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2596 Background: PF-02341066 (PF) is a selective, ATP-competitive, small molecule orally bioavailable inhibitor of the ALK and MET/HGF receptor tyrosine kinases being developed for the treatment of...

10004 Background: Palifosfamide (ZIO-201) references a novel DNA cross-linking composition that comprises the functional active metabolite of ifosfamide (IFOS). It lacks the hemorrhagic cystitis and CNS toxicity of IFOS. Palifosfamide has broad activity in human sarcoma xenograft models and is active in IFOS-resistant xenografts. This study is a randomized Phase II trial evaluating safety and efficacy of the combination of palifosfamide + doxorubicin vs. doxorubicin alone in patients with metastatic/unresectable first- and second-line soft tissue sarcoma (STS). Methods: Multicenter, multinational, stratified for age and histopathologic subtype. Patients may receive up to 6 cycles until progression or toxicity occurs. After 6 cycles, patients who meet protocol criteria may get palifosfamide. The primary end-point is progression-free survival (PFS); the secondary end-points are safety and survival. Results: A total of sixty-seven patients with STS were randomized with 66 treated and 62 eligible. Enrollment ...

10637 Background: Several sarcoma subtypes are characterized by well-defined genetic events such as chromosomal translocations or gene amplification. In translocation-associated sarcomas, the prese...

10098 Background: Clear cell sarcoma is a rare soft tissue sarcoma subtype associated with the characteristic translocation t(12;22)(q13;q12). There have been few studies documenting the response r...

10006 Background: Adjuvant therapy with imatinib mesylate increases recurrence-free survival (RFS) in primary GIST (Lancet 2009). However, the effect of tumor pathologic and molecular factors on outcome in patients treated with or without adjuvant imatinib is uncertain. Methods: 713 patients with a KIT-expressing, ≥3 cm, primary GIST were randomized postoperatively in a double-blind manner to 1 year of 400 mg imatinib or placebo daily. Patients underwent serial (q3 months x8 then q6 months x6) cross-sectional imaging. Mitotic rate (mitoses per 50 high power fields) and KIT and PDGFRA mutation status were determined by central pathologic analyses. Tumor mitotic rate, size, location, and mutation status were available in 513 patients. Results: Tumor size was

10021 Background: Only 30-50% of women with high-grade, uterine leiomyosarcoma (uLMS) limited to the uterus at diagnosis remain progression free at 2 years. Adjuvant pelvic radiation has not improved outcomes. Fixed-dose rate GT, and D, each have achieved objective responses in 25-53% of patients with metastatic uLMS. We sought to determine whether adjuvant treatment of uLMS with GT, followed by D, would result in ≥50% of women remaining progression-free at 2 years. Methods: Women with FIGO stage I, II, or serosa-positive-only III, high-grade uLMS and adequate organ function were eligible. Pts were required to initiate treatment within 12 weeks of complete resection and have no evidence of disease. Treatment: 4 cycles of every 3-week G 900mg/m2 over 90 minutes days 1 and 8, T 75 mg/m2 day 8 with granulocyte stimulating factor support. If disease-free by CT after cycle 4, then: 4 cycles of every 3-week D 60 mg/m2. CT performed 6 weeks after D, then every 3 months for 2 years, then every 6 months for 3 year...

Background: Mesenchymal Stem Cells (MSCs) have been proposed as the cell of origin of sarcoma. In soft tissues, the microvascular pericyte has been recently shown to have MSC properties (Crisan et al, Cell Stem Cell 3:301). We have previously reported the isolation of benign mesenchymal cultures from sarcoma surgical samples. These cells have the immunophenotype (CD45-CD31- CD73+CD105+CD90+) and in vitro differentiation potential characteristic of MSCs. We hypothesized that these sarcoma-associated MSCs (SA-MSCs) are derived from pericytes associated with the sarcoma vasculature. Methods: We examined whether benign SA-MSCs have surface markers characteristic of pericytes and cooperate with endothelial cells in tube formation assays. We also examined expression of CD146, a pericyte marker, and CD105, an MSC marker, in sarcoma archived tissue by IHC. Results: Benign SA-MSCs indeed demonstrated properties of pericytes, such as characteristic cytoplasmic projections at low density, surface expression of pericyte markers (CD146, PDGFR-beta, NG2 and endosialin) and cooperation with endothelial cells in tube formation assays in matrigel. To demonstrate the existence of pericytes in sarcoma archived tissue, we used CD146 that has been shown to be restricted to the vasculature in sarcoma, but thought to be an endothelial marker (Shih et al, CCR 2:569). We demonstrate for the first time the existence of CD146+ pericytes in diverse sarcoma subtypes. In addition, we examined sarcoma archived material for expression of CD105 (endoglin), an MSC marker and a known tumor-specific endothelial marker in carcinoma. We found that CD105 was expressed in sarcoma endothelium similarly to its expression on carcinoma endothelium. In addition, by three-color immunofluorescence, CD105 was expressed in a subset of sarcoma pericytes. It was not expressed in normal surrounding tissues. Conclusion: We show that SA-MSC cultures have properties of pericytes. Using a novel combination of markers we demonstrate that pericytes are abundant in sarcoma. Being in contact with both the endothelial cells and the malignant sarcoma cells, pericytes represent a novel stromal element in sarcoma and a potential therapeutic target. Based on these findings we are evaluating endosialin (CD248) and CD105 antibodies, both of which are in clinical development, for effectiveness in preclinical models of sarcoma. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C250.

