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Research Corner Abstracts by Tom Swartz and Bruce Shriver
In This Issue
Delays in Referral of Soft Tissue Sarcomas It is well established that soft tissue sarcomas (STSs) are more effectively treated in a specialist centre. However, delays in time taken for a patient to be referred to a specialist centre may lead to a poorer prognosis. This study aimed to identify the length of these delays and where they occur. Patients with a proven STS were included. They were recruited from both outpatient clinics and from the surgical ward of the Royal Orthopaedic Hospital (Birmingham, UK). A structured interview was used to take a detailed history of the patients' treatment pathway, before arriving at the specialist centre. Dates given were validated using the case notes. The Results were as follows: The median time for the patient to present to a specialist centre from the onset of symptoms was 40.4 weeks. The median delay until presentation to a medical professional (patient delay) was 1.3 weeks. Median delay in referral to a specialist centre (service delay) was 25.0 weeks. The authors Conclude: Medical professionals rather than patients contribute the greatest source of delay in patients reaching a specialist centre for treatment of STS. Adherence to previously published guidelines could decrease this delay for diagnosis of possible sarcoma. Steps should be taken to refer patients directly to a diagnostic centre if they have symptoms or signs suggestive of STS.
The importance of adult stem cells in the development of cancer is becoming increasingly well appreciated. The investigators of this study hypothesized that sarcomas of soft tissue could be categorized by their developmental/differentiation status from stem cell to mature tissue, similar to the hematological malignancies. They conducted gene expression analyses during in vitro differentiation of human mesenchymal stem cells into adipose tissue, as a representative mature connective tissue, and identified genes whose expression changed significantly during adipogenesis. Gene clustering and distance correlation analysis allowed the assignment of a unique time point during adipogenesis that strongly correlated to each of the four major liposarcoma subtypes. Then using a novel gene expression strategy, in which liposarcomas were compared to their corresponding adipocytic maturing cells, they identified a group of genes overexpressed in liposarcomas that indicate the stage of differentiation arrest, i.e., sharing a similar expression profile to adipocytic cells at a corresponding stage of differentiation, and a distinct set of genes overexpressed in liposarcomas that are not found in the corresponding stage of differentiation. They propose that the latter set is enriched for candidate transformation-associated genes. They conclude that their results indicate that a degree of developmental maturity can be quantitatively assigned to solid tumors, supporting the notion that transformation of a solid tumor stem cell can occur at distinct stages of maturation.
The aim of this study was to evaluate feasibility and toxicity of bevacizumab (Avastin®), a monoclonal antibody directed against the vascular endothelial growth factor in children and young adults. Fifteen patients (male: n = 8; female: n = 7; median age, 14.6 years) received bevacizumab for recurrent or progressive solid tumors (carcinoma: n = 3; neuroblastoma: n = 2; astrocytoma grade III: n = 2; rhabdomyosarcoma: n = 2; nephroblastoma: n = 2; benign vascular tumors: n = 2; synovial sarcoma: n = 1; and malignant hemangiopericytoma: n = 1) on a compassionate basis. Bevacizumab was administered at 5–10 mg/kg body weight intravenously every 2–3 weeks. Most patients received chemotherapy in addition to bevacizumab. Duration of bevacizumab therapy ranged from 1.5 to 23 months. The Results were as follows: Bevacizumab-related side-effects were mild and included hypertonia (n = 2), proteinuria/hematuria (n = 2), epistaxis (n = 2), local erythema (n = 1), and defective wound healing and ascites (n = 1). Radiographic objective responses (partial responses) were observed in two patients with astrocytoma grade III and in one patient each with neuroblastoma and pleomorphic rhabdomyosarcoma, respectively. The investigators Conclude: Bevacizumab seems to have a good acute safety profile and some antitumor activity in heavily pretreated children and young adults with recurrent solid tumors. Prospective clinical trials are urgently needed to further evaluate the safety and efficacy of bevacizumab in pediatric patients.
Recombinant tumor necrosis factor- (TNF-) combined with melphalan is clinically administered through isolated limb perfusion (ILP) for regionally advanced soft tissue sarcomas of the limbs. In preclinical studies, wild-type p53 gene is involved in the regulation of cytotoxic action of TNF- and loss of p53 function contributes to the resistance of tumor cells to TNF-. The relationship between p53 status and response to TNF- and melphalan in patients undergoing ILP is unknown. The investigators of this study examined 110 cases of unresectable limbs sarcomas treated by ILP. Immunohistochemistry was carried out using DO7mAb, which reacts with an antigenic determinant from the N-terminal region of both the wild-type and mutant forms of the p53 protein, and PAb1620mAb, which reacts with the 1620 epitope characteristic of the wild-type native conformation of the p53 protein. The immunohistochemistry data were then correlated with various clinical parameters. The Results were as follows: P53DO7 was found expressed at high levels in 28 patients, whereas PAb1620 was negative in 20. The tumors with poor histological response to ILP with TNF- and melphalan showed significantly higher levels of p53-mutated protein. The investigators Conclude that these results might be a clue to a role of p53 protein status in TNF- and melphalan response in clinical use.
