potently inhibited growth and induced apoptosis in 2 of 8 MM cell lines. RNA from un-treated and DZNep treated cells was profiled by Affymetrix HG-U133 Plus 2.0 microarray and genes with a significant change in gene expression were determined by significance analysis of microarray (SAM) testing. ALOX5 was the most down-regulated gene (5.8-fold) in sensitive cells and was expressed at low level in resistant cells. The results were corroborated by quantitative RT-PCR. Western-blot analysis indicated ALOX5 was highly expressed only in sensitive cell line H929 and greatly decreased upon DZNep treatment. Ectopic expression of ALOX5 reduced sensitivity to DZNep in H929 cells. Furthermore, down-regulation of ALOX5 by RNA interference could also induce apoptosis in H929. Gene expression analysis on MM patient dataset indicated ALOX5 expression was significantly higher in MM patients compared to normal plasma cells. We also found that Bcl-2 was overexpressed in DZNep insensitive cells, and cotreatment with DZNep and ABT-737, a Bcl-2 family inhibitor, synergistically inhibited growth and induced apoptosis of DZNep insensitive MM cells. Taken together, this study shows one of mechanisms of the DZNep efficacy on MM correlates with its ability to down-regulate the ALOX5 levels. In addition, DZNep insensitivity might be associated with overexpression of Bcl-2, and the combination of ABT-737 and DZNep could synergistically induced apoptosis. These results suggest that DZNep may be exploited therapeutically for a subset of MM.

The chromosomal translocation, t(4;11)(q21;q23), is the most common type of 11q23 chromosomal abnormality, being highly prevalent in infant acute leukemias and associated with a poor prognosis. The t(4;11) results in the fusion of an 11q23 gene (MLL, HRX, Htrx-1, or ALL-1) and a 4q21 gene (AF-4 or FEL). To further evaluate the 4q21 gene and its role in t(4;11) acute leukemia, we have cloned a 38-kb genomic region and mapped exons of the AF-4 gene. The 4q21 breakpoints in 19 cases of t(4;11) acute leukemia were analyzed by Southern analysis and pulsed- field gels. Seventeen of the 19 cases had breakpoints on chromosome 4q21 that were scattered in this 38 kb region. Expression of the AF-4 gene was studied in a total of 28 various nonhematopoietic, hematopoietic, and t(4;11) leukemic cell lines. The AF-4 gene was expressed in all cell lines as a major and a minor transcript. In addition to the normal transcripts, two fusion transcripts from the derivative 11 and derivative 4 chromosomes were identified in all t(4;11) cell lines except B1, which had only the der(11) transcript. These findings suggest that the breakpoints on 4q21 cluster over a broader area than do the breakpoints in the 11q23 gene, and that der(11) encodes the fusion RNA found consistently in leukemia cells.

Acute lymphoblastic leukemia (ALL) in infants generally shows distinctive biologic features and has a poor prognosis. Cytogenetic studies indicate that many infant leukemias have chromosome 11q23 translocations. Because of these findings and the distinct clinical features of infant leukemia, we investigated 30 cases of infant ALL for molecular defects of 11q23. Fourteen cases had cytogenetic abnormalities of 11q23, and all of them showed 11q23 rearrangements at the molecular level. An additional seven cases also had 11q23 molecular rearrangements, including one with normal cytogenetic analysis. Molecular abnormalities of 11q23 were significantly correlated with adverse prognostic factors, including age under 6 months, hyperleukocytosis, CD10- phenotype, and early treatment failure. Molecular analysis identified a group of infants with germline 11q23 that had a very good treatment outcome with a projected event-free survival of 80% at median follow-up of 46 months compared to 15% in infants with rearranged 11q23 (P < .001). These findings suggest that a high proportion (70%) of infants with ALL have 11q23 rearrangements and that these rearrangements are not always detectable by cytogenetic analysis. The presence of germline 11q23 DNA may identify a subgroup of infant ALL patients with a good outcome using current therapy and a different etiology for their ALL.

Inactivation of tumor-related genes by promoter hypermethylation is a common epigenetic event in the development of variety of tumors. Dermo1 (also called twist2) is a novel cancer-related gene which belongs to the basic helix-loop-helix (bHLH) transcription factor family. Herein, we report that dermo1 expression was sporadically abrogated in human cancer cells by transcriptional silencing associated with CpG island promoter hypermethylation. Direct sequencing of bisulfite-modified DNA from a panel of seven human cancer cell lines (HL60, Molm14, MV4-11, RS4:11, MDM-BA231, H358, and H1299) revealed that CpG dinucleotides in the dermo1 promoter were methylated. RT-PCR results demonstrated that dermo1 CpG island hypermethylation was accompanied by a low basal dermo1 expression level. Our data implicate dermo1 as a tumor suppressor gene and a valuable molecular marker for human cancer.

Pulse-modulated second harmonic imaging microscopes (PM-SHIMs) exhibit improved signal-to-noise ratio (SNR) over conventional SHIMs on sensitive imaging and quantification of weak collagen signals inside tissues. We quantify the spatial distribution of sparse collagen inside a xenograft model of human acute myeloid leukemia (AML) tumor specimens treated with a new drug against receptor tyrosine kinase (ABT-869), and observe a significant increase in collagen area percentage, collagen fiber length, fiber width, and fiber number after chemotherapy. This finding reveals new insights into tumor responses to chemotherapy and suggests caution in developing new drugs and therapeutic regimens against cancers.

WT1 (Wilms' tumor gene 1) overexpression is implicated in the prognosis of acute leukemia. The purpose of this study was to investigate WT1 expression and its clinical implication in childhood acute leukemia (AL) in Chinese population. Bone marrow specimen from 200 children at different stages of acute leukemia and from 21 children without leukemia were studied. The WT1 expression at diagnostic marrow specimen in both acute myeloid leukemia (AML) and acute lymphoid leukemia (ALL) was higher than control group, whereas WT1 expression in AML was higher than in ALL, and WT1 expression level in relapse in ALL increased more significantly than in AML. The WT1 expression level showed positive correlation with the hypodiploidy and BCR-ABL fusion gene in acute leukemia. A rapidly decrease of WT1 expression level predicted a good response to the induction therapy and low expression of WT1 correlates with remission status. This study suggested that WT1 expression levels in acute leukemia can potentially be a marker for evaluating therapeutic efficacy, correlating with monitoring minimal residue disease, and predicting hematological relapse in children acute leukemia.

