Citations

ObjectiveNumerous factors can impede or facilitate patients’ medication decision‐making and adherence to physicians’ recommendations. Little is known about how patients and physicians jointly view issues that affect the decision‐making process. Our objective was to derive an empirical framework of patient‐identified facilitators to lupus medication decision‐making from key stakeholders (including 15 physicians, 5 patients/patient advocates, and 8 medical professionals) using a patient‐centered cognitive mapping approach.MethodsWe used nominal group patient panels to identify facilitators to lupus treatment decision‐making. Stakeholders independently sorted the identified facilitators (n = 98) based on their similarities and rated the importance of each facilitator in patient decision‐making. Data were analyzed using multidimensional scaling and hierarchical cluster analysis.ResultsA cognitive map was derived that represents an empirical framework of facilitators for lupus treatment decisions from multiple stakeholders’ perspectives. The facilitator clusters were 1) hope for a normal/healthy life, 2) understand benefits and effectiveness of taking medications, 3) desire to minimize side effects, 4) medication‐related data, 5) medication effectiveness for “me,” 6) family focus, 7) confidence in physician, 8) medication research, 9) reassurance about medication, and 10) medication economics.ConclusionConsideration of how different stakeholders perceive the relative importance of lupus medication decision‐making clusters is an important step toward improving patient‐physician communication and effective shared decision‐making. The empirically derived framework of medication decision‐making facilitators can be used as a guide to develop a lupus decision aid that focuses on improving physician‐patient communication.

Abstract Treatment of glioblastoma multiforme (GBM) continues to be a challenge due to its infiltrative nature, tumor heterogeneity, and lack of therapeutic agents that penetrate the blood-brain barrier (BBB). While the primary tumor is permeable to some degree, the ability of GBM cells to invade areas of parenchyma with an intact BBB indicates development of BBB-penetrable compounds is a necessity. The multifunctional protein MDM2 holds promise as a therapeutic target in a variety of cancers and plays a critical role in controlling cell survival, invasion, and DNA repair. MDM2 antagonists such as nutlin3a and RG7112 are being used to interrogate the impact of modulating MDM2 function in combination with front-line therapy. Our objective was to investigate whether MDM2 antagonists, alone or in combination with temozolomide (TMZ), can augment cell death in orthotopic GBM xenograft models. In vitro data indicate that TMZ and nutlin3a are synergistic in decreasing cell viability in wild type (wt) p53 U87-MG, primary human wt p53 GBM10 and mutant (mt) p53 GBM43 cells. Pharmacodynamic studies demonstrated that the mechanism of action for promoting cell death following exposure to TMZ/nutlin3a was multifactorial. Comet assays indicated that repair of TMZ-mediated DNA damage was significantly delayed in wt and mt p53 GBM cells treated with TMZ/nutlin3a compared to TMZ alone and the base excision repair protein Ape1 was downregulated in cells treated with TMZ/nutlin3a. Pharmacokinetic studies guided development of rational dosing regimens in which 2-3 five-day cycles of TMZ followed by nutlin3a 4 hours later were investigated. In ectopic U87-MG xenografts, nutlin3a sensitized xenografts to TMZ-mediated cell death. Orthotopic studies employing U87-MG, GBM10, and GBM43 tumors are being utilized to determine if nutlin3a levels detectable in the brain via HPLC-MS/MS(API 4000) are sufficient to modulate MDM2 function in the context of TMZ and increase survival. In the TMZ-resistant GBM10 (wtp53, MGMTpos, and PTENnull) orthotopic model, there was a modest increase in median survival from 63 days with