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Water deficit is one of the most important causes of decreased yield in cultivated plants. Non-foliar green organs in cotton play an important role in yield formation at the late growth stage. Although better photosynthetic performance was observed in a non-foliar organ (bract) compared with leaves under water deficit. However, the physiological response of each organ in cotton to water deficit has not been comprehensively studied in relation to the water status and photosynthesis characteristics. We studied the maintenance of water status of each organ in cotton by measuring their relative water content, proline content and stomatal characteristics. Water deficit significantly decreased the surface area of each organ, but to a lesser extent in non-foliar organs. Our results showed that the relative contribution of biomass accumulation of non-foliar organs increased under water deficit. Non-foliar organs (bracts and capsule wall) showed less ontogenetic decrease in O2 evolution capacity and in RuBPC activity (per dry weight) as well as better antioxidant systems than leaves at various days after anthesis. We conclude that the photosynthesis from non-foliar organs is important for increasing cotton yield especially under water deficit conditions.

OBJETIVO: Avaliar o desempenho clínico de lentes progressivas (LP) em présbitas amétropes, comparando LP Gradal Top® às LP que usavam. MÉTODOS: Realizou-se um estudo clínico com 40 présbitas satisfeitos com seus óculos com adição PURPOSE: To study the clinical performance of progressive addition lens (PAL) Gradal Top®, Carl Zeiss Vision compared with other PAL lenses previously worn by presbyopic ametropes. METHODS: Forty presbyopes satisfied with their PAL spectacles (addition

Abstract Background Kidney disease is the 9th leading cause of death in Singapore. While preventive effects have focused on early detection and education, little is known about the knowledge level of chronic kidney disease (CKD) locally. We seek to evaluate the knowledge of CKD among primary care patients. Methods We conducted a cross-sectional survey of a convenience sample of 1520 patients from 3 primary care centers. Those with existing CKD or on dialysis were excluded. Knowledge was assessed based on 7 questions on CKD in the self-administered questionnaire. One point was given for each correct answer with a maximum of 7 points. Results 1435 completed all 7 questions on CKD. Mean age was 48.9 ±15.0 (SD) years. 50.9% were male. 62.3% had a secondary and below education and 52.4% had a monthly household income of ≤ $2000. 43.7% had chronic diseases. Mean score was 3.44 ± 1.53 (out of a maximum of 7). Median score was 4. In multivariate logistic regression, being older {>60 years [Odds Ratio (OR) 0.50, 95% Confidence Interval (CI) 0.32-0.79]; 40–60 years (OR 0.62, 95% CI 0.43,0.89)}, less educated [up to primary education (OR 0.33, 95% CI 0.22-0.49)], having a lower monthly household income [ Conclusion This suggests that CKD education should be targeted at older patients with lower education and lower socioeconomic status.

Background: The alkaloid camptothecin analog SN38 is a potent antineoplastic agent, but cannot be used directly for clinical application due to its poor water solubility. Currently, the prodrug approach on SN38 has resulted in 3 FDA-approved cancer therapeutics, irinotecan, ONIVYDE, and Trodelvy. However, only 2-8% of irinotecan can be transformed enzymatically in vivo into the active metabolite SN38, which severely limits the drug's efficacy. While numerous drug delivery systems have been attempted to achieve effective SN38 delivery, none have produced drug products with antitumor efficacy better than irinotecan in clinical trials. Therefore, novel approaches are urgently needed for effectively delivering SN38 to cancer cells with better efficacy and lower toxicity., Methods: Based on the unique properties of human serum albumin (HSA), we have developed a novel single protein encapsulation (SPE) technology to formulate cancer therapeutics for improving their pharmacokinetics (PK) and antitumor efficacy and reducing their side effects. Previous application of SPE technology to doxorubicin (DOX) formulation has led to a promising drug candidate SPEDOX-6 (FDA IND #, 152154), which will undergo a human phase I clinical trial. Using the same SPE platform on SN38, we have now produced two SPESN38 complexes, SPESN38-5 and SPESN38-8. We conducted their pharmacological evaluations with respect to maximum tolerated dose, PK, and in vivo efficacy against colorectal cancer (CRC) and soft tissue sarcoma (STS) in mouse models., Results: The lyophilized SPESN38 complexes can dissolve in aqueous media to form clear and stable solutions. Maximum tolerated dose (MTD) of SPESN38-5 is 250 mg/kg by oral route (PO) and 55 mg/kg by intravenous route (IV) in CD-1 mice. SPESN38-8 has the MTD of 45 mg/kg by IV in the same mouse model. PK of SPESN38-5 by PO at 250 mg/kg gave mouse plasma AUC 0-∞ of 0.05 and 4.5 nmol × h/mL for SN38 and SN38 glucuronidate (SN38G), respectively, with a surprisingly high molar ratio of SN38G:SN38 = 90:1. However, PK of SPESN38-5 by IV at 55 mg/kg yielded much higher mouse plasma AUC 0-∞ of 19 and 28 nmol × h/mL for SN38 and SN38G, producing a much lower molar ratio of SN38G:SN38 = 1.5:1. Antitumor efficacy of SPESN38-5 and irinotecan (control) was evaluated against HCT-116 CRC xenograft tumors. The data indicates that SPESN38-5 by IV at 55 mg/kg is more effective in suppressing HCT-116 tumor growth with lower systemic toxicity compared to irinotecan at 50 mg/kg. Additionally, SPESN38-8 and DOX (control) by IV were evaluated in the SK-LMS-1 STS mouse model. The results show that SPESN38-8 at 33 mg/kg is highly effective for inhibiting SK-LMS-1 tumor growth with low toxicity, in contrast to DOX's insensitivity to SK-LMS-1 with high toxicity., Conclusion: SPESN38 complexes provide a water soluble SN38 formulation. SPESN38-5 and SPESN38-8 demonstrate better PK values, lower toxicity, and superior antitumor efficacy in mouse models, compared with irinotecan and DOX., (© 2023. The Author(s).)

