Immune response to C. novyi-NT immunotherapy.

Clostridium novyi-NT (CVN-NT) spores germinate in hypoxic regions of tumors and have successfully cured induced neoplasia in mouse models and resulted in objective tumor responses in naturally developing neoplasia in the dog. The objective of this pilot, descriptive, prospective, clinical investigation, was to evaluate and describe the immune response to CNV-NT spores to better understand which immune pathways might play a role in the response to this bacteriolytic immunotherapy. Intratumoral injection of CNV-NT spores result in increased phagocytosis and NK cell-like function after treatment. Intravenous injection of CNV-NT spores resulted in increased LPS-induced TNF-α production, LTA-induced IL-10 production and NK cell-like function post-treatment. Increased NK cell-like function was sustained to 28 (intratumoral) or 56 (intravenous) days post-treatment, and increased phagocytic function was sustained to 28 days post-treatment suggesting that CNV-NT spores induce longer-term immune cell function changes. Future investigations evaluating long-term immune system changes and associations between immune function and tumor remission rates should include evaluation of these pathways.

Identification of immunologic and clinical characteristics that predict inflammatory response to C. Novyi-NT bacteriolytic immunotherapy.

BACKGROUND: Clostridium novyi-NT (CNV-NT), has shown promise as a bacterolytic therapy for solid tumors in mouse models and in dogs with naturally developing neoplasia. Factors that impact the immunologic response to therapy are largely unknown. The goal of this pilot study was to determine if plasma immune biomarkers, immune cell function, peripheral blood cytological composition and tumor characteristics including evaluation of a PET imaging surrogate of tumor tissue hypoxia could predict which dogs with naturally developing naïve neoplasia would develop an inflammatory response to CNV-NT. RESULTS: Dogs that developed an inflammatory response to CNV-NT had a higher heart rate, larger gross tumor volume, greater tumor [64Cu]ATSM SUVMax, increased constitutive leukocyte IL-10 production, more robust NK cell-like function and greater peripheral blood lymphocyte counts compared to dogs that did not develop an inflammatory response to CNV-NT. Of these, unstimulated leukocyte IL-10 production, heart rate, and gross tumor volume appeared to be the best predictors of which dogs will develop an inflammatory response to CNV-NT. CONCLUSIONS: Development of inflammation in response to CNV-NT is best predicted by pretreatment unstimulated leukocyte IL-10 production, heart rate, and gross tumor volume.

Evaluation of 1-(2-deoxy-2-fluoro-1-D-arabinofuranosyl)-5-iodouracil (FIAU) as an ex vivo bacterial detection agent.

The nucleoside analogue, 1-(2-deoxy-2-fluoro-1-D-arabinofuranosyl)-5-iodouracil (FIAU) is a substrate for thymidine kinase (TK), which is commonly expressed in bacteria. It is currently being investigated in clinical studies as an in vivo bacterial infection detection agent. In developing countries where imaging facilities are not readily available, deploying such technology can be a big hurdle. However, a portable ex vivo system might provide a good alternative. In an in vitro system, [(125)I]-FIAU incubated with bacteria is phosphorylated by TK, and is trapped within the bacteria, which can be detected by radioscintography. The suitability of this agent to be utilized as part of an ex vivo bacterial detection system was evaluated. In the first part of this report, the optimization of the incubation and detection condition using E. coli as a test case is described. Samples were incubated in a growth promoting medium containing the label, then after filtering and washing, the amount of radioactivity trapped on the filter was quantitated by a scintillation counter. As a proof of concept demonstration, blinded urine samples from urinary tract infection (UTI) patients and normal donors were tested in the FIAU system. Of the 13 UTI positive and 15 normal urine samples tested, there were 2 false negatives and 1 false positive, respectively. Potential explanations for the false positive and negatives as well as the commercialization possibility of this system will be discussed.

Nanoparticles for improved local retention after intra-articular injection into the knee joint.

