Variety Of Solid Tumor Malignancies Research Articles

Transcriptional Co-Activator P300 Promotes Extra-Medullary Disease Formation and Myeloma Disease Progression

The transcription co-activator P300 regulates gene expression by histone H3 lysine 27 acetylation (H3K27ac) of chromatin and transcriptional regulators. P300 has been suggested to control a variety of biological functions, including differentiation, cell-cycle control, proliferation, apoptosis, and DNA damage response. The clinicopathologic/prognostic importance of P300 in multiple myeloma (MM) is unknown to date. P300 has been linked to the induction of epithelial-to-mesenchymal transition (EMT) and metastasis in a variety of solid tumor malignancies. Like EMT in solid tumors, extra-medullary disease (EMD) in MM relates to a poor prognosis and resistance to treatment. Similarly, the molecular changes concerning EMD in MM are poorly understood, and the role of P300 in MM EMD is yet to be investigated. We have employed chemical and functional genomics to unravel the role of P300 in MM disease progression and EMD formation. Six human MM cell lines-MM1S, KMS-18, H929, U266, OPM-2, and AMO-1-were used to construct lentiviral shRNA-mediated P300 knockdown (K/D) cells. In addition, to analyzing the effect of time-dependent P300 degradation, we have used a proteolysis-targeting chimera (PROTAC) compound termed “JQAD1” that selectively targets P300 for degradation. Treatment with JQAD1 effectively reduced the survival with an IC 50 of 50 nM in MM1S cells treated for 72 h. Furthermore, we wanted to understand the mechanism of P300 degradation-mediated inhibition of cell proliferation. Cell cycle analysis revealed a 40% increase in G0/G1 phase cell cycle arrest following treatment with 50 nM JQAD1 for 72 h. Blocking CD138 receptors is known to enhance MM sensitivity to proteasome inhibitors and significantly reduce tumor size. Immuno-flowcytometry analysis revealed that loss of P300 reduces CD138 expression in MM cells, suggesting their susceptibility to proteasome inhibitors. Next, we wanted to understand if P300 plays a role in developing EMD in MM. Boyden chamber-based migration and invasion assays exhibited reduced migration and invasion of P300 K/D cells with respect to control cells. Furthermore, to understand the distribution and localization of P300 silenced MM cells in vivo, we have used immune deficient NSG mice and studied MM progression after intravenous tumoral injection. In vivo studies revealed that P300 silencing led to reduced MM liver, kidney homing, and EMD formation, as observed through bioluminescence imaging, histology, and flow cytometry analysis, respectively. Further investigations are currently underway to find novel interaction partners and potential targets of P300 in MM and to delineate its mechanism of action. In conclusion, our research indicates that P300 is critical for the development of EMD and the progression of MM. This could potentially serve as a new therapeutic target for MM relapse and dissemination.

Abstract 3325: ERK inhibitor increases cancer stem cell population in NSCLC through Slug-Mediated epithelial-to-mesenchymal transition

Abstract The aberrant activation of the Mitogen-activated protein kinase pathway (MAPK/ERK pathway) has been found to play an important role in many cancers including non-small-cell lung cancer (NSCLC). Multiple inhibitors targeting this pathway have been developed to treat cancer patients harboring the aberrant MAPK pathway genes. ERK inhibitors (ERKi) have been shown to be effective in a variety of solid-tumor malignancies, but like other MAPK inhibitors, they rarely confer complete and durable responses. Therefore, it is important to define the mechanism underlying potential tumor relapse following ERKi treatment. In this study, we found that ERKi treatment expanded the cancer stem cell (CSC) population in various NSCLC cell lines and a NSCLC patient-derived xenograft (PDX) harboring aberrant MAPK signaling. Such expansion of the CSC population is mainly caused by enhanced epithelial-to-mesenchymal transition (EMT)-mediated cancer cell dedifferentiation. Mechanistically, we revealed that ERK inhibition induces EMT via pSTAT3-mediated upregulation of EMT-transcription factor Slug. Specifically, we found that ERK inhibition increased phosphorylation of STAT3 at Y705, which further increased the expression of miR-204. Increased miR-204 then downregulated expression of SPDEF, a transcriptional repressor of Slug, thereby derepressing Slug expression. Moreover, we found that a low expression of Slug is necessary for maintaining stemness properties of CSCs. Thus, ERKi treatment-increased Slug must be downregulated to allow these cells to fully acquire stemness, and this process is mediated by a Slug-miR-200c negative feedback loop. Finally, we demonstrated that the JAK/STAT pathway inhibitor Ruxolitinib can block the ERK inhibitor-induced EMT and CSCs expansion, providing a feasible combination inhibitor approach for treating NSCLC patients. Overall, these studies provide a better understanding of the tumor relapse following ERKi treatment, and define the role of Slug in de novo production of CSCs and their maintenance. Citation Format: Shurui Cai, Na Li, Xuetao Bai, Ananya Banerjee, Lavudi Kousalya, Qi-en Wang. ERK inhibitor increases cancer stem cell population in NSCLC through Slug-Mediated epithelial-to-mesenchymal transition [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3325.