The tumor suppressor p53 is the most frequently mutated protein in human cancer and tops the list of high-value precision oncology targets. p53 prevents initiation and progression of cancer by inducing cell-cycle arrest and various forms of cell death. Tumors have thus evolved ways to inactivate p53, mainly by TP53 mutations or by hyperactive p53 degradation. This review focuses on two types of p53 targeting compounds, MDM2 antagonists and mutant p53 correctors. MDM2 inhibitors prevent p53 protein degradation, while correctors restore tumor suppressor activity of p53 mutants by enhancing thermodynamic stability. Herein we explore both novel and repurposed p53 targeting compounds, discuss their mode of action, and examine the challenges in advancing them to the clinic., Competing Interests: Declaration of interests P.K. is listed as inventor on the following patents, which describe molecules involved in p53 reactivation. Amaro, R.E. et al.: Small molecules to enhance p53 activity. US20160193214 A1. Luecke, H. et al. Small molecules for restoring function to p53 cancer mutants, US20150307519 A1. Kaiser, P. et al. Methods and compositions for treating cancer using small molecules that reactivate p53, provisional patent, UC Case 2020-611 (pending)., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Background: Whether pembrolizumab given both before surgery (neoadjuvant therapy) and after surgery (adjuvant therapy), as compared with pembrolizumab given as adjuvant therapy alone, would increase event-free survival among patients with resectable stage III or IV melanoma is unknown., Methods: In a phase 2 trial, we randomly assigned patients with clinically detectable, measurable stage IIIB to IVC melanoma that was amenable to surgical resection to three doses of neoadjuvant pembrolizumab, surgery, and 15 doses of adjuvant pembrolizumab (neoadjuvant-adjuvant group) or to surgery followed by pembrolizumab (200 mg intravenously every 3 weeks for a total of 18 doses) for approximately 1 year or until disease recurred or unacceptable toxic effects developed (adjuvant-only group). The primary end point was event-free survival in the intention-to-treat population. Events were defined as disease progression or toxic effects that precluded surgery; the inability to resect all gross disease; disease progression, surgical complications, or toxic effects of treatment that precluded the initiation of adjuvant therapy within 84 days after surgery; recurrence of melanoma after surgery; or death from any cause. Safety was also evaluated., Results: At a median follow-up of 14.7 months, the neoadjuvant-adjuvant group (154 patients) had significantly longer event-free survival than the adjuvant-only group (159 patients) (P = 0.004 by the log-rank test). In a landmark analysis, event-free survival at 2 years was 72% (95% confidence interval [CI], 64 to 80) in the neoadjuvant-adjuvant group and 49% (95% CI, 41 to 59) in the adjuvant-only group. The percentage of patients with treatment-related adverse events of grades 3 or higher during therapy was 12% in the neoadjuvant-adjuvant group and 14% in the adjuvant-only group., Conclusions: Among patients with resectable stage III or IV melanoma, event-free survival was significantly longer among those who received pembrolizumab both before and after surgery than among those who received adjuvant pembrolizumab alone. No new toxic effects were identified. (Funded by the National Cancer Institute and Merck Sharp and Dohme; S1801 ClinicalTrials.gov number, NCT03698019.)., (Copyright © 2023 Massachusetts Medical Society.)

Autologous T cells transduced to express a high affinity T-cell receptor specific to NY-ESO-1 (letetresgene autoleucel, lete-cel) show promise in the treatment of metastatic synovial sarcoma, with 50% overall response rate. The efficacy of lete-cel treatment in 45 synovial sarcoma patients (NCT01343043) has been previously reported, however, biomarkers predictive of response and resistance remain to be better defined. This post-hoc analysis identifies associations of response to lete-cel with lymphodepleting chemotherapy regimen (LDR), product attributes, cell expansion, cytokines, and tumor gene expression. Responders have higher IL-15 levels pre-infusion (p = 0.011) and receive a higher number of transduced effector memory (CD45RA- CCR7-) CD8 + cells per kg (p = 0.039). Post-infusion, responders have increased IFNγ, IL-6, and peak cell expansion (p < 0.01, p < 0.01, and p = 0.016, respectively). Analysis of tumor samples post-treatment illustrates lete-cel infiltration and a decrease in expression of macrophage genes, suggesting remodeling of the tumor microenvironment. Here we report potential predictive and pharmacodynamic markers of lete-cel response that may inform LDR, cell dose, and strategies to enhance anticancer efficacy., (© 2022. This is a U.S. Government work and not under copyright protection in the US; foreign copyright protection may apply.)

Therapy for cutaneous melanoma, the deadliest of the skin cancers, is inextricably linked to the immune system. Once thought impossible, cures for metastatic melanoma with immune checkpoint inhibitors have been developed within the last decade and now occur regularly in the clinic. Unfortunately, half of tumors do not respond to checkpoint inhibitors and efforts to further exploit the immune system are needed. Tantalizing associations with immune health and gut microbiome composition suggest we can improve the success rate of immunotherapy. The gut contains over half of the immune cells in our bodies and increasingly, evidence is linking the immune system within our gut to melanoma development and treatment. In this review, we discuss the importance the skin and gut microbiome may play in the development of melanoma. We examine the differences in the microbial populations which inhabit the gut of those who develop melanoma and subsequently respond to immunotherapeutics. We discuss the role of dietary intake on the development and treatment of melanoma. And finally, we review the landscape of published and registered clinical trials therapeutically targeting the microbiome in melanoma through dietary supplements, fecal microbiota transplant, and microbial supplementation., (© 2022. The Author(s).)

Opinion Statement: Clinical trial enrollment should be actively encouraged in all patients diagnosed with advanced, surgically unresectable chondrosarcoma (CS) due to the lack of consensus treatment recommendations. In the absence of an appropriate clinical trial, treatments are determined based on histologic subtype of CS with consideration given to targetable mutations (i.e., IDH1). Conventional CS is inherently resistant to cytotoxic chemotherapy and patients may benefit from antiangiogenic therapy including off-label use of pazopanib. Individuals harboring an IDH1 mutation may derive clinical benefit from ivosidenib, an IDH1 inhibitor. Upon progression and with functional status permitting, alternative options include mTOR inhibitors (sirolimus, temsirolimus) or other tyrosine kinase inhibitors (dasatinib), though no clear sequencing data exists. For dedifferentiated CS, conventional chemotherapies with osteosarcoma-like regimens are upfront options although prospective data is limited with minimal overall benefit. Alternative treatment options include immunotherapy with pembrolizumab or ivosidenib in IDH1-mutant, dedifferentiated CS, but questionable efficacy was observed in small sample sizes with either approach. In mesenchymal CS, treatment with Ewing sarcoma-like chemotherapy regimens may be considered, although data supporting its use is even more limited given its rarity., (© 2022. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Purpose: Soft-tissue sarcomas (STS) are a rare, heterogeneous group of mesenchymal tumors. For decades the mainstay of treatment for advanced, unresectable STS has been palliative chemotherapy. High levels of activated MET receptor have been reported in various sarcoma cell lines, together with elevated vascular endothelial growth factor (VEGF) levels in patients with STS, suggesting that dual targeting of the VEGF and MET pathways with the multi-receptor tyrosine kinase inhibitor cabozantinib would result in clinical benefit in this population., Patients and Methods: We performed an open-label, multi-institution, single-arm phase II trial of single-agent cabozantinib in adult patients with advanced STS and progressive disease after at least 1 standard line of systemic therapy. Patients received 60 mg oral cabozantinib once daily in 28-day cycles, and dual primary endpoints of overall response rate and 6-month progression-free survival (PFS) were assessed. Changes in several circulating biomarkers were assessed as secondary endpoints., Results: Six (11.1%; 95% CI, 4.2%-22.6%) of the 54 evaluable patients enrolled experienced objective responses (all partial responses). Six-month PFS was 49.3% (95% CI, 36.2%-67.3%), with a median time on study of 4 cycles (range, 1-99). The most common grade 3/4 adverse events were hypertension (7.4%) and neutropenia (16.7%). Patients' levels of circulating hepatocyte growth factor (HGF), soluble MET, and VEGF-A generally increased after a cycle of therapy, while soluble VEGFR2 levels decreased, regardless of clinical outcome., Conclusions: Cabozantinib single-agent antitumor activity was observed in patients with selected STS histologic subtypes (alveolar soft-part sarcoma, undifferentiated pleomorphic sarcoma, extraskeletal myxoid chondrosarcoma, and leiomyosarcoma) highlighting the biomolecular diversity of STS., (©2021 The Authors; Published by the American Association for Cancer Research.)