PURPOSE: To evaluate using cationic polymeric nanoparticles that interact with hyaluronate to form ionically cross-linked hydrogels to increase the intra-articular retention time of osteoarthritis drugs in the synovial cavity. METHODS: In vitro tests included nanoparticle release from cross-linked hydrogels using syringe and membrane dissolution tests, viscosity measurement of synovial fluid containing hydrogels, and release-rate measurement for a model active conjugated to a cationically substituted dextran using a hydrolyzable ester linkage in a sink dissolution test. Nanoparticle retention after intra-articular injection into rat knees was measured in vivo using fluorescence molecular tomography. RESULTS: Diffusional and convective transport of cationic nanoparticles from ionically cross-linked hydrogels formed in synovial fluid was slower in vitro than for uncharged nanoparticles. Hydrogels formed after the nanoparticles were mixed with synovial fluid did not appreciably alter the viscosity of the synovial fluid in vitro. In vitro release of a conjugated peptide from the cationic nanoparticles was approximately 20% per week. After intra-articular injection in rat knees, 70% of the nanoparticles were retained in the joint for 1 week. CONCLUSIONS: This study demonstrates the feasibility of using cationic polymeric nanoparticles to increase the retention of therapeutic agents in articular joints for indications such as osteoarthritis.

Novel anti-inflammatory effects of doxazosin in rodent models of inflammation.

BACKGROUND: Doxazosin is an α(1)-adrenergic receptor antagonist for the treatment of high blood pressure and benign prostatic hyperplasia. Peripheral α-adrenergic receptors have been implicated in inflammation. AIM: To examine the anti-inflammatory effects of doxazosin in rodent models of inflammation. METHOD: The anti-inflammatory properties of doxazosin were investigated in 4 models. In all studies, drug treatment was administered 15 min prior to challenge. In the lipopolysaccharide (LPS)-induced systemic inflammation model, LPS was injected systemically at 0.25 mg/kg. At 90 min after challenge, blood samples were collected for analysis. In the LPS-induced pulmonary inflammation model, LPS was instilled intranasally. Four hours after challenge, the lungs were harvested for monocyte chemoattractant protein-1 (MCP-1) analysis. In a delayed-type hypersensitivity model, the mice were injected intravenously with sheep red blood cells, and rechallenged in the left footpad 7 days later. Drug treatment was given on day 6 and 7 just prior to the rechallenge. The thickness of hind footpads was measured at 15 min after rechallenge. In the thioglycollate-induced peritoneal monocyte infiltration model, mice were challenged with 3% thioglycollate, and 2 h later peritoneal lavage fluid was collected for MCP-1 analysis. RESULTS: In animals challenged systemically and intranasally with LPS, doxazosin inhibited TNF-α and MCP-1 production, respectively. In the delayed-type hypersensitivity model, footpad swelling was inhibited by doxazosin. Doxazosin decreased the level of MCP-1 release in the peritoneal cavity of thioglycollate-stimulated animals, though this effect was not statistically significant. CONCLUSION: This is the first set of studies that reports the novel anti-inflammatory effects of doxazosin.

Possible therapeutic effect of trilostane in rodent models of inflammation and nociception.

BACKGROUND: Trilostane was identified in an in vivo screen of compounds in a lipopolysaccharide model of inflammation to support a repurposing effort. There is no previous documentation of any anti-inflammatory effects of trilostane. OBJECTIVE: The aim of this study was to elucidate the novel pharmacologic activity of trilostane in a series of inflammation and nociception signal-finding models. METHODS: Anti-inflammatory effects of trilostane were evaluated in lipopolysaccharide-induced systemic and lung inflammation models and in a 2,4-dinitrofluorobenzene-induced delayed-type hypersensitivity (DTH) model in the mouse ear. The analgesic activities of trilostane were evaluated in a hot plate nociception model as a function of paw-withdrawal latency and in the formalin-induced nociception model with a behavioral end point. In all studies, trilostane was administered 15 minutes before challenge. In the DTH model, the animals were given a second dose 24 hours after the first dose. RESULTS: Trilostane inhibited tumor necrosis factor-α and monocyte chemoattractant protein-1 production in the lipopolysaccharide-induced systemic and pulmonary inflammation models. It also significantly reduced ear swelling in the 2,4-dinitrofluorobenzene-induced DTH model. In the hot plate nociception model, trilostane increased the latency of paw-licking behavior. Trilostane also significantly reduced the duration of pain behaviors in the late phase of the formalin-induced inflammatory pain model. CONCLUSIONS: These signal-finding studies suggest that trilostane has novel anti-inflammatory and analgesic properties.

