Diffuse intrinsic pontine gliomas (DIPG) at recurrence: is there a window to test new therapies in some patients?

Children with diffuse intrinsic pontine glioma (DIPG) need new and more efficient treatments. They can be developed at relapse or at diagnosis, but therefore they must be combined with radiotherapy. Survival of children after recurrence and its predictors were studied to inform the possibility to design early phase clinical trials for DIPG at this stage. Among 142 DIPG patients treated between 1998 and 2014, 114 had biopsy-proven DIPG with histone H3 status available for 83. We defined as long survivors' patients who survived more than 3 months after relapse which corresponds to the minimal life expectancy requested for phase I/II trials. Factors influencing post-relapse survival were accordingly compared between short and long-term survivors after relapse. Fifty-seven percent of patients were considered long survivors and 70% of them had a Lansky Play Scale (LPS) above 50% at relapse. Patients who became steroids-independent after initial treatment for at least 2 months had better survival after relapse (3.7 versus 2.6 months, p = 0.001). LPS above 50% at relapse was correlated with better survival after relapse (3.8 versus 1.8 months, p < 0.001). Patients with H3.1 mutation survived longer after relapse (4.9 versus 2.7 months, p = 0.007). Patients who received a second radiotherapy at the time of relapse had an improved survival (7.5 versus 4 months, p = 0.001). In the two-way ANOVA analysis, steroid-independence and LPS predicted survival best and the type of histone H3 (H3.1 or H3.3) mutated did not improve prediction. Survival of many DIPG patients after relapse over 3 months would make possible to propose specific trials for this condition. Steroid-independence, H3 mutation status and LPS should be considered to predict eligibility.

Characterization of a porcine model of post-operative pain.

BACKGROUND: Management of acute pain related to surgical intervention, termed post-operative pain or POP, continues to be a major healthcare challenge. While the rat plantar incision model provides valuable data to researchers about the mechanisms mediating POP, the development of topical and localized treatments in small animal models is limited. To help address these issues, we describe here the characterization of a large animal model of incisional pain. METHODS: Pigs underwent full-skin incision or full-skin and muscle incision and retraction (SMIR). Withdrawal thresholds were determined using the Von Frey test at baseline, 0.5-12 h post-surgery and on days 1, 2, 3, 5 and 7 post-surgery. The analgesic effects of systemic morphine [0.1 or 1.0 mg/kg intramuscular (i.m.) dose] and local anaesthetic ropivacaine were studied. Spontaneous pain-like behaviours were scored and analysed. The effects on wound healing were evaluated by gross observation and by histopathological examination. RESULTS: Pigs incurring SMIR demonstrated significantly increased mechanical hypersensitivity compared with pigs that underwent full-skin incision only (p < 0.05). Maximal analgesia was achieved with morphine (1 mg/kg i.m. dose) at 0.5 h post-treatment. Local treatment with ropivacaine was effective at increasing the withdrawal threshold to Von Frey filaments compared with saline control (p < 0.05) for a period of at least 6 h. Wounds healed normally with no signs of infection, redness or swelling. CONCLUSIONS: We propose that the pig model of incisional pain can provide an appropriate translational model for validating new topical and localized treatments for POP in humans.

Induction of an antiinflammatory effect and prevention of cartilage damage in rat knee osteoarthritis by CF101 treatment.

OBJECTIVE: Studies have suggested that rheumatoid arthritis (RA) and osteoarthritis (OA) share common characteristics. The highly selective A(3) adenosine receptor agonist CF101 was recently defined as a potent antiinflammatory agent for the treatment of RA. The purpose of this study was to examine the effects of CF101 on the clinical and pathologic manifestations of OA in an experimental animal model. METHODS: OA was induced in rats by monosodium iodoacetate, and upon disease onset, oral treatment with CF101 (100 microg/kg given twice daily) was initiated. The A(3) adenosine receptor antagonist MRS1220 (100 microg/kg given twice daily) was administered orally, 30 minutes before CF101 treatment. The OA clinical score was monitored by knee diameter measurements and by radiographic analyses. Histologic analyses were performed following staining with hematoxylin and eosin, Safranin O-fast green, or toluidine blue, and histologic changes were scored according to a modified Mankin system. Signaling proteins were assayed by Western blotting; apoptosis was detected via immunohistochemistry and TUNEL analyses. RESULTS: CF101 induced a marked decrease in knee diameter and improved the changes noted on radiographs. Administration of MRS1220 counteracted the effects of CF101. CF101 prevented cartilage damage, osteoclast/osteophyte formation, and bone destruction. In addition, CF101 markedly reduced pannus formation and lymphocyte infiltration. Mechanistically, CF101 induced deregulation of the NF-kappaB signaling pathway, resulting in down-regulation of tumor necrosis factor alpha. Consequently, CF101 induced apoptosis of inflammatory cells that had infiltrated the knee joints; however, it prevented apoptosis of chondrocytes. CONCLUSION: CF101 deregulated the NF-kappaB signaling pathway involved in the pathogenesis of OA. CF101 induced apoptosis of inflammatory cells and acted as a cartilage protective agent, which suggests that it would be a suitable candidate drug for the treatment of OA.