Mutation of the p53 gene is the most common genetic alteration in human cancers. Our study proposes to rationally design a p53 antisense oligonucleotide (ASO) repository, which contains a series of ASOs containing single nucleotide differences to discriminate between each mutant and wild type (WT) p53. The Sfold software was used to predict target-accessibility and we designed an initial series of antisense oligonucleotides (ASO) that target the p53 mutants A161T, R175H and R249S. Western-blot analysis indicated that ASOs strongly inhibited the expression of p53 mutants in a panel of human tumor cell lines (SNU-449, SK-BR-3 and PLC/PRF/5) while having little effect on the expression of WT p53 (HepG2 cells). In three cancer lines harboring each of the p53 mutations, mutant-specific ASO treatment led to a dose-dependent inhibition of cell growth, cell viability, colony formation and invasion, and expression of mutant p53-dependent survival proteins. Our preliminary results indicate that a single nucleotide difference in ASOs can discriminate between mutant and WT p53. These observations support the hypothesis that a p53 ASO repository can be a potentially valuable tool to knock down oncogenic mutant p53 and warrant the testing of a p53 ASO repository in in vivo settings.

Background Oxaliplatin is a platinum compound used in the treatment of gastrointestinal malignancies, including colorectal cancer. The incidence of hypersensitivity reaction in patients receiving oxaliplatin is approximately 15%, with severe reaction (grade 3 and 4) occurring in 2% of patients. Case presentation We report two patients with metastatic colorectal cancer who developed de novo hypersensitivity reaction and acute thrombocytopenia after oxaliplatin infusion. Both patients had oxaliplatin treatment several years before and exhibited hypersensitivity on the third dose of oxaliplatin in recent treatment. Oxaliplatin was discontinued when clinical reaction was identified. Both patients were confirmed to have strong oxaliplatin-induced IgG platelet-reactive antibodies. Both patients' thrombocytopenia resolved within two weeks after discontinuation of oxaliplatin. One patient had disease stabilization lasting for three months without chemotherapy. Both patients subsequently received other chemotherapeutic agents without evidence of hypersensitivity reaction or immune-mediated thrombocytopenia. Conclusion We recommend vigilant monitoring of complete blood count and signs and symptoms of bleeding after the occurrence of oxaliplatin-induced hypersensitivity to avoid serious complications of immune-mediated thrombocytopenia.

The authors report a case of orbital plasmacytoma in a 48-year-old man with known multiple myeloma. He presented with proptosis, diplopia, and decreased vision of the left eye for several weeks. He had been previously treated for IgA lambda multiple myeloma with chemotherapy, radiation, and autologous stem cell transplant. After a left orbitotomy, flow cytometry revealed a tumor rich in plasma cells expressing CD138 with equivocal lambda light chain expression. The patient underwent orbital radiation, with improvement of vision and disc edema OS. The patient is currently undergoing salvage chemotherapy for relapse of multiple myeloma. This is the third reported case of IgA myeloma involving the orbit.

Chromosome 11 band q23 is commonly involved in nonrandom chromosomal translocations in hematopoietic malignancies, especially in infant acute leukemias. By using pulsed-field gel electrophoresis (PFGE) with restriction endonuclease digests of DNA from both a leukemia cell line (RS4;11) bearing the t(4;11)(q21;q23) and from human/hamster hybrid cells, we have been able to construct a detailed restriction map of the chromosome 11q23 region and have localized the t(4;11) chromosome 11 breakpoint to a region located approximately 200 to 230 kb telomeric to the CD3 gamma region and approximately 580 kb centromeric to the PBGD gene. PFGE analyses of DNA from clinical leukemia specimens and cell lines indicated a tight clustering of breakpoints in all eight t(4;11) acute leukemias studied. These data strongly suggest that discrete genetic loci are interrupted on both chromosomes 4 and 11 in a manner likely to be critically involved in the pathogenesis of t(4;11) acute leukemias. To our knowledge, these results represent the first evidence of breakpoint clustering in t(4;11) acute leukemias. In contrast to t(4;11), other 11q23 abnormalities studied to date have frequently shown evidence for alternative breakpoint sites in 11q23.

Natural phytoestrogens such as the isoflavones genistein and daidzein, and the flavones quercetin exhibit anti-cancer properties. This study was purpose to investigate the anti-proliferative effect of phytoestrogens on acute myeloid leukemia (AML) and chronic myeloid leukemia (CML) cells, and their synergistic antileukemic effect in combination with chemotherapeutic drugs. Optimal dosage of genistein, quercetin and in combination with chemicals for leukemia cells were determined by experiments. Cell viability, apoptosis induction and cell cycle arrest were detected by trypan blue staining, MTT assay, optical microscopy, flow cytometry (FCM). The schedule treatment of combination of genistein and chemicals was determined. The results showed that genistein exhibited a dose- and time-dependent inhibitory effect on cell proliferation in NB4 and HL-60 cells, induced apoptosis and cell cycle arrest in G2/M phase. Quercetin had evident inhibitory effect on the proliferation of K562 and K562/A cells. The combination of genistein and chemicals exerted a synergistic effect on cell growth inhibition. In conclusion, this study demonstrated the synergistic antileukemic effect of genistein with chemotherapeutic drugs on leukemic cells. This combination appears to be a new idea for the clinical novel treatment of leukemia.

In vitro experiments have shown the PIM1 kinase to have diverse biological roles in cell survival, proliferation and differentiation. In humans, PIM1 is often expressed in both normal and transformed cells. The PIM1 kinase is a true oncogene implicated in early transformation and tumour progression in haematopoietic malignancies and prostate carcinomas. It is associated with aggressive subgroups of lymphoma, is a marker of poor prognosis in prostate carcinomas and has been suggested to have a role in hormone insensitivity of prostate malignancies. PIM1 has a possible role in other carcinomas with 6p21 genomic alterations. On one hand, PIM1 (due to its role in malignancy) appears to be a promising target for drug development programmes but, on the other hand, the complexity of its molecular structure has posed challenges in the development of PIM1 inhibitors. In this review we discuss PIM1 expression in human tissues (including some new data from our laboratory), its role in human malignancies, as well as the possibilities and challenges in the development of target therapy for PIM1.

