Osteosarcoma Flashcards by Bushra Zaidi

Bracha et al. The expression and role of seratonin receptor 5HTR2A in canine osteoblats and an OSA cell line

5HTR2A was _____ in malignant cell line compared to normal cells

In CnOb cells, ERK-P was _____ in response to both seratonin and 5HTR2A antagonist, ritanserin

In COS cells, ERK-P ____ with both treatments

CREB was ____ in CnOb cells and ____ in COS cells

Seratonin treatment promoted ____ in malignant cells but not normal osteeoblasts

Ritanserin ____ cell viability in normal and OSA cells

Ritanserin induced ____

Oveexpressed

Decreased

Increased

undetectable, constitutively

cell viability

Reduced

Apoptosis in COS

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Dailey et al. HES1, a target of Notch signaling, is elevated in canine osteosarcoma, but reduced in the most aggressive tumors

Hairy and enhancer of split 1 (HES1), a transcriptional repressor, is downstream target of Notch signaling

Notch signaling and HES1 have been linked to growth and survival

HES1 mRNA was ___ in tumor samples relative to normal bone, but ___ in tumor samples from dogs with DFI<100 days to those DFI>300d

NOTCH2 and HEY1 mRNA was ___ in tumors relative to normal bone but not differntially expressed between DFI tumor groups

Survival analysis?

Notch signaling occurs and may contribute to canine OSA. Mechanisms that do not alter HES1 expression may drive aggressive tumor

Increased

Increased, decreased

Increased

Decreased HES1 and shorter DFI

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Sternberg et al. Association between absolute tumor burden and serum bone-specific alkaline phosphatase in canine appendicular OSA

96 dogs with appendicular OSA

Quantity of membranous BALP was propotional with?

In dogs devoid of macroscopic metastases there was a positive correlation between serum BALP and ___

Dog with progressive metastases, serum BALP activity ___ and coincided with development of _____

Tumor burden is determinant of serum BALP activity

Cell densitiy, not tumorigenic or metastatic phenotype

Primary tumor size

Increased, macroscopic metastases

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Skorupski et al. Carboplatin versus alternating carboplatin and doxorubicin for the adjuvant treatment of canine appendiculay OSA: a randomized, phase III trial

6 doses of carboplatin or 3 doses of carboplatin and doxorubicin in alternating schedule

DFI for carboplatin vs. carboplatin and doxorubicin?

425 (14 mo) versus 135 days (4.5 mo)

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Sivacolundhu et al. 2013. Ulnar OSA in dogs: 30 cases (1992-2008)

11 dogs treated with partial ulnar ostectomy and 14 with amputation, 5 dogs no resection

Median DFI and ST

Negative prognostic factors on univariable analysis?

Negative prognostic factor on multivariate analysis? MST?

437 (14.6 mo) vs. 463 days (15.4 mo)

Histologic subtype and lung metastasis

Telangiectatic or telengiectatic-mixed subtype, 208 days

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Amsellem et al. 2014. Appendicular OSA in small breed dogs: 51 cases (1986-2011)

Dogs <15kg

9 treated nonsurgically, 16 amputation, 26 curative intent treatment, MST?

MST did not differ significantly between amputation and curative intent group - chemo efficacy?

For dogs in nonsurgical group, MST ____ significantly as tumor ___ increased

For dogs in amputation group, MST ___ as body weight increased

Tumor histologic score and mitotic index were lower and MST following amputation

112 days (3.7 mo), 257 days (8.6), 415 days (13.8 mo)

Decreased, tumor histologic score

Decreased

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Culp et al. 2014. Evaluation of outcome and prognostic factors for dogs living greater than one year after diagnosis of osteosarcoma: 90 cases

Median age

Median weight

Serum ALP activity was high in?

Most common tumor location?

Metastatic disease?

MST beyond 1 year?

Which dogs had significantly improved survival?

8.2 years

38 kg

48%

distal portion of radius 60%

54%

243 days

Surgical site infection after limb sparing surgery

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Batchinski et al. Evaluation of ifosfamide salvage therapy for metastatic canine OSA

19 dogs and 17 evaluated for response

Ifosfamide doses 375-425 mg/m2 with median of 2 doses

Overall response to ifosfamide?

Toxicity?

Median survival duration from first dose to death?

11.8%

2 dogs were hospitalized

95 days

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Alvarez et al. 2014. Postoperative adjuvant combination therapy with doxorubicin and noncytotoxic suramin in dogs with appendicular OSA

Suramin increased tumor sesitivity to chemotherapy

47 dogs received 6.75 mg/kg suramin IV followed by 30 mg/m2 doxorubicin 4 hrs later - repeated q2wk x 5 doses

Median DFI

MST

203 days (6.8 mo)

359 days (1 yr)

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Coyle et al. Biologic behaivour of canine mandibular OSA. A retrospective study of 50 cases (1999-2007)

How many dogs developed metastatic disease?

Median MFI

MST

In univariate analysis what was prognostic?

Grade also influence ST, ___ dogs with grade II/III tumors met free at 1 year vs. ___ for grade I

___ alive with grade II/III vs. ____ alive with grade I

Multivariable analysis, histologic grade and adjuvant chemotherapy were prognostic for MFI and MST

58%

627 days (20.9 mo)

525 days (17.5)

MI >40 decreased MFI and ST

24% vs. 72%

24% vs, 77%

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Bracha et al. Evaluation of toxicities from combined metronomic and MTD chemotherapy in dogs with OSA

Tolerability of piroxicam and cyclophosphamide combined with carboplatin or caboplatin and doxorubicin MTD

All dogs underwent amputation and chemotherapy

14 dogs treated with carbo and 16 treated with carbo and doxo

Toxicities?

DFI

MST

Grade 3 and 4 in carbo group

Not significantly diffrernt 192 d in cabro and metronomic vs. 182 day

217 (7 mo) vs. 189 days (6.3 mo)

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Selmic et al. 2014. Comparison of carboplatin and doxorubicin-based chemotherapy protocols in 470 dogs after amputation for treatment of appendicular OSA

carbo for 4 or 6 cycles (CARBO6), dox q14d or 21d for 5 cycles, alternating carbo and dox q21d for 3 cycles

Median DFI

MST

A lower proportion of dogs prescribed ___ experienced AEs compared to other protocols (___%)

____ was not associated with development of metastasis or death

291 days (10 mo)

284 days (9.5 mo)

CARBO6, 48%

DI - dose intensity

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Duffy et al. Outcome following treatment of soft tissue and visceral extraskeletal OSA in 33 dogs: 2008-2013

Outcome after surgical treatment of non-mammary and non-thyoridal sofy tissue and visceral EOS in dogs

Most common primary tumor site?

