Specific Oncoproteins Research Articles

Combinatorial cancer immunotherapy strategies with proapoptotic small-molecule IAP antagonists.

Members of the inhibitor of apoptosis (IAP) family control several critical aspects of innate immunity, cell death, and tumorigenesis. Small molecule antagonists that target specific IAP oncoproteins, primarily cIAP1 and cIAP2, but potentially also XIAP and Livin, modulate distinct immune signal transduction pathways that can lead to an increased sensitivity of tumors cells to cytokine-mediated apoptosis. These antagonists are based on the structure of an endogenous cellular IAP inhibitor called Smac. Smac is normally sequestered within the mitochondria and is released into the cytoplasm upon cell death stimuli, thereby overcoming the anti-apoptotic action of the IAPs. The therapeutic usefulness of recombinant tumoricidal cytokines to treat cancer patients is principally limited due to their unacceptable adverse side effects. Therefore, investigators have sought to develop alternative regimens that do not rely on exogenously delivered death ligands. These approaches include the stimulation of the immune system with oncolytic virus-based agents or Toll-like receptor agonists in combination with Smac mimetics. Similarly, preclinical combination immunotherapy studies reveal that recombinant interferon synergizes with Smac mimetics to kill cancer. This strategy opens up new therapeutic avenues for anti-cancer therapy by modulating specific immune-mediated death pathways employing unique dual-pronged combinatorial approaches.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Abstract NG02: Rational drug combinations to improve anti-EGFR targeted therapies in metastatic colorectal cancer

Abstract NG02: Rational drug combinations to improve anti-EGFR targeted therapies in metastatic colorectal cancer

Emerging strategies for high-risk and relapsed/refractory acute myeloid leukemia: Novel agents and approaches currently in clinical trials

High-risk acute myeloid leukemia (AML) is defined by clinical and biologic features that predict for poor response to induction chemotherapy and high risk of relapse. Despite even the most aggressive and well-developed strategies for care, most patients succumb to the disease. No currently available treatment has demonstrated consistent efficacy in terms of remission induction or long-term survival. This review will highlight some of the emerging strategies to treat high-risk AML with an emphasis on clinical trials of novel strategies currently enrolling patients. Targeted molecular therapies, novel cytotoxics, and immune-based therapies are under investigation for the management of high-risk AML. Some of the agents covered include tyrosine kinase inhibitors targeted to AML specific oncoproteins, nanoparticle formulations of existing drugs, nucleoside analogs, monoclonal antibodies, chimeric antigen receptors, bispecific T-cell engaging antibodies, and vaccines. As our understanding of the biology of AML has improved, targeted therapy for AML has emerged, offering to change not only response rate, but also the nature of response. Differentiation, rather than necrosis or apoptosis, is often seen in response to targeted agents and may be seen more frequently in the future. Interventions that might be more widely used in the near future include FLT3 inhibitors and nanoparticle formulations of drugs already known to have activity in the disease. Long term immune therapy holds significant promise.

A Phase II Open-Label Study of Ganetespib, a Novel Heat Shock Protein 90 Inhibitor for Patients With Metastatic Breast Cancer

BackgroundGanetespib is a small molecule, nongeldanamycin HSP90 inhibitor with potent inhibitory effects on HSP90-dependent oncoproteins of relevance to breast cancer pathogenesis. We therefore tested ganetespib in an unselected cohort of patients with MBC. Patients and MethodsPatients were treated with single agent ganetespib at 200 mg/m2 once weekly for 3 weeks, on a 28-day cycle. Therapy was continued until disease progression. The primary end point was ORR using Reponse Evaluation Criteria in Solid Tumors version 1.1. ResultsTwenty-two patients were enrolled with a median age of 51(range, 38-70) years and a median Eastern Cooperative Oncology Group performance status of 0 (range, 0-1). Most patients had at least 2 previous lines of chemotherapy in the metastatic setting. Most common toxicities, largely grade 1/2, were diarrhea, fatigue, nausea, and hypersensitivity reaction. The ORR in this unselected population was 9%, with all responses coming from the subset of patients with HER2-positive MBC (2/13; 15%). One patient with TNBC had objective tumor regression in the lung metastases. The clinical benefit rate (complete response + partial response + stable disease > 6 months) was 9%, median progression-free survival was 7 weeks (95% confidence interval [CI], 7-19), and median overall survival was 46 weeks (95% CI, 27-not applicable). ConclusionThe study did not meet the prespecified criteria for ORR in the first stage of the Simon 2-stage model in this heavily pretreated unselected population of MBC. However, activity was observed in trastuzumab-refractory HER2-positive and TNBC. Ganetespib was well tolerated and responses in more targeted populations harboring specific HSP90-dependent oncoproteins justifies its further study, particularly as part of rational combinations.