TMZ to 73 days with TMZ/nutlin3a. Higher and more sustained brain levels of MDM2 antagonists will likely be necessary to improve survival. The MDM2 antagonist RG7112 has an improved PK profile and structural analysis of RG7112 and nutlin3a via QikProp 3.0 (www.schrodinger.com) indicates that RG7112 also has an improved predicted brain/blood partition coefficient (plog BB) compared to nutlin3a (RG7112 = -0.17; nutlin3a = -0.415). Studies are in progress to assess the brain penetration of RG7112 and its effect on GBM growth in vivo. Taken together, our data suggest that modulation of MDM2 function in the context of cytotoxic therapy has the potential to alter mechanisms involved in DNA repair that can promote cell death and improve survival. Citation Format: Haiyan Wang, Shanbao Cai, Barbara J. Bailey, Lawrence M. Gelbert, M. Reza Saadatzadeh, Aaron A. Cohen-Gadol, Jann N. Sarkaria, Paul Territo, Taxiarchis M. Georgiadis, T. Zachary Gunter, Samy Meroueh, Eric C. Long, David R. Jones, Lindsey D. Mayo, Shannon Harlan, Karen E. Pollok. Sensitization of temozolomide-mediated glioblastoma cell death by targeting MDM2: Assessment of PD biomarkers, brain penetration, and efficacy in humanized orthotopic xenograft models. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2741. doi:10.1158/1538-7445.AM2014-2741

Abstract Glioblastoma multiforme (GBM) comprises the largest group of brain tumors which are drug resistant and respond very poorly to the current therapies. In this study, we used sulforaphane (SFN), a multi-targeting agent with cancer preventive and anti-cancer activities and showed that it targets GBM established cell lines, early primary cultures, and CD133+ GBM stem cells as well as in GBM stem-like spheroids. SFN at 5-50 μM triggered significant inhibition of cell survival and induced apoptotic cell death in GBM cells and CD133+ stem cells isolated from four GBM cell lines. SFN induced apoptosis in U87MG cells was associated with caspase-7 activation. Moreover, SFN triggered formation of intracellular reactive oxygen species (ROS) and when the cells were pre-treated with 10 mM N-acetyl cysteine (NAC), ROS production and cell survival in cells treated with 5-10 μM were similar to the control untreated U87MG cells, revealing that SFN-triggered cell death is ROS-dependent. Moreover, SFN-generated ROS in U87MG cells were formed at the Mitochondrial Respiratory Chain (MRC) level. SFN also increased expression of the TRAIL receptor DR5 in GBM cells, U87MG and SF767 cells by 24 h post-exposure. Moreover, as revealed by comet assay, SFN increased single- and double-strand DNA breaks in GBM. Compared to untreated control cells, a significantly higher amount of γ-H2AX foci and as consequence higher number of DNA double-strand breaks (DSBs) breaks were observed in the SFN-treated sample. In vivo studies, using NOD/SCID mice revealed that SFN administration via oral gavage at 100 mg/kg for 3 cycles significantly decreases the growth of ectopic xenografts established from the early passage primary cultures of GBM10. Our results show that SFN robustly inhibits growth of GBM cells in vitro and in vivo and induces cell death in established cell cultures, early passage primary cultures, as well as it is effective in eliminating GBM cancer stem cells, which play a major role in drug resistance and disease recurrence. These results suggest that use of SFN alone or in combination with other agents, may potentially improve survival of brain tumor patients. Citation Format: Khadijeh Bijangi-Vishehsaraei, Mohammad R. Saadatzadeh, Haiyan Wang, Malgorzata M. Kamocka, Wenjing Cai, Aaron A. Cohen-Gadol, Stacey L. Halum, Karen E. Pollok, Jann N. Sarkaria, Ahmad R. Safa. Sulforaphane depresses proliferation and induces cell death in glioblastoma multiforme (GBM) cells, GBM stem cell-like spheroids, and tumor xenografts through modulation of multiple cell signaling pathways. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2267. doi:10.1158/1538-7445.AM2014-2267