The A3 adenosine receptor agonist CF102 induces apoptosis of hepatocellular carcinoma via de-regulation of the Wnt and NF-kappaB signal transduction pathways.

The A3 adenosine receptor (A(3)AR) is highly expressed in tumors and was suggested as a target for cancer treatment. In this study, we show that A(3)AR is highly expressed in tumor tissues and in peripheral blood mononuclear cells (PBMCs) derived from patients with HCC, as well as from HCC tumor-bearing rats. The high expression level of the receptor was directly correlated to overexpression of NF-kappaB, known as a transcription factor of A(3)AR. CF102, a synthetic highly selective agonist to A(3)AR induced a marked dose response inhibition of tumor growth in N1S1 HCC tumor rats, via de-regulation of the NF-kappaB and the Wnt signal transduction pathways, resulting in apoptosis of tumor cells. Taken together, A(3)AR is highly expressed in tumors and PBMCs of HCC patients and tumor-bearing rats. CF102 induced apoptosis and tumor growth inhibition. These data suggest A(3)AR as a novel targeted therapy to treat HCC.

Tissue-specific gene therapy directed to tumor angiogenesis.

Gene therapy directed specifically to the vascular wall, particularly to angiogenic endothelial cells is a prerequisite in vascular disease treatment. Angiogenesis is a major feature in many pathological conditions including wound healing, solid tumors, developing metastases, ischemic heart diseases and diabetic retinopathy. In the present study we developed a tissue-specific gene therapy to the angiogenic blood vessels of tumor metastasis using an adeno-based vector containing the murine preproendothelin-1 (PPE-1) promoter. Genes activated by the PPE-1 promoter were highly expressed in bovine aortic endothelial cells in vitro. Systemic injection of the adenoviral vectors AdPPE-1-luciferase and AdCMV-luciferase to normal C57BL/6 mice, resulted in higher activity of PPE-1 promoter compared with CMV promoter in the aorta and vascularized tissues such as heart, kidney, lung and pancreas. Systemic administration of the adenoviral vector, in mice bearing Lewis lung carcinoma, resulted in high and specific activity of PPE-1 in the new vasculature of primary tumors and lung metastasis. Cellular distribution of the delivered gene revealed highest expression of GFP in angiogenic endothelial cells of the metastasis. We expect that this approach of 'vascular-directed' gene therapy will be applicable to both vascular diseases and cancer.

Intravenous rFVIIa administered for hemorrhage control in hypothermic coagulopathic swine with grade V liver injuries.

BACKGROUND: Intravenous administration of recombinant activated human clotting factor VII (rFVIIa) has been used successfully to prevent bleeding in hemophilia patients undergoing elective surgery, but not in previously normal trauma patients. This study was conducted to determine whether rFVIIa was a useful adjunct to gauze packing for decreasing blood loss from grade V liver injuries in hypothermic and coagulopathic swine. METHODS: All animals (n = 10, 35 +/- 2 kg) underwent a 60% isovolemic exchange transfusion with 6% hydroxyethyl starch and were cooled to 33 degrees C core temperature. The swine then received a grade V liver injury and 30 seconds later, either 180 microg/kg rFVIIa, or saline control. All animals were gauze packed 30 seconds after injury and resuscitated 5.5 minutes after injury with lactated Ringer's solution to their preinjury mean arterial pressure. Posttreatment blood loss, mean arterial pressure, resuscitation volume, and clotting studies were monitored for 1 hour. Histology of lung, kidney, and small bowel were obtained to evaluate for the presence of microvascular thrombi. RESULTS: At the time of injury, core temperature was 33.3 degrees +/- 0.4 degrees C, hemoglobin was 6 +/- 0.7 g/dL, prothrombin time was 19.1 +/- 1.0 seconds, activated partial thromboplastin time was 29.0 +/- 4.8 seconds, fibrinogen was 91 +/- 20 mg/dL, and platelets were 221 +/- 57 x 105/mL, with no differences between groups (p > 0.05). Clotting factor levels confirmed a coagulopathy at the preinjury point. The posttreatment blood loss was less (p < 0.05) in group 1 (527 +/- 323 mL), than in group 2 (976 +/- 573 mL). The resuscitation volume was not different (p > 0.05). One-hour survival in both groups was 100%. Compared with the control group, rFVIIa increased the circulating levels of VIIa and, despite hypothermia, shortened the prothrombin time 5 minutes after injection (p < 0.05). Laboratory evaluation revealed no systemic activation of the clotting cascade. Postmortem evaluation revealed no evidence of large clots in the hepatic veins or inferior vena cava, or microscopic thrombi in lung, kidney, or small intestine. CONCLUSION: rFVIIa reduced blood loss and restored abnormal coagulation function when used in conjunction with liver packing in hypothermic and coagulopathic swine. No adverse effects were identified.

