Combinatorial immune checkpoint blockade increases myocardial expression of NLRP-3 and secretion of H-FABP, NT-Pro-BNP, interleukin-1ß and interleukin-6: biochemical implications in cardio-immuno-oncology.

Background: Immune checkpoint blockade in monotherapy or combinatorial regimens with chemotherapy or radiotherapy have become an integral part of oncology in recent years. Monoclonal antibodies against CTLA-4 or PD-1 or PDL-1 are the most studied ICIs in randomized clinical trials, however, more recently, an anti-LAG3 (Lymphocyte activation gene-3) antibody, Relatlimab, has been approved by FDA in combination with Nivolumab for metastatic melanoma therapy. Moreover, Atezolizumab is actually under study in association with Ipilimumab for therapy of metastatic lung cancer. Myocarditis, vasculitis and endothelitis are rarely observed in these patients on monotherapy, however new combination therapies could expose patients to more adverse cardiovascular events. Methods: Human cardiomyocytes co-cultured with human peripheral blood lymphocytes (hPBMCs) were exposed to monotherapy and combinatorial ICIs (PD-L1 and CTLA-4 or PD-1 and LAG-3 blocking agents, at 100 nM) for 48 h. After treatments, cardiac cell lysis and secretion of biomarkers of cardiotoxicity (H-FABP, troponin-T, BNP, NT-Pro-BNP), NLRP3-inflammasome and Interleukin 1 and 6 were determined through colorimetric and enzymatic assays. Mitochondrial functions were studied in cardiomyocyte cell lysates through quantification of intracellular Ca++, ATP content and NADH:ubiquinone oxidoreductase core subunit S1 (Ndufs1) levels. Histone deacetylases type 4 (HDAC-4) protein levels were also determined in cardiomyocyte cell lysates to study potential epigenetic changes induced by immunotherapy regimens. Results: Both combinations of immune checkpoint inhibitors exert more potent cardiotoxic side effects compared to monotherapies against human cardiac cells co-cultured with human lymphocytes. LDH release from cardiac cells was 43% higher in PD-L1/CTLA-4 blocking agents, and 35.7% higher in PD-1/LAG-3 blocking agents compared to monotherapies. HDAC4 and intracellular Ca++ levels were increased, instead ATP content and Ndufs1 were reduced in myocardial cell lysates (p < 0.001 vs. untreated cells). Troponin-T, BNP, NT-Pro-BNP and H-FABP, were also strongly increased in combination therapy compared to monotherapy regimen. NLRP3 expression, IL-6 and IL-1ß levels were also increased by PDL-1/CTLA-4 and PD-1/LAG-3 combined blocking agents compared to untreated cells and monotherapies. Conclusions: Data of the present study, although in vitro, indicate that combinatorial immune checkpoint blockade, induce a pro- inflammatory phenotype, thus indicating that these therapies should be closely monitored by the multidisciplinary team consisting of oncologists, cardiologists and immunologists.

Cardiotoxicity and pro-inflammatory effects of the immune checkpoint inhibitor Pembrolizumab associated to Trastuzumab.

BACKGROUND: The immunotherapy has revolutionized the world of oncology in the last decades with considerable advantages in terms of overall survival in cancer patients. The association of Pembrolizumab and Trastuzumab was recently proposed in clinical trials for the treatment of Trastuzumab-resistant advanced HER2-positive breast cancer. Although immunotherapies are frequently associated with a wide spectrum of immune-related adverse events, the cardiac toxicity has not been properly studied. PURPOSE: We studied, for the first time, the putative cardiotoxic and pro-inflammatory effects of Pembrolizumab associated to Trastuzumab. METHODS: Cell viability, intracellular calcium quantification and pro-inflammatory studies (analyses of the production of Interleukin 1ß, 6 and 8, the expression of NF-kB and Leukotriene B4) were performed in human fetal cardiomyocytes. Preclinical studies were also performed in C57BL6 mice by analyzing fibrosis and inflammation in heart tissues. RESULTS: The combination of Pembrolizumab and Trastuzumab leads to an increase of the intracellular calcium overload (of 3 times compared to untreated cells) and to a reduction of the cardiomyocytes viability (of 65 and 20-25%, compared to untreated and Pembrolizumab or Trastuzumab treated cells, respectively) indicating cardiotoxic effects. Notably, combination therapy increases the inflammation of cardiomyocytes by enhancing the expression of NF-kB and Interleukins. Moreover, in preclinical models, the association of Pembrolizumab and Trastuzumab increases the Interleukins expression of 40-50% compared to the single treatments; the expression of NF-kB and Leukotriene B4 was also increased. CONCLUSION: Pembrolizumab associated to Trastuzumab leads to strong cardiac pro-inflammatory effects mediated by overexpression of NF-kB and Leukotriene B4 related pathways.