Differential effects of cyclosporin and etanercept treatment on various pathologic parameters in a murine model of irradiation-induced mucositis.

BACKGROUND: Radiation therapy is the most prescribed treatment for many oncologic indications. One of its common side effects is mucositis with hallmark apoptosis in the intestinal crypt and diarrhea. OBJECTIVE: We investigated the potential beneficial effects of etanercept and cyclosporin treatment during radiation exposure. The effects of these drugs on intestinal apoptosis, long-term weight loss, diarrhea severity, and survival were examined. METHODS: For acute observation studies, animals pretreated with phosphate buffer saline (PBS) vehicle, either etanercept, or cyclosporin were challenged with either 1 Gy or 13 Gy irradiation and sacrificed 6 hours later. The animals' small intestines were then harvested for histologic analysis. For chronic survival studies, 14.5 Gy irradiation was applied. Etanercept or cyclosporin treatments were given 15 minutes before the irradiation, followed by daily administration. RESULTS: At 6 hours postirradiation the maximum apoptotic index observed in the small intestine was ∼25% for both 1 Gy and 13 Gy irradiation. Etanercept and cyclosporin pretreatment had no effect on the irradiation-induced apoptosis. During chronic observation, the rate of weight loss was similar in all test groups. At 7 days postirradiation, the weight loss in phosphate buffered saline-treated control, etanercept, and cyclosporin groups reached a maximum at 19%, 24%, and 31.8%, respectively. The weight lost in the cyclosporin group was significantly higher than in the control group. Neither treatment reduced the severity of diarrhea, but cyclosporin increased the survival rate. Sixty percent of cyclosporin-treated animals survived compared with 27% in the PBS-treated control group and 47% in the etanercept-treated group. Serum tumor necrosis factor-α levels, a biomarker for both etanercept's mechanism of action and treatment efficacy, was inhibited by etanercept throughout the study, but cyclosporin only showed an inhibitory effect at 48 hours postirradiation. CONCLUSIONS: Our study demonstrates that cyclosporin increases the survival rate of irradiated animals without affecting parameters such as intestinal histology, weight loss, and diarrhea severity.

Paired 18F-Fluorodeoxyglucose (18F-FDG), and 64Cu-Copper(II)-diacetyl-bis(N(4)-methylthiosemicarbazone) (64Cu-ATSM) PET Scans in Dogs with Spontaneous Tumors and Evaluation for Hypoxia-Directed Therapy.

Hypoxia is associated with neoplastic tissue, protecting cancer cells from death by irradiation and chemotherapy. Identification of hypoxic volume of tumors could optimize patient selection for hypoxia-directed medical, immunological, and radiation therapies. Clostridium novyi-NT (CNV-NT) is an oncolytic bacterium derived from attenuated wild-type Clostridium novyi spores, which germinates exclusively in the anaerobic core of tumors with low-oxygen content. The hypothesis was that 64Cu-ATSM would localize to regions of hypoxia, and that greater hypoxic volume would result in greater germination of Clostridium novyi-NT (CNV-NT). Tumor-bearing companion dogs were recruited to a veterinary clinical trial. Dogs received a CT scan, 18F-FDG PET scan (74 MBq) and 64Cu-ATSM PET scan (74 MBq). Scan regions of interest were defined as the highest 20% of counts/voxel for each PET scan, and regions with voxels overlapping between the two scans. Maximum standardized uptake value (MaxSUV) and threshold volume were calculated. Direct oximetry was performed in select tumors. Tumor types evaluated included nerve sheath tumor (10), apocrine carcinoma (1), melanoma (3) and oral sarcoma (6). MaxSUVATSM ranged from 0.3-6.6. Measured oxygen tension ranged from 0.05-89.9 mmHg. Inverse of MaxSUVATSM had a linear relationship with oxygen tension (R2 = 0.53, P = 0.0048). Hypoxia <8 mmHg was associated with an SUVATSM > 1.0. Hypoxic volume ranged from 0 to 100% of gross tumor volume (GTV) and MaxSUVATSM was positively correlated with hypoxic volume (R = 0.674; P = 0.0001), but not GTV (P = 0.182). Tumor hypoxic volume was heterogeneous in location and distribution. 64Cu-ATSM-avid regions were associated with differential CT attenuation. Hypoxic volume did not predict CNV-NT germination. 64Cu-ATSM PET scanning predicts hypoxia patterns within spontaneously occurring tumors of dogs as measured by direct oxymetry. Total tumor volume does not accurately predict degree or proportion of tumor hypoxia.