Which dogs had longer survival times? MST?

Spleen

Wide or radical tumor excision had longer survival than marginal tumor excision - 90 days vs. 13 days

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Viall et al. Antagonism of seratonin receptor 1B decreased viability and promotes apoptosis in the COS canine OSA cell line

Seratonin receptor 1B (5HTR1B) exhibits anti-proliferative activity in osteoblasts

Equal levels of 5HTR1B gene and protein expression in normal and malignant osteoblasts

Treatment with seratonin enhanced viability OSA cells but not normal osteoblasts

Anpirtoline, 5HTR1B antagonist, caused no change in viability

Specific receptor antagonist SB224289 caused reduction in osteoblast viability, effect more in OSA cells

Aberrant signaling in COS - evaluating ERK and CREB

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Holmes et al. Canine OSA cell lines from patients with differing serum ALP concentration display no behaivoural differences in vitro

Difference in cell proliferation, migration, and cell invasion between cell lines with normal or increased ALP activity

No differnce was noted

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McCleese et al. Met interacts with EGFR and Ron in canine OSA

Met, RTK, is overexpressed in canine OSA

In humans, RTK Met, EGFR, and Ron are coexpressed and engage in heterodimerization

Expression of EGFR and Ron in canine OSA cell lines?

TGF-a and HGF induced amplification of ERK1/2 and STAT3 phosphorylation in OSA cells and Met was phosphorylated following TGF-a stimulation - receptor cross talk

Treatment of OSA cells with gefitinib and crizotinib inhibited cell proliferation

Met, EGFR, Ron interact in OSA cells

Expressed in OSA cell lines and primary tissue, EGFR and Ron are phosphorylated in OSA tumor samples, and Met is co-associated with EGFR and Ron in cell lines

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Maniscalco et al. PDGF and PDGFRs in canine OSA: new targets for innovative therapeutic startegies in comparitive oncology

PDGF and PDGFRs overexpressed in 70-80% human OSA

33 canine samples

Expression of PDGF-A and PDGF-B in OSA?

Expression of PDGFR-A and PDGFR-B in OSA?

qPCR showed all canine OSA cells overexpressed PDGFR-a while 6/7 overexpressed PDGFR-b and PDGF-A compared to normal osteoblastic cell line

PDGFR inhibitor casue dose- and time-dependent decrease in AKT phosphorylation - PDGFRs/PDGFs are co-expressed suggest autocrine/paracrine loop

40% and 60%

78% and 81%

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Bergeron et al. The effect of Zhangfei on the unfolded protein response and growth of cells derived from canine and human OSAs

Protein zhangfei could suppress unfolded protein response (UPR) and growth of OSA cells

Zhangfei expressing D-17 cells displayed large vacuoles and expressed phosphatidylserine on external surface - induced micropinocytosis and apoptosis

Zhagfei inhibited thapsigargin-induced UPC in D17 cells

Ability of Zhangfei to suppress UPR and tumor cell growth may not be linked

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Loftus et al. The 5-lipooxygenase inhibitor tepoxalin induced oxidative damage and altered PTEN status prior to apoptosis in canine OSA cell lines

Tepoxalin treated cells undergo apoptosis through ____ activation

Apoptosis is superceded by _____

5-LOX inhibition also increased ____ activity by preventing its alkylation or oxidation

PTEN modification allows PIP3 heigtening AKT phosphorylation

Target oxidation and LOX inhibition play roles in apoptotic response

Caspace-3

ROS

PTEN

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Piskun et al. B-catenin transcriptional activity is minimal in canine OSA and its targeted inhibition results in minimal changes to cell line behaivour

Impact of inhibiting canonical Wnt signaling in canine OSA - B-catenine siRNA or dominant negative T-cell factor construct

There were no changes to cell lines with either construct

B-catenin activity was ___ in normal canine osteoblasts compared to canine OSA cell lines

Wnt signaling has minimal activity in canine OSA and targeted inhibition is not relevant

3-fold higher

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Portela et al. Pro-tumorigenic effects of transforming growth factor beta 1 in canine OSA

TGFB1 contributes to skeletal remodeling inducing osteoblast proliferation, migration, and angiogeneis

33 dogs with appendicular OSA

Canine OSA cells secrete ___, express ___, and TGFB1 signaling blockade _____

Naturally occuring OS samples abundantly express ______

OS-bearing dogs, circulating TGFB1 concentration correlated with ___

TGFB1 inhibitors may impede OS progression

TGF1B, cognate receptors, TGF1B, decreased proliferation, migration, and VEGF secretion

TGFBR1/II

N-telopeptide excretion

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Shoeneman et al. Survivin inhibition via EZN-3042 in canine lymphoma and OSA

Survivin inhibits apoptosis and drives cell proliferation - elevated in human and canine cancer

Survivin inhibition with EZN-3042 inhibits growth, induced apoptosis, enhances chemosensitivity in vitro

Survivin directed therapies may be effective in treatment of canine LSA and OS

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Szigetvari et al. Wnt5a expression in canine OSA

Wnt5a protein, ligant for non-canonical Wnt, implicated in aggressiveness of cancer cell lines, including those from human OSA origin

Wnt5a expression in canine OSA?

Association with PFS?

54%

Wnt5a was not associayed with PFS

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Neumann et al. 2015. The association of endothelin-1 signaling with bone alkaline phosphatase expression and protumorigenic activities in canine OSA

Endothelin-1 is active during bone repair, and responsible for osteoblast migration, survival, proliferation, and bone alkaline phosphatase expression

45 dogs with appendicular OSA

Expression of endothelin-1 and endothelin-A receptor?

What does the signaling mediate?