Repurposing Itraconazole As an Anti-Cancer Agent Against Diffuse Large B-Cell Lymphoma

Introduction: Results from the Retrospective Non-Hodgkin Lymphoma Research (SCHOLAR-1) study demonstrated that relapsed/refractory Diffuse Large B cell Lymphoma (r/r DLBCL) have a low response rate to third-line therapy (20-30%) and an overall survival of 6 months. It stresses the need to develop novel therapeutic strategies for patients with r/r lymphoma. Itraconazole (ITZ) is a common widely clinical used orally bioavailable azole antifungal drug, with a more than thirty years well-documented safety record. Recently, scientific evidence has highlighted ITZ anti-tumor effects at clinically achievable doses in patients with variety of solid tumor malignancies. However, little is known about ITZ activity in B-cell lymphoma, especially in chemotherapy resistant lymphoma. Here we are the first group to report that potent anti-tumor activity of ITZ as a single agent or combined with other chemotherapeutic drugs in both sensitive and resistant lymphoma pre-clinical models. We also further explored the mechanism of action of ITZ to overcome drug resistance.Methods: A panel of rituximab-chemotherapeutic therapy-sensitive (TSCL), -resistant (TRCL) cell lines, developed in our laboratory, and primary tumor cells isolated from B-cell lymphoma patients were exposed to escalating doses of ITZ (0-20uM). IC50 values were calculated using the GraphPad Prism6 software. Changes in cell viability, cell number and cellular ATP levels were evaluated using the Presto Blue, Trypan Blue and Titer Glo assay, respectively. Mitochondrial respiration rate indicated as oxygen consumption rate (OCR) were measured by an XFe96 flux analyzer. Mitochondrial membrane potential (MMP) were quantified by DiOC6 staining and flow cytometry. Subsequently, TSCL and TRCL cells were exposed to ITZ alone or in combination with various chemotherapy agents or small inhibitors and differences in cell viability was determined. Coefficient of synergy was calculated using the CalcuSyn software. HKII-VDAC complex disruption by ITZ was identified by co-immunoprecipitation and western blot. HKII knockdown was generated by using ON-TARGET plus SMART pool siRNA containing a mixture of 4 siRNAs designed to specifically target HKII. Standard 51Cr release assays were conducted to study the effects of ITZ on the anti-tumor activity of monoclonal antiCD20 antibodies against TSCL and TRCL cell lines. For in vivo studies, severe combined immunodeficiency (SCID) mice were inoculated via tail vein injection (iv) with Raji cells (day 0) and assigned to observation, rituximab (at 10mg/kg/dose), obinotuzumab (at 10mg/kg/dose) with or without ITZ. Anti-tumor activity was evaluated by functional imaging and length of survival.Results: We found that ITZ induced a dose-dependent cell death in TSCL, TRCL and primary patient lymphoma cells. Single agent of ITZ decreased cell growth, reduced ATP production, lower the mitochondrial respiration rate and MMP in both TSCL and TRCL. ITZ exhibited additive/synergistic effects when combined with chemotherapeutic drugs (dexamethasone and cisplatin), proteasome inhibitors (bortezomib, carfilzomib, or ixazomib), targeted inhibitors (ABT199 and A-1210477 [an MCL-1 inhibitor]). At the molecular level, we found that, ITZ disrupted HKII/VDAC complex formation. Silencing HKII in TRCL attenuated the killing effect of ITZ. More importantly, ITZ in combination with rituximab or obinutuzumab led to reduced tumor intensity by functional imaging than rituximab or obinotuzumab single agent in vivo.Conclusion: Our findings support the potential benefits for using ITZ as an anti-cancer drug are: 1) reduce costs, 2) clear defined toxicity profile, 3) availability, and 4) pre-clinical activity in rituximab-chemotherapy resistant lymphomas. ITZ can be an inexpensive and effective agent to enhance standard chemotherapy in B-cell lymphoma. Repurposing of ITZ can result in novel therapeutic options for lymphoma patients. (Research supported by Roswell Park Cancer Institute Alliance Grant, National Natural Science Foundation of China (NSFC) 81573063) DisclosuresNo relevant conflicts of interest to declare.