Synthesis and Characterization of Lipid Immuno-Nanocapsules for Directed Drug Delivery: Selective Antitumor Activity against HER2 Positive Breast-Cancer Cells

Lipid nanocapsules (LNC) are usually developed as nanocarriers for lipophilic drug delivery. The surface characteristics of these colloidal particles are determinant for a controlled and directed delivery to target tissues with specific markers. We report the development of immuno-nanocapsules, in which some antibody molecules with different immuno-specificity are conjugated to the nanocapsule surface, offering the standardization of a simple method to obtain vectorized nanosystems with specific recognition properties. Nanocapsules were prepared by a solvent-displacement technique, producing an oily core coated by a functional shell of different biocompatible molecules and surface carboxylic groups. Three different antibodies (one a specific HER2 oncoprotein antibody) were conjugated with these nanoparticles by the carbodiimide method, which allows the covalent immobilization of protein molecules through carboxylic surface groups. The immuno-nanocapsules were completely characterized physico-chemically via electrokinetic and colloidal stability experiments, confirming the correct immobilization of these antibody molecules on the colloidal nanoparticles. Also, additional immunological analyses verified that these IgG-LNC complexes showed the expected specific immuno-response. Finally, different healthy and tumoral breast-cell lines were cultured in vitro with Nile-Red-loaded and docetaxel-loaded HER2 immuno-nanocapsules. The results indicate that our immuno-nanocapsules can increase their uptake in HER2 overexpressing tumoral cell lines.

Synthetic Lethality through Combined Notch–Epidermal Growth Factor Receptor Pathway Inhibition in Basal-Like Breast Cancer

Basal-like breast cancers (BLBC) are highly aggressive, yet selective therapies targeting the specific oncoproteins driving these tumors have not been developed. These cancers frequently express epidermal growth factor receptor (EGFR), with resistance to its inhibition being well documented, albeit poorly understood. Notch pathway activation is also common in this breast cancer subtype and can be suppressed by gamma-secretase inhibitors, which effectively block receptor cleavage and activation. Herein, we show that although inhibition of either EGFR or Notch signaling alone is insufficient to suppress basal-like breast tumor cell survival and proliferation, simultaneous inhibition uncovers a synthetic lethal relationship between these two oncogenic pathways. This lethality is due in part to significant decreases in AKT activation caused by combined EGFR and Notch inhibition. Expression of the activated form of Notch1 restores AKT activity and enables cells to overcome cell death after dual-pathway blockade. Combined pathway inhibition is also dramatically more effective at suppressing tumor growth in mice than blocking EGFR or Notch signaling alone. Thus, we show that Notch pathway activation contributes to resistance to EGFR inhibition, and provide a novel treatment strategy for BLBCs.

NF-κB as a target for oncogenic viruses.

NF-κB is a pivotal transcription factor that controls cell survival and proliferation in diverse physiological processes. The activity of NF-κB is tightly controlled through its cytoplasmic sequestration by specific inhibitors, IκBs. Various cellular stimuli induce the activation of an IκB kinase, which phosphorylates IκBs and triggers their proteasomal degradation, causing nuclear translocation of activated NF-κB. Under normal conditions, the activation of NF-κB occurs transiently, thus ensuring rapid but temporary induction of target genes. Deregulated NF-κB activation contributes to the development of various diseases, including cancers and immunological disorders. Accumulated studies demonstrate that the NF-κB signaling pathway is a target of several human oncogenic viruses, including the human T cell leukemia virus type 1, the Kaposi sarcoma-associated herpesvirus, and the Epstein-Bar virus. These viruses encode specific oncoproteins that target different signaling components of the NF-κB pathway, leading to persistent activation of NF-κB. This chapter will discuss the molecular mechanisms by which NF-κB is activated by the viral oncoproteins.

