Bernard Séguin

A new approach for prediction of tumor sensitivity to targeted drugs based on functional data

BackgroundThe success of targeted anti-cancer drugs are frequently hindered by the lack of knowledge of the individual pathway of the patient and the extreme data requirements on the estimation of the personalized genetic network of the patient’s tumor. The prediction of tumor sensitivity to targeted drugs remains a major challenge in the design of optimal therapeutic strategies. The current sensitivity prediction approaches are primarily based on genetic characterizations of the tumor sample. We propose a novel sensitivity prediction approach based on functional perturbation data that incorporates the drug protein interaction information and sensitivities to a training set of drugs with known targets.ResultsWe illustrate the high prediction accuracy of our framework on synthetic data generated from the Kyoto Encyclopedia of Genes and Genomes (KEGG) and an experimental dataset of four canine osteosarcoma tumor cultures following application of 60 targeted small-molecule drugs. We achieve a low leave one out cross validation error of <10% for the canine osteosarcoma tumor cultures using a drug screen consisting of 60 targeted drugs.ConclusionsThe proposed framework provides a unique input-output based methodology to model a cancer pathway and predict the effectiveness of targeted anti-cancer drugs. This framework can be developed as a viable approach for personalized cancer therapy.

Biologic Behavior and Clinical Outcome of 25 Dogs with Canine Appendicular Chondrosarcoma Treated by Amputation: A Veterinary Society of Surgical Oncology Retrospective Study

OBJECTIVEnTo characterize biologic behavior, clinical outcome, and effect of histologic grade on prognosis for dogs with appendicular chondrosarcoma treated by amputation alone.nnnSTUDY DESIGNnCase series.nnnANIMALSnDogs (n=25) with appendicular chondrosarcoma.nnnMETHODSnMedical records were searched to identify dogs with appendicular chondrosarcoma treated by limb amputation alone. Information recorded included signalment, anatomic location, radiographic appearance, and development of metastasis. Histopathologic diagnosis was confirmed and graded (1, 2, or 3). Survival curves were generated by the Kaplan-Meier method and the association between covariates (gender, age, weight, and tumor grade) and survival were evaluated using the univariate proportional hazards model.nnnRESULTSnHistopathology slides were available for 25 dogs. Rates of pulmonary metastasis were as follows: grade 1-0%, grade 2-31%, and grade 3-50%. Overall median survival time (MST) was 979 days. Age, weight, and sex were not significantly associated with survival (P=.16; .33; and .31, respectively). Survival was significantly associated with tumor grade (P=.008), with dogs with tumor grade of 1, 2, and 3 having MSTs of 6, 2.7, and 0.9 years, respectively.nnnCONCLUSIONnCanine appendicular chondrosarcoma can be treated effectively with amputation alone. Low to intermediate grade chondrosarcoma has a good prognosis, whereas high-grade tumors appear to behave aggressively.nnnCLINICAL RELEVANCEnThe overall prognosis for appendicular chondrosarcoma is better than that of appendicular osteosarcoma treated by amputation alone or in combination with chemotherapy.

A case study of personalized therapy for osteosarcoma.

Effective targeted therapies are needed in sarcomas, but the biological heterogeneity of these tumors has presented a challenge to clinical integration of small molecule inhibitors in sarcoma treatment. Here we outline a process to personalize therapy for sarcomas through a case study of a canine with spontaneous osteosarcoma.

Frequent alteration of the tumor suppressor gene APC in sporadic canine colorectal tumors.

Sporadic canine colorectal cancers (CRCs) should make excellent models for studying the corresponding human cancers. To molecularly characterize canine CRC, we investigated exonic sequence mutations of adenomatous polyposis coli (APC), the best known tumor suppressor gene of human CRC, in 23 sporadic canine colorectal tumors, including 8 adenomas and 15 adenocarcinomas, via exon-resequencing analysis. As a comparison, we also performed the same sequencing analysis on 10 other genes, either located at human 5q22 (the same locus as APC) or 18q21 (also frequently altered in human CRC), or known to play a role in human carcinogenesis. We noted that APC was the most significantly mutated gene in both canine adenomas and adenocarcinomas among the 11 genes examined. Significantly, we detected large deletions of ≥10 bases, many clustered near the mutation cluster region, as well as single or two base deletions in ∼70% canine tumors of both subtypes. These observations indicate that like in the human, APC is also frequently altered in sporadic colorectal tumors in the dog and its alteration is an early event in canine colorectal tumorigenesis. Our study provides further evidence demonstrating the molecular similarity in pathogenesis between sporadic human and canine CRCs. This work, along with our previous copy number abnormality study, supports that sporadic canine CRCs are valid models of human CRCs at the molecular level.