Caudal additives for postoperative pain management in children: S(+)-ketamine and neostigmine.

BACKGROUND: The aim of the present pilot study was to compare the analgesic efficacy of S(+)-ketamine either alone or in combination with neostigmine for caudal blockade in pediatric surgery. METHODS: A total of 40 children were randomly assigned to receive after induction of general anesthesia either caudal S(+)-ketamine 1 mg.kg(-1) (group K, n = 20) or caudal S (+)-ketamine 0.5 mg.kg(-1) plus neostigmine 10 microg.kg(-1) (group KN, n = 20). Anesthesia was maintained with sevoflurane and a laryngeal mask airway (LMA), no additional analgesics were administered. Postoperative pain and sedation were assessed by the Children's Hospital of Eastern Ontario Pain Score and Ramsay scale for 24 h. RESULTS: No statistical difference in duration of analgesia and sedation was found. Mean duration of postoperative analgesia was 18 +/- 9.4 h in group K and 21.8 +/- 6.7 h in group KN. There was a significantly higher incidence of postoperative vomiting after administration of caudal ketamine with neostigmine (30% group KN Vs 0% group K; P < 0.05). CONCLUSIONS: This pilot study demonstrates equianalgesic effects on postoperative pain relief in children with both caudal S(+)-ketamine 1 mg.kg(-1) and caudal S(+)-ketamine 0.5 mg.kg(-1) plus neostigmine 10 microg.kg(-1). Further studies are required to confirm adoption of caudal neostigmine into routine clinical practice.

Caudal analgesia in children: S(+)-ketamine vs S(+)-ketamine plus clonidine.

BACKGROUND: The aim of this study was to evaluate postoperative analgesia provided by caudal S(+)-ketamine and S(+)-ketamine plus clonidine without local anesthetic. METHODS: Forty-four children aged 1-5 years consecutively scheduled for inguinal hernia repair, hydrocele repair or orchidopexy were randomly assigned to receive a caudal injection of either S(+)-ketamine 1 mg x kg(-1) (group K) or S(+)-ketamine 0.5 mg x kg(-1) plus clonidine 1 microg x kg(-1) (group KC). Postoperative analgesia and sedation were evaluated by CHEOPS and Ramsay scale from emergence from general anesthesia for 24 h. RESULTS: No statistical difference was observed between study groups with respect to pain and sedation assessment. A slight trend toward a reduced requirement for rescue analgesia in group KC was observed, although not statistically significant. CONCLUSIONS: Caudal S(+)-ketamine 1 mg x kg(-1) and S(+)-ketamine 0.5 mg x kg(-1) plus clonidine 1 microg x kg(-1) are safe and provide effective postoperative analgesia in children without adverse effects.

Lipid peroxidation, circulating cytokine and endothelin 1 levels in healthy volunteers undergoing hyperbaric oxygenation.