Battling the Hematological Malignancies: The 200 Years' War

The delineation of the hematological malignancies began near the end of the first third of the 19th century with the recognition of the similarity among cases with lymph node tumors and an enlarged spleen (Hodgkin's disease). Descriptions of chronic and acute leukemia and myeloma followed thereafter. In the first years of the 20th century the discovery of x-radiation permitted palliative orthovoltage radiation therapy of Hodgkin's disease. Following World War II, legitimate drug therapy for the hematological malignancies was introduced: nitrogen mustard, adrenocorticotropic hormone and cortisone acetate, and anti-folic acid derivatives, initially aminopterin. Today, about 14 classes of drugs (different mechanisms of action) and >50 individual agents are being used, with others under study. Several examples of agents targeting specific transcription factors or oncoproteins have been introduced. Despite remarkable progress, including the ability to cure acute leukemia in about 70% of children, cure several genetic variants of acute myelogenous leukemia in younger adults, cure some cases of lymphoma in children and younger adults, and induce prolonged remission in many affected persons, the majority of patients face an uncertain outcome and shortened life. Thus, we have much to do in the next several decades. The significant hurdles we must overcome include: the apparent infrequency of an exogenous cause that can be avoided, the exponential increase in incidence rates with age and the dramatic negative effect of aging on the results of treatment, the challenge of one trillion or more disseminated cancer cells among which are a smaller population of cancer stem cells, the profound genetic diversity of the hematological malignancies (apparently hundreds of unique genetic primary lesions), the redundant growth and survival pathways defining the cancer phenotype, the decreasing market for pharmaceutical companies as therapy becomes more specific (fewer target patients) and drug development costs become more expensive, and the significant negative long-term effects of current therapy on both children and adults. These challenges will be gradually overcome, if we (a) develop new models of cooperation among academia, industry, and government, (b) continue the growth of international participation in cancer research (more keen minds to the task), and (c) convince the governments of the world, including that of the U.S., that an investment in minimizing the effects of cancer is as important as defending against other threats to the welfare and longevity of their citizens.

Effects of Integrin α5β1-Mediated Signal TransductionPathways on the Proliferation and Apoptosis of K562 and CML Cells.

Objective and methods: More than 90% of chronic myeloid leukemia (CML) patients have the Philadelphia (Ph) chromosome which produces a specific oncoprotein known as BCR/ABL (P210 protein). Having strong tyrosine kinase activity, the BCR/ABL oncoprotein can trigger several signal transduction pathways and inhibit the normal signal transduction pathways of CML progenitor cells. Presenting on cell surface, Integrins belong to the adhesive molecule family. They mediate adhesion of cell to cell or cell to extra cellular matrix (ECM) and take part in many physiological and pathological processes such as signaling transduction, cell differentiation, proliferation and apoptosis. Many studies showed that abnormal circulating of CML Ph+ cells in peripheral blood and unregulated proliferation of CML Ph+ cells in bone marrow were related closely to the abnormal adhesion function of integrins. It is also well known that Interferon (IFN)-α is one of the effective therapeutic agents for CML. Therefore the present study was designed, by cell culture, FCM, RT-PCR and Western Blot, to observe the effects of IFNα-2b on the proliferation of k562 cells and fresh leukemia cells from patients with CML-in blast crisis (CML cells), before and after integrin α5β1 having been neutralized by Anti-integrin α5β1 monoclonal antibody (Anti-α5β1), and to investigate the changes of mRNA and protein of several key molecules in integrin-mediated pathways, such as FAK, ERK, AKT, PI3K and Cychin D1, p21 and survivin before and after the treatment of Anti-α5β1 with or without IFNα-2b.Results: The expression level of integrin α5β1 on K562 cells and CML cells was much higher than that on health volunteers' BMMNC. And the adhesion capability of K562 cells and CML cells to fibronectin (FN) decreased significantly than that of health volunteers' BMMNC. IFNα-2b improved the adhesion capability of K562 cells and CML cells to FN. Anti-α5β1 inhibited proliferation but induced apoptosis and G0/G1 arresting of K562 cells and CML cells. Anti-α5β1 downregulated the expression level of p-FAK-, p-ERK-, PI3K- and p-AKT-mRNA and protein. Anti-α5β1 also downregulated the expression level of Cyclin D1 mRNA, but upregulated the expression level of P21 mRNA. Moreover, the effect of Anti-α5β1 is stronger than that of IFNα-2b.Conclusions: Anti-α5β1 inhibited integrin α5β1-mediated signal transduction pathways: FAK-ras-ERK pathway and PI3K-AKT-survivin pathway. This might be the mechanisms of Anti-α5β1 inhibiting proliferation and inducing apoptosis of K562 cellsand CML cells. This indicated that the dysfunction of integrin α5β1 pathway might have important meanings in the leukemogenesis of CML and might become a new target for the treatment of CM.