Hemipelvectomy in Dogs and Cats: Technique Overview, Variations, and Description

OBJECTIVEnTo review and describe the surgical procedure of hemipelvectomy in dogs and cats.nnnSTUDY DESIGNnDescriptive report.nnnANIMALSnDogs and cats.nnnMETHODSnAnatomic surgical technique description and PubMed literature search.nnnRESULTSnHemipelvectomy is an involved, aggressive surgical procedure that must be tailored to each specific clinical case. Thorough preoperative assessment and planning, as well as considerations for closure options are critical for a successful outcome. Although outcome data are scant, the prognosis after hemipelvectomy correlates to the underlying disease process. Proactive postoperative analgesia and slower return to function are important factors in patient management.nnnCONCLUSIONSnWith careful pre- and postoperative planning, hemipelvectomy can be used successfully in dogs and cats.nnnCLINICAL RELEVANCEnHemipelvectomy in the dog and cat is a viable treatment option for selected neoplastic and traumatic conditions.

The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine osteosarcoma in vitro.

BackgroundOsteosarcoma (OS) affects over 8000 dogs/year in the United States. The disease usually arises in the appendicular skeleton and metastasizes to the lung. Dogs with localized appendicular disease benefit from limb amputation and chemotherapy but most die within 6–12 months despite these treatments. Taurolidine, a derivative of taurine, has anti-tumor and anti-angiogenic effects against a variety of cancers. The following in vitro studies tested taurolidine as a candidate for adjuvant therapy for canine OS. Tests for p53 protein status and caspase activity were used to elucidate mechanisms of taurolidine-induced cell death.ResultsTaurolidine was cytotoxic to osteosarcoma cells and increased the toxicity of doxorubicin and carboplatin in vitro. Apoptosis was greatly induced in cells exposed to 125 μM taurolidine and less so in cells exposed to 250 μM taurolidine. Taurolidine cytotoxicity appeared caspase-dependent in one cell line; with apparent mutant p53 protein. This cell line was the most sensitive to single agent taurolidine treatment and had a taurolidine-dependent reduction in accumulated p53 protein suggesting taurolidine’s effects may depend on the functional status of p53 in canine OS.ConclusionTaurolidine’s cytotoxic effect appears dependent on cell specific factors which may be explained, in part, by the functional status of p53. Taurolidine initiates apoptosis in canine OS cells and this occurs to a greater extent at lower concentrations. Mechanisms of cell death induced by higher concentrations were not elucidated here. Taurolidine combined with doxorubicin or carboplatin can increase the toxicity of these chemotherapy drugs and warrants further investigation in dogs with osteosarcoma.

Comparative analysis of the surface exposed proteome of two canine osteosarcoma cell lines and normal canine osteoblasts

BackgroundOsteosarcoma (OSA) is the most common primary bone tumor of dogs and carries a poor prognosis despite aggressive treatment. An improved understanding of the biology of OSA is critically needed to allow for development of novel diagnostic, prognostic, and therapeutic tools. The surface-exposed proteome (SEP) of a cancerous cell includes a multifarious array of proteins critical to cellular processes such as proliferation, migration, adhesion, and inter-cellular communication. The specific aim of this study was to define a SEP profile of two validated canine OSA cell lines and a normal canine osteoblast cell line utilizing a biotinylation/streptavidin system to selectively label, purify, and identify surface-exposed proteins by mass spectrometry (MS) analysis. Additionally, we sought to validate a subset of our MS-based observations via quantitative real-time PCR, Western blot and semi-quantitative immunocytochemistry. Our hypothesis was that MS would detect differences in the SEP composition between the OSA and the normal osteoblast cells.ResultsShotgun MS identified 133 putative surface proteins when output from all samples were combined, with good consistency between biological replicates. Eleven of the MS-detected proteins underwent analysis of gene expression by PCR, all of which were actively transcribed, but varied in expression level. Western blot of whole cell lysates from all three cell lines was effective for Thrombospondin-1, CYR61 and CD44, and indicated that all three proteins were present in each cell line. Semi-quantitative immunofluorescence indicated that CD44 was expressed at much higher levels on the surface of the OSA than the normal osteoblast cell lines.ConclusionsThe results of the present study identified numerous differences, and similarities, in the SEP of canine OSA cell lines and normal canine osteoblasts. The PCR, Western blot, and immunocytochemistry results, for the subset of proteins evaluated, were generally supportive of the mass spectrometry data. These methods may be applied to other cell lines, or other biological materials, to highlight unique and previously unrecognized differences between samples. While this study yielded data that may prove useful for OSA researchers and clinicians, further refinements of the described techniques are expected to yield greater accuracy and produce a more thorough SEP analysis.