Understanding the molecular etiology and heterogeneity of disease has a direct effect on cancer therapeutics. To identify novel molecular changes associated with breast cancer progression, we conducted phosphoproteomics of the MCF10AT model comprising isogenic, ErbB2- and ErbB3-positive, xenograft-derived cell lines that mimic different stages of breast cancer. Using in vitro animal model and clinical breast samples, our study revealed a marked reduction of epidermal growth factor receptor (EGFR) expression with breast cancer progression. Such diminution of EGFR expression was associated with increased resistance to Gefitinib/Iressa in vitro. Fluorescence in situ hybridization showed that loss of EGFR gene copy number was one of the key mechanisms behind the low/null expression of EGFR in clinical breast tumors. Statistical analysis on the immunohistochemistry data of EGFR expression from 93 matched normal and breast tumor samples showed that (a) diminished EGFR expression could be detected as early as in the preneoplastic lesion (ductal carcinoma in situ) and this culminated in invasive carcinomas; (b) EGFR expression levels could distinguish between normal tissue versus carcinoma in situ and invasive carcinoma with high statistical significance (P < 0.001, n = 81). However, no significant correlation of EGFR expression with disease-free survival and overall survival was observed. This is the first time EGFR expression has been tracked meaningfully and developmentally from the normal condition through disease progression using in vitro, xenograft, and matched normal and tumor samples. Thus, our study provides a new insight into the role of EGFR in breast cancer development. Although no value of EGFR expression in prognosis was found, our findings are likely to have implications in the design of clinical trials targeting the EGFR family of proteins in breast cancer. [Mol Cancer Ther 2007;6(11):2828–42]

Neoangiogenesis plays an important role in leukemogenesis. We investigated the in vivo anti-leukemic effect of ABT-869 against AML with wild-type FLT3 using RFP transfected HL60 cells with in vivo imaging technology on both the subcutaneous and systemic leukemia xenograft models. ABT-869 showed a five-fold inhibition of tumor growth in comparison with vehicle control. IHC analysis revealed that ABT-869 decreased p-VEGFR1, Ki-67 labeling index, VEGF and remarkably increased apoptotic cells in the xenograft models. ABT-869 also reduced the leukemia burden and prolonged survival. Our study supports the rationale for clinically testing an anti-angiogenesis agent in AML with wild-type FLT3.

More effective therapeutic strategies are required for patients with poor-prognosis systemic sclerosis (SSc). A phase 2 single-arm study of high-dose immunosuppressive therapy (HDIT) and autologous CD34-selected hematopoietic cell transplantation (HCT) was conducted in 34 patients with diffuse cutaneous SSc. HDIT included total body irradiation (800 cGy) with lung shielding, cyclophosphamide (120 mg/kg), and equine antithymocyte globulin (90 mg/kg). Neutrophil and platelet counts were recovered by 9 (range, 7 to 13) and 11 (range, 7 to 25) days after HCT, respectively. Seventeen of 27 (63%) evaluable patients who survived at least 1 year after HDIT had sustained responses at a median follow-up of 4 (range, 1 to 8) years. There was a major improvement in skin (modified Rodnan skin score, −22.08; P < .001) and overall function (modified Health Assessment Questionnaire Disability Index, −1.03; P < .001) at final evaluation. Importantly, for the first time, biopsies confirmed a statistically significant decrease of dermal fibrosis compared with baseline (P < .001). Lung, heart, and kidney function, in general, remained clinically stable. There were 12 deaths during the study (transplantation-related, 8; SSc-related, 4). The estimated progression-free survival was 64% at 5 years. Sustained responses including a decrease in dermal fibrosis were observed exceeding those previously reported with other therapies. HDIT and autologous HCT for SSc should be evaluated in a randomized clinical trial.

Myelodysplastic syndromes (MDSs) and myeloproliferative neoplasms (MPNs) are subgroups of myeloid neoplasms with chronic clinical courses. MDSs are characterized by simultaneous proliferation and apoptosis of hematopoietic cells and peripheral blood cytopenias. MDS with isolated del(5q) is a specific subtype of MDS that occurs primarily in older women, and is characterized by macrocytic anemia, increased platelet count, micromegakaryocytes, and favorable clinical course. …

We have recently reported that accumulation of misfolded nuclear hormone receptor corepressor (N-CoR) as insoluble protein aggregates in acute promyelocytic leukemia (APL) cells induces endoplasmic reticulum (ER) stress and activates unfolded protein response (UPR). Although accumulation of misfolded proteins is known to trigger UPR-induced cytotoxic cell death in several neurodegenerative disorders, APL cells are notably resistant to UPR-induced apoptosis. The molecular basis for the paradoxical response of APL cells to UPR is not known. Here, we report that a glycoprotease, selectively expressed in APL cells, regulates the response of APL cells to UPR-induced apoptosis through processing of misfolded N-CoR protein. Results show that misfolded N-CoR is cleaved selectively in APL cells, and cellular extracts of APL cells and human primary APL cells contain activity that cleaves N-CoR protein. Purification and spectrometric analysis of N-CoR cleaving activity from an APL cell line reveals that it is a glycoprotein endopeptidase known as OSGEP. Furthermore, the cleavage of N-CoR in APL cells could be blocked by the broad-spectrum protease inhibitor AEBSF and by RNA interference–mediated down-regulation of OSGEP expression. AEBSF selectively inhibits growth and promotes apoptosis of APL cells possibly through a mechanism involving AEBSF-induced accumulation of insoluble N-CoR protein and by triggering ER stress. Taken together, these findings suggest that selective induction of protease activity in APL cells may represent a novel cytoprotective component of UPR, which could be exploited by tumor cells to survive the toxic insult of misfolded protein(s). (Cancer Res 2006; 66(20): 9903-12)

