Robert L. Saylors

STAT3 Activation in Stromal/Osteoblastic Cells Is Required for Induction of the Receptor Activator of NF-κB Ligand and Stimulation of Osteoclastogenesis by gp130-utilizing Cytokines or Interleukin-1 but Not 1,25-Dihydroxyvitamin D3 or Parathyroid Hormone

Interleukin (IL)-6-type cytokines stimulate osteoclastogenesis by activating gp130 in stromal/osteoblastic cells and may mediate some of the osteoclastogenic effects of other cytokines and hormones. To determine whether STAT3 is a downstream effector of gp130 in the osteoclast support function of stromal/osteoblastic cells and whether the gp130/STAT3 pathway is utilized by other osteoclastogenic agents, we conditionally expressed dominant negative (dn)-STAT3 or dn-gp130 in a stromal/osteoblastic cell line (UAMS-32) that supports osteoclast formation. Expression of either dominant negative protein abolished osteoclast formation stimulated by IL-6 + soluble IL-6 receptor, oncostatin M, or IL-1 but not by parathyroid hormone or 1,25-dihydroxyvitamin D3. Because previous studies suggested that IL-6-type cytokines may stimulate osteoclastogenesis by inducing expression of the tumor necrosis factor-related protein, receptor activator of NF-κB ligand (RANKL), we conditionally expressed RANKL in UAMS-32 cells and found that this was sufficient to stimulate osteoclastogenesis. Moreover, dn-STAT3 blocked the ability of either IL-6 + soluble IL-6 receptor or oncostatin M to induce RANKL. These results establish that STAT3 is essential for gp130-mediated osteoclast formation and that the target of STAT3 during this process is induction of RANKL. In addition, this study demonstrates that activation of the gp130-STAT3 pathway in stromal/osteoblastic cells mediates the osteoclastogenic effects of IL-1, but not parathyroid hormone or 1,25-dihydroxyvitamin D3.

Cyclophosphamide Plus Topotecan in Children With Recurrent or Refractory Solid Tumors: A Pediatric Oncology Group Phase II Study

PURPOSE To determine the response rate of the combination of cyclophosphamide and topotecan in pediatric patients with recurrent or refractory malignant solid tumors. PATIENTS AND METHODS A total of 91 pediatric patients, 83 of whom were fully assessable for response and toxicity, received cyclophosphamide (250 mg/m2/dose) followed by topotecan (0.75 mg/m2/dose), each given as a 30-minute infusion daily for 5 days. All patients received filgrastim (5 mcg/kg) daily until the absolute neutrophil count (ANC) was > or = 1,500 microL after the time of the expected ANC nadir. RESULTS A total of 307 treatment courses were given to the 83 fully assessable patients. Responses (complete response plus partial response) were seen in rhabdomyosarcoma (10 of 15 patients), Ewings sarcoma (six of 17 patients), and neuroblastoma (six of 13 patients). Partial responses were seen in two of 18 patients with osteosarcoma and in one patient with a Sertoli-Leydig cell tumor. Twenty-three patients had either minor responses (n = 6) or stable disease (n = 17); the median number of courses administered to patients with partial or complete response was six (range, two to 13 courses), and the median administered to those with stable disease was three (range, one to 11 courses). The toxicity of the combination was limited principally to the hematopoietic system. Of 307 courses, 163 (53%) were associated with grade 3 or 4 neutropenia, 84 (27%) with grade 3 or 4 anemia, and 136 (44%) with grade 3 or 4 thrombocytopenia. Despite the severe myelosuppression, only 34 (11%) of 307 courses were associated with grade 3 or 4 infection. Nonhematopoietic toxicity of grades > or = 3 was rare and consisted of nausea and vomiting (two courses), perirectal mucositis (one course), transaminase elevation (one course), and hematuria (two courses). CONCLUSION The combination of cyclophosphamide and topotecan is active in rhabdomyosarcoma, neuroblastoma, and Ewings sarcoma. Stabilization of disease was seen in osteosarcoma, although objective responses were rare in this disease. The therapy can be given with acceptable hematopoietic toxicity with the use of filgrastim support.

Neuroblastoma: Effect of genetic factors on prognosis and treatment

Background and Methods. Genetic analysis of tumor tissue has provided considerable insight into mechanisms of malignant transformation and progression. Neuroblastomas have been studied by cytogenetics, flow cytometry, and molecular genetic techniques, and these studies have identified several specific abnormalities that allow subclassification of these tumors into genetic/ clinical subtypes.

Secondary acute myelogenous leukemia following safe exposure to etoposide.

