[Show abstract][Hide abstract] ABSTRACT: KSHV ORF57 (MTA) promotes RNA stability of ORF59, a viral DNA polymerase processivity factor. Here, we show that the integrity of both ORF59 RNA ends is necessary for ORF57-mediated ORF59 expression and deletion of both 5' and 3' regions, or one end region with a central region, of ORF59 RNA prevents ORF57-mediated translation of ORF59. The ORF59 sequence between nt 96633 and 96559 resembles other known MTA-responsive elements (MREs). ORF57 specifically binds to a stem-loop region from nt 96596-96572 of the MRE, which also binds cellular RBM15. Internal deletion of the MRE from ORF59 led to poor export, but accumulation of nuclear ORF59 RNA in the presence of ORF57 or RBM15. Despite of being translatable in the presence of ORF57, this deletion mutant exhibits translational defect in the presence of RBM15. Together, our results provide novel insight into the roles of ORF57 and RBM15 in ORF59 RNA accumulation and protein translation.
[Show abstract][Hide abstract] ABSTRACT: Biliary tract carcinoma is a rare malignancy. We performed a comprehensive analysis of published prospective clinical trials in advanced biliary tract carcinoma in an attempt to identify active regimens in this setting. We searched PubMed and abstracts presented at the American Society of Clinical Oncology, Gastrointestinal Cancer Symposium, European Society of Medical Oncology and European Cancer Organization conferences for clinical trials in this disease. We found 83 trials. The effect of gemcitabine on overall survival benefit showed a strong trend (p = 0.014) and an improvement in progression-free survival (p = 0.003). Gemcitabine-based regimens containing 5-fluorouracil showed a trend toward an improved overall survival (p = 0.047) relative to platinum agents. Our findings support gemcitabine as the chemotherapy backbone for the treatment of patients with cholangiocarcinoma. Gemcitabine plus 5-fluorouracil combinations warrant further investigations.
[Show abstract][Hide abstract] ABSTRACT: The downstream targets of hypoxia inducible factor-1 alpha (HIF-1alpha) play an important role in tumor progression and angiogenesis. Therefore, inhibition of HIF-mediated transcription has potential in the treatment of cancer. One attractive strategy for inhibiting HIF activity is the disruption of the HIF-1alpha/p300 complex, as p300 is a crucial coactivator of hypoxia-inducible transcription. Several members of the epidithiodiketopiperazine (ETP) family of natural products have been shown to disrupt the HIF-1alpha/p300 complex in vitro; namely, gliotoxin, chaetocin, and chetomin. Here, we further characterized the molecular mechanisms underlying the antiangiogenic and antitumor effects of these ETPs using a preclinical model of prostate cancer. In the rat aortic ring angiogenesis assay, gliotoxin, chaetocin, and chetomin significantly inhibited microvessel outgrowth at a GI50 of 151, 8, and 20 nM, respectively. In vitro co-immunoprecipitation studies in prostate cancer cell extracts demonstrated that these compounds disrupted the HIF-1alpha/p300 complex. The downstream effects of inhibiting the HIF-1alpha/p300 interaction were evaluated by determining HIF-1alpha target gene expression at the mRNA and protein levels. Dose-dependent decreases in levels of secreted VEGF were detected by ELISA in the culture media of treated cells, and the subsequent downregulation of VEGFA, LDHA, and ENO1 HIF-1alpha target genes were confirmed by semi-quantitative real-time PCR. Finally, treatment with ETPs in mice bearing prostate tumor xenografts resulted in significant inhibition of tumor growth. These results suggest that directly targeting the HIF-1alpha/p300 complex with ETPs may be an effective approach for inhibiting angiogenesis and tumor growth.
Molecular Cancer 04/2014; 13(1):91. DOI:10.1186/1476-4598-13-91 · 5.40 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Brain metastases of breast cancer cause neurocognitive damage and are incurable. We evaluated a role for temozolomide in the prevention of brain metastases of breast cancer in experimental brain metastasis models.
Temozolomide was administered in mice following earlier injection of brain-tropic human epidermal growth factor receptor 2 (HER2)-positive Jimt1- BR3 and triple negative 231-BR-EGFP sublines, the latter with and without expression of 06-methylguanine-DNA methyltransferase (MGMT). Additionally, the percentage of MGMT-positive tumor cells in 62 patient-matched sets of breast cancer primary tumors and resected brain metastases was determined immunohistochemically.