Introduction Acute Myeloid Leukemia (AML) is a heterogeneous disease of myeloid progenitor cells where clonal expansion of these cells overwhelms the bone marrow leading to hematopoietic failure. Interest in identifying disruption in key regulatory pathways has led to potential for novel therapies. One particular cell signaling pathway, namely that of AKT (PKB), has been shown to correlate with AML prognosis. It has been shown previously that Protein Phosphatase 2A (PP2A) is responsible for the dephosphorylation of AKT at Thr 308 leading to decreased AKT activity. PP2A is a heterotrimeric complex consisting of a catalytic (C), scaffolding (A) and one of many regulatory (B) subunits. Studies have demonstrated that the regulatory subunit, B55α, is responsible for targeting the PP2A holoenzyme to AKT to dephosphorylate Thr 308 (Kuo et al, 2008). In fact, B55α expression has been shown to be inversely correlated with AML prognosis and Thr 308 phosphorylation of AKT (Ruvolo et al, 2011). Despite this fact, no studies have specifically demonstrated a mechanism whereby B55 Alpha expression is regulated in AML. In addition, although the potential exists for targeted therapy in patients through either activation of B55α-PP2A or disruption of AKT activity, the effectiveness of these therapies in AML has yet to be defined. In the following study we demonstrate novel loss of function mutations in the B55α gene identified in 3 AML patient samples out of 12 tested, and their effect on downstream cellular signaling pathways, including AKT, showing a possible mechanism by which B55α may act as a tumor suppressor in myeloid cells. Results Both genomic and cDNA were analyzed from leukophoresis samples of 12 unselected patients with known diagnosis of AML whose clinical data remained unknown to the primary investigator until sequencing results were completed. Two novel B55α mutations were identified in 3 of the patient samples. In 2 samples, the same nonsense mutation was identified which led to a premature stop codon at codon 77. Interestingly both of these patients had M1 AML with FLT3 ITD positivity and NPM1 mutation positivity as well as >90% blasts on marrow smear at presentation. The other mutation was a single base pair deletion leading to a frame shift causing a premature stop codon at codon 25. This patient had residual AML NOS after partial treatment and was negative for both FLT3 and NPM1 mutations. B55α protein expression was undetectable in these 3 samples, but present in the other non-mutant samples (Figure 1A). Another regulatory B subunit, B56γ, was unaffected in all samples. Lack of B55α expression correlated with loss of B55α-PP2A complexes and loss of PP2A-AKT interaction as well as increased levels in AKT Thr308 phosphorylation (Figure 1B). Samples containing B55α mutations demonstrated enhanced sensitivity to the AKT inhibitor MK2206 (Figure 2), a compound currently undergoing evaluation as a cancer chemotherapeutic in clinical trials (Yap et al, 2011). In addition, these samples were less sensitive to the PP2A activator FTY720 (Neviani et al, 2007). Conclusion Novel AML-associated mutations were identified in the B55α subunit of PP2A. The mutations led to loss of B55α protein expression and loss of B55α specific PP2A complexes within the cell. Loss of B55α PP2A disrupted interaction between PP2A and AKT and led to increased phosphorylation of AKT at Thr 308, a residue important for AKT activation. It has been previously shown that decrease in B55α expression correlates with recurrence of AML, however here we demonstrate novel loss of function mutations within the B55α gene that have the significant consequence of activating a known oncogenic protein, AKT. These samples demonstrate enhanced sensitivity and responsiveness to AKT inhibition by MK2206 compared to cells with intact B55α protein. Taken together our data illustrate a significant pathway in leukemogenesis involving one specific subset of PP2A holoenzymes. In addition, our data suggest that the presence of B55α loss of function mutations may enhance tumor responsiveness to targeted therapy by AKT inhibition. Further studies are required to evaluate the clinical efficacy of such therapies in vivo, however the results indicate a novel molecular targeting approach for optimizing AML therapy. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.

High-dose immunosuppressive therapy followed by autologous hematopoietic stem cell transplantation (HSCT) is currently being evaluated for the control of severe autoimmune diseases. The addition of antithymocyte globulin (ATG) to high-dose chemoradiotherapy in the high-dose immunosuppressive therapy regimen and CD34 selection of the autologous graft may induce a higher degree of immunosuppression compared with conventional autologous HSCT for malignant diseases. Patients may be at higher risk of transplant-related complications secondary to the immunosuppressed state, including Epstein-Barr virus (EBV)-associated posttransplantation lymphoproliferative disorder (PTLD), but this is an unusual complication after autologous HSCT. Fifty-six patients (median age, 42 years; range, 23–61 years) with either multiple sclerosis (n = 26) or systemic sclerosis (n = 30) have been treated. The median follow-up has been 24 months (range, 2–60 months). Two patients (multiple sclerosis, n = 1; systemic sclerosis, n = 1) had significant reactivations of herpesvirus infections early after HSCT and then developed aggressive EBV-PTLD and died on days +53 and +64. Multiorgan clonal B-cell infiltrates that were EBV positive by molecular studies or immunohistology were identified at both autopsies. Both patients had positive screening skin tests for equine ATG (Atgam) and had been converted to rabbit ATG (Thymoglobulin) from the first dose. Of the other 54 patients, 2 of whom had partial courses of rabbit ATG because of a reaction to the intravenous infusion of equine ATG, only 1 patient had a significant clinical reactivation of a herpesvirus infection (herpes simplex virus 2) early after HSCT, and none developed EBV-PTLD. The T-cell count in the peripheral blood on day 28 was 0/μL in all 4 patients who received rabbit ATG; this was significantly less than in patients who received equine ATG (median, 174/μL; P = .001; Mann-Whitney ranked sum test). Although the numbers are limited, the time course and similarity of the 2 cases of EBV-PTLD and the effect on day 28 T-cell counts support a relationship between the development of EBV-PTLD and the administration of rabbit ATG. The differences between equine and rabbit ATG are not yet clearly defined, and they should not be considered interchangeable in this regimen without further study.