BACKGROUND: The aim of the present study was to evaluate the effects of hyperbaric oxygenation on lipid peroxidation, on the release of circulating cytokines (TNFa, IL6, IL1b) and endothelin-1 (ET1). EXPERIMENTAL DESIGN: single arm, prospective study. SETTING: ICU hyperbaric division of a University Hospital. PATIENTS: fifteen healthy volunteers (10 male and 5 female, mean age 32+/-7 years) studied during hyperbaric oxygenation divided at random into two groups: group A (7 subjects) and group B (8 subjects). INTERVENTIONS: Both groups were consecutively pressurized at 2 atmospheres (2 atm abs) and 2.8 atm abs, with a constant descending rate of 1 m/min; in accordance with the experimental design, group A breathed pure oxygen continuously through facial masks and group B breathed chamber air during pressurization. MEASURES: Twenty millilitres of blood were drawn from all individuals at the following times: 1) basal, before HBO; 2) after 10 min at 2 atm abs; 3) after 10 min at 2.8 atm abs; 4) 30 min after the end of HBO. In all collected samples thiobarbituric reacting substances were evaluated, using the spectrophotometric technique, IL1 TNF and IL6 serum levels by ELISA and endothelin 1 plasma levels by radioimmunoassay. RESULTS: In both groups, TBARS levels showed a twofold increase (p<0.05) in relation to the baseline, during and after hyperbaric oxygenation. Serum IL6 and IL1b values did not significantly change over the study in any of the volunteers. TNFa amounts significantly increased (p<0.05) during HBO, at 2 atm abs and 2.8 atm abs in both groups, with almost twofold increments. ET1 plasma values increased (p<0.05) in all volunteers during and after HBO: at 2 atm abs (range 7 to 24 pg/ml), 2.8 atm abs (range 7 to 19 pg/ml) and 30 min after (range 8 to 17 pg/ml) in relation to baseline (range 4 to 12 pg/ml). All the studied compounds had a similar trend in the two groups. CONCLUSIONS: Hyperbaric oxygenation in healthy volunteers can induce not only lipid peroxidation, but also liberation of compounds such as TNFa and endothelins, no matter whether pure oxygen is breathed or not. These results suggest that the phenomenon behind this release might be leukocyte activation as induced by HBO. The possible role of ET1 in determining vasoconstriction occurring during HBO is also suggested.

Inhaled nitric oxide administration during one-lung ventilation in patients undergoing thoracic surgery.

OBJECTIVE: To evaluate the effects of inhaled nitric oxide (iNO) on hemodynamics and oxygenation during one-lung ventilation (OLV) in the lateral decubitus position in patients undergoing elective thoracic surgery. DESIGN: Prospective study. SETTING: University hospital. PARTICIPANTS: Thirty consecutive patients scheduled for thoracotomy. INTERVENTIONS: Anesthesia consisted of thoracic epidural analgesia combined with general anesthesia (isoflurane, fentanyl, and vecuronium bromide). Systemic and pulmonary circulations were monitored with a radial artery catheter and a pulmonary artery catheter. Inhaled NO, 40 ppm, was administered during OLV, and the inhaled gas mixture was monitored for NO and nitrogen dioxide (NO2). Hemodynamic and oxygenation data were collected before and during inhaled NO administration. MEASUREMENTS AND MAIN RESULTS: Inhaled NO caused a reduction of pulmonary vascular resistance index from 249 +/- 97.6 dyne. sec. cm(-5) to 199.3 +/- 68.9 dyne. sec. cm(-5) (p < 0.05), without effects on systemic hemodynamics or impairment of oxygenation. A stratification of the patients according to values of QS/QT (< 30%, 30% to 44%, > or = 45%), PaO(2)/fraction of inspired oxygen (> or = 200, 100 to 199, < 100), and pulmonary hypertension (mean pulmonary arterial pressure < 24 or > or = 24 mmHg) showed that inhaled NO causes a significant reduction of mean pulmonary artery pressure in patients with pulmonary hypertension, mainly as a result of a reduction of pulmonary vascular resistance index, and improves oxygenation by reducing intrapulmonary shunt in patients with severe hypoxemia during OLV. CONCLUSIONS: Inhaled NO administration neither significantly decreased mean pulmonary arterial pressure in patients with normal pulmonary artery pressure nor improved oxygenation in nonhypoxic patients. Nevertheless, inhaled NO is effective in patients with pulmonary hypertension and hypoxemia during OLV.