It has been previously demonstrated that bone marrow (BM) cells migrate to Ewing's tumors and differentiate into endothelial cells within the tumor vasculature. Recent evidence suggests that the roles of BM cells in tumors are more diverse. The investigators of this study investigated whether non-endothelial cell types critical for tumor vessel development are also derived from migrated BM cells. They utilized BM transplantation with GFP(+) transgenic mice as BM donors and nude mice as recipients to track the fate of migrated BM cells. After engraftment, they injected recipient mice either subcutaneously or intramuscularly with Ewing's sarcoma cells. They labeled functional tumor vessels using intravascular perfusion staining with tomato lectin. They assessed BM cell recruitment/differentiation within the tumor microenvironment using immunohistochemistry. They found that Ewing's tumors contained BM-derived cells that had differentiated into endothelial cells lining perfused tumor vessels. A substantial fraction of recruited BM cells also resided in the vessel vicinity and expressed desmin and PDGFR-beta, indicating smooth muscle cell differentiation. In order to further characterize the role of stem/progenitor cells in Ewing's sarcoma, they sorted Tie2(-) BM cells from Tie2-GFP transgenic mice and then injected them intravenously into Ewing's tumor-bearing mice. Tie2(-) BM progenitors migrated to Ewing's tumors and differentiated into Tie2(+) cells occupying a perivascular residence and expressing alpha-smooth muscle actin, desmin and PDGFR-beta, as well as VEGFR-2. They did not observe differentiation of Tie2(-) cells into Tie2(+) perivascular cells in VEGF(165)-inhibited TC/siVEGF(7-1) tumors. The differentiation of Tie2(-) BM cells into Tie2(+) cells in parental but not VEGF(165)-inhibited tumors indicates that the tumor microenvironment may influence the differentiation pathway.
Sonodynamic therapy (SDT) is a novel and promising cancer therapy that uses a combination of ultrasound and hematoporphyrin to induce apoptosis in some cancer cells. However, the mechanism(s) of SDT-induced cell apoptosis is not well understood. This study investigated SDT-induced apoptosis in sarcoma 180 cells. Cell suspension were treated by 1.75-MHz continuous focused ultrasound in the presence of hematoporphyrin for 3 minutes, and apoptosis was assessed by flow cytometry, scanning electron microscopy, terminal deoxynucleotidyl transferase–mediated deoxyuridine triphosphate–biotin nick end labeling, confocal microscopy, and apoptosis-related protein analysis. The Results were as follows: DNA breaks, apoptotic bodies, and cleaved poly (adenosine triphosphate-ribose) polymerase were observed 1 hour after SDT. By using laser-scanning confocal microscopy, the investigators found that the Fas-associated death domain and caspase 8 translocated from the cytoplasm to the plasma membrane. Activities of caspase 8 and caspase 3 were detected by an immunohistochemical assay. The results suggested that SDT led to activation of caspase 8, which in turn activated downstream caspase 3. In addition, Z-Ile-Glu-Thr-Asp-fluoromethylketone, a specific inhibitor for caspase 8, was used to confirm the effect of caspase 8 in apoptosis. The investigators Conclude: that their data primarily show that SDT can induce apoptosis in sarcoma 180 cells in vitro, and caspase 8 may play an important role in SDT-induced apoptosis.
Human sarcomas are rare but diverse malignant tumors derived from mesenchymal tissue. Clinical response to therapy is currently determined by the modified World Health Organization (WHO) criteria or the Response Evaluation Criteria in Solid Tumors (RECIST), but these standards correlate poorly with sarcoma patient outcome. The investigators of this study introduced ligand-directed particles with elements of AAV and phage (AAVP) to enable integration of tumor targeting to molecular imaging. They report drug-response monitoring and prediction in a nude rat model of human sarcoma by AAVP imaging. As a proof-of-concept, they imaged Herpes simplex thymidine kinase in a clinic-ready setting with PET to show that one can a priori predict tumor response to a systemic cytotoxic. They believe given the target expression in patient-derived sarcomas, this platform may be translated in clinical applications. And Sarcoma-specific ligands and promoters may ultimately lead to an imaging transcriptome.
Treatment of Ewing's sarcoma using an antisense oligodeoxynucleotide to regulate the cell cycle Ewing's sarcoma (ES) is one of the most malignant tumors of bone and soft tissue in children and young adults. ES belongs to a group of small round cell tumors (SRCTs) that also includes neuroblastoma, rhabdomyosarcoma, and malignant lymphoma. However, ES exhibits several specific chimeric genes (EWS-FLI1, EWS-ERG, EWS-ETV1, EWS-E1AF, and EWS-FEV) caused by chromosomal translocations that are not shared by other SRCTs. These chimeric genes regulate the expression of various other genes; that is, they activate inhibitors of DNA binding 2 (Id2) gene expression or they suppress transforming growth factor beta II (TbetaRII) receptor gene expression. The regulation of these chimeric genes may affect critical cell signal transductions, such as signals involved in cell cycle and apoptosis in ES tumor cells. Using an antisense oligodeoxynucleotide against a sequence containing the ATG initiation codon of the EWS-FLI1 chimeric gene that specifically reacts with the EWS-FLI1 and EWS-ERG chimeric genes, the investigators were able to regulate the cell cycle through the down-regulation of Id2. They report in this paper that treatment with an antisense oligodeoxynucleotide against this chimeric gene was very useful for inducing the regression of ES tumor growth. Thus, they conclude that this chimeric gene may be an important target for the treatment of ES patients.