Temozolomide, when dosed at 50, 25, 10 or 5 mg/kg, 5 days/week, beginning 3 days after inoculation, completely prevented the formation of experimental brain metastases from MGMT-negative 231-BR-EGFP cells. At a 1 mg/kg dose, temozolomide prevented 68% of large brain metastases, and was ineffective at a dose of 0.5 mg/kg. When the 50 mg/kg dose was administered beginning on days 18 or 24, temozolomide efficacy was reduced or absent. Temozolomide was ineffective at preventing brain metastases in MGMT-transduced 231-BR-EGFP and MGMT-expressing Jimt-1-BR3 sublines. In 62 patient-matched sets of primary breast tumors and resected brain metastases, 43.5% of the specimens had concordant low MGMT expression, while in another 14.5% of sets high MGMT staining in the primary tumor corresponded with low staining in the brain metastasis.
Temozolomide profoundly prevented the outgrowth of experimental brain metastases of breast cancer in an MGMT-dependent manner. These data provide compelling rationale for investigating the preventive efficacy of temozolomide in a clinical setting.
Clinical Cancer Research 03/2014; 20(10). DOI:10.1158/1078-0432.CCR-13-2588 · 8.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Mutant Ras oncogenes produce proteins that are unique to cancer cells and represent attractive targets for vaccine therapy. We have shown previously that vaccinating cancer patients with mutant ras peptides is feasible and capable of inducing a specific immune response against the relevant mutant proteins. Here, we tested the mutant ras peptide vaccine administered in combination with low dose interleukin-2 (IL-2) or/and granulocyte-macrophage colony-stimulating factor (GM-CSF) in order to enhance the vaccine immune response.
5000mug of the corresponding mutant ras peptide was given subcutaneously (SQ) along with IL-2 (Arm A), GM-CSF (Arm B) or both (Arm C). IL-2 was given SQ at 6.0 million IU/m2/day starting at day 5, 5 days/week for 2 weeks. GM-CSF was given SQ in a dose of 100mug/day one day prior to each ras peptide vaccination for 4 days. Vaccines were repeated every 5 weeks on arm A and C, and every 4 weeks on arm B, for a maximum of 15 cycles or until disease progression.
We treated 53 advanced cancer patients (38 with colorectal, 11 with pancreatic, 1 with common bile duct and 3 with lung) on 3 different arms (16 on arm A, 18 on arm B, and 19 on arm C). The median progression free survival (PFS) and overall survival (OS) was 3.6 and 16.9 months, respectively, for all patients evaluable for clinical response (n = 48). There was no difference in PFS or OS between the three arms (P = 0.73 and 0.99, respectively). Most adverse events were grade 1-2 toxicities and resolved spontaneously. The vaccine induced an immune response to the relevant ras peptide in a total of 20 out of 37 evaluable patients (54%) by ELISPOT, proliferative assay, or both. While 92.3% of patients on arm B had a positive immune response, only 31% of patients on arm A and 36% of patients on arm C had positive immune responses (P = 0.003, Fisher's exact test).
The reported data showed that IL-2 might have a negative effect on the specific immune response induced by the relevant mutant ras vaccine in patients with advanced cancer. This observation deserves further investigations. Trial registration: NCI97C0141.
Journal of Translational Medicine 02/2014; 12(1):55. DOI:10.1186/1479-5876-12-55 · 3.99 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: We recently reported the clinical results of a Phase I trial combining ipilimumab with a vaccine containing transgenes for prostate-specific antigen (PSA) and for a triad of costimulatory molecules (PROSTVAC) in patients with metastatic castration-resistant prostate cancer. Thirty patients were treated with escalating ipilimumab and a fixed dose of vaccine. Of 24 chemotherapy-naïve patients, 58 % had a PSA decline. Combination therapy did not exacerbate the immune-related adverse events associated with ipilimumab. Here, we present updated survival data and an evaluation of 36 immune cell subsets pre- and post-therapy. Peripheral blood mononuclear cells were collected before therapy, at 13 days and at 70 days post-initiation of therapy, and phenotyped by flow cytometry for the subsets of T cells, regulatory T cells, natural killer cells, and myeloid-derived suppressor cells. Associations between overall survival (OS) and immune cell subsets prior to treatment, and the change in a given immune cell subset 70 days post-initiation of therapy, were evaluated. The median OS was 2.63 years (1.77-3.45). There were trends toward associations for longer OS and certain immune cell subsets before immunotherapy: lower PD-1(+)Tim-3(NEG)CD4EM (P = 0.005, adjusted P = 0.010), higher PD-1(NEG)Tim-3(+)CD8 (P = 0.002, adjusted P = 0.004), and a higher number of CTLA-4(NEG) Tregs (P = 0.005, adjusted P = 0.010). We also found that an increase in Tim-3(+) natural killer cells post- versus pre-vaccination associated with longer OS (P = 0.0074, adjusted P = 0.015). These results should be considered as hypothesis generating and should be further evaluated in larger immunotherapy trials.