S100A8 and S100A9 are two members of the S100 calcium-binding protein family, preferentially form functional heterodimers of S100A8/S100A9, and have been increasingly recognized as biomarkers in malignancies. Recent proteomic studies revealed that S100A8 and S100A9 played pivotal roles in hematologic malignancies and elevated expression of S100A8/S100A9 implicated in glucocorticoid resistance in MLL-rearranged infant acute lymphoblastic leukemia (ALL). In addition, S100A8 proteomic expression in leukemic cells was reported to predict survival in AML patients. However, information on the gene expression level of S100A8 and S100A9 and their clinical correlation in acute myeloid leukemia (AML) is lacking. Using the real-time quantitative RT-PCR, we analyze the transcription levels of S100A8 and S100A9 in AML patient leukemic specimens underwent pre-planned induction chemotherapy. A total of 189 patient cases at different stages of AML (excluding acute promyelocytic leukemia [APL]), including 91 newly diagnosed AML, 64 patient remission marrow specimens and 34 patient specimens as well as 20 controls without leukemia were included in the study collected from the period between 2007 and 2011. Among the cohort of 91 newly diagnosed AML, the over all median OS was 20 months, and the median follow-up for survivors was 24 months (range: 17 to 60 months). There were significant positive correlations of transcription levels between S100A8 and S100A9 in AML patients from different stages. The expression levels of S100A8 and S100A9 in newly diagnosed and relapsed AML patients revealed no significant difference, but were both lower than those in complete remission and control group. Patients with high transcription level of S100A8 and S100A9 were predominantly in AML with myelo-monocytic differentiation (M4, M5) whereas those with low transcription level of S100A8 and S100A9 often showed more immature cytomorphology (M0, M1), erythrocytic or megakaryocytic differentiation. The subgroup of patient with high transcription level of S100A8 could be a predictor for inferior overall survival (OS) (P = 0.0012). High levels of transcription for both S100A8 and S100A9 in de novo AML patients could predict shorter OS than those with low levels after adjustment on their ages at diagnosis (P = 0.003). In a multivariate analysis for OS, high S100A8 transcription was a significant prognostic factor (P =0.001) after analysis adjustment for age (P = 0.019), bone marrow blast percentage (P = 0.04) and cytogenetic classification (P = 0.05) at diagnosis. Using a combination of S100A8 transcription level and cytogenetic risk classification, survival analysis gave result that the new stratification was highly correlated with the OS (P < 0.0001). With significantly different OS, patients from the intermediate-risk group can be divided into two subgroups (IH = cytogenetically intermediate-risk with S100A8 high transcription and IL = cytogenetically intermediate-risk with S100A8 low transcription). Patients from group IL emerged with a probability of OS similar to the cytogenetically favorable-risk group, whereas the survival curve of IH subgroup was close to the unfavorable-risk group. (Figure)FigureRisk stratification of de novo AML patients by a combination of age and cytogenetic characteristics with S100A8 expression levels. 1H = cytogenetically favorable-risk with S100A8 high expression, 1L = cytogenetically favorable-risk with S100A8 low expression, 2H = cytogenetically intermediate-risk with S100A8 high expression, 2L = cytogenetically intermediate-risk with S100A8 low expression, 3H = cytogenetically unfavorable-risk with S100A8 high expression, 3L = cytogenetically unfavorable-risk with S100A8 low expression.Figure. Risk stratification of de novo AML patients by a combination of age and cytogenetic characteristics with S100A8 expression levels. 1H = cytogenetically favorable-risk with S100A8 high expression, 1L = cytogenetically favorable-risk with S100A8 low expression, 2H = cytogenetically intermediate-risk with S100A8 high expression, 2L = cytogenetically intermediate-risk with S100A8 low expression, 3H = cytogenetically unfavorable-risk with S100A8 high expression, 3L = cytogenetically unfavorable-risk with S100A8 low expression. In conclusion, the transcription levels of S100A8 and S100A9 were significantly associated with development and prognosis of AML. Both S100A8 and S100A9 expression levels provided useful clinical information, and more importantly, S100A8 expression level significantly correlates with prognosis in addition to well-known cytogenetic risk factors, and could potentially further refine current stratification of de novo AML patients. Disclosures: No relevant conflicts of interest to declare.

Abstract 1447 Insulin-like growth factor binding protein 7 (IGFBP7) has been ascribed properties of both tumor suppressor and enhancer of cell proliferation. In solid tumors the important role of IGFBP7 as a tumor suppressor was revealed in several studies. In acute T-lymphoblastic leukemia (T-ALL), high IGFBP7 expression is associated with a more immature phenotype of early T-ALL, inferior survival, and predicts primary chemotherapy resistance. In a separate study, IGFBP7 acts as a positive regulator of ALL and bone marrow stromal cells growth, and significantly enhances in-vitro resistance to asparaginase. Higher IGFBP7 mRNA levels were associated with lower leukemia-free survival (P=0.003) in precursor B-cell Ph negative ALL patients (n=147) treated with a contemporary polychemotherapy protocol. In acute myeloid leukemia, the role of IGFBP7 is largely unknown. In our previous published study [Hu et al, 2011], we demonstrated that IGFBP7 overexpressed in majority of childhood AML (n=66) at diagnosis and upon relapsed, but not at remission stage. We now further explore its mechanism in promoting AML cells proliferation. Compared with control, transfection of full length IGFBP7 in K562 cells [V-BP7] resulted in 23% increased of proliferation in 48 hours. Cell cycle analysis by flow cytometry showed decreased G0/G1 phase and increased S phase in V-BP7 comparing with control, suggesting enhanced cell cycle progression. While transfection of IGFPB7 siRNA produced an opposite effect of reducing the cell growth in K562 cells. In consistent with the nature of a secretory protein, the extracellular IGFBP7 level in the condition media from v-BP7 was significantly higher than that from vector control or parental K562 cells measured by ELISA. Incubation parental K562 cells in V-BP7 derived conditioned medium resulted in significant growth enhancement. Gene expression profiling (GEP) was performed on V-BP7 in contrast to parental K562 cells. Genes which were up-regulated or down-regulated more than 2 folds were regarded as significant difference. Among 10 verified genes, AKT3 showed the highest extent of up-regulation and IGFBP7 siRNA transfection reduced its expression. Cyclin D1 (CCND1) expression was also significantly up-regulated and validated by RT-PCR and Western blot. V-BP7 treated with an AKT inhibitor (Triciribine) at 2.5μM for 72 hours showed 50% reduction of cell viability. The cell cycle analysis indicated that triciribine reversed cell cycle progression in V-BP7, by increasing cells in G0/G1 phase and reducing cells in S phase. Western blot demonstrated that both phospho-AKT3 and CCND1 were down regulated after treatment with triciribine. Using real time RT-PCR, we further identified that IGFBP7 and AKT3 expression were significantly correlated (p=0.001; r=0.255) in 39 newly diagnosed childhood AML. Conclusions IGFBP7 aberrantly overexpressed in majority of childhood AML. IGFBP7 promotes proliferation of K562 cells and AML via overexpression/activation of AKT3 and CCND1. Disclosures: No relevant conflicts of interest to declare.

A chromosome band 4q21 gene (MLLT2, formerly called AF-4/FEL) involved in a reciprocal translocation with chromosome band 11q23 in t(4;11) acute leukemia has been cloned. To provide better definition of gene order and relationships in this region where MLLT2 resides, we used pulsed field gel electrophoresis (PFGE) to investigate 13 genes (including MLLT2) with physical locations in bands 4q11→q25. Somatic cell hybrids derived from RS4; 11 a leukemic cell line carrying the t(4; 11 )(q21 ;q23), were also used to localize genes in relation to MLLT2. Linkage of the interleukin 8 (IL8), albumin (ALB), and platelet factor 4 (PF4) genes was confirmed by NotI, SalI and SacIIdigests. The maximum distance between PF4 and ALB is 210 kb and between ALB and IL8 is 420 kb. The alcohol dehydrogenase, class I (ADH2, ADH3) gene cluster can be linked to the alcohol dehydrogenase, class III gene (ADH5) by SacII, NruI, and EagI digests. The maximum distance between them is 590 kb. Our study indicated that ALB, α-fetoprotein (AFP), PF4, β-thromboglobulin (PPBP), GROl (encoding a cytokine also called melanoma growth-stimulatory activity), and IL8 genes can be physically linked. In this study the γ-interferon induced protein 10 (INP10), bone morphogenetic protein 3 (BMP3), annexin III (ANX3), KIT, amphiregulin (AREG), immunoglobulin J polypeptide (IGJ), deoxycytidine kinase (DCK) and MLLT2 genes were not linked to one another or to the above two groups of genes. Our analysis using somatic cell hybrids combined with previous reports demonstrated that the ADH gene cluster is telomeric to MLLT2 and KIT, ALB, AFP, PF4, βTG, GROl, IL8, ANX3, AREG and DCK are centromeric to MLLT2.