The Clinical Approach Towards Chondrosarcoma This review article provides an overview of the histopathology, classification, diagnostic procedures, and therapy of skeletal chondrosarcoma. Rare subtypes of chondrosarcoma, including dedifferentiated, mesenchymal, and clear cell chondrosarcoma, are discussed as well. Potential new systemic treatment targets are also discussed.
Genetic variation and response to morphine in cancer patients Pain is a common symptom for patients with cancer, and opioids are the treatment of choice for moderate or severe cancer-related pain. Side effects, such as drowsiness, confusion, and hallucinations, can limit the use of opioids in clinical practice. The authors of this study prospectively recruited 228 cancer patients who received morphine. Clinical data, including pain and side-effect scores, were correlated with genotype data. The authors found that genetic variation in the multidrug resistance-1 gene (MDR-1) was associated with moderate or severe drowsiness and confusion or hallucinations. Patients who carried the common guanosine (G) allele at position 2677 in exon 26 were less likely to experience drowsiness and confusion or hallucinations than patients who carried the variant thymidine or adenosine alleles, which code for alternate amino-acid substitutions (chi-square statistic, 13.3; P = .0003). In addition, genetic variation in the catechol-O-methyltansferase (COMT) enzyme was associated independently with these central side effects. Single nucleotide polymorphisms (SNPs) in intron 1 were associated significantly with central side effects; the most significant was at position -4873G (chi-square statistic, 9.1; P = .003). SNPs in intron 1, defined as haplotype, were present in 10.4% of the population and were associated significantly with central side effects (chi-square statistic, 7.7; P = .005). Genotype data did not correlate with morphine dose or serum morphine or metabolite concentrations in this study. Thus, the authors Conclude: COMT and MDR-1 genotypes are correlated with morphine-related central side effects. The authors believe that this work adds significantly to the current understanding of genetic variants that may influence an individual's response to opioids.
Sarcomas of soft tissues account for more than 10% of diagnosed cancer in adults. Once the sarcomas have metastasized, current treatment methods are generally ineffective. The investigators of this study identified a nutrient mixture (NM) containing lysine, proline, ascorbic acid and green tea extract that demonstrates a broad spectrum of antitumor activity. They investigated whether the antitumor effect of a NM on sarcoma cell lines was due to apoptosis. Chondrosarcoma (SW-1353), synovial sarcoma (SW-982), liposarcoma (SW-872) and fibrosarcoma (HT-1080) cell lines (ATCC) were cultured in the recommended media and treated with NM in different doses: 0, 100, 500 and 1000 µg/ml. Cell proliferation was measured by MTT assay, morphology by H&E staining, and apoptosis by Live Green Caspase Detection Kit. The sarcoma cell lines studied showed no toxicity at 100 µg/ml NM, moderate at 500 µg/ml and significant at 1000 µg/ml. When treated with NM at 500 and 1000 µg/ml, obvious apoptotic cells were observed by H&E. The apoptotic cells showed shrinkage with condensed and darkly stained nuclei and strongly acidophilic cytoplasm. Slight apoptosis was observed at 100 µg, moderate at 500 µg/ml and significant at 1000 µg/ml NM for all sarcoma cell lines. The investigators thus believe these results suggest that NM induces apoptosis in sarcoma cell lines and raises the possibility of its use as a new therapeutic agent.
Ewing's sarcoma family tumors (ESFT) express the EWS-FLI-1 fusion gene generated by the chromosomal translocation t(11;22)(q24;q12). Expression of the EWS-FLI-1 fusion protein in a permissive cellular environment is believed to play a key role in ESFT pathogenesis. However, EWS-FLI-1 induces growth arrest or apoptosis in differentiated primary cells, and the identity of permissive primary human cells that can support its expression and function has until now remained elusive. In this study, the investigators show that expression of EWS-FLI-1 in human mesenchymal stem cells (hMSC) is not only stably maintained without inhibiting proliferation but also induces a gene expression profile bearing striking similarity to that of ESFT, including genes that are among the highest ESFT discriminators. Expression of EWS-FLI-1 in hMSCs may recapitulate the initial steps of Ewing's sarcoma development, allowing identification of genes that play an important role early in its pathogenesis. Among relevant candidate transcripts induced by EWS-FLI-1 in hMSCs, the investigators found the polycomb group gene EZH2, which they show to play a critical role in Ewing's sarcoma growth. These observations are consistent with the investigators other recent findings using mouse mesenchymal progenitor cells and provide compelling evidence that hMSCs are candidate cells of origin of ESFT.
V5N2 ESUN Copyright © 2008 Liddy Shriver Sarcoma Initiative. |
and melphalan 
