Evaluation of intraoperative fluorescence imaging-guided surgery in cancer-bearing dogs: a prospective proof-of-concept phase II study in 9 cases.

The objective was to prospectively evaluate the application of intraoperative fluorescence imaging (IOFI) in the surgical excision of malignant masses in dogs, using a novel lipid nanoparticle contrast agent. Dogs presenting with spontaneous soft-tissue sarcoma or subcutaneous tumors were prospectively enrolled. Clinical staging and whole-body computed tomography (CT) were performed. All the dogs received an intravenous injection of dye-loaded lipid nanoparticles, LipImage 815. Wide or radical resection was realized after CT examination. Real-time IOFI was performed before skin incision and after tumor excision. In cases of radical resection, the lymph nodes (LNs) were imaged. The margin/healthy tissues fluorescence ratio or LN/healthy tissues fluorescence ratio was measured and compared with the histologic margins or LN status. Nine dogs were included. Limb amputation was performed in 3 dogs, and wide resection in 6. No adverse effect was noted. Fluorescence was observed in all 9 of the tumors. The margins were clean in 5 of 6 dogs after wide surgical resection, and the margin/healthy tissues fluorescence ratio was close to 1.0 in all these dogs. Infiltrated margins were observed in 1 case, with a margin/healthy tissues fluorescence ratio of 3.2. Metastasis was confirmed in 2 of 3 LNs, associated with LN/healthy tissues fluorescence ratios of 2.1 and 4.2, whereas nonmetastatic LN was associated with a ratio of 1.0. LipImage 815 used as a contrast agent during IOFI seemed to allow for good discrimination between tumoral and healthy tissues. Future studies are scheduled to evaluate the sensitivity and specificity of IOFI using LipImage 815 as a tracer.

Phase-0/phase-I study of dye-loaded lipid nanoparticles for near-infrared fluorescence imaging in healthy dogs.

Near-infrared (NIR) fluorescence imaging using FDA-approved indocyanine green (ICG) has been the subject of numerous studies during the past few years. It could constitute a potentially exciting new paradigm shift in veterinary oncology, especially to develop in vivo fluorescence imaging diagnostics and surgery guidance methods. The objective of this study was to evaluate the pharmacologic and toxicological characteristics in healthy beagle dogs of LipImage™ 815, a formulation made of NIR-dye-loaded lipid nanoparticles. The initial dosage for the evaluation of biodistribution was extrapolated from data in mice and then adapted to define the more adapted dose (MAD) according to the fluorescence results obtained in 5 dogs using a Fluobeam® 800 imaging device (phase 0 study). A single dose acute toxicity study was then performed (3 dogs, phase I study). Before the systemic administration of LipImage™ 815, the dogs presented a very mild residual fluorescence, particularly in the liver and kidneys. After injection, the plasma fluorescence continuously decreased, and the signal was relatively homogeneously distributed throughout the different organs, though more pronounced in the liver and to a lesser extent in the steroid-rich organs (adrenal, ovaries), intestines, lymph nodes and kidneys. A MAD of 2.0µg/kg was found. No evidence of acute or delayed general, hepatic, renal or hematologic toxicity was observed at 1-fold, 5-fold or 10-fold MAD. The results of this phase-0/phase-I study showed that an optimal dosage of LipImage™ 815 of 2.0µg/kg allowed the achievement of a fluorescence signal suitable for surgery guidance application without any acute side effects.

Characterization of a novel canine T-cell line established from a spontaneously occurring aggressive T-cell lymphoma with large granular cell morphology.

Dogs with lymphoma are established as good model for human non-Hodgkin lymphoma studies. Canine cell lines derived from lymphomas may be valuable tools for testing new therapeutic drugs. In this context, we established a canine T-cell line, PER-VAS, from a primary aggressive T-cell lymphoma with large granular morphology. Flow cytometric analysis revealed a stable immunophenotype: PER-VAS cells were positively labelled for CD5, CD45, MHC II and TLR3, and were negative for CD3, CD4 and CD8 expression. Although unstable along the culture process, IL-17 and MMP12 proteins were detectable as late as at passages 280 and 325i.e. respectively 24 and 29 months post isolation. At passage 325, PER-VAS cells maintained the expression of IL-17, CD3, CD56, IFNγ and TNFα mRNAs as shown by RT-PCR analysis. Stable rearrangement of the TCRγ gene has been evidenced by PCR. PER-VAS cells have a high proliferation index with a doubling time of 16.5h and were tumorigenic in Nude mice. Compared to the canine cell lines already reported, PER-VAS cells display an original expression pattern, close to NKT cells, which makes them valuable tools for in vitro comparative research on lymphomas.

Composite tissue allotransplantation in newborns: a swine model.

BACKGROUND: Management of congenital limb aplasia or facial malformations could be improved by composite tissue allotransplantation (CTA), a technique that has never been performed in newborns. For this, however, the induction of donor-specific tolerance would be mandatory, as long-term immunosuppression is not acceptable in this non-lifesaving procedure. Induction of tolerance has been shown to be possible in a newborn CTA rat model but has never been tested in large-animal models. Our goals were to establish a model of CTA in newborn swine to see if tolerance could be obtained without immunosuppression and to assess rejection or tolerance properties via clinical and histologic examinations. MATERIALS AND METHODS: We applied a CTA heterotopic knee swine model. We performed two series of surgical procedures: Series 1 was 20 autografts in 6-day-old (1-10) 2,544 kg (1,140-4,060 kg) piglets; Series 2 was 10 allografts without immunosuppression between outbred animals aged 7.8 d (6-10) and weighing 2,770 kg (2,200-3,550 kg). RESULTS: In Series 1, six early deaths and two cases of vascular failure were observed. In Series 2, no spontaneous deaths were observed and all piglets presented clinical and histologic rejection. CONCLUSIONS: Our findings strongly suggest that newborn immunologic status is not sufficient for the development of tolerance in large animals without immunologic intervention. Complications and animal death after transplantation correlate with age and weight. Low rates for both vascular failure and postoperative death permit the use of this model in piglets weighing over 2 kg and aged more than 6 d for research on newborn CTA.