Circulating bone ALP activities were positively correlated with primary tumor bone mineral densities

Expressed in canine OS cells

Survival, proliferation, and bone alkaline phosphatase activities

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**Rodrigues et al. OSA tissues and cell lines from patients with differing serum ALP concentration display minimal difference in gene expression patterns** Microarray analysis in 18 OSA samples and 6 OSA cell lines from dogs with normal and increased ALP No difference in gene expression patterns were noted. **ALP concentration is not associated with intrinsic differences of OSA cells**

**Oblak et al. The impact of pamidronate and chemotherapy on survival times with appendiculat primary bone tumors treated with palliative RT** RT alone, RT + chemotherapy, RT + pamidronate, and RT + chemotherapy + pamidronate MST was longest for which dogs? MST shortest for which dogs? Difference in MST for dogs who receieved ___ and those that did not was statistically signifcant in univariate analysis The addition of ___ into any protocol improved survival?

RT and chemotherapy - 307 days (10 mo) RT and pamidronate - 69 days Pamidronate Chemotherapy

**Helmerick et al. The effects of baicalein on canine osteosarcoma cell proliferation and death** Baicelein is a flavanoid with anti-inflammatory and antineoplastic properties Canine OSA cell lines were treated with Baicelein Modest ____ changes were observed in 1 cell line Effective in inducing ___ and did not prevent doxorubicin cell proliferation inhibition

Cell cycle Apoptosis

Thayanithy et al. 2012. Combinatorial treatment of DNA and chromatin-modifying drugs cause cell death in human and canine osteosarcoma cell lines Downregulation of microRNAs (miRNAs) at the 14q32 locus stabilizes the expression of cMYC, thus significantly contributing to osteosarcoma (OS) pathobiology. Here, we show that downregulation of 14q32 miRNAs is epigenetically regulated. The predicted promoter regions of miRNA clusters at 14q32 locus showed no recurrent patterns of differential methylation, but Saos2 cells showed elevated histone deacetylase (HDAC) activity. Treatment with 4-phenylbutyrate increased acetylation of histones associated with 14q32 miRNAs, but interestingly, robust restoration of 14q32 miRNA expression, attenuation of cMYC expression, and induction of apoptosis required concomitant treatment with 5-Azacytidine, an inhibitor of DNA methylation. These events were associated with genome-wide gene expression changes including induction of pro-apoptotic genes and downregulation of cell cycle genes. Comparable effects were achieved in human and canine OS cells using the HDAC inhibitor suberoylanilide hydroxamic acid (SAHA/Vorinostat) and the DNA methylation inhibitor Zebularine (Zeb), with significantly more pronounced cytotoxicity in cells whose molecular phenotypes were indicative of aggressive biological behavior. These results suggested that the combination of these chromatin-modifying drugs may be a useful adjuvant in the treatment of rapidly progressive OS.

**won Seo et al. Fluoroquinolone-mediated inhibition of cell growth, S-G2/M cell cycle arrest, and apoptosis in canine OSA cell lines** Most patients are treated with flouriquinolone who develop post-operative infections Ciprofloxacin or enrofloxacin ____ expression leading to \_\_\_\_? Flouroquinolone exposure induced ____ of canine OSA cells as demonstrated by ______ and \_\_\_\_, and activation of \_\_\_\_\_

Inhibited p21^WAF1, decreased proliferation and increased S-G2/M accumulation Apoptosis, cleavage of caspace-3, PARP, caspace 3/7

**Simcock et al. 2012. Evaluation of a single subcutaneous infusion of carboplatin as adjuvant chemotherapy for dogs with OSA: 17 cases** Limb amputation or limb-sparing surgery followed by single SC infusion of carboplatin (total dose 300 mg/m2 over 3, 5, 7 days) MST? AEs? No significant difference was found in ST of dogs grouped to tumor location, type of surgery, duration of carboplatin infusion, or postoperative infection

365 days 9 had hematological effects, 1 GI, 7 infections at surgical site

**Maeda et al. Genomic instability and telomere fusion of canine OSA cells** All 8 cell lines increased abnormal metacentric chromosome and telomere fusion and interstitial telomeric signals Evidence of unstable telomeres, colocalization of g-H2AX and telomere signals in interphase cells All canine OSA samples were telomerase positive **Telomere fusion may serve as potential marker for canine OSA**

**Bongiovanni et al. IHC investigation of cell cycle apoptosis regulators (Survivin, B-catenin, p53, caspace 3) in canine appendicular OSA** B-catenin staining? Nuclear survivin and p53? Moderate/high B-catenin expression was significantly associated with \_\_\_\_\_\_? Moderate/high nuclear p53 expression was associated with \_\_\_? Moderate/high nuclear survivin showed tendency toward \_\_\_? **p53 was negative prognostic marker and survivin potential positive prognostic indicator (confusing)**

Nuclear staining detected in normal osteoblasts adjacent to tumor in 47% of cases. Cytoplasmic and membranous staining was also observed Positive in all cases Development of metastasis Moderate/high histologic grade and shorter OS Longer OS

**Ryseff et al. Detection of ALP in canine cells previously stained with Wright-Giemsa and its utility in differentiating OSA from other mesenchymal tumor** 83 dogs, 17 OSA and 66 other tumors (amelonotic melanoma, GIST, collision tumor, and anaplastic sarcoma) ALP expression? Sensitivity and specificity?

15/17 OSA and 4/66 other tumors expressed ALP 88% and 94%

**Boerman et al. Prognostic factors in canine appendicular OSA - a meta-analysis** What were negative prognostic factors? What was associated with shorter ST and DFI?

Elevated ALP and proximal humerus for both ST and DFI in dogs with appendicular OSA Increasing age, however was not statistically significant

**Selvarajah et al. Expression of EGFR in canine OSA: association with clinicopathological parameters and prognosis** EGFR mRNA and protein expression in OS cell lines, normal bones, frozen primary OS and tissue microarrays EGFR expression in primary OS? Tissue microarray analysis revealed thay subset of canine OS with high EGFR was associated with ___ ? Cytoplasmic expression of EGFR? **Subset of dogs may benefit from anti-EGFR therapies**

Significantly elevated in OS compared to normal bones and in OS metastasis to the lung in comparison to extrapulmonary sites Significantly shoter survival times and DFI 75% of metastasis and similar in primary tumors

**Karnik et al. 2012. Accuracy of CT in determining lesion size in canine appendicular OSA** Statistically significant association with good correlation between true lenght of OSA compared to lenght of intramedullary/endosteal abnormalities on CT Not significant association between tumor lenght and lenght of periosteal proliferation Statistical significant association but poor correlation between tumor lenght compared to lenght of abnormal contrast enhancement