Targeted biological therapies that selectively interfere with cancer cell growth signals may improve patient survival by enhancing the effects of radiation, with the added benefit of reduced systemic toxicity. Head and Neck Squamous Cell Carcinoma (HNSCC) is the most common epithelial malignancy arising in the upper aerodigestive tract. It is the sixth most common cancer worldwide. Signal transducers and activators of transcription 3 (STAT3), is present in a number of different cancer cells including head and neck cancer and is related to chemoresistance and radioresistance, therefore is a potential novel molecular target for therapy to improve survival of the patients. In the present study, we report that linifanib (ABT-869), is a novel multi-targeted receptor tyrosine kinase (RTK) inhibitor of members of vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF) receptor families, has anti-proliferative effects in HNSCC cell lines in a dose dependent manner. It inhibits STAT3 signaling and related proteins and arrests cell cycle at G2/M phase, induces apoptosis, inhibits colony formation efficiency of the cells. In addition, when this multi-targeted receptor tyrosine kinase inhibitor is combined with radiation, significant antitumor effects are achieved in the cell lines. Thus, it may provide a therapeutic strategy and improves efficacy of radiation against HNSCC.

Advancements in rheumatoid arthritis (RA) treatment protocols and introduction of targeted biological therapies have markedly improved patient outcomes, despite this, up to 50% of patients still fail to achieve a significant clinical response. In veterinary medicine, stem cell therapy in the form of autologous stromal vascular fraction (SVF) is an accepted therapeutic modality for degenerative conditions with 80% improvement and no serious treatment associated adverse events reported. Clinical translation of SVF therapy relies on confirmation of veterinary findings in targeted patient populations. Here we describe the rationale and preclinical data supporting the use of autologous SVF in treatment of RA, as well as provide 1, 3, 6, and 13 month safety outcomes in 13 RA patients treated with this approach.

Abstract 1435 Abstract ABT-737 is a small molecule antagonist of BCL-2, BCL-XL, and BCL-w currently under evaluation in clinical trials in the oral form of ABT-263 (Navitoclax). Acquired resistance to BCL-2 inhibitors will inevitably emerge. Published pre-clinical data showed increased levels of BFL-1 and/or MCL-1 proteins in lymphoma, and concurrent up-regulation of BCL-XL and BCL2A1 in chronic lymphocytic leukemia, which are not targeted by ABT-737. To investigate potential mechanisms of resistance to BCL-2 inhibitors in acute myeloid leukemia, we developed resistant cell lines by long-term culture of HL-60 and MV4-11 cells with ABT-737, designated as HL-60R and MV4-11R. Parental lines, HL-60 and MV4-11 which were highly sensitive to ABT-737 with IC50 values of 30 nM and 90nM respectively, whereas those for HL-60R and MV4-11R were 10μM and 4.7μM respectively. Annexin V binding assay revealed that both resistant lines were resistant to ABT-737 induced apoptosis as compared to the parental HL-60 and MV4-11 cells. To explore the common pathway mediating acquired resistance to ABT-737, genes differentially expressed 2-fold or greater between parental and resistant lines on HL-60 and MV4-11 were studied. Interestingly, significant changes for BCL-2 family members were not found, suggesting that non-BCL2 family genes could play important roles in mediating the drug resistance to ABT-737. Among them, C3aR1 [Complement component 3a receptor 1] was significantly over-expressed in both resistant cell lines when compared to parental lines and verified with real-time RT-PCR. C3aR1 inhibitor, SB 290157, showed preferential cytotoxicity to both HL-60R and MV4-11R in a dose-dependent way and spared parental cells during 48 hrs in vitro MTT assay. Our findings suggest C3aR1 plays a key role in the resistance phenotype to Bcl-2 inhibitor and blocking C3aR1 revert the resistance. C3aR1 encodes a G-protein coupled seven trans-membrane receptor for C3a, an important inflammatory mediator. The C3a-C3aR axis modulated SDF-1–CXCR4 axis-dependent responses and regulated the homing of hematopoietic stem/progenitor cells into bone marrow. C3aR1 over expression has been associated with FLT3 and D835/I836 mutation cytogenetically normal acute myeloid leukemia, renal cancer and melanoma. We further investigate the role of C3aR1 in 138 cases of AML including 50 cases in initial diagnosis, 68 cases in remission and 20 cases in relapse with real-time RT-PCR. The result showed that C3aR1 expression is highest in initial stage and lowest in remission (p=0.006), there was no significant difference between initial and relapse stages (p=0.384). [Table 1] For newly diagnosed AML patients, high C3aR1 is statistically associated with younger age [median: 34 vs 43 years old], higher presenting WBC [median: 39K vs 27K], higher marrow % blasts [70 vs 55%], but not related to efficacy of first induction treatment, FAB classification or conventional chromosome risk groups.Table 1C3aR1 expression in AML clinical casesAML casesAge (year)C3AR1malefemalemedianrangeM-estimatorrangepInitial2822367-79199.777.49-2514.6P1 = 0.006Remission37313412-7092.2523.45-2286P2 = 0.046Relapse11944.513-54142.7512.4-2372.04P3 = 0.384Note: M-estimator was used for evaluating the differential expression of AML samples and the value was amplified by 104p1: initial vs remission; p2: relapse vs remission; p3 initial vs relapse The biological significant of C3a-C3aR axis in AML and resistance phenotype is intriguing and may suggest a novel mechanism of evading the apoptotic regulation by Bcl-2 gene family as suggested by our current study. Ongoing studies will further elucidate relationship of C3a-C3aR axis in AML. Disclosures: No relevant conflicts of interest to declare.

4575 Background: The efficacy of therapies for prostate cancer following failure of androgen deprivation and docetaxel is limited. SRC dysregulation may be important in prostate cancer progression....