Background: This single-arm, multicenter, phase 2 study evaluated the safety and antitumor activity of pazopanib in patients with unresectable, pulmonary metastatic osteosarcoma. Patients and Methods . Patients with pulmonary metastatic osteosarcoma unresponsive to chemotherapy were eligible. Patients who received prior tyrosine kinase inhibitor therapy were excluded. Pazopanib at 800 mg once daily was administered for 28-day cycles. Tumor responses were evaluated by local radiology assessment 1 month prior to and after initiation of treatment to calculate tumor doubling time and after every even numbered cycle. The primary endpoints were progression-free survival at 4 months, concomitant with a demonstrated 30% increase in tumor doubling time relative to the pretreatment growth rate., Results: 12 patients (7 female) were enrolled. The study was terminated prematurely due to withdrawal of financial support by the sponsor. 8 subjects were eligible for the primary analysis, whereas 4 patients were in a predefined exploratory "slow-growing" cohort. In the "fast-growing" cohort, 3 of the 8 patients (37.5%) eligible for first-stage analysis were deemed "success" by the preplanned criteria, adequate to proceed to second-stage accrual. In addition, 1 of the 4 patients in the "slow-growing" cohort experienced a partial remission. Grade 1-2 diarrhea was the most common adverse event, and grade 3 events were infrequent., Conclusion: This study illustrates a novel method of demonstrating positive drug activity in osteosarcoma by increasing tumor doubling time, and this is further supported by a partial response in a patient with "slow-growing" disease. This trial is registered with NCT01759303., Competing Interests: The authors declare they have no conflicts of interest., (Copyright © 2022 Paul Frankel et al.)

Small-molecule chemotherapeutics are potent and effective against a variety of malignancies, but common and severe side effects restrict their clinical applications. Nanomedicine approaches represent a major focus for improving chemotherapy, but have met limited success. To overcome the limitations of chemotherapy drugs, we have developed a novel Single Protein Encapsulation (SPE)-based drug formulation and delivery platform and tested its utility in improving doxorubicin (DOX) treatment. Using this methodology, a series of SPEDOX complexes were generated by encapsulating various numbers of DOX molecules into a single human serum albumin (HSA) molecule. UV/fluorescence spectroscopy, membrane dialysis, and dynamic light scattering techniques showed that SPEDOXs are stable and uniform as monomeric HSA and display unique properties distinct from those of DOX and DOX-HSA mixture. Furthermore, detailed procedures to precisely monitor and control both DOX payload and binding strength to HSA were established. Breast cancer xenograft tumor studies revealed that SPEDOX-6 treatment displays improved pharmacokinetic profiles, higher antitumor efficacy, and lower DOX accumulation in the heart tissue compared with unformulated DOX. This SPE technology, which does not involve nanoparticle assembly and modifications to either small-molecule drugs or HSA, may open up a new avenue for developing new drug delivery systems to improve anticancer therapeutics., Competing Interests: Conflict of Interest The author C. J. Yu is a named inventor for patent applications regarding “Single Protein-Encapsulated Pharmaceutics for Enhancing Therapeutic Effects” in pending. The other authors declare no conflict of interest.

Background: Desmoplastic small round cell tumor (DSRCT) is a rare, highly aggressive, translocation-associated soft-tissue sarcoma that primarily affects children, adolescents, and young adults, with a striking male predominance. It is characterized by t(11;22) generating a novel EWSR1-WT1 fusion gene. Secondary genomic alterations are rarely described., Methods: Tumor tissue from 83 DSRCT patients was assayed by hybrid-capture based comprehensive genomic profiling, FoundationOne® Heme next generation sequencing analysis of 406 genes and RNA sequencing of 265 genes. Tumor mutation burden was calculated from a minimum of 1.4 Mb sequenced DNA. Microsatellite instability status was determined by a novel algorithm analyzing 114 specific loci., Results: Comprehensive genomic profiling identified several genomically-defined DSRCT subgroups. Recurrent genomic alterations were most frequently detected in FGFR4, ARID1A, TP53, MSH3, and MLL3 genes. With the exception of FGFR4, where the genomic alterations predicted activation, most of the alterations in the remaining genes predicted gene inactivation. No DSRCT were TMB or MSI high., Conclusions: In summary, recurrent secondary somatic alterations in FGFR4, ARID1A, TP53, MSH3, and MLL3 were detected in 82% of DSRCT, which is significantly greater than previously reported. These alterations may have both prognostic and therapeutic implications.

Background: Gene-modified autologous T cells expressing NY-ESO-1 c259 , an affinity-enhanced T-cell receptor (TCR) reactive against the NY-ESO-1-specific HLA-A*02-restricted peptide SLLMWITQC (NY-ESO-1 SPEAR T-cells; GSK 794), have demonstrated clinical activity in patients with advanced synovial sarcoma (SS). The factors contributing to gene-modified T-cell expansion and the changes within the tumor microenvironment (TME) following T-cell infusion remain unclear. These studies address the immunological mechanisms of response and resistance in patients with SS treated with NY-ESO-1 SPEAR T-cells., Methods: Four cohorts were included to evaluate antigen expression and preconditioning on efficacy. Clinical responses were assessed by RECIST v1.1. Engineered T-cell persistence was determined by qPCR. Serum cytokines were evaluated by immunoassay. Transcriptomic analyses and immunohistochemistry were performed on tumor biopsies from patients before and after T-cell infusion. Gene-modified T-cells were detected within the TME via an RNAish assay., Results: Responses across cohorts were affected by preconditioning and intra-tumoral NY-ESO-1 expression. Of the 42 patients reported (data cut-off 4June2018), 1 patient had a complete response, 14 patients had partial responses, 24 patients had stable disease, and 3 patients had progressive disease. The magnitude of gene-modified T-cell expansion shortly after infusion was associated with response in patients with high intra-tumoral NY-ESO-1 expression. Patients receiving a fludarabine-containing conditioning regimen experienced increases in serum IL-7 and IL-15. Prior to infusion, the TME exhibited minimal leukocyte infiltration; CD163 + tumor-associated macrophages (TAMs) were the dominant population. Modest increases in intra-tumoral leukocytes (≤5%) were observed in a subset of subjects at approximately 8 weeks. Beyond 8 weeks post infusion, the TME was minimally infiltrated with a TAM-dominant leukocyte infiltrate. Tumor-associated antigens and antigen presentation did not significantly change within the tumor post-T-cell infusion. Finally, NY-ESO-1 SPEAR T cells trafficked to the TME and maintained cytotoxicity in a subset of patients., Conclusions: Our studies elucidate some factors that underpin response and resistance to NY-ESO-1 SPEAR T-cell therapy. From these data, we conclude that a lymphodepletion regimen containing high doses of fludarabine and cyclophosphamide is necessary for SPEAR T-cell persistence and efficacy. Furthermore, these data demonstrate that non-T-cell inflamed tumors, which are resistant to PD-1/PD-L1 inhibitors, can be treated with adoptive T-cell based immunotherapy., Trial Registration: ClinicalTrials.gov, NCT01343043 , Registered 27 April 2011.

