A protective erythropoietin evolutionary landscape, NLRP3 inflammasome regulation, and multisystem inflammatory syndrome in children.

The low incidence of pediatric severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection and the associated multisystem inflammatory syndrome (MIS-C) lack a unifying pathophysiological explanation, impeding effective prevention and therapy. Activation of the NACHT, LRR, and PYD domains-containing protein (NLRP) 3 inflammasome in SARS-CoV-2 with perturbed regulation in MIS-C, has been reported. We posit that, early age physiological states and genetic determinants, such as certain polymorphisms of renin-angiotensin aldosterone system (RAAS) molecules, promote a controlled RAAS hyperactive state, and form an evolutionary landscape involving an age-dependent erythropoietin (EPO) elevation, mediating ancestral innate immune defenses that, through appropriate NLRP3 regulation, mitigate tissue injury and pathogen invasion. SARS-CoV-2-induced downregulation of angiotensin-converting enzyme (ACE)2 expression in endothelial cells (EC), impairment of endothelial nitric oxide (NO) synthase (eNOS) activity and downstream NO bioavailability, may promote a hyperactive RAAS with elevated angiotensin II and aldosterone that, can trigger, and accelerate NLRP3 inflammasome activation, while EPO-eNOS/NO abrogate it. Young age and a protective EPO evolutionary landscape may successfully inhibit SARS-CoV-2 and contain NLRP3 inflammasome activation. By contrast, increasing age and falling EPO levels, in genetically susceptible children with adverse genetic variants and co-morbidities, may lead to unopposed RAAS hyperactivity, NLRP3 inflammasome dysregulation, severe endotheliitis with pyroptotic cytokine storm, and development of autoantibodies, as already described in MIS-C. Our haplotype estimates, predicted from allele frequencies in population databases, are in concordance with MIS-C incidence reports in Europeans but indicate lower risks for Asians and African Americans. Targeted Mendelian approaches dissecting the influence of relevant genetic variants are needed.

POS-250 A RANDOMIZED TRIAL OF EFFECT OF CHOLECALCIFEROL SUPPLEMENTATION ON CARDIOVASCULAR DISEASE, INFLAMMATION, AND BONE METABOLISM MARKERS IN CKD: BASELINE CHARACTERISTICS OF FEASIBILITY PHASE

Introduction: Studies have linked lower vitamin D levels with cardiovascular disease (CVD) and mortality in general population and chronic kidney disease (CKD). The preliminary evidence of vitamin D supplementation is encouraging but there is a huge void with respect to good quality long term data supporting the use of this promising intervention for translation into better outcomes for CVD in CKD. This study is exploring the effect of cholecalciferol supplementation on cardiovascular disease, markers of inflammation and bone metabolism in CKD. We present the baseline characteristics of feasibility phase of the trial. Methods: The study is a multicentric, prospective, randomized, placebo controlled, double blind trial in two parallel groups and feasibility phase is being done at Postgraduate Institute of Medical Education and Research, Chandigarh, India. The trial is registered at Clinical Trials Registry of India (CTRI/2019/05/019211). After a run-in period of 2 weeks, the enrolled subjects are randomized in 1:1 to receive either 60,000 IU/2 weeks of cholecalciferol or matching placebo. The subjects will be then followed up every three month till 3 years. The primary outcome of the study is a composite of major adverse cardiac events (MACE). Secondary outcome measures include all-cause mortality, need of RRT, change in hsCRP, IL-6, iPTH, FGF-23, bone specific alkaline phosphatase, and CTX-1. Results: A total of 720 subjects have been screened till date. Out of 119 enrolled, 86 subjects have been randomized over 24 months period. 76% subjects have completed annual follow up at 12 months, 66% subjects - 15thmonths follow up, 40%- 18 months follow up, 26% subjects - 21 months follow up, 6% subjects – 24 months follow up. Baseline characteristics and serum biomarkers levels has been analysed in 80 subjects. Mean age of the subjects were 51.3 ± 12.2 years and 58.8 % were males. Serum haemoglobin levels were 11.6 ±1.7 g/dl. Mean eGFR was 26.3 (17.4, 35.1) ml/min/1.73m2. Outcome events were;MACE: 1 (due to CVD), death other than due to MACE: 1 (due to COVID 19), subjects with composite of all-cause death and non-fatal MACE: 2, subject with need of RRT:1 and subjects with composite of 50% decline in GFR or need of RRT: 3. 2 serious adverse events unrelated to study drug were reported during the course of study. Table: Baseline levels of various serum biomarkers [Formula presented] Conclusions: Despite COVID 19 related restrictions being in place for most of the last 18 months, the study has been able to screen and enrol participants. The follow ups have been ensured either through physical or remote (mobile/telephonic) means. Once in the multi-centric phase, the study will be able to test a relatively inexpensive intervention in the form of vitamin D supplementation for CVD in CKD. No conflict of interest

POS-252 THE DIFFERENTIAL IMPACT OF TWO NOVEL INTRAVENOUS IRON AGENTS ON FIBROBLAST GROWTH FACTOR 23, PHOSPHATE AND OTHER CLINICAL AND FUNCTIONAL MARKERS: METHODOLOGY AND BASELINE DATA