Novel anti-inflammatory effects of repaglinide in rodent models of inflammation.

BACKGROUND: Repaglinide is an FDA-approved treatment for type 2 diabetes mellitus. The anti-inflammatory effect of repaglinide in the absence of diabetes has not been reported previously. It is the objective of this set of studies to investigate the potential anti-inflammatory effects of repaglinide. METHOD: The in vivo anti-inflammatory effects of repaglinide were studied in two different models of delay type hyperreactivity (DTH) response induced by sheep red blood cells (sRBC) and 2,5'-dinitrofluorobenzene (DNFB), and in two different rodent models of lipopolysaccharide (LPS) challenge. RESULTS: In mice systemically sensitized with sRBC, which subsequently received a local injection of sRBC in the footpad, local swelling occurred within 24 h after challenge. Repaglinide was efficacious in attenuating this response. In an orthogonal DTH model using DNFB as the antigen, the animals received topical sensitization with DNFB on their shaved backs, followed by topical challenge on the left ears. Repaglinide efficaciously downregulated the resulting ear swelling response. In mice challenged systemically or intratracheally with LPS, repaglinide significantly decreased serum tumor necrosis factor α level and bronchial alveolar lavage fluid MCP-1 levels, respectively. CONCLUSION: This set of data suggests novel anti-inflammatory effects of repaglinide in nondiabetic animals. However, the high dose required for an efficacious effect would make this application impractical in the clinic.

Anti-inflammatory and immunomodulatory effects of bortezomib in various in vivo models.

Bortezomib (Velcade®) is a proteasome inhibitor that has been approved for the treatment of multiple myeloma and mantle cell lymphoma. It has been shown to inhibit the expression of cell adhesion molecules, co-stimulatory molecules, and NFκB activation, to deplete alloreactive T lymphocytes, and to decrease Th1 cytokine production. The anti-inflammatory effects of bortezomib were further investigated in this current set of studies. Systemic treatment with bortezomib was efficacious in the thioglycolate-induced MCP-1 production model, and the dinitrofluorobenzene-induced delayed-type hypersensitivity model. Psoriasis is an autoimmune disease that affects about 2% of the world population. Many treatments have been reported with varying degrees of efficacy. A topical bortezomib formulation was developed to minimize systemic exposure. Its tolerability was investigated in a topical imiquimod (IMQ)-induced psoriasis model. Daily application of IMQ on mouse skin induced inflamed scaly skin lesions resembling plaque-type psoriasis. Fatality was observed in the 1-mg/ml dose group. At 0.1 and 0.01 mg/ml, bortezomib potentiated IMQ-induced erythema, scaling, skin thickening, and caused necrotic lesions. Lower doses had no effect on the clinical observations. Histologically, bortezomib dose-dependently increased parakeratosis, hyperkeratosis, acanthosis, and inflammatory cell infiltration. This study demonstrated that topical bortezomib is not suitable for the treatment of psoriasis.

In vivo effects of immunomodulators in a murine model of Fluorouracil-induced mucositis.