Chondrosarcomas constitute a heterogeneous group of primary bone cancers characterized by hyaline cartilaginous neoplastic tissue. They are the second most common primary bone malignancy. The vast majority of chondrosarcomas are conventional chondrosarcomas, and most conventional chondrosarcomas are low- to intermediate-grade tumors (grade 1 or 2) which have indolent clinical behavior and low metastatic potential. Recurrence augurs a poor prognosis, as conventional chondrosarcomas are both radiation and chemotherapy resistant. Recent discoveries in the biology, genetics, and epigenetics of conventional chondrosarcomas have significantly advanced our understanding of the pathobiology of these tumors and offer insight into potential therapeutic targets., Competing Interests: No competing interests were disclosed.No competing interests were disclosed.No competing interests were disclosed.

Importance: Gastrointestinal stromal tumors (GISTs) are life-threatening when metastatic or not amenable to surgical removal. In a few patients with advanced GISTs refractory to imatinib mesylate, treatment with sunitinib malate followed by regorafenib provides tumor control; however, additional active treatments are needed for most patients., Objective: To evaluate the 6-month progression-free survival (PFS), tumor objective response, and overall survival rates in patients with GISTs treated with dasatinib., Design, Setting, and Participants: This single-arm clinical trial used a Bayesian design to enroll patients 13 years or older with measurable imatinib-refractory metastatic GISTs treated at 14 sarcoma referral centers from June 1, 2008, through December 31, 2009. A control group was not included. Patients were followed up for survival for a minimum of 5 years from date of enrollment. Tumor imaging using computed tomography or magnetic resonance imaging was performed every 8 weeks for the first 24 weeks and every 12 weeks thereafter. Tumor response was assessed by local site using the Choi criteria. Treatment was continued until tumor progression, unacceptable toxic effects after reduction in drug dose, or patient or physician decision. Archival tumor tissue was evaluated for expression of the proto-oncogene tyrosine-protein kinase Src (SRC), phosphorylated SRC (pSRC), and succinate dehydrogenase complex iron sulfur subunit B (SDHB) proteins and for mutation in the V-Kit Hardy-Zuckerman 4 feline sarcoma viral oncogene homolog (KIT) and platelet-derived growth factor receptor α (PDGFRA) genes. Data analysis was performed from May 19, 2017, through December 20, 2017., Interventions: Dasatinib, 70 mg orally twice daily., Main Outcomes and Measures: The primary end point was the 6-month PFS estimate using greater than 30% as evidence of an active drug and less than 10% as evidence of inactive treatment., Results: In this study, 50 patients were enrolled (median age, 60 years; age range, 19-78 years; 31 [62%] male and 19 [38%] female; 41 [82%] white), and 48 were evaluable for response. The estimated 6-month PFS rate was 29% in the overall population and 50% in a subset of 14 patients with pSRC in GISTs. Objective tumor response was observed in 25%, including 1 patient with an imatinib-resistant mutation in PDGFRA exon 18., Conclusions and Relevance: Dasatinib may have activity in a subset of patients with imatinib-resistant GISTs. Further study is needed to determine whether pSRC is a prognostic biomarker.

SK-UT-1 uterine leiomyosarcomas (Ut-LMS) cells were transduced with a fatty acid synthase (FASN)-containing retroviral vector to recapitulate the "lipogenic phenotype of cancer." Consistent with this model, forced expression of FASN enhanced SK-UT-1 proliferation, migration, and cellular motion. Further investigation showed FASN promotes trimethylation of H3K9 (H3K9me3) and acetylation of H3K27 (H3K27ac) in SK-UT-1 cells. In contrast, siRNA targeting of FASN in high endogenous FASN expressing SK-LMS-1 Ut-LMS cells inhibits trimethylation of H3K9 and acetylation of H3K27. Palmitate, the predominant fatty acid product of FASN, increased H3K9me3, H3K27ac and H3K27me3 detection in SK-UT-1 cells. FASN promoted histone 3 methylation and acetylation through alteration of histone 3-modifying enzymatic activities (HDAC, HDM, HMT and HAT). ChIP-seq in SK-UT-1-FASN cells with anti-H3K9me3 antibody identified regions of enriched binding compared to vector-only cells. One differentially-enriched gene, CRISP1, was investigated further by ChIP-PCR. The transcriptionally repressive function of H3K9me3 was confirmed in CRISP1. Our results provide mechanistic insight into the pathobiology of the "lipogenic phenotype of cancer." Here, FASN reprograms the Ut-LMS epigenome through chromatin remodeling to promote the "malignant phenotype."

Background: Alveolar soft part sarcoma (ASPS), chondrosarcoma (CS), chordoma, epithelioid sarcoma, and solitary fibrous tumor (SFT) are malignant tumors that are relatively resistant to chemotherapy and for which more effective drug therapy is needed., Methods: The 5 listed subtypes were enrolled into a single indolent sarcoma cohort in a phase 2 study of dasatinib using a Bayesian continuous monitoring rule for enrollment. The primary objective was to estimate the 6-month progression-free survival (PFS) rate according to the Choi criteria with a target of ≥50%. Cross-sectional imaging was performed before the start of treatment, every 2 months for 6 months, and then every 3 months during treatment. The 2- and 5-year survival rates were determined., Results: One hundred sixteen patients were enrolled within 45 months, and 109 began treatment with dasatinib. The 6-month PFS rate and the median PFS were 48% and 5.8 months, respectively. The PFS rate at 6 months was highest with ASPS (62%) and lowest with SFT (30%). More than 10% of the patients with ASPS, CS, or chordoma had stable disease for more than 1 year. Collectively, for all 5 subtypes, the 2- and 5-year overall survival rates were 44% and 13%, respectively. An objective response was observed in 18% of the patients with CS or chordoma., Conclusions: Dasatinib failed to achieve control of sarcoma growth for at least 6 months in more than 50% of the patients in this trial according to the Choi tumor response criteria. An objective tumor response and prolonged stable disease was observed in >10% of patients with CS or chordoma. Cancer 2017;90-97. © 2016 American Cancer Society., (© 2016 American Cancer Society.)