Cancer Immunology and Immunotherapy 02/2014; 74(19 Supplement). DOI:10.1007/s00262-014-1524-0 · 3.94 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Objectives Anti-CD20 (rituximab), anti-CD52 (alemtuzumab), anti-CD22 (BL22, HA22), and anti-CD25 (Oncotac) are therapeutic options that are the mainstay or in preclinical development for the treatment of chronic lymphocytic leukemia (CLL). Studies suggest that levels of surface antigen expression may affect response to monoclonal antibody-based therapy. Methods Using the flow cytometric Quantibrite method (BD Biosciences, San Jose, CA) to determine antibodies bound per cell, we quantified the levels of surface expression of CD20, CD22, CD25, and CD52 in CLL cells from 28 untreated patients. Results The CLL cells in all cases expressed CD20, CD22, and CD52 but 4 (14%) cases were negative for CD25. Although the ranking of levels of expression from highest to lowest was CD52, CD20, CD22, and CD25, the level of antigen expression on any specific case could not be accurately predicted. Conclusions Quantification of antigens might be useful in evaluating new antigens to target for therapy and may provide a systematic approach to selecting individualized therapy in CLL.
American Journal of Clinical Pathology 12/2013; 140(6):813-8. DOI:10.1309/AJCPYFQ4XMGJD6TI · 3.01 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Eradication of minimal residual disease (MRD) after allotransplantation in persons with chronic lymphocytic leukemia (CLL) is associated with lower rates of relapse. Rapid engraftment of donor lymphocyte elements may contribute to MRD control but it remains unclear if this strategy will benefit patients. Here we report incidence of MRD eradication and graft versus host disease (GvHD) in persons with rapid versus later donor T-lymphocyte engraftment after lymphodepleting chemotherapy and reduced intensity conditioning (RIC) allotransplantation. Twenty-seven subjects received lymphodepleting chemotherapy to facilitate donor engraftment followed by fludarabine and cyclophosphamide RIC and a blood cell allograft. MRD was monitored by multicolor flow cytometry post transplantation. Complete donor T-lymphoid (TLC) and myeloid (MC) chimerism were achieved in 25 subjects at a median of 28 days (range 14-60 days) and 21 days (range 14-180 days). Achieving complete donor TLC by day 14 versus day ≥28 correlated with occurrence of ≥grade-2 acute GvHD (90% [95% confidence interval (CI), 78-100%] versus 35% [95% CI, 16-54%], P=0.014) and better control of minimal residual disease in the bone marrow at day 100, median 0% (range, 0-0.1%) versus 8.5% (range, 0-92%; P=0.016). Among 11 persons with early donor TLC none had progressive disease (PD) and 7 died of treatment related mortality (TRM). In persons with later development of TLC 8 of 16 had PD and 2 died of TRM. Time to donor myeloid chimerism had no impact on outcomes. Rapid establishment of donor TLC results in more complete eradication of early MRD but greater incidence of acute GvHD and TRM in persons with CLL undergoing RIC allotransplantation.
[Show abstract][Hide abstract] ABSTRACT: The interaction between CD27 and its ligand, CD70, has been implicated in regulating cellular immune responses to cancer. In this article, we report on the role of soluble CD27 (sCD27) in T cell activation and its elevation in the serum of cancer patients after immunotherapy. In vitro, sCD27 is preferentially derived from activated CD4(+) T cells. Adding sCD27 to stimulated PBMCs increases T cell activation and proliferation, and is associated with the immunologic synapse-related proteins myosin IIA, high mobility group box 1, and the TCR Vβ-chain. The pool of serum sCD27 is shown to be greater in healthy donors than in cancer patients. However, metastatic cancer patients treated with immunotherapy showed a significant increase in the serum sCD27-pool posttherapy (p < 0.0005); there was also an increased trend toward an association between enhanced sCD27-pool posttherapy and overall survival (p = 0.022). The identification of sCD27 as an immune modulator associated with enhanced human T cell activation in vitro and in vivo provides a rationale for developing new immunotherapeutic strategies aimed at enhancing sCD27 for treating cancer and potentially other diseases.
The Journal of Immunology 05/2013; 190(12). DOI:10.4049/jimmunol.1300022 · 5.36 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: BACKGROUND: There is currently no standard of care for the second-line treatment of advanced pancreatic cancer. The aim of this analysis was to compare the different therapeutic approaches in this setting. METHODS: We carried out a systematic analysis of second-line studies in advanced pancreatic cancer that have progressed on or following gemcitabine and published or presented from 2000 to 2012. RESULTS: Forty-four clinical trials (t) were identified; of which 34 met the inclusion criteria treating an aggregate total of 1503 patients (n). Patients who received treatments (t: 33; n: 1269) had a median overall survival (OS) of 6 months compared with 2.8 months for patients who received best supportive care only (t: 2; n: 234) (P = 0.013). The gemcitabine and platinum-based combination (t: 5; n: 154) provided a median progression-free survival and OS of 4 and 6 months compared with 1.6 and 5.3 for the rest of the regimens (t: 29; n: 1349) (P = 0.059 and 0.10, respectively) and 2.9 and 5.7 for the combination of 5-fluorouracil and platinum agents (t: 12; n: 450) (P = 0.60 and 0.22, respectively). CONCLUSION(S): Although not conclusive, these data showed that the advantage of second-line chemotherapy in pancreatic cancer is very limited and there is a need for more studies.