PURPOSE To present two patients as illustrations of the risk of developing secondary acute myelogenous leukemia (sAML) when theoretically safe doses of etoposide (VP-16) are used. PATIENTS AND METHODS Patient no. 1 was a 15-year-old white girl diagnosed with stage IIa Hodgkins disease. She was treated with a combination of vincristine, doxorubicin, bleomycin, and VP-16 (2 g/m2 total) over 4 months, followed by 25.5 Gy of involved-field radiotherapy. Patient no. 2 was an 11-year-old white boy diagnosed with virus-associated hemophagocytic syndrome (VAHS). He was treated with VP-16 intravenously (IV) and orally (0.3 g and 2.8 g/m2, respectively). RESULTS Patient no. 1 developed AML 16 months from the diagnosis of Hodgkins disease. Patient no. 2 developed AML 26 months from diagnosis. Both bone marrows were consistent with French-American-British (FAB) M4 disease. Both patients had abnormalities of the long arm of chromosome 11. CONCLUSION The use of low-dose or oral VP-16 can be associated with the development of sAML. Clinicians should be cautious in the use of VP-16 in low-risk diseases.

2-Chlorodeoxyadenosine (2-CDA) for the treatment of refractory or recurrent Langerhans cell histiocytosis (LCH) in pediatric patients

BACKGROUND Pediatric patients with Langerhans cell histiocytosis (LCH) may become refractory to conventional therapy or present with repeated recurrences over several years. Current therapeutic options such as prednisone, vinblastine, etoposide, and cyclosporine are associated with significant acute toxicities and late effects. Recent reports suggested that 2-chlorodeoxyadenosine (2-CDA) may be an effective agent in adults with LCH. The purpose of this study was to determine the safety and efficacy of 2-CDA in children with LCH. METHODS This report presents the data collected from the first three patients that have completed this trial. Patients were enrolled in a prospective study after informed consent was obtained. Patients had a confirmed diagnosis of LCH that had recurred several times or not responded to standard therapy. Patients were given a starting dose of 5 mg/M2 of daily continuous infusion for three days duration. Two patients had their dose increased to 6.5 mg/M2/ day. A total of 4-6 courses were given, and courses were repeated every 3-4 weeks. Thirteen of fifteen courses were given as outpatients at home. RESULTS Each patient completed therapy with myelosuppression the primary toxicity. Pt. 1 initially received a higher dose of 2-CDA and developed sepsis. The dose was reduced to current study levels and no other incidence of infection, fever, and neutropenia, or blood product transfusion was required. All three patients are free of active disease 10-18 months after completing 2-CDA. CONCLUSION Three patients with LCH refractory to standard therapy had CR to 2-CDA, given at 5-6.5 mg/M2/day for 3 days, without significant toxicity.

Pediatric Craniospinal Axis Irradiation With Helical Tomotherapy: Patient Outcome and Lack of Acute Pulmonary Toxicity

PURPOSE To present the patient outcomes and risk of symptomatic acute radiation pneumonitis (ARP) in 18 pediatric patients treated with helical tomotherapy to their craniospinal axis for a variety of neoplasms. METHODS AND MATERIALS A total of 18 patients received craniospinal axis irradiation with helical tomotherapy. The median age was 12 years (range, 2.5-21). The follow-up range was 3-48 months (median, 16.5). Of the 18 patients, 15 received chemotherapy in the neoadjuvant, adjuvant, or concomitant setting. Chemotherapy was tailored to the particular histologic diagnosis; 10 of 18 patients underwent surgical removal of the gross primary tumor. The patients were followed and evaluated for ARP starting at 3-6 months after completion of craniospinal axis irradiation. ARP was graded using the Common Toxicity Criteria, version 3. RESULTS At the last follow-up visit, 14, 2, and 2 patients were alive without disease, alive with disease, and dead of disease, respectively. The cause-specific survival rate was 89% (16 of 18), disease-free survival rate was 78% (14 of 18), and overall survival rate was 89% (16 of 18). No patient had treatment failure at the cribriform plate. No patient developed symptoms of ARP. CONCLUSION Craniospinal axis irradiation using helical tomotherapy yielded encouraging patient outcomes and acute toxicity profiles. Although large volumes of the lung received low radiation doses, no patient developed symptoms of ARP during the follow-up period.