Background: Dasatinib exhibited activity in preclinical models of sarcoma. The Sarcoma Alliance for Research through Collaboration (SARC) conducted a multicenter, phase 2 trial of dasatinib in patients with advanced sarcoma., Methods: Patients received dasatinib twice daily. The primary objective was to estimate the clinical benefit rate (CBR) (complete response or partial response within 6 months or stable disease duration of ≥6 months) with a target of ≥25%. Patients were enrolled into 1 of 7 different cohorts and assessed by imaging every 8 weeks using Choi criteria tumor response and a Bayesian hierarchical design. For each subtype, enrollment was stopped after a minimum of 9 patients were treated if there was a <1% chance the CBR was ≥25%., Results: A total of 200 patients were enrolled. Accrual was stopped early in 5 cohorts because of low CBR. The leiomyosarcoma (LMS) and undifferentiated pleomorphic sarcoma (UPS) cohorts fully accrued and 6 of 47 and 8 of 42 evaluable patients, respectively, exhibited clinical benefit. The probability that the CBR was ≥25% in the LMS and UPS cohorts was 0.008 and 0.10, respectively. The median progression-free survival ranged from 0.9 months in patients with rhabdomyosarcoma to 2.2 months in patients with LMS. The median overall survival was 8.6 months. The most frequent adverse events were constitutional, gastrointestinal, and respiratory, and 36% of patients required dose reduction for toxicity. Serious adverse events attributed to therapy occurred in 11% of patients., Conclusions: Dasatinib may have activity in patients with UPS but is inactive as a single agent in the other sarcoma subtypes included herein. The Bayesian design allowed for the early termination of accrual in 5 subtypes because of lack of drug activity., (© 2015 American Cancer Society.)

Background: The MEK inhibitor, selumetinib, suppresses soft-tissue sarcoma (STS) cell proliferation in vitro. Mammalian target of rapamycin inhibitors possess modest activity against STS; however, resistance develops via MAPK pathway feedback activation. The combination of selumetinib and temsirolimus synergistically inhibits STS cell line growth. Therefore, a randomized phase II trial of selumetinib vs selumetinib plus temsirolimus was conducted., Methods: Seventy-one adults with advanced STS who received ⩽ 2 prior chemotherapeutics were randomized to selumetinib 75 mg p.o. bid and allowed to crossover upon progression, or to selumetinib 50 mg p.o. bid plus temsirolimus 20 mg i.v. weekly, with primary endpoint of progression-free survival (PFS)., Results: There was no difference in PFS between the two arms for the overall cohort (median 1.9 vs 2.1 months); an improved median PFS was observed in the combination arm (N = 11) over single agent (N = 10) in the prespecified leiomyosarcoma stratum (median 3.7 vs 1.8 months; P = 0.01). Four-month PFS rate was 50% (95% confidence interval 0.19-0.81) with the combination vs 0% with selumetinib alone in the leiomyosarcoma cohort. Most common grade 3/4 adverse events with the combination were mucositis (29%), lymphopenia (26%), neutropenia and anaemia (20% each)., Conclusions: While single-agent selumetinib has no significant activity in STS, the combination may be active for leiomyosarcomas.

Nelfinavir and its analogs inhibit proliferation and induce apoptosis of castration-resistant prostate cancer through inhibition of site-2 protease (S2P) activity, which leads to suppression of regulated intramembrane proteolysis. Western blotting in nelfinavir and its analog treated cells confirms accumulation of precursor SREBP-1 and ATF6. Nelfinavir and its analogs inhibit human homolog M. jannaschii S2P cleavage of an artificial protein substrate CED-9 in an in vitro proteolysis assay in a dose-dependent manner. Nelfinavir and its analogs are more potent inhibitors of S2P cleavage activity than 1,10-phenanthroline, a metalloprotease-specific inhibitor. Further, cluster analysis of gene expression from treated DU145 and PC3 cell lines demonstrate a close similarity of nelfinavir, its analogs, and 1,10-phenanthroline. These results show nelfinavir and its analogs inhibit castration-resistant prostate cancer proliferation by blocking regulated intramembrane proteolysis through suppression of S2P cleavage activity. This leads to accumulation of precursor SREBP-1 and ATF6, and development of insufficient reserves of their transcriptionally-active forms. The present results validate S2P and regulated intramembrane proteolysis as novel therapeutic targets for castration-resistant prostate cancer therapeutics. A clinical trial of nelfinavir or its analogs should be developed for castration-resistant prostate cancer.

Background: Insulin-like growth factor-1 receptor (IGF-1R) is implicated in the pathogenesis of rhabdomyosarcoma (RMS), osteosarcoma (OS), and synovial sarcoma (SS). The authors conducted a multi-institutional phase 2 trial of the monoclonal antibody R1507 in patients with various subtypes of recurrent or refractory sarcomas., Methods: Eligibility criteria included age ≥ 2 years and a diagnosis of recurrent or refractory RMS, OS, SS, and other soft tissue sarcomas. Patients received a weekly dose of 9 mg/kg R1507 intravenously. The primary endpoint was the best objective response rate using World Health Organization criteria. Tumor imaging was performed every 6 weeks × 4 and every 12 weeks thereafter., Results: From December 2007 through August 2009, 163 eligible patients from 33 institutions were enrolled. The median patient age was 31 years (range, 7-85 years). Histologic diagnoses included OS (n = 38), RMS (n = 36), SS (n = 23), and other sarcomas (n = 66). The overall objective response rate was 2.5% (95% confidence interval, 0.7%-6.2%). Partial responses were observed in 4 patients, including 2 patients with OS, 1 patient with RMS, and 1 patient with alveolar soft part sarcoma. Four additional patients (3 with RMS and 1 with myxoid liposarcoma) had a ≥ 50% decrease in tumor size that lasted for <4 weeks. The median progression-free survival was 5.7 weeks, and the median overall survival was 11 months. The most common grade 3/4 toxicities were metabolic (12%), hematologic (6%), gastrointestinal (4%), and general constitutional symptoms (8%)., Conclusions: R1507 is safe and well tolerated but has limited activity in patients with recurrent or refractory bone and soft tissue sarcomas. Additional studies to help identify the predictive factors associated with clinical benefit in selected histologies such as RMS appear to be warranted., (© 2014 American Cancer Society.)