BACKGROUND: Fluorouracil (5-FU) is a pyrimidine analogue used as a cancer treatment. Its toxic side effects, including mucositis, are reported to occur in 40% of the treated patients. Because of the inflammatory component of mucositis, we explored the possibility of modulating this condition with an immunomodulatory agent and a tumor necrosis factor-α inhibitor. OBJECTIVE: The aim of this study was to evaluate the effect of 2 immunosuppressive agents, etanercept and cyclosporine, in a murine model of 5-FU-induced mucositis. METHODS: To study the short-term effects of 5-FU on mucositis, cyclosporine and etanercept were administered to mice after an injection of 5-FU. The animals (n = 8) were euthanized at 6 hours post-challenge. Hematoxylin and eosin-stained histologic sections of the small intestine were examined for signs of apoptosis. To further examine the potential of cyclosporine in the treatment of 5-FU-induced mucositis in a longer duration, the animals (N = 15) were given 2 challenges of 5-FU within 6 hours. All mice were dosed daily until day 9 with either cyclosporine (100 mg/kg) or phosphate-buffered saline (PBS). RESULTS: Six hours after 5-FU challenge, 25 mg/kg etanercept and 50 mg/kg cyclosporine had no effect on 5-FU-induced apoptosis (P > 0.05). However, 100 mg/kg cyclosporine significantly reduced the cumulative level of apoptosis >41.6% of the intestinal crypt surface (P < 0.05). During long-term observation, all mice began to lose weight at a rate of approximately 0.8 g/day after 5-FU exposure. The rates of weight loss and survival were not affected by cyclosporine treatment. The diarrhea onset began on day 4 with 46.7% of the PBS-treated mice showing signs of diarrhea compared with 53.3% in the cyclosporine group. The diarrhea score for both groups plateaued on day 7, with a cumulative score of 41 for the PBS group and 50 for the cyclosporine group. Cyclosporine treatment did not affect the diarrhea onset day or severity compared with the PBS-treated group (P > 0.05). CONCLUSIONS: Our data indicated that etanercept is not a suitable treatment for 5-FU-induced mucositis. Despite decreased apoptosis in the gut, cyclosporine did not affect the severity of the diarrhea or survival. Therefore, we concluded that cyclosporine treatment was only effective in mediating the short-term apoptotic events in the intestines but has no long-term effect on the animals' survival and diarrhea.

Intratumoral Injection of Clostridium novyi-NT Spores in Patients with Treatment-refractory Advanced Solid Tumors.

PURPOSE: Intratumorally injected Clostridium novyi-NT (nontoxic; lacking the alpha toxin), an attenuated strain of C. novyi, replicates within hypoxic tumor regions resulting in tumor-confined cell lysis and inflammatory response in animals, which warrants clinical investigation. PATIENTS AND METHODS: This first-in-human study (NCT01924689) enrolled patients with injectable, treatment-refractory solid tumors to receive a single intratumoral injection of C. novyi-NT across 6 dose cohorts (1 × 104 to 3 × 106 spores, 3+3 dose-escalation design) to determine dose-limiting toxicities (DLT), and the maximum tolerated dose. RESULTS: Among 24 patients, a single intratumoral injection of C. novyi-NT led to bacterial spores germination and the resultant lysis of injected tumor masses in 10 patients (42%) across all doses. The cohort 5 dose (1 × 106 spores) was defined as the maximum tolerated dose; DLTs were grade 4 sepsis (n = 2) and grade 4 gas gangrene (n = 1), all occurring in three patients with injected tumors >8 cm. Other treatment-related grade ≥3 toxicities included pathologic fracture (n = 1), limb abscess (n = 1), soft-tissue infection (n = 1), respiratory insufficiency (n = 1), and rash (n = 1), which occurred across four patients. Of 22 evaluable patients, nine (41%) had a decrease in size of the injected tumor and 19 (86%) had stable disease as the best overall response in injected and noninjected lesions combined. C. novyi-NT injection elicited a transient systemic cytokine response and enhanced systemic tumor-specific T-cell responses. CONCLUSIONS: Single intratumoral injection of C. novyi-NT is feasible. Toxicities can be significant but manageable. Signals of antitumor activity and the host immune response support additional studies of C. novyi-NT in humans.