Abstract 1817: C/EBPβ antagonist peptide, ST101, attenuates oncogenic gene transactivation in cancer cells to drive antitumor activity

Abstract Transcription factor dysregulation is common in cancer, resulting in aberrant gene expression that drives oncogenesis. Antagonism of oncogenic transcription factor activity, by disrupting essential protein-protein interactions needed for activation of downstream effector molecules or association with DNA, represents a powerful approach to target this previously ‘undruggable' class of proteins. CCAAT/Enhancer Binding Protein Beta (C/EBPβ) is a transcription factor overexpressed in many cancers that regulates expression of factors that promote tumor survival, proliferation and inhibit differentiation. Here, we decribe the anti-tumor activity of ST101, a cell-penetrating peptide antagonist of C/EBPβ. To demonstrate ST101 disruption of the interaction of C/EBPβ with co-factor activating transcription factor 5 (ATF5), a competition ELISA assay was performed. Recombinant ATF5 associated with plate-bound C/EBPβ with a binding affinity of ∼1 nM. Addition of ST101 inhibited this interaction in dose-dependent manner, resulting in an IC50 of 25 nM. To demonstrate ST101 disruption of C/EBPβ phosphorylation and gene transactivation in cancer cells, western blot analysis and quantitative polymerase chain reaction (qPCR) were performed on U251 glioblastoma, MCF7 breast adenocarcinoma and A549 lung adenocarcinoma cells. Administration of ST101 resulted in a dose-dependent decrease in C/EBPβ activation, as evidenced by a decrease in Thr189 phosphorylation. Anatgonism of C/EBPβ activity resulted in a dose-dependent decrease in mRNA expression of genes involved in survival (BCL2 and the baculoviral inhibitor of apoptosis factors BIRC3, BIRC5), inhibition of differentiation (Inhibitor of DNA binding proteins ID1, ID2 and ID3) and proliferation (cyclins CCNB1 and CCNA2 and cyclin-dependent kinases CDK1 and CDK2). Finally, in a mouse xenograft model, 25mg/kg ST101 administered three times per week for three weeks resulted in significant and sustained tumor growth inhibition in U251 subcutaneous tumors, both when ST101 administration was initiated early (day 2, 200 mm3 tumors, p<0.05) or late (day 16, >500mm3 tumors, p<0.05). These data demonstrate the therapeutic potential of systemic administration of ST101 and support clinical development of ST101 as a potent peptide therapeutic for a variety of solid tumor malignancies. Citation Format: Jim A. Rotolo, Rick Ramirez, Mark Koester, Erin Gallagher, Siok Leong, Lila Ghamsari, Gene Merutka, Barry J. Kappel. C/EBPβ antagonist peptide, ST101, attenuates oncogenic gene transactivation in cancer cells to drive antitumor activity [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1817.

Oligometastatic Disease and Local Therapies: A Medical Oncology Perspective.

