William C. Eward

A mouse-human phase 1 co-clinical trial of a protease-activated fluorescent probe for imaging cancer

A first-in-human phase 1 clinical trial of the PEGylated protease-activated fluorescent probe, LUM015, enables tumor imaging at a safe and tolerable dose in humans. Protease probe tested in humans Cancer cells secrete more of the protease cathepsin than healthy cells, partly as a way to enzymatically remodel their surroundings for tumor growth and metastasis. Whitley et al. developed an imaging probe that could be activated in the presence of these cathepsins, thus allowing surgeons to distinguish tumor margins intraoperatively. Their probe, called LUM015, was able to signal the presence of cancer in vivo in a mouse sarcoma model, and in a so-called “co-clinical trial” in 15 patients, it was safe and cleaved as expected in different types of tumor tissues. With favorable biodistribution and pharmacokinetics also demonstrated, protease-activated probes are now poised for further adaptation to tumor resections, signaling the presence of residual cancer. Local recurrence is a common cause of treatment failure for patients with solid tumors. Intraoperative detection of microscopic residual cancer in the tumor bed could be used to decrease the risk of a positive surgical margin, reduce rates of reexcision, and tailor adjuvant therapy. We used a protease-activated fluorescent imaging probe, LUM015, to detect cancer in vivo in a mouse model of soft tissue sarcoma (STS) and ex vivo in a first-in-human phase 1 clinical trial. In mice, intravenous injection of LUM015 labeled tumor cells, and residual fluorescence within the tumor bed predicted local recurrence. In 15 patients with STS or breast cancer, intravenous injection of LUM015 before surgery was well tolerated. Imaging of resected human tissues showed that fluorescence from tumor was significantly higher than fluorescence from normal tissues. LUM015 biodistribution, pharmacokinetic profiles, and metabolism were similar in mouse and human subjects. Tissue concentrations of LUM015 and its metabolites, including fluorescently labeled lysine, demonstrated that LUM015 is selectively distributed to tumors where it is activated by proteases. Experiments in mice with a constitutively active PEGylated fluorescent imaging probe support a model where tumor-selective probe distribution is a determinant of increased fluorescence in cancer. These co-clinical studies suggest that the tumor specificity of protease-activated imaging probes, such as LUM015, is dependent on both biodistribution and enzyme activity. Our first-in-human data support future clinical trials of LUM015 and other protease-sensitive probes.

Cross Species Genomic Analysis Identifies a Mouse Model as Undifferentiated Pleomorphic Sarcoma/Malignant Fibrous Histiocytoma

Undifferentiated pleomorphic sarcoma/Malignant Fibrous Histiocytoma (MFH) is one of the most common subtypes of human soft tissue sarcoma. Using cross species genomic analysis, we define a geneset from the LSL-KrasG12D; Trp53Flox/Flox mouse model of soft tissue sarcoma that is highly enriched in human MFH. With this mouse geneset as a filter, we identify expression of the RAS target FOXM1 in human MFH. Expression of Foxm1 is elevated in mouse sarcomas that metastasize to the lung and tissue microarray analysis of human MFH correlates overexpression of FOXM1 with metastasis. These results suggest that genomic alterations present in human MFH are conserved in the LSL-KrasG12D; p53Flox/Flox mouse model of soft tissue sarcoma and demonstrate the utility of this pre-clinical model.

The effect of the setting of a positive surgical margin in soft tissue sarcoma

The objectives of this study were to evaluate the risk of local recurrence and survival after soft tissue sarcoma (STS) resection with positive margins and to evaluate the safety of sparing adjacent critical structures.