Abstract There is a significant unmet medical need to develop more effective treatments for cancers of the central and peripheral nervous systems. Glioblastoma multiforme (GBM) is the most common form of brain cancer, and successful long-term treatment has not been realized. Neuroblastomas (NB) arise in the sympathetic nervous system and are the most common extracranial solid tumor of childhood. Survival of children with disseminated, high-risk NB tumors is less than 50%, even with intensive multimodal treatment. To improve treatment outcome, inhibition of murine double minute 2 (MDM2) function in the context of front-line therapy is being explored. MDM2 is a multi-functional protein that plays a critical role in controlling cell growth and migration. MDM2 is a negative regulator of p53 and p73 and, under low stress conditions, sequesters them in the cytoplasm. However, inhibition of MDM2 interactions with p53 and p73 by the MDM2 antagonist nutlin3a can lead to activation of p53/p73-mediated cell-death in tumor cells. The objective of the present study was to evaluate the extent to which blockade of MDM2-p53/p73 interactions by nutlin3a, alone or in combination with front-line therapies, could augment cell death in GBM and NB cells with a variety of molecular profiles. Using the median effect model, we found that temozolomide (TMZ) and nutlin3a were synergistic in increasing cell death in wtp53 U87 and U373 but were predominantly additive in mtp53 U118 cells. Primary patient GBM cells were typically resistant to TMZ relative to established cell lines. However, resistance to TMZ was decreased in the presence of nutlin3a. Further, TMZ and nutlin3a were synergistic in primary GBM10 and GBM43 but not in recurrent primary IU-GBM16, IU-GBM23, IU-GBM27, and IU-GBM32 cells. Moreover, in NB cells with wtp53, nutlin3a and cisplatin were synergistic in MYCN nonamplified SK-N-SH and additive in MYCN amplified IMR5. In mtp53 NB cells, nutlin3a and cisplatin were synergistic in MYCN nonamplified SK-N-FI as well as in MYCN amplified SK-N-DZ. Nutlin3a alone produced dose-related increases in cell death of all GBM and NB cell lines irrespective of p53 status, suggesting that p53-independent mechanisms play a role in inhibition of cell growth produced by blockade of MDM2-mediated signaling. Consistent with this hypothesis, nutlin3a increased p73 protein levels in mtp53 GBM43 cells that were exposed to TMZ. Inhibition of MDM2 function also inhibited repair of TMZ-mediated DNA damage in GBM. In comet assays, nutlin3a+TMZ decreased DNA repair relative to TMZ alone in wtp53 U87 and GBM10 and in mtp53 GBM43 cells. The present results demonstrate that MDM2 is a valid therapeutic target in GBM and NB cells with varying genetic profiles. In conclusion, inhibition of MDM2 signaling by nutlin3a is generally synergistic to additive in combination with front-line chemotherapeutics used to treat cancers of the central and peripheral nervous systems. Citation Format: Harlan E. Shannon, Haiyan Wang, Shanbao Cai, Wei Michael Liu, Lawrence M. Gelbert, Aaron A. Cohen-Gadol, Jann N. Sarkaria, Lindsey D. Mayo, Karen E. Pollok. MDM2 as a therapeutic target: improving upon front-line chemotherapy for nervous system tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3435. doi:10.1158/1538-7445.AM2013-3435

Abstract Glioblastoma (GBM) is the most common and malignant form of primary brain tumor. Average survival rate of GBM is 12-15 months from diagnosis. Due to its highly invasive nature, current treatment options have not been able to improve the survival rate in past 20 years. In order to discover GBM therapeutic targets, omics technologies have been widely used in order to identify potential biomarkers. Identification of cancer biomarkers for diagnosis and prognosis continues to evolve as clinical specimen pipelines and technologies are optimized. This research study focused on investigating