10585 Background: Desmoids are rare, slow growing, histologically benign tumors without metastatic potential. Methods: We examined outcomes of patients with desmoid tumors receiving systemic therapy at a single institution, to provide a basis for examination of newer agents. Retrospective chart review of 682 patients with desmoid tumors (1982–2006) from a prospectively collected sarcoma database. The activity of NSAIDs was not addressed. Patients without measurable disease, those receiving therapy we could not document, and those receiving prophylactic therapy were excluded. Results: A total of 70 patients received 163 lines of well-documented systemic therapy starting before 1/1/2007. Nine patients died, 7 of progressive disease/surgical complications, and two with Gardner syndrome-related malignancies. Demographics: 45 females (65%), median age 30 (range 15–75), 21 with Gardner syndrome (30%), median follow-up 68 months, and median of 2 lines of therapy (1–7). Intra-abdominal primary location was most common (29/70=41%). Tamoxifen, doxorubicin (including Caelyx/Doxil), imatinib, and methotrexate combinations were most commonly used. Five partial responses were observed with anthracyclines (43 courses administered), 2 with hormonal therapy (32 courses), and 1 with imatinib (35 courses), for 8 PR s. Conclusions: Anthracycline-containing regimens, hormonal therapy, and tyrosine kinase inhibitors have modest activity against desmoid tumors. The choice of therapy for these morbid and potentially fatal tumors should balance the efficacy of treatments with their short- and long-term side effects. No significant financial relationships to disclose.

10503 Background: The IGF1 system has been implicated in sarcoma development and inhibition of IGF1R function has been shown to induce clinical responses in select sarcomas. Methods: Objectives included response rate (RR) and progression-free survival (PFS) to R1507 in patients with recurrent or refractory Ewing's (ES, 2 cohorts- primary refractory vs. others) osteo (OS), synovial (SS), rhabdomyosarcoma (RMS), and other sarcomas. Eligibility included recurrent/refractory measurable disease, age ≥ 12 yrs, life expectancy ≥ 6 weeks, Karnofsky PS ≥ 70, adequate renal, hepatic and bone marrow function. R1507 was administered i.v. at 9 mg/kg over one hour weekly. Response was assessed by WHO criteria every 6 wks X 4 and every 12 wks thereafter. A two-stage design (Green and Dahlberg) was used. The endpoint for the primary refractory ES cohort was PFS at week 18 (planned n=65). RR was the primary endpoint for the remaining cohorts (planned n=240). Results: From 12/07–12/08, 203 eligible patients from 29 centers across the US, Europe and Australia were enrolled. Age ranged from 12–85 yrs (median=27 yrs) and 126 were male. Verified histologic subtypes were ES (n=71), OS (n=43), RMS (n=28), SS (n=25), and others (n=25). 15 severe adverse events were reported in 9 patients, the most common being fatigue (n=2), thrombocytopenia (n=2), dehydration (n=2), and hyperglycemia (n=2). Clinically significant activity has been observed in ES, RMS and OS with several dramatic responses seen in ES and RMS. Independent radiologic review is currently ongoing and updated data will be presented. Conclusions: The rapid accrual amongst many centers in diverse geographical locations demonstrates the feasibility of collaborative research in sarcomas. R1507 is well tolerated and a promising new agent for the treatment of various sarcomas. SARC and Roche are collaborating in additional clinical trials to better define the role of R1507 in the treatment of selected sarcomas. [Table: see text]

10526 Background: The Notch pathway directs normal fat cell development, but its aberrant activation may promote the development of sarcomas. The expression of the Notch pathway in liposarcoma (LPS), however, is unknown. We examined Notch signaling components in LPS's and suppressed Notch activation with drug targeting in LPS cell lines. Methods: RNA was isolated from 18 normal fat and 140 LPS tissue samples from five LPS subtypes: well-differentiated (33%), de-differentiated DD (25%), myxoid (12%), round cell (6%), and pleomorphic (13%), and were hybridized to Affymetrix U133A arrays. Microarray data were normalized with the RMA method. Correlation analysis identified genes expressed between sample classes, using Empirical Bayes t-test, and genes associated with survival, using Cox regression. The Notch pathway in two LPS lines, DDLS and LS141, was suppressed with a novel gamma-secretase inhibitor (GSI) or with siRNA to Notch receptors. Viability was assessed by colony formation, apoptosis by DAPI staining, and Notch expression by immunoblotting. Results: Expression of Notch-3 and its targets, Hes-1, Hey-1, and survivin, was increased in LPS subtypes, compared to fat tissue (p No significant financial relationships to disclose.

3509 Background: PF-02341066 (PF) is a selective, ATP-competitive, small molecule oral inhibitor of the c-Met/HGFR and ALK receptor tyrosine kinases that has not previously been tested in humans. A Phase 1 dose-escalation trial evaluating PF as an oral single agent was conducted to investigate safety, PK and PD in patients (pts) with advanced cancer (excluding leukemias). Methods: PF was administered under fasting conditions QD or BID on a continuous schedule to pts in successive dose-escalating cohorts at doses ranging from 50 mg QD to 300 mg BID. Pts with advanced cancer were enrolled in the study. Results: Thirty-seven pts were enrolled into the dose escalation part of the study. Tumor types included colorectal, pancreatic, sarcoma, ALCL and NSCLC. The MTD was 250 mg BID. Three DLTs were observed: grade 3 increase in ALT (1 pt at 200 mg QD) and grade 3 fatigue (2 pts at 300 mg BID). The most common AEs were nausea, emesis, fatigue and diarrhea. Nausea and emesis were independent of dose or duration of treatment. Mean AUC (30–57% CV) and Cmax (36–69% CV) increased proportionally with dose from 100 mg QD to 300 mg BID. The median terminal half-life was 46 hours. A 2- to 4-fold increase in the oral midazolam (MDZ) AUC was observed following 28-days of PF dosing at 100 mg QD (n = 3) and 300 mg BID (n = 2), respectively, suggesting PF to be an inhibitor of CYP3A. Ten pts have entered an enriched RP2D cohort of pts with tumors harboring c-Met amplification/gene mutation or ALK fusion genes. There has been 1 confirmed PR in a sarcoma pt with ALK rearrangement (inflammatory myofibroblastic tumor). Among 10 NSCLC pts whose tumors harbor EML4-ALK rearrangement, 1 pt has had a PR, 2 pts have achieved unconfirmed PR and 4 pts have had SD (3 have experienced reduction in tumor burden by ∼20% in measurable lesions and 1 has been treated for 28 weeks). Conclusions: The MTD of PF is 250 mg BID. The major AEs were fatigue or GI-related, and all AEs were manageable and reversible. There was no evidence of non-linear PK at PF doses >100 QD. Treatment with PF-02341066 resulted in promising clinical activity against tumors carrying activating ALK gene rearrangements. Further study of PF in pts with ALK-dependent tumors is warranted. [Table: see text]