Pseudotumor with superimposed periprosthetic infection following metal-on-metal total hip arthroplasty: A case report

Second-generation metal-on-metal bearing surfaces made of modern cobalt-chromium-molybdenum alloys are widely used for total hip arthroplasty in the United States1. While these bearing surfaces offer considerable advantages over conventional metal-on-polyethylene articulations, metal hypersensitivity reactions to these implants are an important, although uncommon, cause of failure2. This unique mode of failure, which has been reported with first-generation implants2,3 and subsequently in association with second-generation metal-on-metal bearing surfaces, appears to be caused by an immunologic delayed hypersensitivity response to metal particles4-6. This local tissue reaction, described as an aseptic lymphocyte-dominated vasculitis-associated lesion (ALVAL), was characterized by Willert et al.7 as having histologic features that include diffuse perivascular infiltrates of T and B lymphocytes and plasma cells, high endothelial venules, massive fibrin exudation, accumulation of macrophages with droplike inclusions, and infiltrates of eosinophilic granulocytes and necrosis. Histologically, high endothelial venules are characterized as vascular and/or lymphatic vessels with reactive endothelium, often seen in association with an inflammatory response. Willert et al. suggested that the possibility of such a reaction be considered when, following primary implantation of metal-on-metal implants, a patient reports the recurrence of preoperative pain and there is a marked joint effusion or the development of osteolysis in the absence of a suspicion of deep periprosthetic infection. A subsequent report, however, described this hypersensitivity reaction mimicking infection in a patient with elevated levels of inflammatory biomarkers who was later found to have no deep infection on revision8. We describe a patient with a failure of a metal-on-metal total hip replacement consistent with a metal hypersensitivity reaction who presented with a pseudotumor and massive lower-extremity edema secondary to extrinsic femoral vein compression and subsequently developed a superimposed deep periprosthetic infection prior to revision. To our knowledge, this has not …

Intraoperative detection and removal of microscopic residual sarcoma using wide-field imaging

The goal of limb‐sparing surgery for a soft tissue sarcoma of the extremity is to remove all malignant cells while preserving limb function. After initial surgery, microscopic residual disease in the tumor bed will cause a local recurrence in approximately 33% of patients with sarcoma. To help identify these patients, the authors developed an in vivo imaging system to investigate the suitability of molecular imaging for intraoperative visualization.

Immunohistochemistry of the canine vomeronasal organ

The canines olfactory acuity is legendary, but neither its main olfactory system nor its vomeronasal system has been described in much detail. We used immunohistochemistry on paraffin‐embedded sections of male and female adult dog vomeronasal organ (VNO) to characterize the expression of proteins known to be expressed in the VNO of several other mammals. Basal cell bodies were more apparent in each section than in rodent VNO and expressed immunoreactivity to anticytokeratin and antiepidermal growth factor receptor antibodies. The thin layer of neurone cell bodies in the sensory epithelium and axon fascicles in the lamina propria expressed immunoreactivity to neurone cell adhesion molecule, neurone‐specific beta tubulin and protein gene product 9.5. Some neurones expressed growth‐associated protein 43 (GAP43): and a number of those also expressed neurone‐specific beta tubulin‐immunoreactivity. Some axon fascicles were double labelled for those two proteins. The G‐protein alpha subunits Gi and Go, involved in the signal transduction pathway, showed immunoreactivity in the sensory cell layer. Our results demonstrate that the canine vomeronasal organ contains a population of cells that expresses several neuronal markers. Furthermore, GAP43 immunoreactivity suggests that the sensory epithelium is neurogenic in adult dogs.

NF1 deletion generates multiple subtypes of soft-tissue sarcoma that respond to MEK inhibition

Soft-tissue sarcomas are a heterogeneous group of tumors arising from connective tissue. Recently, mutations in the neurofibromin 1 (NF1) tumor suppressor gene were identified in multiple subtypes of human soft-tissue sarcomas. To study the effect of NF1 inactivation in the initiation and progression of distinct sarcoma subtypes, we have developed a novel mouse model of temporally and spatially restricted NF1-deleted sarcoma. To generate primary sarcomas, we inject adenovirus containing Cre recombinase into NF1flox/flox; Ink4a/Arfflox/flox mice at two distinct orthotopic sites: intramuscularly or in the sciatic nerve. The mice develop either high-grade myogenic sarcomas or malignant peripheral nerve sheath tumor (MPNST)-like tumors, respectively. These tumors reflect the histologic properties and spectrum of sarcomas found in patients. To explore the use of this model for preclinical studies, we conducted a study of mitogen-activated protein kinase (MAPK) pathway inhibition with the MEK inhibitor PD325901. Treatment with PD325901 delays tumor growth through decreased cyclin D1 mRNA and cell proliferation. We also examined the effects of MEK inhibition on the native tumor stroma and find that PD325901 decreases VEGFα expression in tumor cells with a corresponding decrease in microvessel density. Taken together, our results use a primary tumor model to show that sarcomas can be generated by loss of NF1 and Ink4a/Arf, and that these tumors are sensitive to MEK inhibition by direct effects on tumor cells and the surrounding microenvironment. These studies suggest that MEK inhibitors should be further explored as potential sarcoma therapies in patients with tumors containing NF1 deletion. Mol Cancer Ther; 12(9); 1906–17. ©2013 AACR.