Combination with other small molecule drugs represents a promising strategy to improve therapeutic efficacy of FLT3 inhibitors in the clinic. We demonstrated that combining ABT-869, a FLT3 inhibitor, with SAHA, a HDAC inhibitor, led to synergistic killing of the AML cells with FLT3 mutations and suppression of colony formation. We identified a core gene signature that is uniquely induced by the combination treatment in 2 different leukemia cell lines. Among these, we showed that downregulation of PTP4A3 (PRL-3) played a role in this synergism. PRL-3 is downstream of FLT3 signaling and ectopic expression of PRL-3 conferred therapeutic resistance through upregulation of STAT (signal transducers and activators of transcription) pathway activity and anti-apoptotic Mcl-1 protein. PRL-3 interacts with HDAC4 and SAHA downregulates PRL-3 via a proteasome dependent pathway. In addition, PRL-3 protein was identified in 47% of AML cases, but was absent in myeloid cells in normal bone marrows. Our results suggest such combination therapies may significantly improve the therapeutic efficacy of FLT3 inhibitors. PRL-3 plays a potential pathological role in AML and it might be a useful therapeutic target in AML, and warrant clinical investigation.

Abstract 2798 Background: Non-Hodgkin's lymphoma is the most common hematologic malignancy, with an estimated >65,000 new cases in the US in 2010. The indolent follicular lymphoma (FL) and marginal zone lymphoma (MZL) subtypes account for approximately 22% and 10% of cases, respectively. Rituximab, cyclophosphamide, vincristine, and prednisone, with doxorubicin (R-CHOP) or without (R-CVP) are the most common regimens used for frontline and relapsed/refractory FL. Bortezomib (VELCADE®) has shown activity alone and with rituximab in relapsed/refractory FL/MZL, and also in combination with chemo-immunotherapy regimens such as R-CHOP. However, overlapping peripheral neuropathy (PN) with vincristine may limit bortezomib dosing. Thus, the aim of this phase 2, two-arm, non-randomized, multicenter study was to evaluate the safety and efficacy of administering bortezomib in place of vincristine in the R-CHOP and R-CVP regimens. Methods: Patients aged ≥18 years with relapsed or refractory FL (including transformed FL) or MZL following ≥1 prior chemotherapy, ≥1 measurable tumor mass, ECOG performance status ≤2, no grade ≥2 residual toxicity from previous therapy/surgery, no grade ≥2 PN, and adequate hematologic, renal, and hepatic function were eligible. Based on investigator preference, patients received up to six 21-day cycles of bortezomib 1.6 mg/m2 IV on days 1 and 8, rituximab 375 mg/m2 IV on day 1, and prednisone 100 mg PO on days 1–5, plus either cyclophosphamide 1000 mg/m2 IV (VR-CP arm), or cyclophosphamide 750 mg/m2 IV and doxorubicin 50 mg/m2 IV (VR-CAP arm) on day 1. All patients then received rituximab 375 mg/m2 IV every 3 months for ≤2 years as maintenance. The primary endpoint was complete response (CR) rate, determined after all patients completed the 12-week follow-up visit. Secondary endpoints included overall response rate (ORR; CR + partial response [PR]), duration of response (DOR), progression-free survival (PFS), and safety/tolerability. Response was assessed by investigators using modified International Workshop Response Criteria. Toxicities were graded using National Cancer Institute Common Terminology Criteria for Adverse Events version 3.0. Results: A total of 55 patients have been enrolled, 48 to VR-CP and 7 to VR-CAP; reasons for selecting VR-CP included extent of prior anthracycline therapy (56%), study center preference (25%), response to prior R-CVP (13%), and age (10%). Of the 48 VR-CP patients, 50% were male, 88% were white, median age was 63.5 years (range 40–85), 47 had FL (52/33/15% grade 1/2/3; 19/43/38% low/intermediate/high risk by FL International Prognostic Index [FLIPI]), and 1 had MZL. Median number of prior lines of therapy was 2 (range 1–10), which included R-CHOP in 27 (56%), rituximab in 19 (40%), and R-CVP in 7 (15%) patients. Of 47 evaluable patients, 14 (30%) achieved CR and 23 (49%; including 1 MZL) PR (ORR 79%). Median DOR and PFS have not been reached. Patients received a median of 6 (range 2–6) cycles of VR-CP; 40 (83%) received all 6 cycles, and 28 (58%) have continued into the rituximab maintenance phase. All patients reported ≥1 adverse event (AE), including 65% with grade ≥3 AEs, and 25% with serious AEs. Most common AEs included diarrhea (52%), nausea (46%), fatigue (44%), neutropenia (31%), constipation, and vomiting (each 23%); grade ≥3 AEs included neutropenia (27%), lymphopenia (10%), and thrombocytopenia (6%). The incidence of PN was 21% (2% grade ≥3). Four (8%) patients discontinued due to AEs. Of 7 VR-CAP patients (2 male, 7 white, median age 57 years [range 55–73]), 4 had FL (3 grade 1, 1 unknown; all FLIPI high-risk), 2 MZL, and 1 chronic lymphocytic leukemia. Median number of prior lines of therapy was 1 (range 1–8). Of 6 response-evaluable patients, 0/4 achieved CR/PR (3 FL, 1 MZL; ORR 67%). Five patients received all 6 cycles; 4 have continued into rituximab maintenance. AEs included fatigue (n=5), alopecia, neutropenia (each n=4), anemia, constipation, nausea, rigors, and thrombocytopenia (each n=3). Neutropenia (n=4) was the only grade ≥3 AE reported in >1 patient. PN was seen in 2 patients (no grade 3). Two patients had serious AEs. There have been no on-study deaths in either arm. Conclusion: These results indicate that VR-CP is active, with a 30% CR rate, in patients with relapsed or refractory FL/MZL, including those who had received prior chemo-immunotherapy. Prolonged follow-up is required to evaluate DOR and PFS; updated results will be presented. Disclosures: Craig: Genentech: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Off Label Use: Use of bortezomib in combination in follicular lymphoma and marginal zone lymphoma. Cabanillas:Celgene: Consultancy, Research Funding; Bayer: Research Funding; Pfizer: Research Funding. Parasuraman:Millennium Pharmaceuticals, Inc.: Employment, Equity Ownership. Neuwirth:Millennium Pharmaceuticals, Inc.: Employment. O'Connor:Millennium Pharmaceuticals, Inc.: Membership on an entity's Board of Directors or advisory committees, Research Funding.