Purpose: HIV protease inhibitors are associated with HIV protease inhibitor-related lipodystrophy syndrome. We hypothesized that liposarcomas would be similarly susceptible to the apoptotic effects of an HIV protease inhibitor, nelfinavir., Methods: We conducted a phase I trial of nelfinavir for liposarcomas. There was no limit to prior chemotherapy. The starting dose was 1,250 mg twice daily (Level 1). Doses were escalated in cohorts of three to a maximally evaluated dose of 4,250 mg (Level 5). One cycle was 28 days. Steady-state pharmacokinetics (PKs) for nelfinavir and its primary active metabolite, M8, were determined at Levels 4 (3,000 mg) and 5., Results: Twenty subjects (13 males) were enrolled. Median (range) age was 64 years (37-81). One subject at Level 1 experienced reversible, grade 3 pancreatitis after 1 week and was replaced. No other dose-limiting toxicities were observed. Median (range) number of cycles was 3 (0.6-13.5). Overall best responses observed were 1 partial response, 1 minor response, 4 stable disease, and 13 progressive disease. Mean peak plasma levels and AUCs for nelfinavir were higher at Level 4 (7.3 mg/L; 60.9 mg/L × h) than 5 (6.3 mg/L; 37.7 mg/L × h). The mean ratio of M8:nelfinavir AUCs for both levels was ~1:3., Conclusions: PKs demonstrate auto-induction of nelfinavir clearance at the doses studied, although the mechanism remains unclear. Peak plasma concentrations were within range where anticancer activity was demonstrated in vitro. M8 metabolite is present at ~1/3 the level of nelfinavir and may also contribute to the anticancer activity observed.

Nelfinavir induces apoptosis in liposarcoma by inhibiting site-2 protease (S2P) activity, which leads to suppression of regulated intramembrane proteolysis. We postulate similar effects in castration-resistant prostate cancer because it exhibits a lipogenic phenotype. Nelfinavir inhibited androgen receptor activation in androgen-sensitive prostate cancer and the nuclear translocation of the fusion proteins sterol regulatory element binding protein-1 (SREBP-1)-enhanced green fluorescence protein (EGFP) and activating transcription factor 6 (ATF6)-EGFP in castration-resistant prostate cancer cells, viewed under confocal microscopy. Nelfinavir and site-1 protease (S1P) and S2P small interfering RNAs (siRNAs) reduced the proliferation of castration-resistant prostate cancer and induced apoptosis, which was opposed by autophagy. Inhibition of autophagy with hydroxychloroquine was additive to the apoptotic effect of nelfinavir. Western blotting of S1P and S2P siRNA knockdown and/or nelfinavir-treated cells confirmed the accumulation of precursor SREBP-1 and ATF6. 3,4-Dichloroisocoumarin, an S1P inhibitor, did not affect SREBP-1 processing. In contrast, 1,10-phenanthroline, an S2P inhibitor, reproduced the nelfinavir-treated molecular and biological phenotype. Nelfinavir-mediated inhibition of regulated intramembrane proteolysis led to the accumulation of unprocessed SREBP-1 and ATF6. This resulted in sequential endoplasmic reticulum stress, inhibition of the unfolded protein response, reduced fatty acid synthase expression and apoptosis, which was countered by autophagy. Inhibition of autophagy was at least additive to this pro-apoptotic effect. These findings provide new insights into nelfinavir-induced endoplasmic reticulum stress and cancer cell death, and lead us to propose investigating its clinical activity in castration-resistant prostate cancer. This report validates S2P as a therapeutic target in castration-resistant prostate cancer., (© 2012 City of Hope and Beckman Research Institute. Journal compilation © 2012 FEBS.)

As a primary malignant bone tumor, osteosarcoma is second only to chondrosarcoma. Although it commonly metastasizes and is aggressive in nature, it rarely colonizes the skin. This is a report of a 22-year-old male with osteosarcoma of the pelvis and metastasis to the lungs and chest wall who developed a clinically unsuspected solitary cutaneous metastasis in the scalp. Instead of the expected cyst, incisional biopsy disclosed a solid tan nodule of chondro-osseous sarcoma. Although rare, cutaneous metastases from osteosarcoma may appear in skin, especially the scalp, or in skin over the primary tumor. New skin lesions in a patient with a history of osteosarcoma warrant investigations including imaging and biopsy.

Purpose: We previously reported that nelfinavir (NFV) induces G(1) cell-cycle block and apoptosis selectively in liposarcoma cell lines due to increased SREBP-1 (sterol regulatory element binding protein-1) expression in the absence of increased transcription. We postulate that NFV interferes with regulated intramembrane proteolysis of SREBP-1 and ATF6 (activating transcription factor 6)., Experimental Design: Time-lapse, confocal microscopic studies show that NFV inhibits the nuclear translocation of full-length SREBP-1-EGFP and ATF6-EGFP fusion proteins. siRNA-mediated knockdown of site-1 protease (S1P) and/or site-2 protease (S2P) leads to inhibition of SREBP-1 intracellular trafficking to the nucleus and reduces liposarcoma cell proliferation. Treatment of LiSa-2 liposarcoma cells with 3,4-dichloroisocoumarin, a serine protease inhibitor of S1P, did not affect SREBP-1 processing. In contrast, 1,10-phenanthroline, an S2P-specific inhibitor, reproduces the molecular and biological phenotypes observed in NFV-treated cells, which implicates S2P as a target of NFV. In vivo evaluation of NFV in a murine liposarcoma xenograft model leads to inhibition of tumor growth without significant toxicity., Results: NFV-induced upregulation of SREBP-1 and ATF6 results from inhibition of S2P, which together with S1P mediates regulated intramembrane proteolysis from their precursor to their transcriptionally active forms. The resulting endoplasmic reticulum (ER) stress and concurrent inhibition of the unfolded protein response induce caspase-mediated apoptosis., Conclusions: These results provide new insight into the mechanism of NFV-mediated induction of ER stress and cell death in liposarcomas and are the first to report targeting S2P for cancer therapy.

Systemic therapy options for sarcomas historically have been limited once these tumors become resistant to traditional cytotoxic chemotherapy. Ongoing preclinical research into their biology and clinical trials based on rational biologic targeting have identified novel therapies. For example, the success of imatinib in gastrointestinal stromal tumor has led to the use of other tyrosine kinase inhibitors in other sarcoma subtypes. Other novel therapies include targeting of the mTOR pathway, and IGF-1 receptor. The heterogeneity of these tumors demands intelligently designed protocols in recognizing efficacy that may be restricted to certain histologic subtypes. This article will cover recent trials of new biologic agents in sarcomas that have exhibited promising activity.

Background: We have previously shown that the receptor tyrosine kinases, KIT and PDGFRalpha, are expressed on ESFT cell lines, and that imatinib induces dose-dependent apoptosis (1). We conducted a Phase II trial to evaluate the effectiveness of imatinib for patients with recurrent ESFT or DSRCT expressing KIT and/or PDGFRalpha., Patients and Methods: Patients were selected for tumor immunohisto-chemical expression > or =2+/4+ for KIT or PDGFRalpha. Imatinib was administered orally 400 mg twice/day for 28 days/course. Primary endpoint was response., Results: Seven patients were enrolled and evaluated. One patient with 3+/4+ PDGFRalpha and 3+/4+ KIT expression had a partial response through 8 courses. 4 patients had progression after 1 cycle. Two patients were not evaluable due to one early death and one refusing treatment., Conclusion: This study intended to enrich for molecular factors that potentially predict response. Given the poor prognosis with recurrent ESFT, further studies with other novel KIT and PDGFRalpha inhibitors are needed.