Chemically induced tumors of inbred mice elicit immunity in animals in which the tumors are induced and in other animals of the same inbred stock. The immunity is specific for each tumor: even two tumors induced in one animal with the same carcinogen are not cross-reactive. Immunity to cancer has since been observed in the case of sarcomas and carcinomas induced by a number of chemical and physical carcinogens and in several species, including mice, rats, and guinea pigs. The nature of molecules which mediate immunity to tumors is a central question in cancer immunology. A small number of such molecules have been biochemically defined. Of these, some are viral antigens expressed in tumor cells, while the relationship of some others to viral antigens is unclear. A surprising majority of nonviral tumor antigens have turned out to bear homology with stress-induced proteins. Four families of such molecules are discussed: the gp96 (hsp100) and p84/86 (hsp90) antigens of chemically induced mouse sarcomas, hsp70 antigens of tumors obtained by transfection of normal rat fetal fibroblasts with an H-ras oncogene, and the albuminoid antigens of murine melanomas and a rat histiocytoma. (Albumin-like antigens are included among the stress-induced proteins because albumin, though constitutively expressed in adult tissues, is heat shock inducible in fetal liver.) Each of these antigens is a moderately abundant protein, present not only in tumors but also in normal tissues. Administration of each of these antigen preparations from the tumor, but not from normal tissue, renders the animal immune to challenge with live cells of the tumor from which the antigens are prepared. And yet, no structural differences in the antigens have been observed between normal tissues and tumors. It is suggested that these stress-induced proteins may not be tumor antigens per se, but may be carriers of immunogenic moieties such as short peptides. The stress-induced proteins may therefore serve either as antigen-presenting molecules like the MHC-encoded molecules or as accessory molecules in the presentation of antigens by MHC molecules. The ability of stress-induced proteins to bind to a variety of molecules, including peptides, is consistent with this possibility.

10531 Background: The efficacy of sorafenib in sarcomas was suggested in a phase I study in solid tumors. We conducted a multicenter, phase II study of sorafenib in sarcoma patients (pts) with six ...

10529 Background: Sunitinib malate is a multitargeted tyrosine kinase inhibitor of KIT, PDGFRs, VEGFRs, FLT3, CSF-1R and RET approved for the treatment of imatinib-resistant/intolerant GIST and adv...

10533 Background: SU is an oral multitargeted tyrosine kinase inhibitor of VEGFRs, PDGFRs, KIT, FLT3, CSF-1R and RET approved multinationally for treatment of imatinib-resistant/intolerant GIST. SU demonstrated early evidence of activity in sarcomas other than GIST during phase I testing. The current phase II study was designed to examine the clinical activity of CDD of SU in pts with advanced STS other than GIST. Methods: Pts with metastatic and/or locally advanced STS, measurable disease, ECOG PS 0–1 and ≤3 prior chemotherapy regimens were eligible. Pts are enrolled in 1/2 study arms: histologies that have shown responses to TKIs (Arm A) or histologies that have not (Arm B). Using a Simon 2-stage design, 21+20 patients will be recruited per arm based on ORR. The treatment regimen is administered as CDD of SU (37.5 mg) in 4-wk cycles. The primary endpoint is ORR per arm based on RECIST. Secondary endpoints include progression-free rate (CR+PR+SD) at 16 and 24 wks and OS. FDG-PET response before and after...

We report a method, Expression-Microarray Copy Number Analysis (ECNA) for the detection of copy number changes using Affymetrix Human Genome U133 Plus 2.0 arrays, starting with as little as 5 ng input genomic DNA. An analytical approach was developed using DNA isolated from cell lines containing various X-chromosome numbers, and validated with DNA from cell lines with defined deletions and amplifications in other chromosomal locations. We applied this method to examine the copy number changes in DNA from 5 frozen gastrointestinal stromal tumors (GIST). We detected known copy number aberrations consistent with previously published results using conventional or BAC-array CGH, as well as novel changes in GIST tumors. These changes were concordant with results from Affymetrix 100K human SNP mapping arrays. Gene expression data for these GIST samples had previously been generated on U133A arrays, allowing us to explore correlations between chromosomal copy number and RNA expression levels. One of the novel aberrations identified in the GIST samples, a previously unreported gain on 1q21.1 containing the PEX11B gene, was confirmed in this study by FISH and was also shown to have significant differences in expression pattern when compared to a control sample. In summary, we have demonstrated the use of gene expression microarrays for the detection of genomic copy number aberrations in tumor samples. This method may be used to study copy number changes in other species for which RNA expression arrays are available, e.g. other mammals, plants, etc., and for which SNPs have not yet been mapped.

10061 Background: Sorafenib (BAY 43–9006) is an oral multi-targeted kinase inhibitor. Sorafenib causes dermatologic toxicity and hypertension, although the mechanisms are poorly understood. We observed significant toxicity requiring dose reductions in our phase II study of sorafenib in STS. Methods: 120 patients (40 M, 80 F; median age, 55 years) were treated between 10/05 and 12/06; accrual continues. All patients initially received sorafenib 400 mg BID. 118/120 registered patients were evaluable for toxicity. Clinical and laboratory variables were analyzed for association with dose reduction, using Kruskal-Wallis rank sum test for continuous variables and Fisher exact test for categorical variables. Variables significant at the p=0.05 level were further analyzed with a multivariate logistic regression model for their effects on dose reduction. Results: 53% of patients (63/118) required dose reductions, 53/63 for grade 2 or greater skin toxicity. Most common grade 3–4 drug-related toxicities included lymphopenia (14%), rash (12%), and hand-foot skin reaction (10%). Sex, height, weight, BSA and serum creatinine (Cr) were significantly associated with dose reduction by univariate analysis. Adjusting for sex and/or low serum Cr, BSA was not significantly associated with dose reduction. Sex and low serum Cr were borderline statistically significant predictors of dose reductions when both variables were included in a multivariate model. Only female sex remained a significant predictor when eliminating one outlier with BSA 2.83 (p=0.04). Conclusions: Female gender appears associated with skin toxicity, requiring dose reductions. Based on this multivariate analysis, a starting dose of 400 mg oral daily in women may limit side effects. Correlation with trough serum sorafenib levels is pending. This study is funded in part by NCI Grant P01-CA47179. No significant financial relationships to disclose.