Numerous studies in a variety of solid tumor malignancies have demonstrated prolonged progression-free and overall survival with the addition of definitive local therapies to systemic therapies in patients with a limited number of metastases. A subset of patients with oligometastases (1-5 metastases) may experience long-term disease remission or cure after local therapies such as surgery or stereotactic body radiation therapy to metastatic sites. This article reviews the literature in oligometastatic disease and considers a theoretical rationale for a curative approach in a subset of oligometastatic solid tumor patients. In oligometastatic colorectal cancer patients with liver-only metastases and in non-small cell lung cancer patients with disease control after primary therapy and with limited nodal involvement, aggressive local therapies should be considered. Clinical trials and further biomarker validation across disease types are necessary to clarify which subsets of patients may define a theorized "oligometastatic state" and therefore benefit from aggressive local therapies.

B cell regulation of the anti-tumor response and role in carcinogenesis

The balance between immune effector cells such as T cells and natural killer cells, and immunosuppressive Treg cells, dendritic, myeloid and monocytic sub-populations in the tumor microenvironment acts to calibrate the immune response to malignant cells. Accumulating evidence is pointing to a role for B cells in modulating the immune response to both solid tumors and hematologic cancer. Evidence from murine autoimmune models has defined B regulatory cell (Breg) subsets that express cytokines such as IL-10, TGF-β, and/or express immune regulatory ligands such as PD-L1, which can suppress T cell and/or natural killer cell responses. Multiple murine tumor models exhibit decreased tumor growth in B cell deficient or B cell depleted mice. In several of these models, B cells inhibit T cell mediated tumor immunity and/or facilitate conversion of T cells to CD4+CD25+FoxP3+ T regs, which act to attenuate the innate and/or adaptive antitumor immune response.Mechanisms of suppression include the acquisition of inhibitory ligand expression, and phosphorylation of Stat3, and induction of IL-10 and TGF-β, resulting in a Breg phenotype. Breg suppressive activity may affect diverse cell subtypes, including T effector cells, NK cells, myeloid derived suppressor cells (MDSC) and/or tumor associated macrophages. B cells may also directly promote tumorigenesis through recruitment of inflammatory cells, and upregulation of pro-angiogenic genes and pro-metastatic collagenases.Breg infiltration has now been identified in a variety of solid tumor malignancies including but not limited to ovarian, gastric, non-small cell lung cancer, pancreatic, esophageal, head and neck, and hepatocellular carcinomas. Increasing evidence suggests that recruitment of B cells and acquisition of suppressive activity within the tumor bed may be an important mechanism through which B cells may modulate innate and/or adaptive anti-tumor immunity. B cell depletion in the clinic using anti-CD20 antibodies and/or inhibitors of BTK and/or other signaling pathways, may be a useful strategy for augmenting the anti-tumor immune response.

Irreversible multitargeted ErbB family inhibitors for therapy of lung and breast cancer.

Overactivation of the ErbB protein family, which is comprised of 4 receptor tyrosine kinase members (ErbB1/epidermal growth factor receptor [EGFR]/HER1, ErbB2/HER2, ErbB3/HER3, and ErbB4/HER4), can drive the development and progression of a wide variety of malignancies, including colorectal, head and neck, and certain non-small cell lung cancers (NSCLCs). As a result, agents that target a specific member of the ErbB family have been developed for the treatment of cancer. These agents include the reversible EGFR tyrosine kinase inhibitors (TKIs) erlotinib and gefitinib; the EGFR-targeting monoclonal antibodies cetuximab and panitumumab; and the HER2-targeting monoclonal antibody trastuzumab. Lapatinib is a dual TKI that targets both EGFR and HER2. In addition, TKIs that inhibit multiple members of the ErbB family and also bind their targets irreversibly are under evaluation for the treatment of cancer. Three such compounds have progressed into clinical studies: the EGFR, HER2, and HER4 inhibitors afatinib, dacomitinib, and neratinib. Phase I studies of these agents have shown clinical activity in NSCLC, breast cancer, and other malignancies. Currently, afatinib is approved for EGFR mutation-positive NSCLC and is in development for squamous NSCLC, and dacomitinib is in phase III of clinical development for NSCLC, neratinib is in phase III of clinical development for the treatment of breast cancer, and afatinib is also in phase III development in head and neck cancer. Final results from clinical trials may lead to the potential approval of these agents in a variety of solid tumor malignancies.