**Wasserman et al. Suppression of canine myeloid cells by soluble factors from cultured canine tumor cells** Canine BM-DCs and DH82 cell line shared? Myeloid cells exposed to tumor derived soluble factors (TDSF) showed? **Soluble factors produced by tumor cells suppress activation and function of myeloid cells.**

CD11b, CD11c, and MHCII expression Decreased MHCII expression and CD80 - reduced phagocytic activity and suppressed proliferation of responder immune cells

**Schmit et al. Cathespin K expression and actvitity in canine OSA** CatK is lysosomal protease with collagenolytic activity, and secretion by osteocalts is responsible for degrading bone matrix Canine OS cells contained ___ CatK in cytoplasmic vesicles In OS cells, ____ induced secretion CatK, which degraded \_\_\_ CatK concentration in OS compared to healthy dogs? In subset of dogs, CatK decreased after palliatve RT and antiresorptive treatment

preformed TGFB1, collagen type I higher

**Shoeneman et al. 2011. Expression and function of survivin in canine OSA** Elevated survivin correlated with? Survivin attenutation lead to?

Increased histologic grade and MI and decreased DFI Inhibited cell cycle progression, increased apoptosis, mitotic arrest, and chemosensitivity, and cooperated with chemotherapy to improve tumor control

**Marley et al. The effects of taurolidine alone and in combination with doxorubicin or carboplatin in canine OSA in vitro** Taurolidine is derivative of taurine and has anti-tumor and anti-angiogenic effects against variety of cancers Effect on OSA cells and in combination with dox or carbo? Taurolidine cytotoxicity was ____ in 1 cell line **Effects may depend on functional status of p53**

Cytotoxic and increased toxicity of dox and carbo in vitro Caspace dependent with mutant p53 protein

**Kruse et al. Evaluation of clinical and histopathologic prognostic factors for survival of canine OSA of extracranial flat and irregular bones** What does not appear to be correlated with survival? What was associated with decreased survival time?

Histologic grade Increased ALP and tumor location in scapula was associated with decreased survival times

**Wells et al. Arginase treatment prevents the recovery of canine LSA and OSA cells resistant to the toxic effects of prolonged arginine deprivation** Effects of arginine deprication in canine LSA and OSA cell lines Cell lines unable to recover 6 days of arginine deprivation but full recovery on return to arginine rich culture Cell death after arginase Adult progenitor cells from BM of healthy dog able to recover after 9 days of culture in arginase Over 3 days in culture, arginase was more effective than asparaginase in inducing cell death in lymphoid cell lines

**Kozicki et al. Adjuvant therapy with carboplatin and pamidronate for canine appendicular OSA** 17 dogs treated with amputation Median DFI compared to historical controls? MST? **Addition of pamidronate to carboplatin is safe and does not impair efficacy of carboplatin treatment**

185 days (6 mo) vs. 172 days 311 (10 mo) vs. 294 days (9.8 mo)

**Sacornrattana et al. Abdominal ultrasonographic findings at diagnosis of OSA in dogs and association with treatment outcomes** How many had abnormalties? Organ with most changes? Metastases identified in how may dogs? Dogs with abnormalties were less likley to receive definitive therapy, exhibited shorter survival (latter not significant) Identifications of lesions in liver or kidney were statistically associated with shorter survival

57% Spleen 2.5%

**Cannon et al. Canine OSA cells exhibit resistance to aurora kinase inhibitors** Effects of aurora kinase inhibitors AZD1152 and VX680 on OSA cells Cytotoxicity was observed but inhibitor concentrastion were higher in human leukemia and canine LSA cells AZD1152 reduced aurora kinase phophorylation - resisatnce not due to target recognition Efflux mediated by ABCB1 and ABCG2 was the not the mechanism

**Marley et al. 2013. PK study and evaluation of safety of taurolidine for dogs with OSA** Taurolidine infusion safe in 6 dogs Taurolidine with PVP (polyvinylpyrrolidine) had allergic reaction but recovered when infusion was stopped 3 dogs with OSA received dox and taurolidine - DCM, PLN, renal insufficiency, vasculopathy 1 dog develop ototoxicity with carbo and taurolidine GI and BM toxicities were not increased over dox and carbo alone **Combo with doxo not recommend but with carbo appears safe**

**Stoewen et al. Factors influencing veterinarian referral to oncology specialists for treatment of dogs with LSA and OSA in Ontario, Canada** 56. 3% recommended referral for LSA and OSA 84. 3% practised \<2hr, 94% were confident in oncology service 22-33% were experienced with use of chemotherapeutics. 60% were experienced with amputation Referral was associated with practioners perception of health status, interaction between clients bond with the dogs and clients financial status, practioner experience with treating cancer, how worthwhile practioner consider treatment to be, and confidence in referral center

Caro et al. 2013. Markers of ironmetabolism in retired racing urinse Greyhounds with and without OSA

Milovancev et al. Comparative analysis of the surface exposed proteome of 2 canine OSA cell lines and normal canine osteoblasts

**Wallace et al. 2013. Diagnostic utility of AUS for routine staging at diagnosis of skeletal OSA in dogs** 23 abnormalites were evaluated and 19 were bening 5% had another primary neoplasia 2/9 had AUS due to palpable abdominal mass was diagnosed with another neoplasia compared with only 1 of the 49 that had US for routine staging Do US in animals with palpable abdominal mass - influence treatment decisions

**Scharf et al. Effect of bevacizumab and growth of canine OSA cells xenografted in athymic mice** Mice in high-dose bevacizumab had significantly delayed tumor grwoth compared to low-dose group **VEGF inhibitors may be useful in treatment of OSA in dogs**

**Schwartz et al. 2013. Orthotopic model of canine OSA in athymic rats for evaluation of SRT** Tumors developed in 10/12 tibial sites and 14 femoral sutes Administration of SRT was feasible Mean tumor necrosis 95% and minimal AE on skin

**Oblak et al. 2015. Comparison of concurrent imaging modalities for staging of dogs with appendicular primary bone tumors** No definitive lesions were identified on radiographs or CT Lesions were identified on thoracic CT that were not visible radiographically Ground glass pulmonary lesions progressed in 3/4 cases - suspicious for metastasis Whole body CT was not suitable alternative to bone scintigraphy however useful as adjunctive diagnostic modality