9522 Background and Methods: The sensitivity to trabectedin (Yondelis, ET-743) in experimental cancer models correlates with functional transcription-coupled nucleotide excision repair (TC-NER) and with deficient homologous recombination repair (HRR) activity. In order to assess the clinical relevance of this observation we have characterized the mRNA expression levels of ERCC1, XPD, BRCA1 and BRCA2 by RT-PCR in historical tumor samples from 92 sarcoma patients (pts) treated with the agent. Results: The overall objective response (CR+PR) rate to Yondelis in this group was 9%, which confirms the previous clinical experience with the compound in unselected, pre-treated patients with sarcoma. The Kaplan-Meier estimates for progression-free survival at 6 months (PFS6) was 23% and for median survival 8 months (mo). 26% of pts were alive at 24 mo. Correlative Study: Pts with high expression levels (> median) of ERCC1 had better PFS6 (32% vs 15%, p = 0.07) and better median survival (12 vs 7 mo) as compared to pts with low expression ( ≤ to median). Pts with low expression levels of BRCA1 had better PFS6 (33 vs 11%, p = 0.02) and superior median survival (15 vs 5 mo, p = 0.0003) than pts with high expression. Expression levels of XPD and BRCA2 (58 pts) had no significant impact on PFS and survival. The analysis of co-expression of ERCC1-BRCA1 identified a highly sensitive group of pts, characterized by high ERCC1 and low BRCA1 expression levels, with a PFS6 of 50% (p=0.0003) and median survival of 20.4 mo (p = 0.0005). A Cox regression analysis demonstrated that ERCC1 (HR=0.52, p = 0.02) and BRCA1 (HR=2.73, p = 0.0006) are independent variables for PFS, while BRCA1 (HR= 2.57, p = 0.0005) is also an independent variable for survival. Conclusion: This exploratory retrospective study supports the hypothesis that the sensitivity to Yondelis in pts with sarcoma correlates with a functional TC-NER and with deficient HRR. Prospective studies in sarcoma and other potentially sensitive tumor types are required to confirm and validate these findings. [Table: see text]

9502 Background: IM resistance in advanced GIST represents a major clinical problem. SU (SU11248) is an oral multitargeted tyrosine kinase inhibitor with antitumor and antiangiogenic activities related to KIT, PDGFRs, VEGFRs, RET and FLT3 inhibition. SU has demonstrated efficacy in clinical trials of pts with IM-R GIST. Methods: This study examined the relationship between tumor kinase genotypes and SU clinical activity in 97 pts with metastatic IM-R GIST treated as part of a phase I/II trial. Tumors were imaged by CT or MRI for RECIST-defined response assessment. Tumor specimens were obtained prior to (n=76) and following (n=64) IM therapy and analyzed for primary or secondary KIT and PDGFRA mutations, respectively. Results: Clinical benefit (CB; defined as PR or SD >6 months) with SU was observed for all major molecular GIST subtypes: KIT exon 11 (42 pts, CB 36%), KIT exon 9 (19 pts, CB 42%), PDGFRA (4 pts, CB 25%), no KIT or PDGFRA mutation (wild-type [WT]; 9 pts, CB 56%). Notably, the PR rate for GISTS with primary KIT exon 9 mutations was 37%, vs 5% for KIT exon 11 mutations (P=0.003). PFS and OS were significantly longer for pts with either a primary KIT exon 9 mutation or WT KIT/PDGFRA compared with pts with a KIT exon 11 mutation (exon 9 vs 11: PFS P=0.0007, OS P=0.005; WT vs exon 11: PFS P=0.03, OS P=0.01). Secondary KIT mutations (in exons 13, 14, 17 or 18) were found in 62% of GISTs with a primary KIT exon 11 mutation, but only in 16% with a primary KIT exon 9 mutation (P=0.002). No secondary mutations were found in GISTs lacking a primary KIT/PDGFRA mutation. Biochemical profiling of secondary kinase mutations revealed in-vitro sensitivity of KIT exon 13 and 14 mutations to SU, while secondary KIT exon 17 and 18 mutations were resistant to SU in vitro. These in-vitro results were consistent with clinical results: CB was observed in 65% of pts with secondary KIT mutations in exon 13 or 14 and in only 9% of pts with secondary KIT exon 17 or 18 mutations (P=0.006). Conclusions: Our findings suggest that, similar to prior results observed using imatinib, CB of SU following failure of IM treatment is significantly influenced by both primary and secondary mutations in the predominant pathogenic kinase, the uncontrolled activity of which is a critical etiologic factor for GIST. [Table: see text]