Abstract 4865: Lack of efficacy of aerosolized resveratrol on B[A]P induced pulmonary adenoma formation in A/J mice.

Abstract Natural products constitute an emerging class of compounds for carcinogenesis prevention in a variety of solid tumor malignancies. Resveratrol is a compound that has outstanding efficacy on cancer prevention in in vitro carcinogenesis models but rapid solubility and metabolism limits its efficacy in animal models when delivered in the diet. However, if regional or topical delivery could be achieved effects of first pass metabolism might be avoided. In the present study, we sought to use aerosol resveratrol to examine whether it could inhibit Benzpyrene inducd pulmonary adenoma formation in A/J mice. Eight groups of 24 mice were given Benzpyrene and aerosol resveratrol was started either one week or 8 weeks after the conclusion of the Benzpyrene. Aerosolized resveratrol (1.0, 0.3, and 0.1 mg/kg) was delivered 5x per week Monday through Friday for weeks 2- 15. At the conclusion of the experiment animals were sacrificed and adenomas counted. We found that the animals tolerated the aerosol resveratrol without toxicity and without weight differences between all groups. Animals formed between 9 and 12 adenomas, on average throughout all groups. There were no effects of aerosol resveratrol on either early stage or late stage carcinogenesis, nor were there any trends between the groups. We conclude that the high water solubility of resveratrol likely allowed for its rapid transit and metabolism through the bronchioles and lung parenchyma and that possibly chemical modifications of resveratrol that allow for its deposition in the lung fields in a way to increase its residence time would allow for better efficacy. Citation Format: Donna Seabloom, Art Gailbraith, Jenny Antonides, Beverly Wuertz, Lee Wattenberg, Frank G. Ondrey. Lack of efficacy of aerosolized resveratrol on B[A]P induced pulmonary adenoma formation in A/J mice. [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 4865. doi:10.1158/1538-7445.AM2013-4865

Irinotecan combined with co-stimulatory molecule blockade prolongs survival of cardiac allografts in alloantigen-primed mice

Memory T cells play an important role in graft rejection. In this study, we investigated the potential effect of Irinotecan (CPT-11), a topoisomerase I inhibitor used in the treatment of a variety of solid tumor malignancies, on memory T cells. CPT-11 treatment alone or combined with blocking monoclonal antibodies (mAb) against co-stimulatory molecules (LFA-1 and CD154) was evaluated in the prevention of heart transplant rejection in alloantigen-primed mice. Our data suggest that CPT-11 reduced the expression of IL-2/IFN-γ and increased IL-10/TGF-β expression in both peripheral blood and within the grafts. CPT-11 could also inhibit alloresponses of memory T cells, while decreasing the proportion of CD4+ memory T cells in the spleen of the recipients and significantly reducing serum alloantibody levels. Our study highlights obvious synergistic effects of CPT-11 when combined with co-stimulatory molecule blockade in prolonging the survival of cardiac allografts in alloantigen-primed mice.

Abstract A42: Sensitivity and resistance to a p-AKT inhibitor in an erlotinib-sensitive model for lung cancer

Abstract Activating mutations in EGFR, such as the L858R mutation, are effective predictors of response to EGFR tyrosine kinase inhibitor (TKI) therapy. While most patients initially respond to EGFR TKIs, the majority relapse within 6-12 months, underscoring the importance of identifying therapies that enhance and prolong EGFR TKI activity in patients. Agents targeting additional nodes in the EGFR pathway have the potential to overcome such resistance but single-agent activity must be understood first. We utilized mice engineered to develop erlotinib-sensitive lung adenocarcinomas driven by the human EGFR L858R mutation to evaluate the efficacy of a p-AKT inhibitor, MK-2206, currently in clinical trials for a variety of solid tumor malignancies. MK-2206 concentration and target activity were assessed in short-term PK/PD studies in both the lungs and blood of tumor-bearing mice. MK-2206 was then evaluated in an efficacy study using a twice-weekly dosing regimen, both as a single agent and in combination with other TKIs. MK-2206 alone resulted in tumor regression or tumor growth inhibition in ∼50% of the mice, but exhibited greater efficacy in combination with other upstream pathway inhibitors. Temporal evaluation of tumor growth inhibition indicated that MK-2206 was most effective at early time points and did not produce a sustained pathway and growth response as treatment continued. Analysis of gene expression changes in mice treated with MK-2006 alone vs. MK-2206 in combination with EGFR TKIs highlighted the pathways perturbed by both drugs.