Annals of Oncology 05/2013; 24(8). DOI:10.1093/annonc/mdt166 · 6.58 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: PURPOSE: Romidepsin is a histone deacetylase inhibitor (HDI) approved for the treatment of both cutaneous and peripheral T cell lymphoma (CTCL and PTCL). During development, a thorough assessment of cardiac toxicity was performed. EXPERIMENTAL DESIGN: A phase II single-agent non-randomized study of romidepsin was performed in patients with CTCL and PTCL who had progressed after at least one prior systemic therapy. RESULTS: Results for the first 42 patients enrolled on the NCI 1312 Phase II study of romidepsin in CTCL or PTCL demonstrated no cardiac toxicity based on serial electrocardiograms, troponins and MUGA scans/echocardiograms. The cardiac assessments reported herein confirm the safety of romidepsin among 131 enrolled patients, while supporting a role for electrolyte replacement. Heart rate increased an average 11 bpm following romidepsin infusion; there was no evidence of increased arrhythmia. Criteria for potassium/magnesium replacement were met prior to 55% of 1365 romidepsin doses; an association with hypoalbuminemia was confirmed. We propose a mechanism for ST segment flattening and depression, the most common electrocardiogram abnormalities observed: HDI-induced alteration of the activity or expression of KATP channels. In addition, examination of the variants of the active transporter of romidepsin, ABCB1, demonstrated a trend towards smaller heart rate changes in the peri-infusion period among wild type compared to variant diplotypes. CONCLUSIONS: We conclude that in the context of appropriate attention to electrolyte levels, the data support the cardiac safety of romidepsin.
Clinical Cancer Research 04/2013; 19(11). DOI:10.1158/1078-0432.CCR-13-0109 · 8.19 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Brain metastases of breast cancer occur in more than one-third of metastatic breast cancer patients whose tumors overexpress HER2 or are triple negative. Brain colonization of cancer cells occurs in a unique environment, containing microglia, oligodendrocytes, astrocytes, and neurons. Although a neuroinflammatory response has been documented in brain metastasis, its contribution to cancer progression and therapy remains poorly understood. Using an experimental brain metastasis model of breast cancer, we characterized the brain metastatic microenvironment of brain tropic, HER2-transfected MDA-MB-231 human breast carcinoma cells (231-BR-HER2). A previously unidentified subpopulation of metastasis-associated astrocytes expressing phosphorylated platelet-derived growth factor receptor β (at tyrosine 751; p751-PDGFRβ) was identified around perivascular brain micrometastases. This subpopulation of p751-PDGFRβ(+) astrocytes was also identified in human brain metastases from eight different craniotomy specimens and in primary cultures of astrocyte-enriched glial cells. Previously, we reported that pazopanib, a multispecific tyrosine kinase inhibitor, prevented the outgrowth of 231-BR-HER2 large brain metastases by 73%. Here, we evaluated the effect of pazopanib on the brain neuroinflammatory microenvironment. Pazopanib treatment resulted in 70% (P = 0.023) decrease of the p751-PDGFRβ(+) astrocyte population, at the lowest dose of 30 mg/kg, twice daily. Collectively, the data identify a new subpopulation of activated astrocytes in the subclinical perivascular stage of brain metastases and show that they are inhibitable by pazopanib, suggesting its potential to prevent the development of subclinical perivascular micrometastases in breast cancer patients.
American Journal Of Pathology 04/2013; 182(6). DOI:10.1016/j.ajpath.2013.02.043 · 4.60 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Flow cytometric (FC) enumeration of abnormal plasma cells (APCs) for diagnosis and prognostication of plasma cell dyscrasias (PCD) is challenging. We studied antigen expression in normal plasma cells (NPC) (N = 34) and APC in a series of unselected PCD (N = 59). NPC subpopulations often demonstrated CD19(-), CD20(+), CD45(-) or dim and CD56(+), an immunophenotype observed in PCD. However abnormal CD81 was only observed in APCs (APC detection sensitivity 95%; specificity 100%). We evaluated differences in antigen expression patterns among MGUS (N = 14), SMM (N = 35) and MM (N = 10), finding the combination of CD45 and CD56 helpful in differentiating MGUS from SMM and MM (p = 0.0002).
Leukemia research 01/2013; DOI:10.1016/j.leukres.2013.12.007 · 2.69 Impact Factor