Pharmacokinetic/pharmacodynamic (PK/PD) differentiation of native and PEGylated recombinant human growth hormone (rhGH and PEG-rhGH) in the rat model of osteoarthritis.

Osteoarthritis (OA) is a degenerative joint disease that has no FDA-approved treatment. The current standard of care does not address the regeneration of the damaged cartilage. Human growth hormone (hGH) is part of the insulin-like growth factor (IGF)-1 axis. There has been preclinical data that suggest its potential regenerative property in the joint. However, unformulated recombinant hGH (rhGH) is short-lived in the joint, and does not provide a desirable pharmacokinetic (PK) profile to support a clinical treatment paradigm. Polyethylene glycol (PEG)ylation is a potential method to extend the half-life of rhGH in the joint. The purpose of this study was to delineate the PK/PD profile of PEG-rhGH in the knee joint in a rat preclinical model of OA. After intra-articular (IA) injection of 100 microg into a rat knee joint that underwent medial meniscectomy, PEG-rhGH exhibits 2-fold longer half-lives in joint than native hGH. However, PEG-rhGH has a much longer systemic exposure. IA injections of PEG-rhGH also resulted in higher levels of IGF-1 in the joint and serum when compared with native rhGH. In order to develop PEG-rhGH as an IA therapeutic treatment for OA, careful dose selection is necessary to avoid systemic effects while retaining its anabolic efficacy in the joint.

Regional citrate anticoagulation for PrismaFlex continuous renal replacement therapy.

BACKGROUND: Since Mehta et al. reported the first successful use of regional citrate anticoagulation (RCA) for continuous renal replacement therapy (CRRT) in 1990, RCA is increasingly used for CRRT because it provides filter patency with minimal risk of bleeding. However, RCA has been associated with significant metabolic complications including hypocalcemia, hypernatremia, metabolic alkalosis, and citrate toxicity. OBJECTIVE: To describe our experience with a newly implemented RCA protocol with acid citrate dextrose formula A (ACD-A) and intravenous calcium gluconate, for use with PrismaFlex CRRT in critically ill patients with acute kidney injury. METHODS: A retrospective chart review was conducted from May 1, 2006, until May 1, 2007, in a 16-bed medical-surgical university-affiliated intensive care unit. Data collected included dialysis filter life, patient and circuit metabolic parameters, and units of packed red blood cells transfused. RESULTS: Forty-eight patients received dialysis with citrate (n = 178 filters). Circuit clotting occurred in 24% of all filters. Mean +/- SD filter life was 38.4 +/- 25.9 hours, and filter survival at 48 hours was 38.2%. Persistent metabolic alkalosis while on CRRT was identified in 6 of 45 (13.3%) patients. Mild hypocalcemia (ionized calcium <3.6 mg/dL) occurred in 11 (23%) patients, but no patient had an ionized calcium level less than 2.8 mg/dL. Six patients, 3 with acute leukemia, required transfusion of 2 or more units of packed red blood cells in 24 hours. CONCLUSIONS: We found that anticoagulation of PrismaFlex CRRT with ACD-A and intravenous calcium gluconate provided reasonable filter patency, but with minor metabolic complications. Close monitoring of electrolyte and acid-base balance is required to minimize metabolic derangements.

Beneficial effect of insulin-like growth factor-1 on hypoxemic renal dysfunction in the newborn rabbit.

Acute normocapnic hypoxemia can cause functional renal insufficiency by increasing renal vascular resistance (RVR), leading to renal hypoperfusion and decreased glomerular filtration rate (GFR). Insulin-like growth factor 1 (IGF-1) activity is low in fetuses and newborns and further decreases during hypoxia. IGF-1 administration to humans and adult animals induces pre- and postglomerular vasodilation, thereby increasing GFR and renal blood flow (RBF). A potential protective effect of IGF-1 on renal function was evaluated in newborn rabbits with hypoxemia-induced renal insufficiency. Renal function and hemodynamic parameters were assessed in 17 anesthetized and mechanically ventilated newborn rabbits. After hypoxemia stabilization, saline solution (time control) or IGF-1 (1 mg/kg) was given as an intravenous (i.v.) bolus, and renal function was determined for six 30-min periods. Normocapnic hypoxemia significantly increased RVR (+16%), leading to decreased GFR (-14%), RBF (-19%) and diuresis (-12%), with an increased filtration fraction (FF). Saline solution resulted in a worsening of parameters affected by hypoxemia. Contrarily, although mean blood pressure decreased slightly but significantly, IGF-1 prevented a further increase in RVR, with subsequent improvement of GFR, RBF and diuresis. FF indicated relative postglomerular vasodilation. Although hypoxemia-induced acute renal failure was not completely prevented, IGF-1 elicited efferent vasodilation, thereby precluding a further decline in renal function.