Osteoprotegerin activates osteosarcoma cells that co-express RANK and RANKL

BACKGROUNDnOsteosarcoma (OS) is an aggressive and often fatal cancer that afflicts over 1000 humans and 10,000 dogs per year in the United States. Recent evidence suggests deregulation in the signaling triad, receptor activator of nuclear factor kappa B (RANK), its activating ligand (RANKL), and the RANKL inhibitor, osteoprotegerin (OPG) plays a key role in the pathogenesis of OS. This study investigated the expression of RANK and RANKL in osteosarcoma tumors and cell lines and describes an activating effect of OPG on OS cells in vitro.nnnRESULTSnCanine OS tumors and cell lines co-express mRNA for both RANK and RANKL. Expression of these proteins in OS cell lines was confirmed by Western blot and immunofluorescence microscopy. Expression of the soluble form of RANKL was not detected in media from OS cells. OPG-Fc incubation increased the phosphorylation status of ERK, AKT and the p65 subunit of nuclear factor kappa B (NFκB) and induced NFκB translocation from the cytoplasm to the nucleus in canine OS cells. OPG increased proliferation in both canine and human derived OS cell lines.nnnCONCLUSIONnRANKL is produced by OS tumors and cell lines that also express RANK. This data provides preliminary evidence for a potential autocrine and or paracrine activation pathway in canine OS. An activating effect of exogenous OPG on signal transduction proteins, NFκB and proliferation in OS is described. These data provide new information concerning aberrant signaling in OS and could be important to those considering OPG as a therapeutic agent for osteosarcoma.

Long-term outcome of dogs treated with ulnar rollover transposition for limb-sparing of distal radial osteosarcoma: 27 limbs in 26 dogs

OBJECTIVEnTo determine outcomes in dogs with distal radial osteosarcoma treated with ulnar rollover transposition (URT) limb-sparing surgery including: viability of the ulnar graft, complications, subjective limb function, disease-free interval (DFI), and survival time (ST).nnnSTUDY DESIGNnRetrospective case series.nnnANIMALSnTwenty-six client-owned dogs with distal radial osteosarcoma and no involvement of the ulna.nnnMETHODSnData of dogs treated with URT were collected at the time of surgery and retrospectively from medical records and by contacting owners and referring veterinarians.nnnRESULTSnURT technique was performed on 27 limbs in 26 dogs. The ulnar graft was determined to be viable in 17 limbs, nonviable in 3, and unknown in 7. Complications occurred in 20 limbs. Infection was diagnosed in 12 limbs. Biomechanical complications occurred in 15 and local recurrence in 2 limbs. Limb function graded by veterinarians or owners was poor in 2 limbs, fair in 4, good in 14, excellent in 3, and unknown in 4. Median DFI was 245 days and median ST was 277 days.nnnCONCLUSIONnThe URT technique maintained the viability of the ulnar graft. The complication rate was high but limb function appeared acceptable. Although sufficient length of the distal aspect of the ulna must be preserved to perform this technique, local recurrence was not increased compared to other limb-sparing techniques when cases were appropriately selected.

Partial resection of the hyoid apparatus during surgical treatment of ectopic thyroid carcinomas in dogs: 5 cases (2011–2013)

OBJECTIVEnTo assess perioperative findings and postoperative complications and outcomes in dogs that had ectopic thyroid carcinomas with invasion into the hyoid apparatus and underwent tumor excision with partial hyoidectomy.nnnDESIGNnRetrospective case series.nnnANIMALSn5 client-owned dogs.nnnPROCEDURESnMedical records of dogs that had an ectopic neuroendocrine tumor with invasion into the hyoid apparatus and underwent tumor excision with partial hyoidectomy were reviewed for information regarding perioperative and postoperative findings and outcome. During surgery in each case, the thyrohyoid and ceratohyoid or epihyoid bones (depending on degree of hyoid apparatus involvement) were sharply transected, allowing en bloc removal of the tumor. The ipsilateral cut ends of the thyrohyoid and ceratohyoid or epihyoid bones (depending on which was cut) were sutured together with polypropylene suture in a simple interrupted pattern.nnnRESULTSnAll partial hyoidectomy procedures were completed without surgical or anesthetic complications. All 5 dogs were able to eat and drink between 7 and 24 hours after surgery, with no signs of dysphagia, ptyalism, or abnormal tongue carriage. Follow-up information was obtained over a period of 173 to 587 days after surgery for all 5 dogs; 4 dogs were still alive at last follow-up. One dog was euthanized 587 days after surgery because of lethargy, inappetence, and hypercalcemia.nnnCONCLUSIONS AND CLINICAL RELEVANCEnFrom this limited series of cases, results suggested that partial resection of the hyoid apparatus during removal of ectopic thyroid carcinoma may be tolerated well and be associated with very good functional outcomes in dogs.