Long-term safety of intramuscular gene transfer of non-viral FGF1 for peripheral artery disease

Peripheral artery disease is a progressive disease. Primary ischemic leg symptoms are muscle fatigue, discomfort or pain during ambulation, known as intermittent claudication. The most severe manifestation of peripheral artery disease is critical limb ischemia (CLI). The long-term safety of gene therapy in peripheral artery disease remains unclear. This four center peripheral artery disease registry was designed to evaluate the long-term safety of the intramuscular non-viral fibroblast growth factor-1 (NV1FGF), a plasmid-based angiogenic gene for local expression of fibroblast growth factor-1 versus placebo in patients with peripheral artery disease who had been included in five different phase I and II trials. Here we report a 3-year follow-up in patients suffering from CLI or intermittent claudication. There were 93 evaluable patients, 72 of them in Fontaine stage IV (47 NV1FGF versus 25 placebo) and 21 patients in Fontaine stage IIb peripheral artery disease (15 NV1FGF versus 6 placebo). Safety parameters included rates of non-fatal myocardial infarction (MI), stroke, death, cancer, retinopathy and renal dysfunction. At 3 years, in 93 patients included this registry, there was no increase in retinopathy or renal dysfunction associated with delivery of this angiogenic factor. There was also no difference in the number of strokes, MI or deaths, respectively, for NV1FGF versus placebo. In the CLI group, new cancer occurred in two patients in the NV1FGF group. Conclusions that can be drawn from this relatively small patient group are limited because of the number of patients followed and can only be restricted to safety. Yet, data presented may be valuable concerning rates in cancer, retinopathy, MI or strokes following angiogenesis gene therapy in the absence of any long-term data in angiogenesis gene therapy. It may take several years until data from larger patient populations will become available.

Local pain and spreading hyperalgesia induced by intramuscular injection of nerve growth factor are not reduced by local anesthesia of the muscle

Injections with local anesthesia for therapeutic and diagnostic purposes are common clinical practice. This double-blind placebo controlled study explores the rational of local anesthetic blocks for the detection of muscle pain as the primary generator in spreading hyperalgesic conditions.

Acute compartment syndrome complicating an intramuscular haemangiosarcoma in a dog

Acute compartment syndrome in dogs is a rare complication of muscle trauma, but it has not been previously reported as a consequence of neoplasia in dogs. This case report describes the occurrence of a compartment syndrome of the femoral compartment in an 11-year-old, male, mixed-breed dog caused by acute bleeding of an intramuscular haemangiosarcoma. The compartment syndrome was relieved by immediate fasciotomy. The dog was euthanased following acute recurrence of clinical signs seven weeks after surgery.

Anaesthesia for castration of piglets: comparison between intranasal and intramuscular application of ketamine, climazolam and azaperone

The aim of this study was to compare the effect of an anaesthetic combination given either intramuscularly (IM) or intranasally (IN) for castration of piglets. Forty piglets aged 4 to 7 days were randomly assigned to receive a mixture of ketamine 15 mg kg-1, climazolam 1.5 mg kg-1 and azaperone 1.0 mg kg-1, IN or IM, 10 minutes prior to castration. Physiological parameters were measured. Castration was videotaped for evaluation by 3 independent observers using a scoring system. Reaction and vocalization to the skin incision and cutting of spermatic cord was evaluated and scored (0 = no reaction, 16 = strong reaction). The IN group had a significantly higher (P < 0.01) castration score, compared to the IM group. There was an association between castration score and room temperature in the IN group (with temperatures below 18 "C associated with a higher castration scores (P < 0.001). Heart rate was significantly higher 10 minutes after castration in the IN group (P < 0.05). Respiratory rate was significantly higher in the IM group at time points -5, -1, 10, 20 and 30 (P < 0.05).The IN group was walking significantly (P < 0.0001) faster than the IM group. In conclusion, this combination provides effective anaesthesia for routine castration of newborn piglets when administered IM. IN administration provided shorter recovery times but had significantly higher castration scores.

