The Oncolytic Activity of Myxoma Virus against Soft Tissue Sarcoma Is Mediated by the Overexpression of Ribonucleotide Reductase.

BACKGROUND: Myxoma virus (MYXV) is an oncolytic poxvirus that lacks the gene for 1 of the subunits of ribonucleotide reductase (RR), a crucial DNA synthesis and repair enzyme. The overexpression of RR has been implicated in the invasiveness of several cancers, including soft tissue sarcomas (STS). The purpose of the study was to investigate the oncolytic efficacy of MYXV in STS with different levels of RR expression. METHODS: The oncolytic effect of recombinant MYXV was evaluated in 4 human STS cell lines, LS141 (a dedifferentiated liposarcoma), DDLS8817 (a dedifferentiated liposarcoma), RDD2213 (recurrent dedifferentiated liposarcoma), and HSSYII (a synovial sarcoma) using infectivity and cytotoxicity assays. Following the overexpression of RRM2 by cDNA transfection and silencing of RRM2 by siRRM2 in these STS cell lines, the RRM2 expression levels were analyzed by Western blot. RESULTS: We observed a direct correlation between viral oncolysis and RRM2 mRNA levels (R = 0.96) in STS. Higher RRM2 expression was associated with a more robust cell kill. Silencing the RRM2 gene led to significantly greater cell survival (80%) compared with the control group (P = .003), whereas overexpression of the RRM2 increased viral oncolysis by 33% (P < .001). CONCLUSIONS: Our results show that the oncolytic effects of MYXV correlate directly with RR expression levels and are enhanced in STS cell lines with naturally occurring or artificially induced high expression levels of RR. Myxoma virus holds promise in the treatment of advanced soft tissue cancer, especially in tumors overexpressing RR.

A Single institution's randomized double-armed prospective study of lumpectomy margins with adjunctive use of the MarginProbe in nonpalpable breast cancers.

Breast conservation surgery (BCS) aims to excise all cancerous tissue while minimizing the amount of healthy breast tissue removed. Up to 30% of patients undergoing BCS require a second operation for re-excision to obtain negative margins. Previous studies reported a lower re-excision rate with intraoperative use of the MarginProbe device (Dune Medical Devices). This device utilizes radiofrequency spectroscopy to detect differences between cancerous and normal tissue. From July 2009 to January 2010, our institution enrolled 46 patients electing for BCS in a prospective double-arm randomized controlled trial and had a significantly lower re-excision rate than that reported in the multicenter trial. Intraoperatively, after performing conventional lumpectomy with excision of any additional shavings deemed necessary based on palpation and visual inspection alone, patients were then randomized. In the device arm, the surgeon used the MarginProbe to interrogate the lumpectomy specimen, taking additional shavings from the cavity surfaces corresponding to the parts of the specimen read as positive by the device. In the control arm, only standard intraoperative assessments were performed. All specimens were evaluated by pathologists who were blinded to the study arm. In this population, 72% had invasive ductal carcinoma (IDC), 20% had ductal carcinoma in situ (DCIS), and 8% had invasive lobular carcinoma (ILC). Average age was 64 years old. The average size of the specimen was 5.6 cm, the average volume was 37.8 cm3 , and the average weight was 32.7 g. The mean size of DCIS was 1.4 cm. For invasive specimens, 32 were T1 and 7 were T2. Prior to randomization, 43 patients were thought to have positive or close margins and therefore underwent additional shavings. Twenty-three patients were randomized to the device arm and 23 to the control arm. In the device arm, 14 (60%) patients had IDC, 7 (30%) had DCIS, and 2 (8%) had ILC, vs the control arm where 19 (82%) patients had IDC, 2 (8%) had DCIS, and 2 (8%) had ILC. Eight (35%) patients in the control group vs 1 (4%) in the device group underwent re-excision for margin involvement (P < .05). The use of the MarginProbe device at our institution significantly improved the ability of our surgeons to obtain clear margins during initial BCS. Our results show a lower re-excision rate (4%) than those published in the multicenter trial (19.8%). We postulate that in the face of more patients having DCIS in our device group (30%), our surgeons responded by taking thicker shavings when the MarginProbe device reported margin involvement during the initial lumpectomy, resulting in greater success achieving clear final margins on the shaved tissue and a significantly lower re-excision rate than previously reported with the MarginProbe device.