proteins, metabolites and lipids biomarkers and patterns of human GBM xenografts employing mass spectrometry. Human tumor cell lines GBM 10 and GBM 43 were injected in the right cerebral hemisphere and flank sites in NOD/SCID mice (n = 10 mice per group). After harvesting brain and flank tumors and control brain from mouse models, protein, metabolites, and lipids of tumor samples were collected through one simple extraction procedure, which may solve sample preparation difficulties in omics studies, developed by Dr. Jiri Adamec. These samples were analyzed by reverse phase high performance liquid chromatography - Fourier transform ion cyclotron resonance mass spectrometry (LC-FTMS). FTMS analyzers have the highest resolving power of all MS instruments, which is ideal for complex mixtures such as GBM tissue. Spectra obtained from the FTMS analysis were analyzed using Student t-tests to detect significant differences in tissue profiles at a level of p = 0.05. Compounds below this threshold were identified through a database using the m/z ratio. Lipidomic analysis indicated the possible up and down-regulation of several lipids classes in GBM tissue connected to metabolic pathways, tumor proliferation and immunodepression, such as glycerophospholipids, both up- and down-regulated (e.g. PC38:4 or PG 35:2, respectively), and down-regulated glycosphingolipids (Galβ1-3GlcNAcβ1-3Galβ1-4Glc-Cer, Galβ1-4GlcNAcβ1-3Galβ1-4Glc-Cer), and triglycerides (TG 55:9, 53:4,52:8). We expect that integrated studies of proteomics, metabolomics, and lipidomics increase the confidence of identification of potential biomarkers. Accompanying these studies is a collaborative effort to improve the effectiveness and efficiency of computational pipelines that are imperative to the analytics, visualization, identification, and interpretation of the omics data. Only by carefully integrating the computational pipelines can we successfully perform the types of integrative studies needed to advance the identification of cancer biomarkers for diagnosis and prognosis, and our integrative studies serve as a case study for our pipeline advancement efforts. Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B37. Citation Format: Soo Jung Ha, Gordon Showalter, Jenna Rickus, Shanbao Cai, Haiyan Wang, Wei Michael Liu, Jann N. Sarkaria, John Springer, Jiri Adamec, Karen E. Pollok, Kari L. Clase. Proteomic and metabolomic analyses of glioblastoma using mass spectrometry. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B37.

SummaryDexamethasone ± thalidomide with infusion of cisplatin, doxorubicin, cyclophosphamide, and etoposide [D(T)PACE] is generally reserved as salvage therapy for aggressive multiple myeloma (MM) or plasma cell leukaemia (PCL) resistant to conventional therapies. The efficacy and durability of this potentially toxic regimen in this setting is unclear. We identified 75 patients who received D(T)PACE for relapsed/refractory MM at two tertiary care centres: Princess Margaret Hospital, Toronto and Mayo Clinic Arizona. At time of D(T)PACE, 16 patients had PCL and three patients had leptomeningeal disease. Patients were heavily pretreated (median three prior regimens, range 1–12; prior autologous stem cell transplant [ASCT] 33%). Overall response rate was 49% (very‐good partial response 16%, partial response 33%) with stable disease in an additional 36%. Median progression‐free survival (PFS) was 5·5 months (95% confidence interval [CI]:4·3–9·8); overall survival (OS) 14·0 months (95% CI:8·7–19·3). Thirty‐five patients proceeded to ASCT or clinical trial, with median PFS for this subset of 13·4 months (95% CI:7·7–20·1) and OS 20·5 months (95% CI:14·8–63·8). D(T)PACE is an effective salvage therapy for heavily pretreated MM patients. Although the overall response rate of 49% in this poor prognosis cohort is reasonable, the PFS is short, suggesting the best role for D(T)PACE is in bridging to definitive therapy, such as transplantation.