9514 Background: We sought to compare the response of metastatic STS to G vs. G+D in a phase III study using a novel Bayesian adaptive randomization strategy. Methods: Entry criteria included non-GIST STS diagnosis, age > 10, measurable disease, ≤ 3 prior regimens, normal organ function, and Grade (Gr) ≤ 1 neuropathy. Pts were stratified by histology (leiomyosarcoma [LMS] vs. other), and history of prior pelvic radiation (PPR) or not, yielding 4 diagnostic subgroups. Therapy was G 1200 mg/m2 (over 120 min, d1+d8) or G 900 mg/m2 (over 90 min, d1+d8) and docetaxel (100 mg/m2 d8) q21d; all pts received (peg)-filgrastim starting d9. 25% dose reductions were employed for PPR or toxicity. The primary endpoint was tumor response (CR, PR, or 24+ weeks stable), using a double-blind Bayesian adaptive randomization procedure to incrementally assign more pts to the superior treatment arm, while accounting for possible treatment-subgroup interactions. Although this study is not based on a 1:1 randomized phase III design, enrolling ∼120 pts would show a difference between a response rate (RR) of 10% for G and 30% with G+D with a power of 0.8 and two-sided alpha of 0.05. Results: 122 pts were randomized; 119 pts had evaluable outcomes. Median number of prior regimens was 1; median number of cycles was 4 (range 1–26). 49 pts were adaptively randomized to G and 73 to G+D, indicating superiority of G+D. Of 119 evaluable pts, 27% (G) and 32% (G+D) showed response as defined. The odds of pts with LMS receiving G+D instead of G increased from 1:1 at the start of the study to ∼6:1 at its completion. RR was 10% (G) vs. 16% (G+D) (p=0.15, Fisher exact test). Consistent with the RR outcomes, progression free survival (PFS) was 6.2 m (G+D, K-M 95%CI 3.6–8.8) vs. 2.6 m (G, 95%CI 2.0–3.2). Overall survival (OS) was 18.0 m (G+D, K-M 95%CI 11.7–24.1) vs. 11.2 m (G, 95%CI 6.8–15.5). Pts receiving G+D had a higher rate of discontinuation due to toxicity than those receiving G (p [Table: see text]

9523 Background: Flavopiridol, a potent cyclin dependent kinase inhibitor, has been shown in vitro to enhance doxorubicin induced apoptosis. Using the BWH dediffentiated liposarcoma xenograft model we have shown that sequential therapy with doxorubicin (D) followed 1 hour later by flavopiridol (F) is superior to doxorubicin alone (p=0.006). Single agent flavopiridol was also active in this xenograft model, but less so than the combination. Methods: We have commenced an ongoing phase I study of the combination of D and F on the basis of this preclinical data in soft tissue sarcoma with fixed dose D (60 mg/m2), followed 1 hour later by escalating doses of F (40–70 mg/m2) administered over one hour, every 3 weeks. Standard phase I eligibility criteria apply. No prior anthracycline therapy is allowed. At 300 mg/m2 D, dexrazoxane is added to D F. Patients with responsive or stable disease after cumulative D dose of 600mg/m2 can receive single agent flavopiridol. Results: Median characteristics of 7 evaluable patients: age 52 (31–65), KPS 80% (70–90), 3 males / 4 females, 0 prior regimens (range 0–2). The combination has been well-tolerated, with no dose limiting toxicity (DLT) yet. There has been 1 grade 3 bleed in the setting of progressive disease and 1 grade 4 neutropenia without fever. Grade 1 and 2 diarrhea, related to flavopiridol have been observed. Pharmacokinetic (PK) studies are ongoing. We have observed no partial responses (PR). Prolonged stable disease (SD) has been seen in 3 patients for 6, 6 and 11 months. Two patients with SD have liposarcoma. One patient with liposarcoma continues on single agent flavopiridol with stable disease after having reached the prescribed limit of D of 600 mg/m2. Conclusions: The combination of doxorubicin and flavopiridol is safe, with no unexpected toxicities. Disease stabilization in STS has been observed. This appears to be an encouraging combination in liposarcoma and is consistent with our preclinical model. Flavopiridol dose escalation is continuing. No significant financial relationships to disclose.

9503 Background: Chemotherapy options for palliative treatment of advanced sarcomas are limited and studies are underway to identify new classes of active agents. Second generation inhibitors of the mammalian target of rapamycin (mTOR) are being tested in phase II trials. Objective responses of sarcomas to the mTOR inhibitor AP23573 have been reported. Sirolimus (rapamycin) is a macrocyclic triene antibiotic that inhibits activation of S6 kinase, the cdk2/cyclinE complex and phosphorylation of retinoblastoma protein and causes cell cycle arrest in G1-S phase. Methods: We report the cases of four (out of 20) patients (pts) with progressive, metastatic sarcoma failing 2–6 chemotherapies treated with sirolimus and followed for clinical benefit. Sirolimus was self-administered daily with (n=2) or without (n=2) daily oral cyclophosphamide 200 mg every other week. Patients were followed in clinic for disease and toxicity assessments every 2 weeks. Molecular analysis of mTOR targets will be performed. Results: Patients with MFH, leiomyosarcoma, angiosarcoma and osteosarcoma were treated using 4–8 mg sirolimus daily for a median of 16 weeks (range 8–28 wks). Three pts were symptomatic from disease and had reduced performance status prior to treatment. All 3 pts had subjective improvement in tumor-related symptoms and 2 had improvement in performance status. LDH and bilirubin fell from 1,712 u/L and 5.6 mg/dl to 388 u/L and 1.1 mg/dl, respectively, in the pt with angiosarcoma during 3 months of sirolimus and cyclophosphamide. Alkaline phosphatase fell from 791 u/L to 120 u/L in the pt with osteosarcoma over 6 weeks of simolimus and cyclophosphamide. Three pts had minor radiographic improvement in tumor and remain on treatment. One pt has died of disease progression. Treatment was well tolerated. Conclusions: Sirolimus treatment was associated with improvement in tumor-related symptoms, performance status and biochemical markers of disease activity and inhibited tumor growth in pts with advanced sarcoma failing multiple prior therapies. Phase I studies of sirolimus in advanced cancer are underway at the University of Chicago. Formal study of sirolimus in advanced sarcoma is contemplated. No significant financial relationships to disclose.