Contemporary reappraisal of the efficacy of adjuvant chemotherapy in resected retroperitoneal sarcoma: Evidence from a nationwide clinical oncology database and review of the literature.

BACKGROUND: While margin-negative resection remains the cornerstone of therapy for retroperitoneal sarcoma (RPS), the impact of adjuvant chemotherapy (AC) on overall survival (OS) remains poorly understood. METHODS: The National Cancer Data Base was queried for patients undergoing curative-intent resection of primary non-metastatic RPS (2004-2013). Multivariable modeling identified factors associated with AC receipt. Cox regression identified covariates associated with OS, and AC and surgery alone (SA) cohorts were matched 1:1 by propensity scores based on these covariates. In the propensity-score matched cohort, OS was compared by Kaplan-Meier estimates. Results from this analysis were presented in the context of a review of the existing literature on the impact of AC in resected RPS. RESULTS: Of 3892 resected RPS patients, 90.0% and 10.0% received SA and AC, respectively. Predictors of AC receipt included younger age, non-Caucasian race, hospital location, histologic grade, adjacent organ invasion, and histologic subtype. The propensity score-matched cohort comprised 767 patients (SA n = 377; AC n = 390); at a median follow-up of 59.2 (IQR 35.0-85.3) months, median OS of the propensity-matched cohort was 53.6 (IQR 22.4-119.5) months. Utilization of AC was associated with significantly worse long-term survival (median OS: 47.8 vs. 68.9 months, p = 0.017; HR 1.30, 95% CI 1.05-1.61). AC was not associated with improved OS in margin-positive (R1/R2) resection, high-grade (G2/G3) and larger (>10 cm) tumors, or in any histologic subtype. Albeit not statistically significant, there was a trend toward improved OS with AC in spindle cell (HR 0.37, 95% CI 0.10-1.38), giant cell (HR 0.82, 95% CI 0.32-2.13), and synovial (HR 0.26, 95% CI 0.05-1.33) sarcoma. CONCLUSIONS: Data from a large nationwide oncology database and review of the existing literature do not support adjuvant chemotherapy regimens following curative-intent resection of RPS, even in subgroups at high risk of failure (e.g., R1/R2 resection, high-grade or large tumors). The possible benefit of conventional adjuvant regimens in spindle cell, giant cell, and synovial sarcoma should be explored in prospective studies.

A cost-effective handheld breast scanner for use in low-resource environments: a validation study.

BACKGROUND: With the incidence of breast cancer rising worldwide, we are evaluating the iBreastExam (iBE) (UE LifeSciences Inc.), a handheld breast scanning device that can be utilized by community health workers to screen for breast abnormalities. The purpose of this study is to determine the sensitivity of the iBE in a population undergoing diagnostic breast imaging. METHODS: Adult patients presenting to a breast imaging center for a diagnostic workup were eligible. Patients underwent an iBE exam performed by a trained ultrasound technician followed by their indicated imaging. Demographic, imaging, and biopsy data were recorded. RESULTS: Seventy-eight iBE exams were completed, 77 females and one male with a mean age of 42 (21-79). All patients were evaluated by ultrasound, 52 had diagnostic mammography and 39 had biopsies. Imaging and/or biopsy confirmed a mass (fibroadenoma, cyst, papilloma, myofibroblastoma, fat necrosis, DCIS, or cancer) in 60 patients. Twelve patients had a cancer diagnosed. In total, 342 quadrants were scanned, 77 quadrants had lesions confirmed on imaging, and iBE correctly identified 66 lesions for a sensitivity of 86 % and specificity of 89 %. CONCLUSIONS: This validation study demonstrated excellent sensitivity of iBE for the identification of clinically significant lesions in patients presenting for diagnostic imaging. TRIAL REGISTRATION: A Cost-Effective Handheld Breast Scanner for Use in Low Resource Environments: A Validation Study: NCT02814292 .