8134 Background: PLD and bortezomib (BORT) are FDA-approved for MM patients who have received one prior therapy. We have previously demonstrated in laboratory studies that lower doses of PLD administered more frequently are better tolerated than higher doses given less often. The combination of BORT and dexamethasone (DEX) is effective for previously untreated MM patients. Prior studies by our group have shown that intravenous administration of DEX 40 mg, BORT 1.0 mg/m2 and PLD 5 mg/m2 (DVD) on days 1, 4, 8, and 11 of a 28-day cycle is effective for relapsed/refractory MM patients and well tolerated. Methods: We conducted a single-arm multi-center phase II study for previously untreated MM patients to evaluate the DVD regimen. Patients were treated to a maximum response plus two additional cycles or completed a maximum of eight cycles of therapy without disease progression. Results: Thirty-five patients were enrolled with a median age of 65 years (range, 42-82 years). Thirty and 33 patients are currently ...

Abstract 4936 Despite recent advances in the treatment of MM, the disease remains incurable and many of the most effective, newer combination therapies are accompanied by significant side effects that have a major negative impact on the patient's quality of life. Pegylated liposomal doxorubicin (PLD) and bortezomib have shown anti-MM efficacy in the laboratory and for the treatment of previously treated MM patients, leading to FDA approval for patients who have failed one prior therapy. Using our severe combined immunodeficiency-hu murine models of human MM, we have previously demonstrated that lower doses of PLD administered daily are more effective and better tolerated than higher amounts given weekly. Moreover, the combination of bortezomib and dexamethasone has been shown to be effective for previously untreated MM patients. Prior studies by our group have shown that combining chemotherapy including PLD with bortezomib administered at 1.0 mg/m2 on days 1, 4, 8, and 11 of a 28-day cycle rather than the standard 1.3 mg/m2 on the same days of a 21-day schedule is effective for MM patients with relapsed or refractory disease and associated with a reduction in the incidence and severity of peripheral neuropathy. Thus, we conducted a single-arm multi-center phase II study for previously untreated MM patients to evaluate the combination of intravenously administered dexamethasone, bortezomib and PLD (DVD). The treatment consisted of intravenous administration of 40 mg dexamethasone followed by 1.0 mg/m2 bortezomib and finally 5.0 mg/m2 PLD on days 1, 4, 8, and 11 of a 28-day cycle. Patients were treated to a maximum response plus two additional cycles or completed a maximum of eight cycles of therapy without disease progression. To date, 22 (of 35 planned) patients have been enrolled with a median age of 64 years (range, 42-79 years). The majority of those on study (68 %) were diagnosed with International Staging System II or III MM. Four patients are too early to assess for response. To date, among the 18 evaluable patients, 16 (89%) have shown objective responses to the DVD regimen, including 2 complete responses (11%), 8 partial responses (44%) and 6 minimal responses (33%). The other 2 patients (11%) had stable disease, with one of these subjects showing a continuing reduction in M-protein after 2 cycles of therapy to date. Thus, disease control was achieved in all patients. To date, no patient has shown progressive disease after 2+ - 12+ months of follow-up. Six patients experienced grade 3 adverse events and one patient with a prior history of pulmonary interstitial fibrosis developed a grade 4 toxicity (shortness of breath). Grade 3 adverse events in three of the six patients were judged not to be related to the study treatment. The most common grade 3 adverse event was reversible neutropenia (n=2). To date, only 2 patients (9%) have developed peripheral neuropathy (grade 1). Notably, there have been no cases of stomatitis or hand-foot syndrome. Thus, these results suggest that the DVD regimen using a modified schedule and doses of the combination of intravenous dexamethasone, bortezomib and PLD is a well tolerated treatment that produces high response rates for previously untreated patients with multiple myeloma. Disclosures Berenson: Millennium Pharmaceuticals: Consultancy, Research Funding, Speakers Bureau; Centocor Ortho Biotech: Consultancy, Speakers Bureau. Hilger:Millennium Pharmaceutcals: Employment.

e16053 Background: We conducted a single arm, phase II trial of single-agent bortezomib, a proteasome inhibitor, in patients with prostate cancer with biochemical recurrence (PSA relapse) after definitive local therapy. Eligible patients had previous radical prostatectomy and/or radiation therapy (including brachytherapy), and a rising PSA over 1 ng/mL without bone scan evidence of metastasis. The primary goal of this study was to determine the PSA response to single agent bortezomib. Secondary goal was to identify the time to PSA relapse after bortezomib therapy defined as time from nadir to PSA >1ng/ml. Methods: Treatment was composed of bortezomib 1.3mg/m2 IV on days 1,4,8,11 then 10 days off for the first two cycles. The third cycle was comprised of weekly bortezomib therapy for three out of four weeks. Sixteen patients were enrolled and treated at 4 sites with13 evaluable patients. Results: Median baseline PSA was 12.0 ng/ml. 3 of 13 patients (23%) achieved complete responses and 1 a partial response (PR) for a response rate of 30% (95% CI: 0.09 –0.61). Median time to PSA relapse in these 3 patients was 6.0 months. Linear longitudinal modeling was used to compare the rate of increase in PSA prior to treatment versus during treatment with bortezomib and found that treatment significantly decreased the slope of log PSA during treatment, slowing PSA rise (p = 0.003). The major toxicity associated with bortezomib therapy was peripheral neuropathy (7 pts, grade 3 in 3 pts) requiring dose reductions in 3 cases and discontinuation of therapy in 4 patients. Other toxicities included non-neutropenic infections (shingles in 2 pts), 1 small bowel ileus (grade 3), and 1 case of thrombocytopenia (grade 3). Conclusions: In a small number of patients there was a 30% PSA response rate to single agent bortezomib therapy. Further investigations using bortezomib as a single-agent or in combination are warranted in this patient population. No significant financial relationships to disclose.

We develop a standardized, fully automated, quantification system for liver fibrosis assessment using second harmonic generation microscopy and a morphology-based quantification algorithm. Liver fibrosis is associated with an abnormal increase in collagen as a result of chronic liver diseases. Histopathological scoring is the most commonly used method for liver fibrosis assessment, where a liver biopsy is stained and scored by experienced pathologists. Due to the intrinsic limited sensitivity and operator-dependent variations, there exist high inter- and intraobserver discrepancies. We validate our quantification system, Fibro-C-Index, with a comprehensive animal study and demonstrate its potential application in clinical diagnosis to reduce inter- and intraobserver discrepancies.