9526 Background: Despite abundant pre-clinical data, the mechanisms of action in humans of multi-targeted small molecule tyrosine kinase inhibitors against sarcomas remain unclear. Herein, we sought evidence from pre- and post-treatment biospies to suggest whether the primary mechanism of action of sunitinib/SU11248 is anti-angiogenic or rather a direct anti-tumor cell effect. Methods: We report data regarding a 59 year old woman with imatinib-resistant GIST who demonstrated a marked tumor response to sunitinib treatment. Her gastric primary GIST tumor was previously resected, followed by recurrence one year later. A complete remission lasting 24 months was then attained on imatinib therapy. After progression of disease, sunitinib was started at 50 mg orally per day. A major clinico-radiographic response was seen after 12 months of therapy, followed by resection. To assess the pharmacodynamic effects of sunitinib, we undertook immunohistochemical analysis of formalin-fixed tissue sampled immediately prior to treatment and just after completing sunitinib, using anti-CD117, anti-CD34, or anti-CD31 antibodies, respectively. Results: We found a dramatic effect of sunitinib on the tumor parenchyma. Whereas the pretreatment biopsy revealed viable, mitotically active, CD117+ and CD34+ tumor cells, the post-treatment specimen was >90% sclerotic and necrotic. The CD31+ blood vessels appeared robust both pre- and post-treatment. Vessel counts did not differ significantly before and after treatment by CD31 staining (3.3±2.0 per field vs. 1.8±1.3, p= 0.09). Thus, the tumor cells were largely eliminated with sunitinib treatment, while blood vessels were still present after 12 months of therapy. Conclusions: This observation supports the assertion that the mechanism of sunitinib in GIST is predominantly cytotoxic against the tumor by directly targeting KIT and that an anti-angiogenic effect, either through the VEGF-R2 in endothelium or through the PDGF-R in pericytes, may be secondary. [Table: see text]

9515 Background: Aggressive fibromatosis (desmoid tumors, AF) are uncommon, locally aggressive, connective tissue neoplasms. Existing literature on systemic treatment of AF is sparse and consists mostly of case reports and small case-series. Based on previous observation of regression of AF treated with IM and tumoral expression of IM targets, SARC (Sarcoma Alliance for Research through Collaboration) included the treatment of AF onto a multi-institution phase II trial of IM in sarcoma. Here we report early clinical and laboratory results of the AF group. Methods: Eligible patients had histologically proven AF, unresectable or difficult to resect without considerable functional impairment. Patients were treated with IM 300 mg po BID (BSA≥1.5m2). The primary endpoint was complete (CR) or partial response(PR) at two months or stable disease (SD) or better at four months. Tumor DNA was extracted from available formalin fixed paraffin embedded tissue specimens and analyzed via allelic PCR and genomic DNA sequence analysis for specific point mutations in PDGFRα exons 12/14/18, PDGFRβ exons 12/18, KIT exons 9/11/13/17, and bRAF. Results: 51 patients were enrolled from 10/02 to 12/05 at 5 institutions, with 45 patients currently evaluable. The median age is 37 (range 14–67), and median number of prior therapies is 1 (range 0–3). 36 patients (80%) reached the primary endpoint of CR/PR at 2 months or SD or better at 4 months. The median time to treatment failure is 6.8 months (95% C.I. 5.8–17.1). Thus far, the maximum change in the largest dimension of the tumor ranged from a 21% increase to a 45% decrease. In 22 available tumor specimens, deletions within PDGFRαE12 and E18 were noted in 1 and 3 patients, respectively, while a wildtype genotype was found in other regions. Conclusions: IM has activity in AF, the mechanism of which remains unclear. While this is the largest reported phase II trial of AF, further improvement in evaluating clinical efficacy in this disease is clearly necessary. We plan an analysis of the maximum change in largest tumor dimension for each patient, which will be particularly beneficial in AF as responses often occur late. We have not as yet identified a laboratory predictor of clinical benefit. Further investigation of other potential targets in fresh tissue is warranted. [Table: see text]

9507 Background: We sought to elucidate the relationship between the human adult mesenchymal stem cell (hMSC), Wnt signaling and sarcomagenesis. Methods: In vitro hMSC differentiation, microarray gene expression analysis, distance correlation analysis, and standard molecular biology techniques were used to explore the role of Wnt in controlling the differentiation of both hMSCs and high grade undifferentiated sarcoma (HGUS; MFH, malignant fibrous histiocytoma), a common form of adult soft tissue sarcoma. Results: We determined that 1) hMSCs appear to be the progenitor cells of HGUS/MFH; 2) Dickkopf-1 (Dkk1), a specific inhibitor of Wnt signaling, is overexpressed in MFH as compared to other sarcoma subtypes and is involved in the proliferation of hMSCs; 3) in hMSCs, Dkk1 levels decline and nuclear β-catenin accumulates as hMSCs reach confluence, a prerequisite for initiation of in vitro differentiation, while in an MFH cell line Dkk1 levels do not decline and there is no nuclear β-catenin accumulation; 3) MFH cells appear to be primed for differentiation and express early markers of mesenchymal differentiation, then undergo apoptosis, if nuclear β-catenin is manipulated to enter the nucleus; 4) Wnt2 signals via the canonical β-catenin pathway and is responsible for “commitment” of hMSC and an MFH cell line to various differentiation pathways, while Wnt5a signals via the non-canonical JNK pathway in preventing apoptosis upon appropriate commitment toward differentiation. Conclusions: We identified the contribution of canonical and non-canonical Wnt signaling in the differentiation of hMSCs and showed that enhancing signaling via these pathways could be exploited as a potential target for therapy for high grade undifferentiated sarcomas. These data implicate Wnt-signaling as a mechanism of tumor suppression in early sarcomagenesis. No significant financial relationships to disclose.

Purpose/Objective: The NIH has identified the relationship between aging and cancer as a subject in urgent need of increased research. Although older age is considered a predictor of poor outcome in soft tissue sarcoma (STS), it remains unclear whether such an outcome could be accounted for by association with other poor prognostic factors, the extent of treatment, or older age per se. The purpose of this study was to attempt to answer these questions in a group of patients with STS of the extremity.

9032 Background: IM benefits the majority of patients with metastatic GIST. Study S0033 aims to evaluate differences between two initial dose levels of IM (400 vs. 800 mg/day), with the crossover possible for pts on lower dose to increase dose in case of disease progression. Dose delays and reductions were required for defined toxicities. The present analysis was performed to analyze the incidence and reasons for dose modification in each arm on this multicenter trial. Methods: The dataset of S0033 (746 pts randomized) was analyzed; recorded dose levels of IM were tabulated and reasons for dose modifications assessed. Results: 350 eligible pts enrolled at 400 mg/day with information on dose modifications available on 335 pts. 126 (62%) had at least one dose delay and 34 (10%) had at least one dose reduction, due most commonly to rash, edema, and GI bleeding. 353 eligible pts enrolled at 800 mg/day IM, 337 with full dosing information. 189 (56%) had at least one dose delay and 148 (44%) had at least one do...