**Covey et al. Stereotactic radiosurgery and fracture fixation in 6 dogs with appendicular OSA** 6 dogs 2 had pathologic fracture at admision and 4 after SRS First 3 fractures repaired using open approach and 3 using minimally invasive percutaneous osteosynthesis Infection occured in 5 dogs (83%) and implant failure in 3 (50%) Survival times ranged from 364-897 days (1-2.5 yr)

**Pang et al. Global gene expression analysis of canine osteosarcoma stem cells reveals a novel role for COX2 in tumor initiation** COX-2 was expressed 141-fold more in CSC than daughter adherent cells COX-2 inhibition had no effect on CSC growth, or resistance to chemotherapy however it inhibited COX-2 in daughter cells prevented sphere formation - potential significance of COX-2 in tumor initiation

**Selmic et al. Outcome and prognostic factors for OSA of the maxilla, mandible, calvarium in dogs: 183 cases** Mean age, Body weight, Breed? Local progression occured in? How many developed metastatic disease MST Dogs that underwent surgery MST? Tumor free margins were associated with decreased hazard of progression or recurrence Dogs with OSA of calvarium had significantly greater hazard of local recurrence or progression

9.3 years, 32kg, purebred 44% 51%, 40% 239 days (8 mo) 329 days (11 mo)

**Maniscalco et al. 2014. Increased expression of insulin life growth factor-1 receptor is correlated with worse survival in canine appendicular OSA** IGF-1R was expressed in ___ of OSA samples and dogs with higher levels had \_\_\_\_ IGF-1R is activated by \_\_\_, in paracrine or endocrine manner - activation of AKT/MAPK signaling, PPP caused p-IGF-1R dephosphorylation with partial blocking of p-MAPK and p-AKT, and apoptosis **IGF-1R expression is correlated with poor prognosis and potential therapeutic target**

71%, decreased survival IGF-1

**Selmic et al. 2014. OSA following tibial plateau leveling osteotomy in dogs: 29 cases** Mean age, weight? Associated clinical signs? Mean interval between TPLO and OSA diagnosis? Cast stainless steel in 18 dods and plates of wrough stainless steel in 4 MST for 10 dogs that underwent treatment with amputation and chemo? **Similar to outcomes reported for dogs with appendicular OSA**

9 yr, 45 kg Swelling 17/21 and lameness 28/29 5.3 years 313 days

**Rodrigues et al. Tumorigenic canine OSA cell lines associated with frizzled-6 upregulation and enhanced population cell frquency** \_\_\_\_ was upregulated in tumorigenic cell lines (7.78 fold) compared to non-tumorigenic cell lines Frizzled-6, coreceptor for Wnt Tumorigenic cell lines had increase in side population of cells compared with non-tumorigenic (6% vs. 1.6%) There was no difference in tumorigenicity, frizzled-6, or percentage of side population cells noted b/t OSA cells lines generated from patient with differing ALP concentrations

Frizzled-6

**Shor et al. 2015. Expression of nociceptive ligands in canine OSA** **10 dogs with appendicular OSA** Canine OS cells express and secrete? OS samples express? Circulating noceiceptive ligand concentration were detectable but failed to correlate with pain status

NGF, endothelin-1, and PGE2 Noceiceptive ligands

**Nagamine et al. 2015. Diversity of histologic patterns and expression of cytoskeletal proteins in canine skeletal OSA** 64% of OS contained multiple subtypes Myxoid, round cell, epitheliod subtypes were observed Epitheloid were observed in OS of the head (OSH) CK was expressed by epitheliod and sarcomatoid tumor cells, and all CK+ tumor cells expressed vimentin Vimentin and SMA were expressed in all subtypes GFAP was expressed in many subtypes, and some showed nerofilament expression

**Mauchle et al. 2014. Identification of anti-proliferative kinase inhibitors as potential therapeutic agents to treat canine OSA** 22 different kinase inhibitors 4 compounds (RO 31-8220, 5-iodorubericidin, BAY 11-7082, and erstatin) whoed significant cell growth inhibitor effect RO 31-8220 and 5-iodorubericidin showed the highest ability to potentiate effects of doxorubicin Targeting PKC, CK1, PKA, ErbB2, mTOR and NF-KB pathways

**Sabattini et al. 2017. Comparative assessment of the accuracy of cytological and histologic biopsies in the diagnosis of canine bone lesions** Accuracy for cytology? Sensitivty and specificity Accuracy for histology? Sensitivty and specificity Tumor was correctly identified cytologically and histologically in?

83% - 83% and 80% 82.1% - 72% and 100% 50% and 55%

**Tuohy et al. 2016. Association of canine OSA and monocyte phenotype and chemotactic function** Percentage of expression of CCR2 and CXCR2 was higher in control dogs compared to OSA PGE2 and TNFa are increased in OSA monocyte culture compared to controls Monocytes from OSA had decreased chemotactic function compared to control dogs **Decreased chemokine receptor expression and chemotaxis - evade immune response**

**Story et al. 2017. Evaluation of weight change during carboplatin therapy in dogs with appendicular OSA** A slight ___ in weight occured over the course of chemotherapy - but not statistically significant Affect on survival?

Increase Weight change did not have affect on survival

**Byrum et al. 2016. Downregulation of CXCR4 expression and functionality after zoledronate exposure in canine OSA** **20 dogs with OSA** All canine OSA cells express CXCR4 and zoledronate ___ expression and functionality by 27% through proteosome degradation and reduced prenylation of heterotrimeric G-proteins in 33% of tumor cell lines In OS dogs, zoledronate reduces CXCR4 expression by 40% and decreases circulating CXCR4 in 18 of 20 dogs

Reduced

**Burton et al. 2015. Implant associated neoplasia in dogs: 16 cases** Median time from implant to diagnosis of neoplasia? Which limbs were most frquently affected? Most common location on bone? Most common tumor?