Outcomes After Oncoplastic Breast-Conserving Surgery in Breast Cancer Patients: A Systematic Literature Review.

BACKGROUND: Surgeons have increasingly performed breast-conserving surgery (BCS) utilizing oncoplastic techniques in place of standard lumpectomy for early-stage breast cancer. We assess oncologic outcomes after oncoplastic BCS for T1-T2 breast cancer. METHODS: A systematic literature review identified peer-reviewed articles in PubMed evaluating BCS with oncoplastic reconstruction. Selected studies reported on positive margin rate (PMR), re-excision rate (RR), conversion to mastectomy rate (CMR), overall survival (OS), disease-free survival (DFS), local recurrence (LR), distant recurrence (DR), complication rate, and/or cosmetic outcomes. RESULTS: The search yielded 474 articles; 55 met the inclusion criteria and collectively evaluated 6011 patients with a mean age 54.6 years over a mean follow-up 50.5 months. T1 (43.8 %) and T2 (39.3 %) invasive ductal carcinoma were the most common tumor histopathologies. PMR, RR, and CMR were 10.8, 6.0, and 6.2 %, respectively, while OS, DFS, LR and DR were 95.0, 90.0, 3.2, and 8.7 %, respectively. Margin widths were heterogeneously defined in studies that included margin assessment. The PMR was not significantly different when positive margins were defined as tumor <10, <5, < 2, and <1 mm from ink margin, or tumor on ink (p = 0.162). Eleven studies reported specific margins for 1455 patients, of whom 143 (9.8 %) had positive margins, including 113 (7.8 %) with tumor on ink. CONCLUSIONS: This study is the largest comprehensive literature review to date on oncoplastic BCS. Our systematic review reveals high rates of OS and DFS with low LR, DR, PMR, RR, CMR and complication rates, thereby confirming the oncologic safety of this procedure in patients with T1-T2 invasive breast cancer.

Loss of MEN1 activates DNMT1 implicating DNA hypermethylation as a driver of MEN1 tumorigenesis.

Multiple endocrine neoplasia type 1 (MEN1) syndrome results from mutations in the MEN1 gene and causes tumor formation via largely unknown mechanisms. Using a novel genome-wide methylation analysis, we studied tissues from MEN1-parathyroid tumors, Men1 knockout (KO) mice, and Men1 null mouse embryonic fibroblast (MEF) cell lines. We demonstrated that inactivation of menin (the protein product of MEN1) increases activity of DNA (cytosine-5)-methyltransferase 1 (DNMT1) by activating retinoblastoma-binding protein 5 (Rbbp5). The increased activity of DNMT1 mediates global DNA hypermethylation, which results in aberrant activation of the Wnt/ß-catenin signaling pathway through inactivation of Sox regulatory genes. Our study provides important insights into the role of menin in DNA methylation and its impact on the pathogenesis of MEN1 tumor development.

Pasireotide (SOM230) is effective for the treatment of pancreatic neuroendocrine tumors (PNETs) in a multiple endocrine neoplasia type 1 (MEN1) conditional knockout mouse model.