Interventional Radiology in Oncology: Clinical Management of Patients Undergoing Transarterial Chemoembolization for Hepatic Malignancies

Interventional radiology procedures treat a variety of solid tumor malignancies. Transarterial chemoembolization (TACE) is a minimally invasive procedure performed under fluoroscopic guidance that is used in treatment of hepatic malignancies because of hepatocellular carcinoma or malignancy that has metastasized to the liver. Patients undergoing a TACE procedure may manifest postembolization symptoms such as fever, nausea, vomiting, and abdominal pain, which may persist after discharge. Patients need to be educated on the importance of compliance with prescribed medications, clinical symptoms to be expected, and signs of potential complications that need to be reported.

Assessment of γ-H2AX levels in circulating tumor cells from patients receiving chemotherapy

Circulating tumor cells (CTCs) are prognostic markers in a variety of solid tumor malignancies. The potential of CTCs to be used as a “liquid biopsy” to monitor a patient’s condition and predict drug response and resistance is currently under investigation. Using a negative depletion, enrichment methodology, CTCs isolated from the peripheral blood of breast cancer patients with stage IV breast cancer undergoing DNA damaging therapy with platinum-based therapy were enriched. The enriched cell suspensions were stained with an optimized labeling protocol targeting: nuclei, cytokeratins 8, 18, and 19, the surface marker CD45, and the presence of the protein γ-H2AX. As a direct or indirect result of platinum therapy, double-strand break of DNA initiates phosphorylation of the histone H2AX, at serine 139; this phosphorylated form is referred to as γ-H2AX. In addition to γ-H2AX staining in specific locations with the cell nuclei, consistent with previous reports and referred to as foci, more general staining in the cell cytoplasm was also observed in some cells suggesting the potential of cell apoptosis. Our study underscores the utility and the complexity of investigating CTCs as predictive markers of response to various therapies. Additional studies are ongoing to evaluate the diverse γ-H2AX staining patterns we report here which needs to be further correlated with patient outcomes.

Abstract A128: A phase I study of liposomal-encapsulated docetaxel (LE-DT) in patients with advanced solid tumor malignancies