Differential cooperation of oncogenes with p53 and Bax to induce apoptosis in rhabdomyosarcoma

BackgroundDeregulated expression of oncogenes such as MYC and PAX3-FKHR often occurs in rhabdomyosarcomas. MYC can enhance cell proliferation and apoptosis under specific conditions, whereas PAX3-FKHR has only been described as anti-apoptotic.ResultsIn order to evaluate how MYC and PAX3-FKHR oncogenes influenced p53-mediated apoptosis, rhabdomyosarcoma cells were developed to independently express MYC and PAX3-FKHR cDNAs. Exogenous wild-type p53 expression in MYC transfected cells resulted in apoptosis, whereas there was only a slight effect in those transfected with PAX3-FKHR. Both oncoproteins induced BAX, but BAX induction alone without expression of wild-type p53 was insufficient to induce apoptosis. Data generated from genetically modified MEFs suggested that expression of all three proteins; MYC, BAX and p53, was required for maximal cell death to occur.ConclusionWe conclude that cooperation between p53 and oncoproteins to induce apoptosis is dependent upon the specific oncoprotein expressed and that oncogene-mediated induction of BAX is necessary but insufficient to enhance p53-mediated apoptosis. These data demonstrate a novel relationship between MYC and p53-dependent apoptosis, independent of the ability of MYC to induce p53 that may be important in transformed cells other than rhabdomyosarcoma.

Developmental stage–selective effect of somatically mutated leukemogenic transcription factor GATA1

Acquired mutations in the hematopoietic transcription factor GATA binding protein-1 (GATA1) are found in megakaryoblasts from nearly all individuals with Down syndrome with transient myeloproliferative disorder (TMD, also called transient leukemia) and the related acute megakaryoblastic leukemia (DS-AMKL, also called DS-AML M7). These mutations lead to production of a variant GATA1 protein (GATA1s) that is truncated at its N terminus. To understand the biological properties of GATA1s and its relation to DS-AMKL and TMD, we used gene targeting to generate Gata1 alleles that express GATA1s in mice. We show that the dominant action of GATA1s leads to hyperproliferation of a unique, previously unrecognized yolk sac and fetal liver progenitor, which we propose accounts for the transient nature of TMD and the restriction of DS-AMKL to infants. Our observations raise the possibility that the target cells in other leukemias of infancy and early childhood are distinct from those in adult leukemias and underscore the interplay between specific oncoproteins and potential target cells.

How thyroid tumors start and why it matters: kinase mutants as targets for solid cancer pharmacotherapy

Treatment of patients with thyroid cancer is usually successful, and most patients are cured of the disease. However, we do not have effective therapies for patients with invasive or metastatic thyroid cancer if the disease is not surgically resectable and does not concentrate radio-iodine. Conventional external beam radiotherapy and chemotherapy are of marginal benefit. In other types of cancer, new therapies are being developed that take advantage of our knowledge of cancer pathogenesis to interfere with the activity of specific oncoproteins believed to be important in disease causation. Because these approaches are being considered for thyroid cancer, I will briefly describe in this review examples of recent breakthroughs in medical therapy of certain hematological malignancies and some solid tumors using drugs that work in this fashion, focusing in particular on compounds that block the enzymatic activity of specific tyrosine kinase oncoproteins. It should be noted, however, that cancers commonly harbor mutations or other disruptions of many genes, each of which could conceivably play a role in disease pathogenesis. This makes the choice of molecular target a difficult and critical decision if these approaches are to succeed. Here I will argue that priority should be given to blocking the function of oncoproteins activated early in tumor development. We have a fairly good understanding of the genetic changes involved in thyroid cancer initiation, and hence these cancers may prove to be particularly well suited for oncoprotein-specific therapies.