BACKGROUND: Pasireotide (SOM230), a long-acting somatostatin analogue (LAR), has improved agonist activity at somatostatin receptors. We tested the effect of SOM230 on insulin secretion, serum glucose concentrations, tumor growth, and survival using an MEN1 transgenic mouse model. METHODS: Eight 12-month-old conditional Men1 knockout mice with insulinoma were assessed. The treatment (n = 4) and control groups (n = 4) received monthly subcutaneous injections of SOM230 or PBS. Serum insulin and glucose levels were determined by enzyme-linked immunosorbent assay and enzymatic colorimetric assay, respectively. Tumor activity, growth, and apoptosis were determined by microPET/CT scan and histologic analysis. RESULTS: On day 7, there was a decrease in serum insulin levels from 1.06 ± 0.28 µg/L to 0.37 ± 0.17 µg/L (P = .0128) and a significant increase in serum glucose from 4.2 ± 0.45 mmol/L to 7.12 ± 1.06 mmol/L (P = .0075) in the treatment group but no change in the control group. Tumor size was less in the treatment group (2,098 ± 388 µm(2)) compared with the control group (7,067 ± 955 µm(2); P = .0024). Furthermore, apoptosis was increased in the treatment group (6.9 ± 1.23%) compared with the control group (0.29 ± 0.103%; P = .002). CONCLUSION: SOM230 demonstrates antisecretory, antiproliferative, and proapoptotic activity in our MEN1 model of insulinoma. Further studies of the effects of SOM230 in PNET patients with MEN1 mutations are warranted.

Restoration of C/EBPα in dedifferentiated liposarcoma induces G2/M cell cycle arrest and apoptosis.

Well-differentiated liposarcoma (WDLS) and dedifferentiated liposarcoma (DDLS) represent the most common biological group of liposarcoma, and there is a pressing need to develop targeted therapies for patients with advanced disease. To identify potential therapeutic targets, we sought to identify differences in the adipogenic pathways between DDLS, WDLS, and normal adipose tissue. In a microarray analysis of DDLS (n = 84), WDLS (n = 79), and normal fat (n = 23), C/EBPα, a transcription factor involved in cell cycle regulation and differentiation, was underexpressed in DDLS when compared to both WDLS and normal fat (15.2- and 27.8-fold, respectively). In normal adipose-derived stem cells, C/EBPα expression was strongly induced when cells were cultured in differentiation media, but in three DDLS cell lines, this induction was nearly absent. We restored C/EBPα expression in one of the cell lines (DDLS8817) by transfection of an inducible C/EBPα expression vector. Inducing C/EBPα expression reduced proliferation and caused cells to accumulate in G2/M. Under differentiation conditions, the cell proliferation was reduced further, and 66% of the DDLS cells containing the inducible C/EBPα expression vector underwent apoptosis as demonstrated by annexin V staining. These cells in differentiation conditions expressed early adipocyte-specific mRNAs such as LPL and FABP4, but they failed to accumulate intracellular lipid droplets, a characteristic of mature adipocytes. These results demonstrate that loss of C/EBPα is an important factor in suppressing apoptosis and maintaining the dedifferentiated state in DDLS. Restoring C/EBPα may be a useful therapeutic approach for DDLS.

Laparoscopic radical cholecystectomy.

Controversy exists as to the role of minimally invasive techniques in the management of early gallbladder cancer. The majority of early gallbladder cancers are diagnosed upon final pathology after laparoscopic cholecystectomy. For stage pT1a tumors, no further surgery is warranted; however, for pT1b or greater lesions, patients usually undergo port-site excisions and completion of open radical cholecystectomy involving a partial hepatectomy of segments IV and V and a lymphadenectomy of the hepatoduodenal ligament. Presented in this paper is a totally laparoscopic radical cholecystectomy performed for suspected early gallbladder cancer. Despite the fact that a preoperative serum IgG4 level was within normal limits, final pathology was consistent with autoimmune cholecystitis. As a result, the laparoscopic radical cholecystectomy may be useful in select patients with a preoperative suspicion of early-stage gallbladder cancer by sparing them the necessity of a second-stage open procedure.

Flavopiridol targets c-KIT transcription and induces apoptosis in gastrointestinal stromal tumor cells.