Efficacy of continuous infusion 2-CDA (Cladribine) in pediatric patients with Langerhans cell histiocytosis

Patients with Langerhans cell histiocytosis (LCH) may behave differently depending on what sites are involved and the response or lack of response to earlier therapies. Therapy for high‐risk patients or those with multiple reactivations continues to be challenging because of variable response rates and frequent toxicities. The goals of this study were to determine the long‐term disease free survival in children with high‐risk or multiply reactivated LCH treated with 2‐CDA, and the toxicity of low dose continuous infusion (CI). Ten children with multiple reactivations or high‐risk disease as defined by the Histiocyte Society were treated with CI 2‐CDA and were evaluable for response and toxicity assessment. The starting dose of 2‐CDA was 5 mg/M2/day for 3 days and escalated to 6.5 mg/M2/day for 3 days if tolerated. The maximum number of courses of 2‐CDA per patient was limited to six. Fifty‐two courses of 2‐CDA were administered without difficulty. After the patient demonstrated no acute toxicity with the first administration of 2‐CDA, the subsequent doses were given at home to all but one patient. All 10 patients had a clinical response, 9 documented by radiographic, or changes in physical exam or review of systems. Toxicity was limited to myelosuppression. Seven of the 10 patients required no additional therapy and remain disease free a median of 50 months from completing therapy. The three remaining patients are currently disease free after receiving other therapy. Further studies are needed to determine the role of 2‐CDA in this patient population. 2‐CDA can be given safely using home therapy, and may effective even in high‐risk patients.

BH3 Inhibitor Sensitivity and Bcl-2 Dependence in Primary Acute Lymphoblastic Leukemia Cells

BH3 mimetic drugs may be useful to treat acute lymphoblastic leukemia (ALL) but the sensitivity of primary tumor cells has not been fully evaluated. Here, B-lineage ALL cell cultures derived from a set of primary tumors were studied with respect to sensitivity to the BH3 mimetics ABT-263 and ABT-199 and to Bcl-2 dependence and function. These ALL cells each expressed high levels of Bcl-2 and exhibited great sensitivity to ABT-263 and ABT-199, which induced rapid apoptotic cell death. BH3 profiling indicated that the ALL cultures were Bcl-2 dependent. Coimmunoprecipitation studies revealed a multifaceted role for Bcl-2 in binding proapoptotic partners including Bax, Bak, Bik, and Bim. ABT-263 disrupted Bcl-2:Bim interaction in cells. Mcl-1 overexpression rendered ALL cells resistant to ABT-263 and ABT-199, with Mcl-1 assuming the role of Bcl-2 in binding Bim. Freshly isolated pediatric ALL blasts also expressed high levels of Bcl-2 and exhibited high sensitivity to Bcl-2 inhibition by the BH3 mimetic compounds. Overall, our results showed that primary ALL cultures were both more sensitive to BH3 mimetics and more uniform in their response than established ALL cell lines that have been evaluated previously. Furthermore, the primary cell model characterized here offers a powerful system for preclinical testing of novel drugs and drug combinations to treat ALL.

Hemophagocytic lymphohistiocytosis secondary to Ehrlichia chaffeensis infection: a case report.

Hemophagocytic lymphohistiocytosis (HLH) is a rare but potentially life-threatening condition that is characterized by fever, splenomegaly, and cytopenia in 2 or more peripheral blood lineages, hypertriglyceridemia, hypofibrinogenemia, and hemophagocytosis. HLH may be primary or may be triggered by numerous etiologies, including infections. Identification of underlying etiology of HLH is important as proper treatment can completely resolve the disease process. We present a patient whose clinical presentation fulfilled the diagnostic criteria for HLH but whose illness was caused by infection with Ehrlichia chaffeensis, emphasizing the need to explore all possible etiologies during evaluation of patients presenting with illnesses consistent with HLH.

Two-drug combinations that increase apoptosis and modulate Bak and Bcl-X L expression in human colon tumor cell lines transduced with herpes simplex virus thymidine kinase

Herpes simplex virus thymidine kinase (HSV-TK) and ganciclovir (GCV) gene therapy can induce apoptosis in tumor cells that are normally resistant to this type of cell death, although the cellular mechanisms by which this occurs remain to be elucidated. Human colon tumor cell lines expressing HSV-TK were treated with GCV or four other inducers of apoptosis: butyrate, camptothecin (CPT), Taxol (paclitaxel), or 7-hydroxystaurosporine (UCN-01). Over a 2–4 day treatment period with GCV or the other four drugs, protein levels of the apoptosis agonist Bak increased 1.5- to 3-fold, whereas a corresponding decrease in the levels of the apoptosis antagonist, Bcl-XL, was observed in butyrate-, CPT-, and 7-hydroxystaurosporine (UCN-01)-treated cells. GCV and paclitaxel treatments resulted in increased levels of Bcl-XL. In two-drug combinations with GCV plus one of the four other drugs, increased tumor cell killing was found with GCV plus UCN-01 or with some GCV/butyrate combinations; the other two tested combinations were largely antagonistic. The GCV/UCN-01 and GCV/butyrate combinations resulted in increased Bak and decreased Bcl-XL protein levels, while the GCV/CPT and GCV/paclitaxel combinations resulted in increased levels of both proteins. The results highlight the potential for new combination therapies of HSV-TK/GCV and chemotherapeutic drugs that result in increased tumor cell apoptosis for future treatments of colon cancer.