Predictors of failure of noninvasive positive pressure ventilation in patients with acute hypoxemic respiratory failure: a multi-center study.

CONTEXT: In patients with hypoxemic acute respiratory failure (ARF), randomized studies have shown noninvasive positive pressure ventilation (NPPV) to be associated with lower rates of endotracheal intubation. In these patients, predictors of NPPV failure are not well characterized. OBJECTIVE: To investigate variables predictive of NPPV failure in patients with hypoxemic ARF. DESIGN: Prospective, multicenter cohort study. SETTING: Eight Intensive Care Units (ICU) in Europe and USA. PATIENTS: Of 5,847 patients admitted between October 1996 and December 1998, 2,770 met criteria for hypoxemic ARF. Of these, 2,416 were already intubated and 354 were eligible for the study. RESULTS: NPPV failed in 30% (108/354) of patients. The highest intubation rate was observed in patients with ARDS (51%) or community-acquired pneumonia (50%). The lowest intubation rate was observed in patients with cardiogenic pulmonary edema (10%) and pulmonary contusion (18%). Multivariate analysis identified age > 40 years (OR 1.72, 95% CI 0.92-3.23), a simplified acute physiologic score (SAPS II) > or = 35 (OR 1.81, 95% CI 1.07-3.06), the presence of ARDS or community-acquired pneumonia (OR 3.75, 95% CI 2.25-6.24), and a PaO2:FiO2 < or = 146 after 1 h of NPPV (OR 2.51, 95% CI 1.45-4.35) as factors independently associated with failure of NPPV. Patients requiring intubation had a longer duration of ICU stay ( P < 0.001), higher rates of ventilator-associated pneumonia and septic complications ( P < 0.001), and a higher ICU mortality ( P < 0.001). CONCLUSIONS: In hypoxemic ARF, NPPV can be successful in selected populations. When patients have a higher severity score, an older age, ARDS or pneumonia, or fail to improve after 1 h of treatment, the risk of failure is higher.

Application of SOFA score to trauma patients. Sequential Organ Failure Assessment.

OBJECTIVE: To assess the ability of the SOFA score (Sequential Organ Failure Assessment) to describe the evolution of organ dysfunction/failure in trauma patients over time in intensive care units (ICU). DESIGN: Retrospective analysis of a prospectively collected database. SETTING: 40 ICUs in 16 countries. PATIENTS: All trauma patients admitted to the ICU in May 1995. MAIN OUTCOME MEASURES AND RESULTS: Incidence of dysfunction/failure of different organs during the first 10 days of stay and the relation between the dysfunction, outcome, and length of stay. Included in the SOFA study were 181 trauma patients (140 males and 41 females). The non-survivors were significantly older than the survivors (51 years+/-20 vs 38+/-16 years, p < 0.05) and had a higher global SOFA score on admission (8+/-4 vs 4+/-3, p < 0.05) and throughout the 10-day stay. On admission, the non-survivors had higher scores for respiratory ( > 3 in 47% of non-survivors vs 17% of survivors), cardiovascular ( > 3 in 24% of non-survivors vs 5.7% of survivors), and neurological systems ( > 4 in 41% of non-survivors vs 16% of survivors); although the trend was maintained over the whole study period, the differences were greater during the first 4-5 days. After the first 4 days, only respiratory dysfunction was significantly related to outcome. A higher SOFA score, admission to the ICU from the same hospital, and the presence of infection on admission were the three major variables associated with a longer length of stay in the ICU (additive regression coefficients: 0.85 days for each SOFA point, 4.4 for admission from the same hospital, 7.26 for infection on admission). CONCLUSIONS: The SOFA score can reliably describe organ dysfunction/ failure in trauma patients. Regular and repeated scoring may be helpful for identifying categories of patients at major risk of prolonged ICU stay or death.