9011 Background: Resistance to IM eventually develops in most pts with metastatic GIST and is often correlated with secondary mutations in the genes encoding KIT or platelet-derived growth factor receptor alpha (PDGFRA) tyrosine kinases. SU11248 is an oral multi-targeted tyrosine kinase inhibitor with both anti-angiogenic and direct anti-proliferative activity mediated by signal blockade of several kinases, including VEGFR2, KIT, PDGFRA, and FLT3. SU11248 can overcome IM resistance in GIST caused by diverse genomic alterations (Demetri et al. Proc ASCO 2004; abstr 3001). We report longer term clinical results of a continuation study accruing pts treated in a prior phase I/II study. Methods: From 07/02 to 07/04, 97 pts with GIST resistant to or intolerant of IM were enrolled in a phase I/II study and received one of three schedules of SU11248 administered by daily oral dosing with intermittent dosing breaks. The phase II schedule chosen was 50 mg/d daily for 4 weeks with a 2 week break. After 6 months on t...

9016 Background: Liposarcoma (LS) is a heterogeneous group of tumors comprising 5 pathologic subtypes: well-differentiated (WD), dedifferentiated (DD), myxoid (MY), round cell (RC), and pleomorphic...

3001 Background: Resistance to Imatinib mesylate (IM) following initial tumor regression and disease control in GIST is increasingly recognized as an unmet medical need. IM resistance can be correlated with the appearance of secondary mutations in the KIT or PDGFRA tyrosine kinases in GIST lesions refractory to IM; activation of alternative signaling pathways and different structural biology of the new mutant kinases contribute to emergence of IM-resistant GIST clones. Methods: Phase I/II clinical trial of SU11248 in pts with progressing IM-resistant GIST. Tumor biopsies were obtained to define the mutational status of the KIT and PDGFRA kinases by dHPLC and sequencing assays. Results: 98 pts with progressive GIST have been enrolled in this ongoing study; with tumor response data available in 48 pts and GIST genotype determined in 41. The phase II regimen chosen was SU11248 50 mg orally once daily for 4 weeks, followed by a 2 week period off drug in each 6 wk cycle. SU11248 therapy induced clinical benefi...

9017 Background: The impact of adjuvant chemotherapy on the survival of patients with soft tissue sarcoma is controversial. Although ifosfamide based chemotherapy (IF) has generated significant responses in the treatment of metastatic synovial sarcoma, its effect on the survival of patients with primary disease remains unclear. The objective of this study is to determine if IF offers a survival benefit to patients with primary extremity synovial sarcoma. Methods: Two prospectively collected sarcoma databases were used to identify a contemporary cohort of 101 adult (≥16yrs) patients with primary, extremity, ≥5cm, deep, synovial sarcomas that underwent surgical treatment for cure from 1990 to 2002. 68 patients were treated with IF and 33 received no chemotherapy (NoC). Clinical, pathologic and treatment variables were analyzed for disease specific survival (DSS) and distant recurrence free survival (DRFS). Results: Median follow-up for the 101 patients was 58 months. 90%(91/101) underwent limb-sparing surge...

9013 Background: Desmoid tumors (aggressive fibromatosis) are rare clonal neoplastic proliferations of connective tissues. Although classified as benign, local aggressiveness can lead to significant impairment. Standard treatment involves wide surgical resection and/or radiation therapy. In cases of unresectable or recurrent disease, tamoxifen, chemotherapy, and NSAIDs have been used with varying success. We and others have previously reported desmoid tumors expressing c-kit, PDGFRα, and/or PDGFR. We reported two patients with extraabdominal desmoid tumors treated with the selective tyrosine kinase inhibitor imatinib (Gleevec) with significant shrinkage. SARC, in association with the Connective Tissue Oncology Society, initiated a prospective phase II trial in patients with desmoid tumors, or one of nine sarcoma subtypes. Here, we report specifically on patients with desmoid tumors. Methods: Patients ≥ 10 years old with desmoid tumors that were not curable by surgical management or in whom curative surger...

T cell hybridomas were established by fusing a CD8+ V beta 8.1+ CTL clone and a CD4+ V beta 8.1+ helper T lymphocyte (HTL) clone to the thymoma cell line BW5147. In contrast to the HTL x BW hybridomas, which retain the same antigen specificity as the original T cell clone, the CTL x BW hybridomas lost the class I MHC-restricted antigen response but acquired a new specificity to Mlsa antigen. Mlsa reactivity of CTL x BW hybridomas was shown to be mediated by the CTL TCR as assayed by inhibition using an anticlonotypic antibody to the CTL clone. Since hybridomas established with BW5147 lose CD8 expression, we have introduced the CD8 molecule into CTL x BW5147 hybridomas by gene transfection. The CD8+ V beta 8.1+ hybridoma was no longer capable of reacting to Mlsa antigen but exhibited the same antigen specificity as the parental CTL clone. Furthermore, the presence of the transfected CD8 molecule in the HTL x BW hybridomas was found to be inhibitory to class II MHC-restricted antigen reactivity. These results demonstrate that, besides its role in increasing the overall avidity of T cell-class I MHC/antigen interaction, the CD8 molecule inhibits T cell-class II MHC gene product/antigen interaction. This negative effect of the CD8 molecule on a class II MHC-restricted response may account for the failure of CD8+ T cells using either V beta 8.1 or V beta 6, which impart reactivity to the Mlsa antigen on CD4+ T cells, to respond to the Mlsa antigen.