ObjectiveFree fatty acids (FFAs) are increased in visceral fat and contribute to insulin resistance through multiple mechanisms, including c‐Jun N‐terminal kinase (JNK) activation and expression of TNFα. Given that insulin‐like growth factor‐1 (IGF‐1)‐mediated proliferation is impaired in omental compared to subcutaneous (SC) preadipocytes, we investigated IGF‐I anti‐inflammatory action in preadipocytes from SC and omental adipose tissue.Design and MethodsPreadipocytes isolated from abdominal SC and omental fat of obese subjects were studied in primary culture. Cells were exposed to FFAs with or without IGF‐I pretreatment followed by analysis of cytokine expression and JNK phosphorylation. Lentivirus infection was used to express a constitutively active AKT (myr‐AKT) in omental preadipocytes.ResultsFFAs increased the expression of tumor necrosis factor (TNF)α, interleukin (IL)‐6, and monocyte chemotactic protein (MCP)‐1 in SC and omental preadipocytes. IGF‐I pretreatment reduced FFA‐induced JNK1 phosphorylation and TNFα expression in SC but not omental preadipocytes. Treatment with the JNK1/2 inhibitor SP600125 reduced FFA‐induced expression of TNFα. FFAs and MALP‐2, a specific TLR2/6 ligand, but not specific ligands for TLR4 and TLR1/2, increased JNK1 phosphorylation. IGF‐I completely inhibited MALP‐2‐stimulated phosphorylation of JNK1. Expression of myr‐AKT in omental preadipocytes inhibited FFA‐stimulated JNK1 phosphorylation.ConclusionsIGF‐I attenuated FFA‐induced JNK1 phosphorylation and TNFα expression through activation of AKT in human subcutaneous but not omental preadipocytes.

Abstract Development of efficacious strategies for eradication of glioblastoma multiforme (GBM) remains a significant challenge. While treatment of GBM with radiation and temozolomide (TMZ) has improved clinical outcome, lack of long-term efficacy can be due to the ability of GBM to acquire resistance to therapy by modulation of signaling pathways that control DNA repair and cell survival. The HDM2 antagonist, Nutlin3, blocks interactions of HDM2 with key signaling molecules such as p53, p73α, and HIF1α, and modulates their downstream effector function. A panel of human GBM cell lines is currently being used to monitor the extent to which Nutlin3 can sensitize cells to TMZ. In vitro survival assays indicate that Nutlin3 potentiates TMZ-mediated cell death; analysis of cell survival data using Dose Effect CalcuSyn software indicates that the combination index value was < 1 and indicative of a synergistic effect between Nutlin3 and TMZ. Western analyses of p53 wild-type GBM indicated increased p53 stabilization and activation of down-stream targets in cells treated with the combination of Nutlin3/TMZ compared to vehicle, TMZ, or Nutlin3. In addition, time-course studies indicated that U87-MG cells treated with Nutlin3/TMZ in comparison to single agent resulted in increased γ-H2AX and DNA-strand breaks. Efficacy studies using ectopic U87-MG xenografts indicated that 2 cycles of 5-consecutive days of Nutlin3/TMZ led to a significant decrease in tumor growth compared to vehicle or single agent (p < 0.001) with no signs of off-target toxicity. Regulation of DNA repair was further analyzed using the primary GBM.10 line that when grown as an ectopic xenograft, expresses the direct-reversal repair protein, O6-methylguanine DNA methyltransferase (MGMT) and proteins involved in base-excision repair (BER). In vivo target-validation studies using GBM.10 xenografts confirmed that repair of TMZ-mediated DNA damage is modulated in the presence of Nutlin3. GBM.10 xenografts were treated with vehicle, TMZ, nutlin3, or TMZ/Nutlin3 and modulation of critical targets determined by Western analyses. Increases in total p53, p53 phosphorylated at residue serine16, p73α, and substantial decreases in MGMT and the BER protein, APE-1, were observed following treatment with Nutlin3/TMZ compared to vehicle or single-agent exposure. Busso et al (Oncogene 2009) recently reported that HDM2 can monoubiquitinate APE-1 leading to proteosomal-mediated degradation. In addition, recombinant activity assays in our laboratory indicate that HDM2 directly monoubiquitinates recombinant MGMT and may play a role in its degradation. These data suggest that DNA repair proteins required for repair of TMZ-mediated DNA damage can be downregulated by altering HDM2-mediated signaling. Combination therapy that targets the p53-HDM2 E3 ligase network and DNA repair represents a novel approach towards improving treatment of GBM. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 5373.