Therapeutic angiogenesis with intramuscular NV1FGF improves amputation-free survival in patients with critical limb ischemia

This study evaluated the efficacy and safety of intramuscular administration of NV1FGF, a plasmid-based angiogenic gene delivery system for local expression of fibroblast growth factor 1 (FGF-1), versus placebo, in patients with critical limb ischemia (CLI). In a double-blind, randomized, placebo-controlled, European, multinational study, 125 patients in whom revascularization was not considered to be a suitable option, presenting with nonhealing ulcer(s), were randomized to receive eight intramuscular injections of placebo or 2.5 ml of NV1FGF at 0.2 mg/ml on days 1, 15, 30, and 45 (total 16 mg: 4 x 4 mg). The primary end point was occurrence of complete healing of at least one ulcer in the treated limb at week 25. Secondary end points included ankle brachial index (ABI), amputation, and death. There were 107 patients eligible for evaluation. Improvements in ulcer healing were similar for use of NV1FGF (19.6%) and placebo (14.3%; P = 0.514). However, the use of NV1FGF significantly reduced (by twofold) the risk of all amputations [hazard ratio (HR) 0.498; P = 0.015] and major amputations (HR 0.371; P = 0.015). Furthermore, there was a trend for reduced risk of death with the use of NV1FGF (HR 0.460; P = 0.105). The adverse event incidence was high, and similar between the groups. In patients with CLI, plasmid-based NV1FGF gene transfer was well tolerated, and resulted in a significantly reduced risk of major amputation when compared with placebo.

Local gene transfer and expression following intramuscular administration of FGF-1 plasmid DNA in patients with critical limb ischemia

NV1FGF is an expression plasmid encoding sp.FGF-1(21-154) currently under investigation for therapeutic angiogenesis in clinical trials. NV1FGF plasmid distribution and transgene expression following intramuscular (IM) injection in patients is unknown. The study involved six patients with chronic critical limb ischemia (CLI) planned to undergo amputation. A total dose of 0.5, 2, or 4 mg NV1FGF was administered as eight IM injections (0.006, 0.25, or 0.5 mg per injection) 3-5 days before amputation. Injected sites (30 cm(3)) were divided into equally sized smaller pieces to assess spatial distribution of NV1FGF sequences (PCR), NV1FGF mRNA (reverse transcriptase-PCR), and fibroblast growth factor-1 (FGF-1)-expressing cells (immunohistochemistry). Data indicated gene expression at all doses. The distribution area was within 5-12 cm for NV1FGF sequences containing the expression cassette, up to 5 cm for NV1FGF mRNA, and up to 3 cm for FGF-1-expressing myofibers. All FGF receptors were detected indicating robust potential for bioactivity after NV1FGF gene transfer. Circulating levels of NV1FGF sequences were shown to decrease within days after injection. Data support demonstration of plasmid-mediated gene transfer and expression in muscles from patients with CLI. FGF-1 expression was shown to be limited to injection sites, which supports the concept of multiple-site injection for therapeutic use.

Combinations of dexmedetomidine and alfaxalone with butorphanol in cats: application of an innovative stepwise optimisation method to identify optimal clinical doses for intramuscular anaesthesia

OBJECTIVES The aim of this study was to optimise dexmedetomidine and alfaxalone dosing, for intramuscular administration with butorphanol, to perform minor surgeries in cats. METHODS Initially, cats were assigned to one of five groups, each composed of six animals and receiving, in addition to 0.3 mg/kg butorphanol intramuscularly, one of the following: (A) 0.005 mg/kg dexmedetomidine, 2 mg/kg alfaxalone; (B) 0.008 mg/kg dexmedetomidine, 1.5 mg/kg alfaxalone; (C) 0.012 mg/kg dexmedetomidine, 1 mg/kg alfaxalone; (D) 0.005 mg/kg dexmedetomidine, 1 mg/kg alfaxalone; and (E) 0.012 mg/kg dexmedetomidine, 2 mg/kg alfaxalone. Thereafter, a modified 'direct search' method, conducted in a stepwise manner, was used to optimise drug dosing. The quality of anaesthesia was evaluated on the basis of composite scores (one for anaesthesia and one for recovery), visual analogue scales and the propofol requirement to suppress spontaneous movements. The medians or means of these variables were used to rank the treatments; 'unsatisfactory' and 'promising' combinations were identified to calculate, through the equation first described by Berenbaum in 1990, new dexmedetomidine and alfaxalone doses to be tested in the next step. At each step, five combinations (one new plus the best previous four) were tested. RESULTS None of the tested combinations resulted in adverse effects. Four steps and 120 animals were necessary to identify the optimal drug combination (0.014 mg/kg dexmedetomidine, 2.5 mg/kg alfaxalone and 0.3 mg/kg butorphanol). CONCLUSIONS AND RELEVANCE The investigated drug mixture, at the doses found with the optimisation method, is suitable for cats undergoing minor clinical procedures.