Gastrointestinal stromal tumors (GIST) are characterized by activating mutations in the c-KIT gene which confers ligand-independent activation of the KIT receptor. Imatinib mesylate has been shown to effectively block constitutively active KIT and delay tumor growth. However, resistance to imatinib mesylate is emerging as a major clinical problem and novel therapies are needed. We report that treatment of GIST cells with the transcriptional inhibitor flavopiridol, initially down-regulates the antiapoptotic proteins bcl-2, mcl-1, and X-linked inhibitor of apoptosis protein which occurs as early as 4 hours after exposure. This is followed at 24 hours by the transcriptional suppression of KIT resulting in poly(ADP-ribose) polymerase cleavage and apoptosis. To separate the apoptotic effect of KIT suppression relative to the down-regulation of antiapoptotic proteins, we used small interfering RNA-directed knockdown of KIT. Results show that focused suppression of KIT alone is sufficient to induce apoptosis in GIST cells, but not to the same extent as flavopiridol. In contrast, imatinib mesylate, which inhibits KIT kinase activity but does not suppress total KIT expression, fails to cause apoptosis. We also show that flavopiridol suppresses KIT mRNA expression through positive transcriptional elongation factor inhibition and decreases KIT promoter activity. This causes a global decrease in the level of functionally mature KIT at the cell surface, resulting in a decrease in autophosphorylation at tyrosine residues 703 and 721, which characterizes activated KIT. Our results indicate that targeting KIT expression and these antiapoptotic proteins with flavopiridol represents a novel means to disrupt GIST cell dependence on KIT signaling and collectively renders these cells sensitive to apoptosis.

High-resolution MAS NMR spectroscopy detection of the spin magnetization exchange by cross-relaxation and chemical exchange in intact cell lines and human tissue specimens.

High-resolution magic-angle-spinning (HR-MAS) NMR spectroscopy detects resolved signals from membrane phospholipids and proteins in intact cell and tissue samples. MAS has the additional advantage of quenching spin-diffusion through a mutual "flip-flop" of neighbor spins by time-independent dipolar coupling as long as the dipolar coupling is "inhomogeneous." Under MAS, significant magnetization transfer (MT) was observed between water and each proton site in membrane phospholipid and between water and the NMR-observable protein proton signals. The MT rates between water and membrane phospholipids are lower than those between water and protein proton signals. The interaction of water to other small molecules is selective with the observation of MT from water to creatine, lactate, taurine, and glycine, but not to triglyceride, phosphocholine, choline, or myo-inositol. HR-MAS NMR allows the detection of a complete MT network between water and each proton group of creatine. Two creatine pools (one motion-restricted and one motion-free) were identified in skeletal muscle.

Role of fecal incontinence in contamination of the environment with vancomycin-resistant enterococci.

BACKGROUND: We tested the hypothesis that patients with vancomycin-resistant Enterococcus (VRE) stool colonization who are continent of feces contaminate the environment less frequently than patients who are colonized and incontinent. METHODS: We prospectively examined the frequency of environmental VRE contamination in the rooms of 15 patients who were continent and 15 who were incontinent and VRE-colonized. Broth-enrichment cultures of bed rails, bedside table, and call buttons were performed at baseline, and 2 and 5 days after environmental disinfection. The numbers of VRE colonies isolated after directly plating environmental swabs onto agar were compared for the continent and incontinent groups. RESULTS: The percentages of patients with 1 or more positive environmental cultures for VRE were not significantly different for the groups of patients who were continent and incontinent at baseline (60% vs 73%, P =.45) or 2 days after disinfection (60% vs 80%, P =.24). The numbers of VRE colonies isolated by direct plating were not significantly different for the continent and incontinent groups (P =.42). CONCLUSIONS: Environmental contamination occurs frequently in the rooms of patients who are continent, and those who are incontinent and VRE-colonized. Our findings suggest that similar infection control measures should be implemented for patients who are continent and incontinent.