5.5 years Hindlimbs - tibia 8/16, femur 5/16, 1 pelvis and femur Diaphysis 15/16 OSA 13/16, HS, FSA, spindle cell sarcoma

**Rubin et al. Factors associated with pathological fractures in dogs with appendicular primary bone neoplasia: 84 cases** Median time from diagnosis to euthanasia? Pathological fractures identified in how many dogs? Most commonly affcted bones? Tumors arising from long bones other than ___ had odds of eventual fracture 5.05 as great as odds for tumors of the radius Lytic tumors had odds eventual fracture 3.22 as great as odds for tumors that appeared blastic or mixed lytic-blastic **Radial tumors less likely to fracture and lytic tumors more likely to fracture**

111 days 38% Femur\>tibia\>humerus\>radius\>ulna Radius

**Meyer et al. 2015. Expression of platelet-derived growth factor BB, erythropoeitin and erythropoeiting receptor in canine and feline OSA** EPO-R is expressed on neoplastic cells - may play a role in tumor progression EPO expression was positive in all tumors and EPO mRNA was detected in 38% of canine and 40% feline samples EPO-R was expressed in all samples EPO-R mRNA was higher in feline tumors, tumor cell lines, and kidney compared to canine tissues All cell lines responded to EPO treatment **Autocrine and paracrine tumor progression cannot be excluded**

**Willcox et al. Serum 25-hydroxyvitamin D concentrations in dogs with OSA do not differ from those age- and weight- matched control dogs** Levels were similar in dogs with OSA and matched controls **25-hydroxyvitamin-D insufficiency may not play a role in pathogenesis**

**Zhang et al. 2015. The effect of Zhangfei/CREBZF on cell growth, differentiation, apoptosis, migration, and the unfolded protein response in several canine OSA cell lines** Like D-17 the 3 cell lines expressed p53 proteins capable of activating with p53 response elements Zhangfei suppressed the growth of UPR-related transcripts Induced activation of osteocalcin expression a marker of osteoblast differentiation and trigged programmed cell death **Induction of Zhangfei, where p53 is functional, may be an effective modality for treatment of OS**

**Herrera et al. 2015. Peroxisome proliferator activated receptor g protein expression is assymmetrically distributed in primary lung tumor and metastatic to lung OSA samples and does not correlate with gene methylation** PPAR-g is a TF plays a role in proliferation and differentiation PPAR-g is expressed in \_\_% of canine NSCLC and __ % of metastatic OSA There was significant loss of 5'PPAR-g methylation from normal lung to primary lung cancer to metastatic OSA Altered PPAR-g promoter methylation at the interrogated locus does not appear to be associated with changes in protein expression

33%, 66%

Fowles et al. Intra- and interspecies gene expression models for predicting drug response in canine OSA

**Mantovani et al. Effects of epidermal growth factor receptor kinase inhibiton on radiation response in canine OSA cells** Effects of small molecule EGFR inhibitor erlotinib on canine OSA radiation response Erlotinib reduced clonogenic survival in 2 canine OSA cell lines and enhacned the impact of RT in 1/3 cell lines Erlotinib enhanced radiation effects and demonstrated single effects Did not alter EGFR nor inhibit PKB/Akt - increased phoshphorylation of Akt VEGF levels increased after erlotinib as single agent and in combination with RT in 2/3 and decreased in 1/3 cell lines

**Levine et al. 2016. Effects and synergy of feed ingredients on canine neoplastic cell proliferation** Examine 5 feed ingredients for antiproliferative effects and interaction with toceranib and doxorubicin Which agents were most effective? \_\_\_ extract was most potent and exhibited synergy with __ extract Combination had additive or synergistic effects with chemo agents

Tea, turmeric, and rosemary extract Turmeric, rosemary

**Murphy et al. 2017. Evaluation of P16 expression in canine appendicular OSA** Lack of canine OSA P16 expression and shorter disease free interval

**Ong et al. Anti-tumor efficacy of etoposide alone and in combination with piroxicam against canine OSA in xenograft model** Etoposide single agent delayed tumor progression with marked reduction of Ki67 in tumor tissue Combination treatment (with piroxicam) did not enhance anti-tumor efficacy of etoposide Both (etoposide alone and combination) downregulated survivin expression but was not followed by increased apoptosis

**Leeper et al. Preliminary evaluation of serum total cholesterol concentrations in dogs with OSA** Total cholesterol was ___ in OSA bearing dogs, \_\_\_% fracture controls, and \_\_% similar age/weight controls Elevated cholesterol was associated with?

elevated 45%, 10%, 6.5% reduced hazard ratio for overall mortality in dogs with OSA

**Kubicek et al. Association between CT characteristics and fractures following stereotactic radiosurgery in dogs with appendicular OSA** MST? Fracture free rates at 3, 6 and 9 month? Bone affected was significantly associated with time to fracture Median time to fracture in dogs with subchondral bone involvement? without subchondral bone involvement? Acute and late skin effects 58% and 16%

9.7 months 73%, 44%, 38% 4.2 months, 16.3 months

**Pagano et al. Safety and toxicity of an accelerated coarsely fractionated radiation protocol for treatment of appendicular OSA in 14 dogs** 10Gy delievered in 2 consecutive days - total of 20Gy 13/14 received adjuvant therapy with pamidronate and NSAID 9 dogs received chemotherapy Median of __ days, % had improved pain control Median DOR? How many developed pathologic fracture? Timing of fracture?

14 days, 93% 80 days 35.7% 24-250 days

**Arthur et al. Risk of OSA in dogs after open fracture fixation** 19,041 fractures, 15 dogs developed OSA affecting the same bone

**Seguin et al. Long term outcome of dogs treated with ulnar rollover transposition for limb-sparing of distal radial OSA: 27 limbs in 26 dogs** Complication occured in how many limbs? Infection? Biomechanical complication? Local recurrence? Limb function graded by veterinarians or owner was poor in 2 limbs, fair in 4, good in 15, excellent in 3, unknown iun 4 Median DFI and ST **High complication rate but limb function was acceptable**

20 (76%) 12 (48%) 15 (58% 2 (8%) 245 (8 mo) and 277 days (9 mo)

**BMI1 is expressed in canine OSA and contributes to cell growth and chemotherapy resistance** BMI1 is stem cell factor BMI1 expression in primary and metastatic OSA? Canine OSA cell lines expressed high levels of BMI1 compared with osteoblasts and knockdown demonstrated resistance to carboplatin and doxorubicin chemotherapy **Inhibition of BMI1 may improve response to chemotherapy**

Strong nuclear expression

**Wehrle-Martinez et al. Osteocalcin and osteonectin expression in canine OSA** 23 OSA, 4 CSA, 4 FSA, 2 HSA, 4 HS OC expression in OSA and CSA? OC in FSA, HSA, HS? sensitivity and specificity for OC in canine OSA? ON?