RNA molecules as anti-cancer agents

The inhibition of cancer growth and progression is one of the major challenges facing modern medicine. Despite significant progress in the development of therapies against cancer, only in a few cases are these therapies effective. Because cancer is a complex disease, agents that target a single oncogenic pathway have low efficacy, in addition to allowing the emergence of drug resistance. There is a need for specific therapy, which can affect a broad range of cancers, with minimal side effects. Here we summarize several novel anti-cancer strategies that answer the above criteria. These strategies utilize the multiple anti-proliferative effects of double stranded RNA (dsRNA): (1) the classical antisense RNA hybridizes with its target mRNA, leading to reduced levels of a specific oncoprotein; (2) short dsRNA of specific sequence, known as small inhibitory RNA (siRNA), can selectively and efficiently inhibit expression of specific oncogenes, expressed in cancer cells but not in normal cells. Shutting down the expression of cancer-promoting genes by siRNA has proven to be an effective approach against several cancers; (3) long dsRNA, frequently expressed in cells infected with viruses, activates mechanisms that efficiently kill the infected cells, thereby preventing spread of the virus. The dsRNA killing strategy (DKS), involving the in situ generation of dsRNA of sufficient length to induce antiviral defenses specifically in cancer cells, is a novel strategy developed in our laboratory. DKS has the potential to be applicable to a wide range of cancers. Thus dsRNA-based anti-cancer strategies could be powerful tools for cancer treatment.

Estrogen-regulated Conditional Oncoproteins: Tools to Address Open Questions in Normal Myeloid Cell Function, Normal Myeloid Differentiation, and the Genetic Basis of Differentiation Arrest in Myeloid Leukemia

Neutrophils, monocytes and dendritic cells are effectors of innate immunity and essential coactivators in the acquired immune response. Understanding the biochemical basis of their mature cell functions, their differentiation from hematopoietic progenitors, and the mechanisms by which myeloid leukemia oncogenes block their differentiation programs, continue to be areas of active research. Four major problems limit progress in these fields. First, the biochemical analysis of mature cells is limited by the time and cost of purifying neutrophils, monocytes, or dendritic cells from wild-type and genetically modified mouse strains. Second, while immortal myeloid cell lines are used to understand the transcriptional basis of normal terminal differentiation following their treatment with differentiation-promoting agents (e.g. G-CSF, IL-6, RA, TPA), these cells contain stable defects responsible for their immortalization, and the degree to which they model normal differentiation is often incomplete. Third, these same inducible cell lines are used as model systems to determine how myeloid oncoproteins prevent differentiation; however, oncoproteins that block differentiation of marrow progenitors cultured in GM-CSF or IL-3 but permit their differentiation in response to G-CSF or RA, do not score effectively in these assays (e.g. Hoxa9, Mll-Enl). Fourth, there is no reproducible method to derive myeloid progenitor lines that execute predictable terminal differentiation to neutrophils, monocytes, or dendritic cells. Developing this type of system is needed to evaluate how myeloid gene inactivation by knockout technologies alters lineage-specific differentiation and mature cell function.Conditional myeloid oncoproteins provide a tool to solve these research problems by providing a predictable and inexpensive means of expanding, in culture, GM-CSF– or IL-3-dependent myeloid progenitors from any genotype, and by permitting their synchronous differentiation to neutrophils, monocytes, or dendritic cells under defined culture conditions following inactivation of the conditional oncoprotein. This system of conditionally immortalizing normal bone marrow precursors provides the large numbers of normal cells required for analysis of cell biology and protein biochemistry, and further provides a model system in which to study the genetic mechanisms controlling terminal differentiation and how specific oncoproteins expressed in the cell lines prevent this differentiation program. The ability to derive conditionally-immortalized progenitor lines from knock-out mice provides cell lines for the reconstitution of knockout gene function and subsequent dissection of knockout protein function by mutational analysis. Finally, conditional myeloid cell lines can be established from both ES cells and from d10 fetal liver cells, allowing for the analysis of embryonic lethal mutants on both the maturation and terminal differentiation of mature myeloid cells. In this review, we summarize the importance and limitations of current approaches in myeloid cell research, and how estrogen-regulated conditional oncoproteins help to solve these problems.