Abstract Background: Docetaxel is a semi-synthetic microtubule disrupting antineoplastic drug indicated for the treatment of a wide variety of solid tumor malignancies. Past clinical trials have shown a dose-response correlation when used in the treatment of prostate and breast cancer. Common dose limiting toxicities are myelosuppression and neuropathy. Liposomes are versatile drug carriers that may increase drug solubility, serve as sustained release systems, provide protection from drug degradation and drug related toxicities, and help overcome multidrug resistance mediated by P-glycoprotein or similar resistant efflux mechanisms. The aims of this phase I study were to determine the maximum tolerated dose (MTD), dose limiting toxicities (DLTs), pharmacokinetics (pK), and clinical response of LE-DT in patients with advanced solid tumor malignancies. Methods: LE-DT was administered using a standard 3+3 dose escalation schema with dose levels of 50, 65, 85, 110, and 132 mg/m2. Drug was infused over 1 hour. Premedication with dexamethasone was not required as it is with standard docetaxel therapy. In patients who experienced infusionrelated reactions pre-medication with steroids, antihistamines, and antipyretics were provided in subsequent cycles. Toxicities were recorded using NCI-CTCAE 3.0, and response was assessed using RECIST criteria. PK samples were drawn during C1 and analyzed using Win NonLin. Results: Twenty-four patients were treated in total. The total number of cycles each patient received ranged from 1 to 21 (median = 4). Dose escalation proceeded until DLTs were experienced by 2 out of 2 patients at the 132mg/m2 dose level (both Grade 4 neutropenia). When additional patients were treated at the 110 mg/m2 dose, 2 patients experienced Grade 4 neutropenia. The dose level was reduced to 85mg/m2, with one patient experiencing Grade 4 neutropenia at this level. The protocol was amended to allow G-CSF growth factor support and dose re-escalation. An additional 4 patients were treated at 110mg/m2 and none of these patients experienced Grade 4 neutropenia. At this dose level, two patients experiencing Grade 3 fatigue. No patient experienced Grade 3/4 neuropathy, even in two patients treated for 10 and 21 cycles. Two patients experienced Grade 2 neuropathy. Additional toxicities included Grade 3 anemia in three patients. Drug pharmacokinetics followed a two-compartment elimination pattern. Cmax and AUCinf were proportional to dose through the 110mg/m2 dose level with a mean clearance of 28.5L/hr/m2. At the 132mg/m2 dose Cl was 45.4 L/hr/m2. Serum half-lives at the differing dose levels ranged from 15.1 to 22.4 hr. There was no correlation between pK measures and toxicity. One patient had a confirmed partial response (PR) and another had an unconfirmed PR (8%). Eight patients (33%) had prolonged stable disease lasting more than 3 months. One patient continues to have stable disease after 21 cycles. Conclusion: LE-DT was tolerable with expected toxicities of neutropenia, anemia, and fatigue, but without water retention (edema). Importantly, in this heavily pretreated population no patient experienced clinically significant neuropathy. Clinical benefit (SD+PR) was observed in 41% of the patients. The recommended phase II dose of LE-DT is 110mg/m2 with growth factor support. Phase II studies of LE-DT in prostate and pancreatic cancers are planned. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A128.

Clinical Utility of the MDASI-BT in Patients with Brain Metastases

Symptom occurrence has been shown to predict treatment course and survival in cancer patients. The M. D. Anderson Symptom Inventory-Brain Tumor Module (MDASI-BT) was recently validated as a tool for primary brain tumor patient self-report of symptoms. This study evaluated the reliability and validity of the MDASI-BT in patients with brain metastases. Data collection included demographic and clinical factors, and the MDASI-BT (0–10 scale). Construct validity was assessed using confirmatory factor analysis, and known-group validity was evaluated by detecting group differences due to disease severity and treatment approach. For reliability, Cronbach's alpha values were computed for each subscale. A sample of 124 patients participated, of which 53.2% were women. Participants were primarily white (79.8%) and married (78.2%), and a variety of solid tumor malignancies were represented. Factor analysis revealed six underlying constructs, including affective symptoms, cognitive dysfunction, focal neurologic deficits, constitutional and gastrointestinal symptoms, and interference with life. The solution with these factors explained 68.4% of the variance. Mean symptom scores were 1.2 and 2.6, and mean interference scores were 1.8 and 4.3 for patients with good and poor Karnofsky scores, respectively ( P < 0.001). These subscales were also sensitive to opioid analgesic use, with group differences of 1.5 and 2.2 ( P < 0.001). Cronbach's alpha was 0.9 for each of the two subscales. Fatigue, sleep disturbance, drowsiness, distress, and dry mouth were the most severe symptoms. The MDASI-BT demonstrated validity and reliability in brain metastases patients and can be used to identify and monitor symptom occurrence in relation to treatment course and survival.

The utility of the MDASI-BT in assessing symptoms in patients with brain metastases