83% OSA and 100% CSA Negative 83% and 71% 100% OSA and 100% non-OSA - 100% sensitivty but no specificty

Massimi et al. 17-AAG and apoptosis, autophagy, and mitophagy in canine OSA cell lines

Roy et al. Comparative proteomic investigation of metastatic and non-metastatic osteosarcoma cells of human and canine origin

**Rebhun et al. Evaluation of optimal water flouridation on the incidence and skeletal distribution of naturally arising OSA in pet dogs** No difference in age, skeletal distrubtion of OSA cases relative to flouride status Dogs with OSA were not more likely to live in area with optimized flouride in water than dogs with LSA or HSA **Does not contribute to OSA**

**Rizzo et al. The effects of sulforaphance on canine OSA proliferation and invasion** SFN found in raw cruciferous vegetables may have utility in chemoprevention - as antineoplastic and free radical scavenger SFN could not induce cell death at physiologic concentrations but diminished cell invasion and downregulation of focal adhesion kianse (FAK) signaling When used with doxorubicin there was a protective effect to dox-induced cytotoxicity

**York et al. Enrofloxacin enhances the effects of chemotherapy in canine OSA cells with mutant and wild-type p53** Moresco and Abrams contained mutations in p53 while no mutation in D17 cell line or normal canine osteoblast cell line The addition of enrofloxacin to either dox or carbo resulted in further reduction in cell viability - regardless of p53 mutational status

**Guiffrida et al. Use of routine histopathology and factor VIII-related antigen/vWF IHS to differentiate primary HSA of bone from telengiectatic OSA in 54 dogs** 20% of tumors were reclassified based on FVIII-RAg/vWF immunoreactivity from original diagnosis of tOSA

**Laver et al. Prospective evaluation of toceranib phosphate in metastatic canine OSA** AEs? SD? Median PFS Plasma VEGF concentration were significantly elevated at 4 weeks of TOC treatment, but no changes in Treg

Common but low grade 18% with remainder having PD 89 days

**Wouda et al. 2017. Safety evaluation of combination carboplatin and toceranib phosphate in tumor bearing dogs: A phase I dose finding study** MTD of carboplatin? Palladia? Dose limiting toxicity? Clinical benefit?

200 mg/m2 IV q 21 days 2.75 mg/kg PO EOD neutropenia Observed in most cases

**Bulla et al. 2017. Platelets inhibit migration of canine OSA cells** Addition of blood leukocytes to platelet samples did not alter the inhibitory effect on migration Platelet treatment downregulated? Interaction between canine platelets or molecules released during platelet activation inhibits their migration - canine pletelets may antagonize metastasis of OSA

SNAI2 and TWIST1 genes - genes involved in EMT

**Frimberger et al. Canine OSA treated by post-amputaion sequential accelerated doxorubicin and carboplatin chemotherapy: 38 cases** MST? 1- and 2- year survival How many dogs hospitalized? What impacted survival time on multivariate analysis? **Outcomes were similar to other chemo protocols**

317 days (10.5 mo), 43% and 14% 5.2% Serum ALP and whether chemo was completed

**Goupil et al. 2012. Prevelence of 5-lipooxygenase expression in canine OSA and the effects of dual 5-LOX/COX inhibitor on OSA cells in vitro and in vivo** 5-LOX inhibitor, tepoxalin, reduced cell proliferation 5-LOX leads to production of lipid mediations and when added to media did not recover cell proliferation Lipid mediation production is not the means by which tepoxalin alters proliferation

**Montinaro et al. 2013. Clinical outcome of 42 dogs with scapular tumors treated by scapulectomy: VSSO retrospective study** Subtotal scapulectomy done in 18 dogs (43%) Limb use was good to excellent overall Only adjunctive chemotherapy had positive effect on DFI

**London et al. 2015. Impact of toceranib/piroxicam/cyclophosphamide maintenace therapy on outcome of dogs with appendicular OSA follwoing amputation and carbopaltin chemotherapy** Median DFI for control and toceranib treated dogs was? Median OS for control and toceranib? 1-year survival rate for control and toceranib? **Addition of toceranib to metronomic piroxicam/cyclophpsphamide therapy following amp + carbo did not improve DFI**

215 (7 mo) and 233 days 242 (8 mo) and 318 days (10.6 mo) 35% and 38%

**Mason et al. 2016. Immunotherapy with Her2-targeting listeria induces HER2-specific immunity and demonstrates potential therapeutic effects in a phase I trial in canine OSA** ADXS31-164 - attenutaed, Listeria monocytogenes expressing human HER2/neu fusion protein to induce immunity Toxicities? ADXS31-164 induced IFN-g response in 15/18 dogs within 6 months ADXS31-164 reduced metastatic disease and increased duration of survival time

Low grade

**Curtis et al. 2016. Combination therapy with zoledronic acid and PTH improves bone architecture and strenght following a clincially relevant dose of SRT for the local tretament of canine OSA in athymic rats** Significant increase in bone volume and increased polar moments of inertia 8 weeks after radiation in the combine ZA/PTH treatment group as compared to RT alone

**Fenger et al. 2016. MIR-9 is overexpressed in spontaneous canine OSA and promotes a metastatic phenmotype inlcusing invasion and migration in osteoblats and OSA cell lines** Mir-9 is overexpressed in OS tumors and cell lines Transduced normal osteoblasts and cell lines with mir9 - invasion and migration but did not affect proliferation or apoptosis Overexpression of mir9 identified alterations in numerous genes - upregulation of GSN, an actin filament severing protein involved in cytoskeletal remodeling Downregulation of mir-9 in OS cell lines reduced GSN with decrease in invasion and migration

**Talbot et al. 2017. Retrospective evaluation of whole body CT for tumor staging in dogs with primary appendicular OSA** 39 dogs Was bone metastasis detected? Pulmonary metastasis? How many dogs had additional masses detected?

No 5% 5%

**Ong et al. 2016. Effects of etoposide alone and in combination with piroxicam on canine OSA cell lines** Effects of etoposide alone on cell line? Effects of etoposide in combination with piroxicam? What caused tha anti-proliferative effect?