Current concepts in treatment of childhood acute lymphoblastic leukemia

Remarkable progress has been made in the treatment of childhood acute lymphoblastic leukemia (ALL). The 5-year event-free survival rates now exceed 70%, reaching 80% or higher in some studies. Current efforts to improve outcome focus on more precise risk classification to avoid over- or under-treatment. Early treatment response, as defined by measurement of minimal residual disease, reflecting both the drug responsiveness of leukemic cells and host pharmacodynamics and pharmacogenomics, is considered the most reliable indicator for gauging the intensity of treatment. Intensive remission induction may not be necessary for patients with standard- or high-risk ALL, provided that they receive post-induction intensification therapy. Several clinical trials suggested that the use of L-asparaginase in the post-induction period only yielded an excellent remission induction rate and long-term event-free survival with low early morbidity. Increasingly, dexamethasone has replaced prednisone as a glucocorticoid of choice to improve outcome. Extended and stronger double-delayed intensification (or reinduction) improved treatment results even in patients with high-risk ALL and a slow early response. A methotrexate dosage of 2.5 g/m2 seems adequate for most patients with B-lineage ALL; 5 g/m2 may be required for those with T-cell ALL. The most successful clinical trials generally feature continuous post-remission therapy; high-dose pulse therapy with long rest period to compensate for myelosuppression was associated with an inferior outcome. Intensity of CNS-directed therapy should be tailored according to the presenting features. Patients with high-risk genetic features, a large leukemic cell burden, T-cell ALL, and the presence of leukemic cells in CSF (even from iatrogenic introduction via traumatic lumbar puncture) require more intensive therapy. In the context of intensive intrathecal and effective systemic therapy, cranial irradiation can now be omitted in over 95% of the patients. Among very high-risk cases, only Philadelphia chromosome-positive ALL cases and perhaps those with T-cell ALL and poor early response benefit from hematopoietic stem cell transplantation with HLA-matched related donor. Ultimately, the curative treatment for very high-risk ALL would depend on the identification and characterization of specific oncoproteins with a pivotal role in leukemogenesis, and the development of molecular-based therapy. Promising agents under evaluation include imatinib mesylate (STI 571) for Ph+ ALL and FLT3 inhibitors forMLL-rearranged leukemia.

Expression of microtubule-associated protein tau by gastrointestinal stromal tumors

The purpose of this work was to study the expression in gastrointestinal stromal tumors (GISTs) of various antigens, including the protein tau associated with enteric neuronal differentiation; to compare their expression with that of c-kit, known to be associated with interstitial cell of Cajal differentiation; and to correlate their expression with the observation of ultrastructural features of gastrointestinal autonomic nerve tumors. Twenty-six GISTs of the stomach and 16 GISTs of the small bowel were included in the study group. Thirty-five tumors served as controls. Tissue sections were immunostained with vimentin, CD34, desmin, specific smooth muscle actin, S100 protein, neuron-specific enolase, PGP9.5, neurofilament, bcl-2 oncoprotein, synaptophysin, chromogranin A, c-kit, and tau. Twenty-one of these tumors were also analyzed ultrastructurally. Of the 42 GISTs, 28 were predominantly spindled, 7 were predominantly epithelioid, and 7 were a mixture of epithelioid and spindle cells. Ten primary GISTs were classified as benign, 9 as borderline, and 23 as malignant. Metastatic dissemination was present at primary surgery in 1 case and eventually developed in 6 patients. Six disease-related deaths were counted. In normal submucous and myenteric plexuses of stomach and small bowel, ganglion cell bodies and nerve fibers strongly expressed tau. Twenty (76.9%) GISTs of the stomach and 12 (75%) of the small bowel expressed tau. Tau often showed intense, diffuse staining patterns in both spindled and epithelioid tumors. Ten (100%) of the 10 benign GISTs, 7 (77.8%) of the borderline GISTs, and 15 (65.2%) of the 23 frankly malignant GISTs expressed tau. Thirty-six GISTs expressed at least 2 different neuronal markers. A coexpression of the neuronal markers and c-kit was observed in 90% of GISTs. The expression of tau was observed in 12 of the 15 GISTs with dense core granules, considered as the definitive finding for a diagnosis of gastrointestinal autonomic nerve tumors. Ten of these also expressed c-kit; 9 were malignant. Tau also immunostained other intra-abdominal tumors, including neuroendocrine carcinomas, paragangliomas and desmoplastic round cell tumors. This immunohistochemical study shows that GISTs are specific tumors of the digestive tract and are nearly always characterized by simultaneous neuronal and interstitial cell of Cajal differentiation. Although the loss of tau expression is observed only in borderline and malignant tumors, its prognostic value is not clear cut. HUM PATHOL 32:1166-1173. Copyright © 2001 by W.B. Saunders Company

Overview of recent topics in clinical pharmacology of anticancer agents.

