A Prospective Randomised, Controlled Trial Switching Sirolimus for Mycophenolate to Enhance Immunological Responses to Third Dose COVID-19 Vaccination in Kidney Transplant Recipients with Poor Baseline Humoral Immunity

Purpose: Kidney transplant recipients (KTR) have inadequate responses to 2-dose COVID vaccination schedules and are at increased risk of severe COVID-19. Formation of T cell memory following vaccination is regulated by mTOR complex 1. mTOR inhibitors have been used in pre-clinical models to boost vaccine-elicited cytotoxic T cell memory responses. In observational studies, KTR receiving mTOR inhibitors had improved serological neutralisation and SARS-CoV-2 reactive T cell responses to 2 doses of COVID-19 vaccine, including cytotoxic T cells and circulating T follicular helper cells. We performed a clinical trial in stable KTR using sirolimus as a substitute for mycophenolate prior to a 3rd dose of COVID-19 vaccine to enhance COVID-19 vaccine responses. Method(s): KTR receiving tacrolimus, mycophenylate and corticosteroid with inadequate response to 2 doses of a COVID vaccine (defined by anti-RBD IgG <100U/ mL) and no history of COVID infection were recruited from 2 Australian transplant centres. Patients were randomised in a 1:1 ratio to continue mycophenolate maintenance or switch to sirolimus (trough level target 6 ng/mL). All patients received a 3rd dose of BNT162b2 COVID-19 vaccine and had immunological responses measured 4-6 weeks later. Result(s): 54 patients were randomised to sirolimus switch (n = 28), or control (n = 26). Patients were 70% male, mean age 57.5 years (SD10.4), with mean graft age 6.2 years (SD 5.4). Mean serum trough concentrations of sirolimus and tacrolimus were 6.4 and 6.1 respectively. There have been no safety or tolerability issues in the sirolimus cohort with stable serum creatinine (mean 117.8 vs 119.3, p=0.6), and mild increase in urinary ACR (mean 5.4 vs 17.4, p=0.1). Final results including immunological testing will be collated March 2022. Conclusion(s): Sirolimus switch is safe and well-tolerated. This trial will determine whether the strategy of mTOR inhibitor therapy peri-vaccination can optimise vaccine immune responses against COVID-19 in KTR.

A Randomized, Controlled, Double-Blind Trial of Inulin to Boost Response to a Third COVID-19 Vaccine Dose in Kidney Transplant Recipients

Purpose: Kidney transplant recipients (KTR) commonly exhibit inadequate responses to 2-dose COVID-19 vaccination schedules and remain at increased risk of severe COVID-19. Gut dysbiosis is common among KTRs and has been associated with poor vaccine responses. We hypothesised that a dietary fibre supplement may correct dysbiosis and enhance responses to a third dose of COVID-19 vaccine in KTRs. Method(s): KTRs who had received 2 doses of a COVID-19 vaccine were recruited from 2 transplant programs in Australia. KTRs with an inadequate response (defined by anti-RBD <100U/mL) were randomised to receive inulin (fibre) or maltodextran (control), 10g dissolved in 200ml water twice daily for 4 weeks prior to, and 4 weeks after a 3rd vaccine, at which time vaccine response was measured by anti-RBD titre, vaccine-specific B and T cell responses, and changes in the gut microbiome. Patients and investigators were blinded to treatment assignment. COVID-19 infection was excluded by measurement of anti-nucleocapsid antigen. Result(s): Of 85 KTRs screened, 71 had baseline anti-RBD<100U/mL and were randomised to inulin (n=37) or control (n=34). Participants were 33% female, mean age 59 yrs (SD 11), with mean eGFR 56 ml/min/1.73m2 (SD 24.8), and were most commonly receiving tacrolimus, mycophenolate and prednisolone. All participants received a third dose of a mRNA COVID-19 vaccine after receiving a dietary supplement for 4 weeks. Week 8 assessment of vaccine response, supplement tolerability and change in microbiome are ongoing. Four participants tested positive for COVID-19 during the study. Conclusion(s): Gut dysbiosis is one potential contributor to the poor COVID-19 vaccine responses exhibited by KTRs. This trial will determine whether a simple dietary fibre supplement is well tolerated and effective in correcting gut dysbiosis and restoring vaccine responsiveness. Improved vaccine responses are urgently required to better protect KTRs from ongoing morbidity and mortality from COVID-19.