Background: GTI-2040 is a 20-mer antisense oligonucleotide targeting the mRNA of ribonucleotide reductase M2. It was combined with oxaliplatin and capecitabine in a phase I trial in patients with advance solid tumors based on previous studies demonstrating potentiation of chemotherapy with ribonucleotide reductase inhibitors., Methods: Patients at least 18 years of age with advanced incurable solid tumors and normal organ function as well as a Karnofsky performance status of > or =60% were eligible. One prior chemotherapy regimen for advanced disease or relapse within 12 months of adjuvant chemotherapy was required. Patients could have received prior fluoropyrimidines, including capecitabine, but not oxaliplatin. Treatment cycles were 21 days. In each cycle, GTI-2040 was given as a continuous intravenous infusion over 14 days, oxaliplatin as a 2-h intravenous infusion on day 1, and capecitabine orally twice a day for 14 days. In cycle 1 only, oxaliplatin and capecitabine were started on day 2 to allow ribonucleotide reductase mRNA levels to be measured with and without oxaliplatin and capecitabine. Doses were escalated in cohorts of three patients using a standard 3 + 3 design until the maximum tolerated dose was established, defined as no more than one first-cycle dose-limiting toxicity among six patients treated at a given dose level., Results: The maximum tolerated dose was estimated to be the combination of GTI-2040 3 mg/kg per day for 14 days, capecitabine 600 mg/m(2) twice daily for 14 days, and oxaliplatin 100 mg/m(2) every 21 days. Dose-limiting toxicities were hematologic. GTI-2040 pharmacokinetics, obtained at steady-state on days 7 and 14, showed the high inter-patient variability previously reported. Two of six patients had stable disease at the maximum tolerated dose and one patient, with heavily pre-treated non-small cell lung cancer, had a partial response at a higher dose level. In samples from a limited number of patients, there was no clear decrease in ribonucleotide reductase expression in peripheral blood mononuclear cells during treatment., Conclusion: A combination of GTI-2040, capecitabine and oxaliplatin is feasible in patients with advanced solid tumors.

Background: Patients with locally advanced or metastatic/recurrent soft tissue and Ewing's sarcoma (EWS) have few treatment options. The purpose of our phase II study was to assess the feasibility, safety and efficacy of tandem high-dose chemotherapy (HDCT) followed by autologous stem cell transplantation (ASCT) in such patients., Patients and Methods: Thirteen patients were enrolled onto this study. The first cycle of HDCT consisted of doxorubicin (150 mg/m(2)) and ifosfamide (14 g/m(2)) mixed with mesna (14 g/m(2)), while the second cycle consisted of melphalan (150 mg/m(2)) and cisplatin (200 mg/m(2))., Results: Eleven out of 13 patients were able to complete both cycles of HDCT. No treatment-related mortality occurred and grade 3 or 4 toxicity was clinically tolerable. The 5-year progression-free survival (PFS) and overall survival (OS) for all patients was 23% (confidence interval, CI: 0-46%) and 31% (CI: 14-70%), respectively. Out of the four patients still alive, two had EWS and measurable disease at the time of ASCT and achieved a complete remission, remaining progression free 126 and 155 months after ASCT., Conclusion: Our study demonstrates the feasibility and safety of tandem HDCT in patients with high-risk or metastatic/recurrent sarcoma, with some patients achieving long-term PFS and OS.

The use of anti-HIV drugs as cancer treatments is not new. Azidothymidine was studied as an antineoplastic in the 1990s, but despite promising in vitro data, clinical trials showed little antitumour activity. HIV protease inhibitors were developed in the early 1990s, and their subsequent incorporation into highly active antiretroviral therapy (HAART) has profoundly changed the natural history of HIV infection. The potential antitumour properties of these drugs have been investigated because of their success in treating HIV-related Kaposi's sarcoma. HAART's effects on Kaposi's sarcoma did not always correlate with immune reconstitution, and activity against other solid and haematological malignancies has been established. Inhibition of tumour-cell invasion and angiogenesis were properties first ascribed to inhibition of HIV protease; however, they have pleiotropic antitumour effects, including inhibition of inflammatory cytokine production, proteasome activity, cell proliferation and survival, and induction of apoptosis. HIV protease inhibitors are thus a new class of anticancer drugs with multiple effects, and other anti-HIV drugs might hold similar promise.

"HIV protease-induced lipodystrophy syndrome" is associated with the use of HIV protease inhibitors for treatment of HIV infection. In-vitro studies suggest that alteration of sterol regulatory element binding protein-1 levels underlie its pathogenesis. We postulated that HIV protease inhibitors may represent a novel class of antiliposarcoma agents. SW872, FU-DDLS-1 and LiSa-2 liposarcoma, and HT1080 and 293 nonliposarcoma cell lines were treated with HIV protease inhibitors (nelfinavir, ritonavir, saquinavir, indinavir and amprenavir), and clonogenic assays were performed. Nelfinavir exhibited the most potent inhibition of clonogenicity, and further assays for proliferation, cell cycle and apoptosis were performed with nelfinavir. Immunoblots were performed for sterol regulatory element binding protein-1, proapoptotic and cell cycle-related protein expression after nelfinavir treatment. Finally, a sterol regulatory element binding protein-1-inducible SW872 cell line was developed to examine the phenotype resulting from upregulated sterol regulatory element binding protein-1. Nelfinavir selectively inhibited clonogenicity and proliferation, and induced G1 cell cycle block and induced apoptosis in a dose-dependent manner in SW872 and LiSa-2 cells, whereas it had minimal or no effect on these parameters in FU-DDLS-1 or nonliposarcoma cells. Nelfinavir induced significant sterol regulatory element binding protein-1 expression in a dose-dependent and time-dependent fashion in sensitive SW872 and LiSa-2 cells, modestly in HT1080 cells, but not in nelfinavir-insensitive FU-DDLS-1 and 293 cells without inducing adipocytic differentiation. Forced expression of sterol regulatory element binding protein-1 in inducible-SW872 cells led to the induction of proapoptotic and antiproliferative proteins, and consequent reduction of cellular proliferation. Our data indicate that nelfinavir represents a novel class of antiliposarcoma agent that acts by selectively upregulating sterol regulatory element binding protein-1 expression in liposarcomas.