A synthetic method has been developed for the iterative preparation of unlabelled and 3H-polyethylene glycol (PEG) oligomers. The strategy involved alkylating the sodium anion of mono-tritylated ethylene glycol oligomers (Ph3C[OCH2CH2]nOH, n=1−28) with O-tosyl-O-allyl-triethylene glycol or O-tosyl-O-allyl-pentaethylene glycol. Subsequent ozonolysis of the terminal olefin followed by reduction with NaBH4 or NaB[3H]4 provided the next higher mono-tritylated ethylene glycol oligomer which could be deprotected to the final glycol oligomer (H[OCH2CH2]nOH, n=4−34) or carried on in the iterative process. The method provides discrete PEG oligomers of high specific activity (20.6–48.6 mCi/mmoL) in 96–98% radiochemical purity.

The title compound was synthesized by the diastereoselective addition of the lithium or sodium anion of dimethyl methylphosphonate to N -trityl-L-phenylalaninal.

Novel modeling and sensitivity analysis techniques are used with experimental data obtained from discharge flow–resonance fluorescence experiments to evaluate the product branching ratio of OH+H2CO. Two channels are considered: the H‐atom abstraction reaction (R2) to form HCO and H2O; and the addition reaction (R17) followed by rearrangement and decomposition to form HCOOH and H. The rate constant values obtained at 298 K are kR2 =(7.75±1.24)×10−12 cm3/molecule s and kR17=(0.2+0.8−0.2) ×10−12 cm3/molecule s. The results demonstrate that the reaction proceeds almost exclusively via the H‐atom abstraction pathway.

Absolute concentrations of H atoms in the absorption region of an atomic resonance photometer have been accurately determined with a chemical kinetic technique that is based on the H + NO2 reaction. Subsequently, the curve of growth for the Lyman- transition (H(SP/sub 3/2,1/2/) reverse arrow H(SS/sub 1/2/)) has been determined with a resonance lamp that is essentially a microwave-driven electrodeless lamp plasma. Additional experiments have been performed in order to measure the temperature and (H) in the resonance lamp plasma. Thus, simplified theoretical calculations of the curve of growth could be made from first principles using no adjustable parameters. These calculations agreed with experiment within experimental error, and therefore, the theoretical oscillator strength, as calculated from the known wave functions for H, is experimentally confirmed for the H(SP/sub 3/2,1/2) reverse arrow H(SS/sub 1/2/) transition. Confidence can now be placed in line absorption calculations and, hence, in measurements of (H), at high absorber temperatures such as those encountered in flames, plasmas, and shock tubes where the atomic resonance absorption spectroscopic (aras) technique is commonly used. 25 references, 9 figures, 4 tables.

The X box is a loosely conserved DNA sequence that is located upstream of all major histocompatibility class II genes and is one of the cis-acting regulatory elements. Despite the similarity between all X-box sequences, each promoter-proximal X box in the mouse appears to bind a separate nuclear factor.

Desmoid tumor (DT; other synonymously used terms: Desmoid-type fibromatosis, aggressive fibromatosis) is a rare and locally aggressive monoclonal, fibroblastic proliferation characterised by a variable and often unpredictable clinical course. Previously surgery was the standard primary treatment modality; however, in recent years a paradigm shift towards a more conservative management has been introduced and an effort to harmonise the strategy amongst clinicians has been made. We present herein an evidence-based, joint global consensus guideline approach to the management of this disease focussing on: molecular genetics, indications for an active treatment, and available systemic therapeutic options. This paper follows a one-day consensus meeting held in Milan, Italy, in June 2018 under the auspices of the European Reference Network for rare solid adult cancers, EURACAN, the European Organisation for Research and Treatment of Cancer (EORTC) Soft Tissue and Bone Sarcoma Group (STBSG) as well as Sarcoma Patients EuroNet (SPAEN) and The Desmoid tumour Research Foundation (DTRF). The meeting brought together over 50 adult and pediatric sarcoma experts from different disciplines, patients and patient advocates from Europe, North America and Japan.

Cancer-associated fibroblasts (CAFs) are a key component of the tumour microenvironment with diverse functions, including matrix deposition and remodelling, extensive reciprocal signalling interactions with cancer cells and crosstalk with infiltrating leukocytes. As such, they are a potential target for optimizing therapeutic strategies against cancer. However, many challenges are present in ongoing attempts to modulate CAFs for therapeutic benefit. These include limitations in our understanding of the origin of CAFs and heterogeneity in CAF function, with it being desirable to retain some antitumorigenic functions. On the basis of a meeting of experts in the field of CAF biology, we summarize in this Consensus Statement our current knowledge and present a framework for advancing our understanding of this critical cell type within the tumour microenvironment., This Consensus Statement highlights the importance of cancer-associated fibroblasts in cancer biology and progression, and issues a call to action for all cancer researchers to standardize assays and report metadata in studies of cancer-associated fibroblasts to advance our understanding of this important cell type in the tumour microenvironment.

A 32-year-old Chinese female never smoker presented with persistent cough in June 2009, and imaging studies revealed a right hilar mass. An endobronchial biopsy of the tumor revealed a moderately differentiated mucinous adenocarcinoma that was cytokeratin 7 (CK7) positive, CK20 negative, transcription tissue factor-1 (TTF-1) positive, and mucicarmine positive. Initial molecular analysis revealed the tumor to be both epidermal growth factor receptor and KRAS wild type. Her staging workup revealed metastasis to the liver and brain. She underwent stereotactic radiosurgery of her brain metastases that was followed by six cycles of cisplatin/pemetrexed/bevacizumab combination chemotherapy followed by bevacizumab maintenance therapy, with a partial response to treatment. In November 2009, she decided to undertake a treatment holiday for 3 months. By February 2010, however, her disease had progressed, and she commenced single-agent erlotinib treatment but had documented disease progression after 6 weeks. She was referred by her treating oncologist for possible enrollment into the phase I crizotinib trial because her clinical profile—young age, adenocarcinoma histology, never-smoking status, and most importantly her tumor is wild type for epidermal growth factor receptor and KRAS—fits the profile of an anaplastic lymphoma kinase (ALK)-positive patient with non-small cell lung cancer (NSCLC).