Hugh B. Black

Erythropoietin (Epo)-responsive anemia is a common and debilitating complication of chronic renal failure and human immunodeficiency virus infection. Current therapy for this condition involves repeated intravenous or subcutaneous injections of recombinant Epo. In this report, we describe the development of a novel muscle-based gene transfer approach that produces long-term expression of physiologically significant levels of Epo in the systemic circulation of mice. We have constructed a plasmid expression vector, pVRmEpo, that contains the murine Epo cDNA under the transcriptional control of the cytomegalovirus immediate early (CMV-IE) promoter, the CMV-IE 5' untranslated region, and intron A. A single intramuscular (i.m.) injection of as little as 10 micrograms of this plasmid into immunocompetent adult mice produced physiologically significant elevations in serum Epo levels and increased hematocrits from preinjection levels of 48 +/- 0.4% to levels of 64 +/- 3.3% 45 days after injection. Hematocrits in these animals remained elevated at greater than 60% for at least 90 days after a single i.m. injection of 10 micrograms of pVRmEpo. We observed a dose-response relationship between the amount of plasmid DNA injected and subsequent elevations in hematocrits. Mice injected once with 300 micrograms of pVRmEpo displayed 5-fold increased serum Epo levels and elevated hematocrits of 79 +/- 3.3% at 45 days after injection. The i.m. injected plasmid DNA remained localized to the site of injection as assayed by the PCR. We conclude that i.m. injection of plasmid DNA represents a viable nonviral gene transfer method for the treatment of acquired and inherited serum protein deficiencies.

OBJECTIVE: The current national opioid epidemic is a public health emergency. We have identified an outbreak of exaggerated opioid toxicity caused by fentanyl adulterated tablets purchased on the street as hydrocodone/acetaminophen. METHODS: Over an 8-day period in late March 2016, a total of 18 patients presented to our institution with exaggerated opioid toxicity. The patients provided a similar history: ingesting their "normal dose" of hydrocodone/acetaminophen tablets but with more pronounced symptoms. Toxicology testing and analysis was performed on serum, urine, and surrendered pills. RESULTS: One of the 18 patients died in hospital. Five patients underwent cardiopulmonary resuscitation, one required extracorporeal life support, three required intubation, and two received bag-valve-mask ventilation. One patient had recurrence of toxicity after 8 hours after naloxone discontinuation. Seventeen of 18 patients required boluses of naloxone, and four required prolonged naloxone infusions (26-39 hours). All 18 patients tested positive for fentanyl in the serum. Quantitative assays conducted in 13 of the sera revealed fentanyl concentrations of 7.9 to 162 ng/mL (mean = 52.9 ng/mL). Pill analysis revealed fentanyl amounts of 600-6,900 μg/pill. The pills are virtually indistinguishable from authentic hydrocodone/acetaminophen tablets and are similar in weight. To date, our county has reported 56 cases of fentanyl opioid toxicity, with 15 fatalities. In our institution, the outbreak has stressed the capabilities and resources of the emergency department and intensive care units. CONCLUSIONS: A serious outbreak of exaggerated opioid toxicity caused by fentanyl-adulterated tablets purchased on the street as hydrocodone/acetaminophen is under way in California. These patients required higher dosing and prolonged infusions of naloxone. Additionally, observation periods off naloxone were extended due to delayed, recurrent toxicity. The outbreak has serious ramifications for public health and safety, law enforcement, and healthcare facilities and resources.

Erythropoietin (Epo)-responsive anemia is a debilitating complication of chronic renal failure and human immunodeficiency virus (HIV) infection that effects more than 150,000 Americans. Patients with Epo-responsive anemias are currently treated with repeated injections of recombinant human Epo. In the studies described in this report, we have examined the safety and efficacy of using a single intramuscular (i.m.) injection of replication-defective adenoviral vectors (RDAd) encoding Epo for the treatment of Epo-responsive anemias in both mice and non-human primates. Our results demonstrate that there is a threshold dose of virus (2.5–8 × 107 pfu/gram of body weight) which is required to obtain long-term Epo expression and polycythemia in both species. A single i.m. injection of mice with 109 pfu of an RDAd encoding murine Epo (AdmEpo) resulted in elevations in hematocrits from control values of 49 ± 0.9% to treated values of 81 ± 3%, which were stable for more than 1 year. Similarly, a single i.m. injection of a monkey with 4 × 1011 pfu of an RDAd-encoding simian Epo (AdsEpo) resulted in elevations of hematocrits from control levels of 40% to treated levels of ≥70%, which were stable for 84 days. Intramuscular injection of monkeys with AdsEpo appeared to be safe in that we did not detect abnormalities in chest X-rays, serum chemistries, hematologic, or clotting profiles (apart from elevated hematocrits) or organ histologies during the 84-day time course of the experiment. Taken together, these results suggest the feasibility of using i.m. injection of RDAd for the treatment of Epo-responsive anemias in humans. We have examined the safety and efficacy of using intramuscular (i.m.) injection of E1- and E3-deleted replication defective adenoviruses (RDAd) encoding erythropoietin (Epo) for the treatment of Epo-responsive anemias in both mice and non-human primates. Our results demonstrate that a single injection of adult immunocompetent mice or cynomologous monkeys with 2.5–8.0 × 107 pfu of RDAd/gm body weight results in the expression of physiological levels of erythropoietin in the systemic circulation of both species. Elevated levels of Epo were detected in the serum for at least 1 year following i.m. injection of mice and for at least 84 days following injection of the monkeys. Intramuscular injection of non-human primates with an RdAd-encoding simian Epo appeared to be safe because we did not detect abnormalities in chest X-rays, serum chemistries, hematologic profiles, or organ histologies. These findings suggest that i.m. injection of RDAd may represent an effective therapy for humans with Epo-responsive anemias.

The preferred assay for measuring and adjusting unfractionated heparin (UFH) infusion to achieve optimal outcomes during extracorporeal membrane oxygenation (ECMO) is not well established. This retrospective cohort study explored safety and efficacy outcome differences between anti-factor Xa (anti-Xa) and activated partial thromboplastin time (aPTT) for UFH in adult venoarterial ECMO. Forty-one patients were included and analyzed. The UFH rate at first goal and time to goal were both higher in the aPTT versus anti-Xa cohort but did not achieve statistical significance (12.14 vs. 9.58 unit/kg/hour (p = 0.29), 20.22 vs. 12.05 hours (p = 0.11)). The aPTT cohort was in target goals 35.0% of the time versus 47.7% in the anti-Xa cohort (p = 0.13), above goal 41.0% vs. 17.3% (p = 0.02), and below-goal 24.0% versus 35.0% of the time (p = 0.34). Minimum heparin rates in the aPTT cohort were 6.28 vs. 3.33 unit/kg/hour in the anti-Xa cohort (p = 0.07), and the maximum UFH rate was 18.77 unit/kg/hour vs. 15.48 unit/kg/hour (p = 0.10). Our findings suggest that aPTT monitoring may result in a delay to target attainment, higher UFH rates, and overall exposure.