2052 Background: Symptom occurrence has been shown to predict treatment course and survival in patients with solid tumors. The M. D. Anderson Symptom Inventory-Brain Tumor Module (MDASI-BT) was recently validated in patients with primary brain. This study evaluated the reliability and validity of the MDASI-BT as well as symptom severity and prevalence in patients with brain metastases. Methods: Patients diagnosed with systemic cancer with brain metastases participated in this cross-sectional study. Data collection included demographic and clinical factors, and the MDASI-BT (0–10 scale). The average for the 22 symptoms and 6 interference items scores the MDASI-BT was computed. Construct validity was assessed using confirmatory factor analysis, and known-group validity was evaluated by detecting group differences due to disease severity and performance status. For reliability, Cronbach’s alpha values were computed for each subscale. Results: A sample of 124 patients participated, of which 53.2% females. Participants were primarily white (79.8%) and married (78.2%), with a variety of solid tumor malignancies represented. Factor analysis revealed six underlying constructs including affective, cognitive, focal neurologic deficit, constitutional, gastrointestinal, and interference with life. This solution explained 68.4% of the variance, and satisfied Harman’s criteria for model fit. Known-group validity was established for the MDASI-BT using the Karnofsky performance status (KPS). Mean symptom scores were 1.2 and 2.6, and mean interference were 1.8 and 4.3, for patients with good (90–100) and poor KPS (80 and below) respectively (p&lt;0.001). These subscales were also sensitive to opioid analgesic use with group differences of 1.5 and 2.2 (p&lt;0.001). Cronbach’s alpha was 0.9 for each of the two subscales. Fatigue, sleep disturbance, drowsiness, distress, and dry mouth were the most severe symptoms. Conclusions: The MDASI-BT demonstrated validity and reliability in patients with brain metastases and can be used to identify symptom occurrence related to the tumor and therapies. No significant financial relationships to disclose.

Cyclin E–Cdk2 Activation Is Associated with Cell Cycle Arrest and Inhibition of DNA Replication Induced by the Thymidylate Synthase Inhibitor Tomudex

Tomudex (ZD1694) is a specific antifolate-based thymidylate synthase inhibitor active in a variety of solid tumor malignancies. Studies were carried outin vitroto evaluate downstream molecular alterations induced as a consequence of the potent and sustained inhibition of thymidylate synthase by Tomudex. Twenty-four hours following the initial 2-h treatment with Tomudex, human A253 head and neck squamous carcinoma cells, not expressing p53 and p21WAF1, were accumulated with DNA content characteristic of early S phase of the cell cycle with a concomitant reduction of cells in G1 and G2/M phases. The changes in cyclin and cdk protein expression and their kinase activities were examined in control and drug-treated A253 cells. Tomudex treatment resulted in the decrease in p27kip1expression, with an increase in cyclin E and cdk2 protein expression and kinase activities 24 h after a 2-h exposure. Although cyclin A protein expression was markedly increased, cyclin A kinase activity was only slightly increased. Cyclin D1, cyclin B, cdk4, and cdc2 protein expression and kinase activities remain constant. Lack of activation of cyclin A– and B–cdc2 was associated with a reduced proportion of cells in G2/M phases. Increased cyclin E–cdk2 protein expression was accompanied by the inhibition of DNA synthesis, with a decrease in E2F-1 expression. These results propose that cyclin E–cdk2 kinase can negatively regulate DNA replication. The studies with dThyd rescue from cyclin E–cdk2 protein overexpression and growth inhibition by Tomudex indicate that increased cyclin E–cdk2 protein expression is associated with effective inhibition of thymidylate synthase and resultant dNTP pool imbalance. Provision of dThyd more than 24 h after exposure to Tomudex allowed cells to replicate DNA for a single cycle back to G1, but did not prevent the profound growth-inhibitory effect manifested in the following 5 days. Tomudex treatment resulted in a time-dependent induction of the megabase DNA fragments, followed by secondary 50- to 300-kb DNA fragmentation. The 50- to 300-kb DNA fragmentation may be derived from the inhibition of DNA synthesis associated with cyclin E–cdk2 activation. These results suggest that the megabase DNA fragmentation is induced as a consequence of inhibition of thymidylate synthase by Tomudex and kilobase DNA fragmentation may correlate with the reduction of p27kip1expression and the increase in cyclin E and cdk2 kinase activities. Activation of cyclin E and cdk2 kinases allows cells to transit from G1 to S phase accompanied by the inhibition of DNA synthesis. The changes in cell cycle regulatory proteins associated with growth inhibition and DNA damage by Tomudex are not p53 dependent.

Monocytosis in malignant disease.

The absolute monocyte count was determined in 100 normal individuals and in 100 patients with a variety of solid tumor malignancies. A significant elevation in the mean value for the latter group was found. Monocytosis is therefore of descriptive and diagnostic value in the evaluation of patients with solid tumor malignancy.