Specific down-regulation of connective tissue growth factor attenuates progression of nephropathy in mouse models of type 1 and type 2 diabetes.

Diabetic nephropathy (DN) remains a major complication in both type 1 and type 2 diabetes. Systemic administration of antitransforming growth factor-beta (TGF-beta) antibody has shown some promise in mouse models of DN. However, chronic blockade of the multifunctional TGB-beta could be problematic. Several downstream effects of TGF-beta are mediated by connective tissue growth factor (CTGF), which is up-regulated in several renal cells and secreted in the urine in the diabetic state. Using murine models of DN (type 1 and type 2) and a CTGF antisense oligonucleotide (ASO) of novel chimeric chemistry, we evaluated the specific role of this target in DN. In the type 1 model of DN, C57BL6 mice were made diabetic using streptozotocin injections and hyperglycemic animals were treated with CTGF ASOs (20 mg/kg/2 qw) for 4 months. ASO, but not mismatch control oligonucleotide, -treated animals showed significant reduction in target CTGF expression in the kidney with a concomitant decrease in proteinuria and albuminuria. Treatment with the CTGF ASO for 8 wk reduced serum creatinine and attenuated urinary albuminuria and proteinuria in diabetic db/db mice, a model of type 2 DN. The ASO also reduced expression of genes involved in matrix expansion such as fibronectin and collagen (I and IV) and an inhibitor of matrix degradation, PAI-1, in the renal cortex, contributing to significant reversal of mesangial expansion in both models of DN. Pathway analyses demonstrated that diabetes-induced phosphorylation of p38 MAPK and its downstream target CREB was also inhibited by the ASO. Our results strongly suggest that blocking CTGF using a chimeric ASO holds substantial promise for the treatment of DN.

Cardiopulmonary resuscitation in prone position: a simplified method for outpatients.

BACKGROUND: The efficacy of cardiopulmonary resuscitation (CPR) is vital for saving lives of victims with sudden cardiac arrest. In 1960, Kuowenhoven and colleagues proposed the method that has become standard for CPR. Despite vast input of resources for public education and training of this procedure, its success rate outside hospitals remains poor to dismal. During CPR, restoration of respiration is as important as circulation. But opening the airway and giving effective mouth-to-mouth respiration is difficult for lay people to learn. Furthermore, most bystanders are reluctant to do mouth-to-mouth respiration because of the risk of infection. Therefore, the general population needs a more simplified CPR method for outpatients. The practice of CPR in the prone position, first proposed by McNeil in 1989, has not been adopted, despite the fact that it meets the desirable requirements of ideal resuscitation: simultaneous restoration of circulation and respiration with a very simple maneuver. METHODS: Part 1 (circulation test): Eleven patients who expired in the intensive care unit (ICU), with arterial lines attached, received standard pre-cordial cardiac massage, and the generated blood pressure (BP) was recorded. They were then turned to the prone position, with the head turned to one side. We compressed the patient's thoracic spine with the same force used in standard CPR (rhythm of approximately 60 per minute each time when the back bounces back), and the BP was also recorded. Part 2 (ventilation test): Ten healthy volunteers (5 doctors and 5 nurses) were enlisted for respiratory assessment during compression on the back. With the nose clipped and spontaneous breathing held, the volunteer's exhaled tidal volume upon compression was measured with a spirometer. RESULTS: Standard external cardiac massage of the cadavers generated BPs of 55 +/- 20/13 +/- 7 mmHg; however, external compression on the back of the cadavers generated higher BP of 79 +/- 20/17 +/- 10 mmHg (p = 0.028, Wilcoxon signed-rank analysis). External compression on the back of the volunteers generated mean tidal volumes of 399 +/- 110 mL. CONCLUSION: Our study revealed that prone CPR provides good respiratory and circulatory support at the same time. It is easy to perform and it may be a good alternative way for bystanders to perform CPR in public surroundings. We recommend that more investigators do further studies on this topic.