The rationale for studying the clinical pharmacology of antineoplastic agents is that the information obtained will result in enhanced drug development and enhanced or improved clinical use. A great deal of effort has been expended in studying the pharmacokinetics and pharmacodynamics of investigational and noninvestigational antineoplastic agents. More recently, a deeper appreciation has developed regarding the importance of the metabolism of antineoplastic agents and the potential role of metabolites in their activity or toxicity, as well as the potential for drug-drug interactions. Investigators studying the clinical pharmacology of antineoplastic agents face an increasingly challenging task as new agents continue to be developed. Some of these challenges arise from the enhanced potency of new agents, resulting in increased difficulty in measuring such agents in biological matrices. Furthermore, as agents have been developed to affect specific biological targets, the necessity of assessing pharmacodynamics at the biochemical or molecular level has become increasingly important. In addition, development of agents with cytostatic, as opposed to cytotoxic, properties poses a further challenge to assessment of pharmacologic effect. In addressing these challenges, a great deal of effort has been expended to develop increasingly sensitive analytical chemical techniques, in evaluating alternative biological matrices, such as saliva, in which to monitor drug concentrations in a less invasive fashion, and in developing limited sampling strategies to assess both the pharmacokinetics and pharmacodynamics of antineoplastic agents. Similarly, a great deal of effort has been expended in providing suitable means for assessing the numerous novel targets for which antineoplastic agents are being developed. These include the assessment of cell cycle kinetics and specific oncoproteins, definition of cell damage such as cleavable complexes, and formation of drug-macromolecular adducts in suitable target cells. Additional effort is being expended to explore nontraditional means of drug delivery. In this regard, the increasing importance of orally administered agents reflects a fundamental change in the approach to antineoplastic drug delivery. Finally, the increased computational power made available by faster personal computers has facilitated a number of innovative modeling techniques involving population modeling, modeling of combination chemotherapy, and assessment of drug-drug interactions.

The gene encoding the granulocyte colony-stimulating factor receptor is a target for deregulation in pre-B ALL by the t(1;19)-specific oncoprotein E2A-Pbx1.

Approximately 25-30% of childhood pre-B cell acute lymphoblastic leukemias (pre-B ALL) is characterized by the presence of a (1;19)(q23;p13.3) translocation. The presence of this translocation is generally accompanied by a poor prognosis. The chimeric gene resulting from this chromosomal rearrangement encodes a hybrid transcription factor, E2A-Pbx1. In an attempt to delineate the genetic cascade initiated by E2A-Pbx1, we sought to identify genes that are deregulated by this transcription factor in t(1;19) pre-B ALL. We show here that the gene encoding the granulocyte colony-stimulating factor receptor (G-CSFr) is specifically upregulated in pre-B cells expressing E2A-Pbx1. G-CSFr is also expressed in cell lines established from t(1;19) pre-B cell leukemia and on primary t(1;19) tumor cells, but not on control cells. These data indicate that G-CSFr gene is a target for deregulation by E2A-Pbx1.

MAP kinase is constitutively activated in gip2 and src transformed rat 1a fibroblasts.

Rat 1a fibroblasts transformed by the Gi2 oncogene, gip2, exhibit a constitutively elevated mitogen-activated protein (MAP) kinase activity that correlates with enhanced tyrosine phosphorylation of the p42 MAP kinase polypeptide. The MAP kinase activity in gip2 transformed cells is 50-60% of the pertussis toxin-sensitive, thrombin-stimulated activity observed in wild-type Rat 1a cells. A similar activation of MAP kinase is observed in src but not ras or raf transformed Rat 1a cells, indicating that the persistent MAP kinase activity results from the action of the specific oncoprotein and is not the consequence of cellular transformation. The enhanced transactivation function of c-Jun characteristic of the transformed phenotype, measured using a collagenase promoter-CAT reporter gene, is observed in gip2, src, ras, and raf transformed Rat 1a cells. The regulatory networks controlled by the four transforming oncogenes therefore alter the activity of specific transcription factors, but only gip2 and src constitutively activate MAP kinase. The findings demonstrate that the catalytic activity of growth factor-regulated cytoplasmic kinases are selectively and stably activated as a consequence of specific oncogene expression.