Attitudes toward physician-assisted suicide among physicians in Vermont

Background: Legislation on physician-assisted suicide (PAS) is being considered in a number of states since the passage of the Oregon Death With Dignity Act in 1994. Opinion assessment surveys have historically assessed particular subsets of physicians. Objective: To determine variables predictive of physicians’ opinions on PAS in a rural state, Vermont, USA. Design: Cross-sectional mailing survey. Participants: 1052 (48% response rate) physicians licensed by the state of Vermont. Results: Of the respondents, 38.2% believed PAS should be legalised, 16.0% believed it should be prohibited and 26.0% believed it should not be legislated. 15.7% were undecided. Males were more likely than females to favour legalisation (42% vs 34%). Physicians who did not care for patients through the end of life were significantly more likely to favour legalisation of PAS than physicians who do care for patients with terminal illness (48% vs 33%). 30% of the respondents had experienced a request for assistance with suicide. Conclusions: Vermont physicians’ opinions on the legalisation of PAS is sharply polarised. Patient autonomy was a factor strongly associated with opinions in favour of legalisation, whereas the sanctity of the doctor–patient relationship was strongly associated with opinions in favour of not legislating PAS. Those in favour of making PAS illegal overwhelmingly cited moral and ethical beliefs as factors in their opinion. Although opinions on legalisation appear to be based on firmly held beliefs, approximately half of Vermont physicians who responded to the survey agree that there is a need for more education in palliative care and pain management.

Impact of liposomal bupivacaine administration on postoperative pain in patients undergoing total knee replacement.

To determine whether liposomal bupivacaine, a long‐acting anesthetic indicated for single‐dose wound infiltration to produce postoperative analgesia, has an impact on postoperative pain in patients undergoing total knee arthroplasties (TKAs).

Mesenchymal-Epithelial Transition in Sarcomas Is Controlled by the Combinatorial Expression of MicroRNA 200s and GRHL2.

ABSTRACT Phenotypic plasticity involves a process in which cells transiently acquire phenotypic traits of another lineage. Two commonly studied types of phenotypic plasticity are epithelial-mesenchymal transition (EMT) and mesenchymal-epithelial transition (MET). In carcinomas, EMT drives invasion and metastatic dissemination, while MET is proposed to play a role in metastatic colonization. Phenotypic plasticity in sarcomas is not well studied; however, there is evidence that a subset of sarcomas undergo an MET-like phenomenon. While the exact mechanisms by which these transitions occur remain largely unknown, it is likely that some of the same master regulators that drive EMT and MET in carcinomas also act in sarcomas. In this study, we combined mathematical models with bench experiments to identify a core regulatory circuit that controls MET in sarcomas. This circuit comprises the microRNA 200 (miR-200) family, ZEB1, and GRHL2. Interestingly, combined expression of miR-200s and GRHL2 further upregulates epithelial genes to induce MET. This effect is phenocopied by downregulation of either ZEB1 or the ZEB1 cofactor, BRG1. In addition, an MET gene expression signature is prognostic for improved overall survival in sarcoma patients. Together, our results suggest that a miR-200, ZEB1, GRHL2 gene regulatory network may drive sarcoma cells to a more epithelial-like state and that this likely has prognostic relevance.