Colchicine overdose with coingestion of nonsteroidal antiinflammatory drugs.

Colchicine has a low therapeutic index. Its toxic effects generally occur at doses ≥ 0.5 mg/kg. We present the case of a 39-year-old female with toxicity following ingestion of 0.28 mg/kg. The patient presented to the emergency department (ED) with severe nausea, vomiting, and abdominal pain following an intentional multidrug ingestion that included colchicine, indomethacin, and zopiclone. Despite toxicologic management and supportive care, admission to the intensive care unit was required for clinical deterioration and symptom management. Shock and multiorgan failure resulted, with death occurring 52 hours postingestion. Although the toxic effects of colchicine are well documented, mortality caused by low doses is relatively uncommon. Management of toxicity consists of early diagnosis, decontamination, and supportive measures. Toxicity may be enhanced by drug interactions inhibiting metabolic enzymes or poor excretion due to renal failure. In this case, the ingestion of a nonsteroidal antiinflammatory drug and the associated volume depletion from the gastrointestinal effects of colchicine may have contributed to renal dysfunction, exacerbating the toxicity of colchicine. This ingestion of a relatively small dose of colchicine led to severe toxicity. Treatment options for colchicine toxicity are limited.

Intratumoral injection of Clostridium novyi-NT spores induces antitumor responses.

Species of Clostridium bacteria are notable for their ability to lyse tumor cells growing in hypoxic environments. We show that an attenuated strain of Clostridium novyi (C. novyi-NT) induces a microscopically precise, tumor-localized response in a rat orthotopic brain tumor model after intratumoral injection. It is well known, however, that experimental models often do not reliably predict the responses of human patients to therapeutic agents. We therefore used naturally occurring canine tumors as a translational bridge to human trials. Canine tumors are more like those of humans because they occur in animals with heterogeneous genetic backgrounds, are of host origin, and are due to spontaneous rather than engineered mutations. We found that intratumoral injection of C. novyi-NT spores was well tolerated in companion dogs bearing spontaneous solid tumors, with the most common toxicities being the expected symptoms associated with bacterial infections. Objective responses were observed in 6 of 16 dogs (37.5%), with three complete and three partial responses. On the basis of these encouraging results, we treated a human patient who had an advanced leiomyosarcoma with an intratumoral injection of C. novyi-NT spores. This treatment reduced the tumor within and surrounding the bone. Together, these results show that C. novyi-NT can precisely eradicate neoplastic tissues and suggest that further clinical trials of this agent in selected patients are warranted.

A new vascular ring connector in surgery for aortic dissection.

OBJECTIVE: To improve the surgical results of aortic dissection, we used a novel vascular ring connector for anastomosis. METHODS: The vascular ring connector is a titanic ring used as a stent in the vascular graft to achieve a quick, blood-sealed, and sutureless anastomosis. From November 2007 to December 2008, 19 consecutive patients (age range 36-77 years; 16 male and 3 female) with aortic dissection underwent open surgery. All patients received aortic reconstruction with vascular grafts (including 5 cases of arch replacement). The combined procedures were 5 Bentall and 4 coronary artery bypass graft operations. RESULTS: There were no significant blood leaks from the anastomotic sites. The time required for each anastomosis was 1 to 2 minutes. All patients were discharged uneventfully and are still doing well after a follow-up period of 1 to 12 months. CONCLUSION: The vascular ring connector may improve the early surgical results of aortic dissection by reducing both the time for anastomosis and the risk of bleeding and may be an alternative technique for aortic reconstruction. Its usefulness in the routine treatment of aortic dissection warrants further evaluation.