Supressed cell growth and viability Concentration dependent cytotoxicity Cdc2-CyclinB1 - increase in G2/M fraction

**Boston et al. 2017. Outcome and complications in dogs with appendicular primary bone tumors treated with stereotactic radiotherapy and concurrent surgical stabilization.** 18 dogs 7 dogs had pathological fracture and 11 were at high risk single dose of SRT (5) or 3 doses (13) Bone plate (15), interlocking nail (3) Complications occured in how many dogs? Most common complication? Post-operative fracture was recored in how many? 9 dogs were amputated at median of? MST? **Treatement with SRT and stabilization associated with high complication rate - consider alternative methods of limb salvage**

16/17, 15/17 being major complicarions Infection - 15/17 3 cases 152 days 344 days (11.5 mo)

**Turner et al. 2017. Prognosis of dogs with stage III OSA following treatment with amputation and chemotherapy with and without metastasectomy** MST for dogs that received no treatment for metastasis? MST for dogs that received metastatectomy and did not? Basis used for metastasectomy?

2 months 232 days (7.7 mo) vs. 49 days \<3 pulmonary nodules on CXR and DFI\>275 days

**Al-Khan et al. 2017. Immunohistochemical validation of spontaneously arising canine OSA as a Model for Human OSA** Markers expressed in canine OSA? Markers variably expressed in canine OSA? **Similar to human OSA, canine OSA may represent model to study the disease**

Vimentin, ALP, Runx2 (runt-related TF4), BMP4 (bone morphogenetic protein 4) Desmin, S100, NSE (neuron specific enolase)

**Swift et al. 2018. Outcome of 9 dogs treated with SRT for primary or metastatic vertebral OSA** 9 dogs treated with SRT - 1 to 5 fractions with total dose 13.5 - 36Gy 6 dogs primary lesions and 3 had metastatic lesions Neurologic signs improved in 4 patients, remained the same in 4 and worsened in 1 5/6 dogs that presented with assessable spinal pain reported improvement in pain MST? Median duration of pain control?

139 days (4.6 mon) 77 days

**Leonardo et al. 2017. miR-1 abd miR-133b expression in canine OSA** Important role in regulation osteogenesis and tumor cell proliferation Expression in tumors compared to normal bones? Expressionof MET and MCL1 **Potential role of miR-1 and miR-133b as biomarkers**

Lower expression in tumors compared to normal bone with higher expression of target genes MET and MCL1 Moderate to strong immunostaining in more than 50% tumor samples

**Schott et al. 2018. Histologic grade does not predict outcome in dogs with appendicular OSA receiving the standard of care** Compare 2 published histologic grading systems with appendicular OSA treated with SOC for curative intent Histologic grade and outcome? What was correlated with outcome?

Histologic grade did not correlate with outcome Increased number of mitotic figures per 3/400x

**Selmic et al. 2018. Association of TPLO with proximal tibial OSA in dogs** TPLO had been performed \>1year ago Dog with TPLO were ___ to develop OSA than dogs without Each 1kg increased in body weight was associated with 11% increase in odds of proximal tibibal OSA **OSA should be differential for new or worsening hindlimb lameness in dogs that underwent TPLO \>1 year previously**

40x

**Matsuyama et al. 2018. Evaluation of metronomic cyclophosphamide chemotherapy as maintenance for dogs with appendicular OSA following limb amputation and carboplatin chemotherapy** Dogs grouped whether they received MC or did not (15 mg/m2 PO q24h and meloxicam 0.1 mg/kg PO Q24hr) 19 dogs received carboplatin and MC chemotherapy and 20 received carboplatin chemotherapy Treatment discontinued in how many dogs? Cause? Median follow up period? Difference in MST and PFS between the 2 groups?

58% due to cystitis 450 days None

**Sprecher et al. 2018. Retrospective analysis of corrosion and ion release from retrieved cast stainless steel tibial plateu leveling osteotomy plates in dogs with and without pre-implant OSA** Greater metal ion release was observed in cast plates than foreged plates Accumulation of metal ions may trigger neoplastic transformation in neighboring cells

**Withers et al. 2018. Association of macrophages and lymphocyte infiltration with outcome in canine OSA** Median number of CD3+ and FOXP3+ infiltrates were 45.8 cells/mm2 and 8.5 mm2 The median area of CD204+ macrophages was 4.7% and dogs with tumors contained greater than 4.7% CD204+ experienced significantly longer DFI **Macrophages may play a role in inhibiting cOSA progression**

**Rici et al. 2018. Combination therapy of canine OSA with bone marrow stem cells, bone morphogenetic proteina and carboplatin in an in vivo model** Combination of mesenchymcal stem cells and rhBMP-2 reduced tumor mass and infiltrastion of neoplastic cells in tissues more efficiently than carboplatin alone

Patatsos et al. 2018. Pre-clionical evaluation of protesosome inhibitors for canine and human OSA

**Gold et al. 2018. Zygomatic arch parosteal OSA in dogs and cats** Parosteal OSA is slow growing tumors arising from surface of long bones 5 dogs and 1 cat Clinical presentation varied from chronic sneezing to facial swelling Histologically - well differentiated fibro-osseous and chondroid components Cellular atypia and mitotic figures were uncommon

**Hans et al. 2018. Effect of surgical site infection on survival after limb amputation in the curative-intent treatment of canine appendicular OSA** 15 dogs with OSA and SSI and 134 dogs with OSA and no SSI Median DFI and MST Difference in DFI dogs with SSI and without SSI? MST with SSI and without SSI? What was associated with decreased DFI and survival? **SSI did not influence survival**

236 days (8 mo) and 283 days (9.4 mo) Not different - 292 days with and 224 days without Not different - 292 days and 280 days Not completeing chemotheray

**Guiffrida et al. 2018. Primary appendicular HSA and tOSA in 70 dogs** Dogs with HSA were more likely than dogs with tOSA to be ___ and have ___ limb tumors \_\_% of HSA occured in hind limbs, particularly \_\_\_ Dogs with tOSA weighed a median of 9.9kg more than dogs with HSA OS for HSA and tOSA? Younger age and more aggressive treatment were associated with longer survival in dogs with HSA but not tOSA 1 year survival rates did not differ between HSA (28%) abd tOSA (7%)

Males and hind limb tumors 78%, tibia 299 days (10 mon) and 213 days (7 mon)