Introduction: Third generation intravenous (IV) iron preparations are increasingly used in the treatment of non-dialysis dependent chronic kidney disease (CKD) associated iron deficiency. Such compounds allow rapid delivery of large concentrations of iron safely at a single sitting. Evidence suggests that their use may lead to improved cardiovascular outcomes. Nonetheless, concerns exist regarding the potential induction of hypophosphatemia via fibroblast-growth-factor 23 (FGF-23) following the use of certain compounds. Raised FGF-23 has been associated with mineral bone and cardiac disorders, alongside prognostic implications. No prior study has provided a head-to-head comparison between iron preparations in CKD. This pilot study is designed to primarily investigate the differential impact of two different IV iron compounds on FGF-23 and phosphate in patients with CKD;secondarily we examine the impact of these compounds on bone markers and functional status, quality of life and cardiovascular function. Methods: This is a randomized controlled double-blinded pilot study recruiting patients with CKD stage 3a – 5 (non-dialysis) and iron deficiency +/- anemia. Patients are randomized to receive either ferric carboxymaltose or ferric derisomaltose over two infusions (one-month apart) to achieve full repletion. The initial dose administered is 1000 mg for both medications, with 500 mg or 1000 mg reserved for the second dose depending on weight and hematinics. Follow up is over a period of three months following the first infusion with measurements of intact FGF-23, phosphate, phosphaturia, vitamin D, parathyroid hormone, bone metabolism markers, functional status and quality of life and cardiac markers (figure 1). [Formula presented] Results: 168 patients were referred to the specialist renal anemia services for pre-screening. Ninety-nine were contacted for interest to participate, with 64 individuals declining to join. The commonest reason for not participating, was the COVID-19 pandemic (43.3%) with patients not keen to travel. Thirty-five patients were screened, and 27 patients enrolled in the study, of which 26 were randomized to receive iron (figure 2). One patient withdrew from the study, as they were unable to attend appointments following successful screening. In our baseline cohort the median age was 67.9 (12.4) and 17 participants were male. Mean hemoglobin was 100.3 (13.5) and hematinic markers consistent were consistent with iron deficiency. Median eGFR was 18.0 (11.3) ml/min/1.73 m2;the population as expected had a raised intact FGF-23 (212.1 (116.4) pg/ml). Serum calcium and phosphate were within normal parameters, while parathyroid hormone and 1,25 (OH)2 Vitamin D were deranged (Table 1). [Formula presented] [Formula presented] Conclusions: ExplorIRON-CKD, like a number of trials, experienced significant disruption in recruitment due to COVID-19. This study will provide further insight to the potential induction of FGF-23 following administration of specific intravenous iron compounds, and identify whether such a differential effect exists in patients with CKD. The effect of such induction in terms of phosphate and other markers of bone metabolism, functional status and cardiac functioning will be observed. The results will aid in hypothesis generation for further studies to identify the potential long-term impact of iatrogenic FGF-23 increase in patients with CKD. Conflict of interest Potential conflict of interest: This study received funding support from Pharmacosmos A/S and the Kidney Research Yorkshire Charity Fund. The funders had no role in the study design, data collection and analysis, and decision to publish or preparation of the . I have no potential conflicts of interest to disclose.

Age and genotype dependent erythropoietin protection in COVID-19.

Erythropoietin (EPO) is the main mediator of erythropoiesis and an important tissue protective hormone that appears to mediate an ancestral neuroprotective innate immune response mechanism at an early age. When the young brain is threatened-prematurity, neonatal hyperbilirubinemia, malaria- EPO is hyper-secreted disproportionately to any concurrent anemic stimuli. Under eons of severe malarial selection pressure, neuroprotective EPO augmenting genetic determinants such as the various hemoglobinopathies, and the angiotensin converting enzyme (ACE) I/D polymorphism, have been positively selected. When malarial and other cerebral threats abate and the young child survives to adulthood, EPO subsides. Sustained high ACE and angiotensin II (Ang II) levels through the ACE D allele in adulthood may then become detrimental as witnessed by epidemiological studies. The ubiquitous renin angiotensin system (RAS) influences the α-klotho/fibroblast growth factor 23 (FGF23) circuitry, and both are interconnected with EPO. Here we propose that at a young age, EPO augmenting genetic determinants through ACE D allele elevated Ang II levels in some or HbE/beta thalassemia in others would increase EPO levels and shield against coronavirus disease 2019, akin to protection from malaria and dengue fever. Human evolution may use ACE2 as a "bait" for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) to gain cellular entry in order to trigger an ACE/ACE2 imbalance and stimulate EPO hypersecretion using tissue RAS, uncoupled from hemoglobin levels. In subjects without EPO augmenting genetic determinants at any age, ACE2 binding and internalization upon SARS-CoV-2 entry would trigger an ACE/ACE2 imbalance, and Ang II oversecretion leading to protective EPO stimulation. In children, low nasal ACE2 Levels would beneficially augment this imbalance, especially for those without protective genetic determinants. On the other hand, in predisposed adults with the ACE D allele, ACE/ACE2 imbalance, may lead to uncontrolled RAS overactivity and an Ang II induced proinflammatory state and immune dysregulation, with interleukin 6 (IL-6), plasminogen activator inhibitor, and FGF23 elevations. IL-6 induced EPO suppression, aggravated through co-morbidities such as hypertension, diabetes, obesity, and RAS pharmacological interventions may potentially lead to acute respiratory distress syndrome, cytokine storm and/or autoimmunity. HbE/beta thalassemia carriers would enjoy protection at any age as their EPO stimulation is uncoupled from the RAS system. The timely use of rhEPO, EPO analogs, acetylsalicylic acid, bioactive lipids, or FGF23 antagonists in genetically predisposed individuals may counteract those detrimental effects.