Chondrosarcomas are the second most common primary malignant tumor of bone. Chemotherapy for conventional chondrosarcomas is generally ineffective. Methylthioadenosine phosphorylase (MTAP) is a ubiquitous enzyme, essential in the salvage pathway of adenine and in methionine synthesis. MTAP-deficient cells are more susceptible than wild-type cells to pharmacologic inhibitors of de novo purine synthesis. Homozygous deletions of MTAP have been reported in hematologic and solid tumor malignancies. Based on these observations, we investigated the frequency of MTAP deletions in conventional, grade II chondrosarcomas by fluorescence in situ hybridization (FISH) analysis: 23 conventional, grade II chondrosarcoma patient samples from the Cleveland Clinic Foundation were analyzed for MTAP deletions. Nuclei were successfully extracted from 14 of 23 samples (61% evaluable) for FISH analysis: 7 of 14 samples (50%) showed either homozygous or hemizygous deletion of the MTAP gene, 6 of 14 (43%) failed to show deletion, and 1 of 14 (7%) was inconclusive. These findings suggest that approximately one-half of conventional, grade II chondrosarcomas may be preferentially sensitive to pharmacologic inhibitors of de novo purine synthesis. The present study led to development by the Intergroup Coalition Against Sarcomas of a phase II trial of pemetrexed, a multitargeted anti-folate, for advanced chondrosarcomas.

Purpose: Dexrazoxane administration prior to short infusion doxorubicin prevents anthracycline-related heart damage. Since delivery of doxorubicin by 96-h continuous intravenous infusion also reduces cardiac injury, we studied delivering dexrazoxane and doxorubicin concomitantly by prolonged intravenous infusion., Methods: Patients with advanced malignancies received tandem cycles of concurrent 96-h infusions of dexrazoxane 500 mg/m2 and doxorubicin 165 mg/m2, and 24 h after completion of chemotherapy, granulocyte-colony stimulating factor (5 microg/kg) and oral levofloxacin (500 mg) were administered daily until the white blood cell count reached 10,000 microl(-1). Plasma samples were analyzed for dexrazoxane and doxorubicin concentrations., Results: Ten patients were enrolled; eight patients had measurable disease. Two partial responses were observed in patients with soft-tissue sarcoma. The median number of days of granulocytopenia (<500 microl(-1)) was nine and of platelet count <20,000 microl(-1) was seven. Six patients received a single cycle because of progression (one), stable disease (four), or reversible, asymptomatic 10% decrease in cardiac ejection fraction (two). Principal grade 3/4 toxicities included hypotension (two), anorexia (four), stomatitis (four), typhlitis (two), and febrile neutropenia (seven), with documented infection (three). One death from neutropenic sepsis occurred. Dexrazoxane levels ranged from 1270 to 2800 nM, and doxorubicin levels ranged from 59.1 to 106.9 nM., Conclusions: These results suggest that tandem cycles of concurrent 96-h infusions of dexrazoxane and high-dose doxorubicin can be administered with minimal cardiac toxicity, and have activity in patients with recurrent sarcomas. However, significant non-cardiac toxicities indicate that the cardiac sparing potential of this approach would be maximized at lower dose levels of doxorubicin.

Aim: We describe the feasibility of combining infusional 5-fluorouracil (5-FU) with intraoperative radiation therapy (IORT)., Methods: Patients with surgically resectable locally advanced gastrointestinal cancers were treated concurrently during surgery with IORT and a 72 h infusion of 5-FU. Patients without previous external beam radiation therapy (EBRT) were subsequently treated with EBRT (40-50Gy) concurrent with a 21-day continuous infusion of 5-FU. Pancreatic, gastric, duodenal, ampullary, recurrent colorectal, and recurrent anal cancer were included., Results: During IORT/5-FU, no chemotherapy-related grade III or IV hematologic or gastrointestinal toxicity was noted. Post-surgical recovery or wound healing was not affected. One of nine patients who received post-operative radiation required a treatment break. During follow-up, there were more complications in patients with pelvic tumours, especially those with previous radiation. Nine patients have had local and/or local regional recurrences, two of these in the IORT field., Conclusions: Treatment with a combination of IORT and 5-FU followed by EBRT and 5-FU is feasible. However, long-term complications may be increased in previously irradiated recurrent pelvic tumours.

TCR engagement leads to the transcriptional activation of cytokine genes and activation-induced cell death. Activated T cells undergo apoptosis upon expression and ligation of Fas ligand (FasL) to Fas/APO-1 (CD95) receptor. FasL expression is under the transcriptional regulation of multiple factors. The present study demonstrates that TCR-inducible FasL expression is also under the direct influence of the IFN regulatory factor (IRF) transcription factor family. Deletion and mutagenesis of a putative IRF-1 binding site in the FasL promoter results in deficient expression of FasL. EMSAs demonstrate specific FasL promoter binding by IRF-1 and IRF-2. Forced expression of either IRF-1 or IRF-2 leads to FasL promoter activation in T cells and FasL expression in heterologous cells. Finally, suppression of IRF-1 expression in T cells results in deficient TCR-induced FasL expression. These results confirm that the IRF family participates in the regulation of FasL gene expression.

Isolated axillary and chest wall soft tissue masses are an uncommon presentation of metastatic cancer. The authors present three patients in whom malignant melanomas metastatic to these sites had been misdiagnosed, leading to inappropriate oncologic treatment planning in all three cases. The presumed diagnoses, even after fine-needle aspiration or trucut biopsies, were soft-tissue sarcoma (n = 2) and undifferentiated breast cancer (n = 1). The combination of taking a thorough history and performing proper immunohistochemical analysis of the biopsy material would have suggested the presence of malignant melanoma in all cases. As the disease appeared locoregionally limited in all patients, radical surgical resection with extended lymphadenectomy was performed without significant dysfunction of the upper extremity. One patient agreed to postoperative immunotherapy with interferon-alpha. Two patients are currently alive 17 and 14 months after operation. One patient was found to have systemic recurrence at 5 months, one experienced two isolated local recurrences in a prior operative site that were amenable to reresection and presently has no evidence of disease 12 months after resection, and one patient remains free of disease at 14 months. Clinical presentation, suggested diagnostic workup, and therapeutic implications are discussed to avoid misdiagnoses in this setting of possible clinical presentations of metastatic melanoma.

We have synthesized and tested a cis-cleaving ribozyme designed to have thermodynamically stable stem-loop structures. This cis-ribozyme cleaves very efficiently in vitro, with a cleavage rate of about 0.5/min. Surprisingly, during the course of in vitro transcription and cleavage of our ribozyme, a product of unusual mobility accumulates and coincides with a sharp decline in the rate of formation of cleavage products. Analyses of this electrophoretic variant demonstrated that it is formed by interactions of the cleavage products. Despite the fact that the products and ribozyme transcript are of identical sequence, the cleavage products interact only with one another and not with the uncleaved precursor. This suggests a significant structural difference between the cleaved and uncleaved ribozyme transcripts. Testing of this cis-ribozyme in both yeast and mammalian cells shows no significant cleavage activity in vivo. We conclude that the structure of the ribozyme flanking sequences is important for optimizing the rate of ribozyme cleavage, but this enhanced rate does not necessarily correlate with enhanced in vivo function.