S-nitrosoderivative of a recombinant fragment of von Willebrand factor (S-nitroso-AR545C) inhibits thrombus formation in guinea pig carotid artery thrombosis model.

Antiplatelet drugs are the mainstays of therapy for acute and chronic cardiovascular diseases. S-nitroso-AR545C - an S-nitrosoderivative of a recombinant von Willebrand factor fragment AR545C spanning Ala 444 to Asp 730 and containing an Arg 545 Cys mutation, was previously found to inhibit ristocetin- and ADP-induced platelet aggregation and the interaction of platelets with extracellular matrix (ECM). In the current study we tested the antithrombotic properties of S-nitroso-AR545C on guinea pig platelets and in a platelet-rich thrombosis model in the guinea pig. Preincubation of guinea pig platelets with 0.1 microM of S-nitroso-AR545C decreased ristocetin-induced agglutination by 40% (p = 0.009) and completely abolished ADP-induced aggregation (p <0.0001). At concentration of 1.0 microM, S-nitroso-AR545C completely inhibited platelet adhesion (represented by surface coverage - SC) and decreased aggregate formation (represented by average aggregate size - AS) by more than 50%. Treatment of guinea pigs with 1.0 mg/kg S-nitroso-AR545C resulted in a significantly delayed time to arterial occlusion (31.7+/-6.0 min vs. 13.9+/-3.2 min, p <0.02). Similarly, total patency time was longer in the group injected with S-nitroso-AR545C compared to the control group. However, the difference was not statistically significant (33.8+/-6.3 min vs. 20.2+/-3.3 min, p = 0.07). No change in platelet count, hematocrit and bleeding time was observed 60 min after injection compared to baseline. In contrast, a significant decrease in SC (p <0.0001) and AS (p <0.01) were observed 60 min after the injection of S-nitroso-AR545C, whereas no change in these parameters was observed in the control group. These observations indicate that S-nitroso-AR545C exhibits significant antiadhesive and antiaggregating effects in-vitro and inhibits clot formation in-vivo suggesting that this compound may have potential therapeutic advantages.

Rate of temperature increase in human muscle during 1 MHz and 3 MHz continuous ultrasound.

To achieve the thermal effects of ultrasound, the tissue temperature must be raised from 1 to > or = 4 degrees C, depending on the desired outcome of the treatment. In the past 25 years, there have been no in vivo studies that have measured rate of change in temperature during 1-MHz ultrasound treatments, and none have ever been performed with the 3-MHz frequency. Thus, we are left to pure speculation regarding how long to administer an ultrasound treatment. We performed this study to plot the rate of temperature increase during ultrasound treatments delivered at various intensities and frequencies. We inserted two 23-gauge thermistors into each subjects' medial triceps surae at the following depths: 1 MHz at depths of 2.5 and 5.0 cm (12 subjects) and 3 MHz at depths of .8 and 1.6 cm (12 subjects). Each subject received a total of four 10-minute treatments, one each at .5, 1.0, 1.5, and 2.0 W/cm2, and temperature was measured every 30 seconds. No significant difference was found in the rate of heating at the two depths (p = .987) within the same frequency and dose levels. The 3-MHz frequency heated significantly faster than the 1-MHz frequency at all doses tested (p < .001). On average, the rate of temperature increased per minute at the two depths of the 1-MHz frequency was: .04 degrees C at .5 W/cm2; .16 degrees C at 1.0 W/cm2; .33 degrees C at 1.5 W/cm2; and .38 degrees C at 2.0 W/cm2. The rate of temperature increase per minute at the two depths of the 3-MHz frequency was: .3 degrees C at .5 W/cm2; .58 degrees C at 1.0 W/cm2; .89 degrees C at 1.5 W/cm2; and 1.4 degrees C at 2.0 W/cm2. The results of this research should enable clinicians to choose the correct frequency, intensity, and treatment time when using thermal ultrasound.