A multi-centre national study of COVID-19 infection in cancer patients

Background & aims: COVID19 disproportionately affects the immunosuppressed, but its epidemiology over time is incompletely characterised. We describe Australian experiences of COVID19 in a national observational study of patients with malignancy. Methods: An ongoing multisite prospective cohort study of adult COVID19 patients with active cancer was conducted. Clinical and laboratory data over 28 months (1/3/20-22/7/22) was collated from 15 hospitals. Results: There were 491 patients included. Patients were a median of 63(IQR:50-71) years with majority male (254,52%). Solid organ malignancy was most common (296,60%), followed by haematological malignancy (180,37%), then both (15,4%). Most common solid tumour was breast cancer (74/296,25%);most common haematological cancer was lymphoma (102/180,57%). Majority (275,56%) were undergoing cancer treatment at COVID19 diagnosis. From 2020-2022, patients presented less with lower respiratory tract infections (57%,36%,5%) with increasing outpatient management (26%,50%,67%). Improved mortality was seen (27%,19%,11%). Median inpatient length of stay was 8(4-11) days. Intensive care admission was low (21,4%). For patients who had repeated respiratory PCR testing, median time from first to last positive test was 17(7-25, n = 123) days. Cancer treatment modification occurred in 18(4%) and delay in 74(15%). Conclusion: Despite improvements in outcomes, COVID19 still results in morbidity with impacts on cancer treatment. This preliminary data shows that cancer patients remain a vulnerable group and should be prioritised for public health interventions.

Safety and efficacy of a three-dose regimen of Plasmodium falciparum sporozoite vaccine in adults during an intense malaria transmission season in Mali: a randomised, controlled phase 1 trial.

BACKGROUND: WHO recently approved a partially effective vaccine that reduces clinical malaria in children, but increased vaccine activity is required to pursue malaria elimination. A phase 1 clinical trial was done in Mali, west Africa, to assess the safety, immunogenicity, and protective efficacy of a three-dose regimen of Plasmodium falciparum sporozoite (PfSPZ) Vaccine (a metabolically active, non-replicating, whole malaria sporozoite vaccine) against homologous controlled human malaria infection (CHMI) and natural P falciparum infection. METHODS: We recruited healthy non-pregnant adults aged 18-50 years in Donéguébougou, Mali, and surrounding villages (Banambani, Toubana, Torodo, Sirababougou, Zorokoro) for an open-label, dose-escalation pilot study and, thereafter, a randomised, double-blind, placebo-controlled main trial. Pilot study participants were enrolled on an as-available basis to one group of CHMI infectivity controls and three staggered vaccine groups receiving: one dose of 4·5 × 105, one dose of 9 × 105, or three doses of 1·8 × 106 PfSPZ via direct venous inoculation at approximately 8 week intervals, followed by homologous CHMI 5 weeks later with infectious PfSPZ by direct venous inoculation (PfSPZ Challenge). Main cohort participants were stratified by village and randomly assigned (1:1) to receive three doses of 1·8 × 106 PfSPZ or normal saline at 1, 13, and 19 week intervals using permuted block design by the study statistician. The primary outcome was safety and tolerability of at least one vaccine dose; the secondary outcome was vaccine efficacy against homologous PfSPZ CHMI (pilot study) or against naturally transmitted P falciparum infection (main study) measured by thick blood smear. Combined artesunate and amodiaquine was administered to eliminate pre-existing parasitaemia. Outcomes were analysed by modified intention to treat (mITT; including all participants who received at least one dose of investigational product; safety and vaccine efficacy) and per protocol (vaccine efficacy). This trial is registered with ClinicalTrials.gov, number NCT02627456. FINDINGS: Between Dec 20, 2015, and April 30, 2016, we enrolled 56 participants into the pilot study (five received the 4·5 × 105 dose, five received 9 × 105, 30 received 1·8 × 106, 15 were CHMI controls, and one withdrew before vaccination) and 120 participants into the main study cohort with 60 participants assigned PfSPZ Vaccine and 60 placebo in the main study. Adverse events and laboratory abnormalities post-vaccination in all dosing groups were few, mainly mild, and did not differ significantly between vaccine groups (all p>0·05). Unexpected severe transaminitis occured in four participants: one participant in pilot phase that received 1·8 × 106 PfSPZ Vaccine, one participant in main phase that received 1·8 × 106 PfSPZ Vaccine, and two participants in the main phase placebo group. During PfSPZ CHMI, approximately 5 weeks after the third dose of 1·8 × 106 PfSPZ, none of 29 vaccinees and one of 15 controls became positive on thick blood smear; subsequent post-hoc PCR analysis for submicroscopic blood stage infections detected P falciparum parasites in none of the 29 vaccine recipients and eight of 15 controls during CHMI. In the main trial, 32 (58%) of 55 vaccine recipients and 42 (78%) of 54 controls became positive on thick blood smear during 24-week surveillance after vaccination. Vaccine efficacy (1-hazard ratio) was 0·51 per protocol (95% CI 0·20-0·70; log-rank p=0·0042) and 0·39 by mITT (0·04-0·62; p=0·033); vaccine efficacy (1-risk ratio) was 0·24 per-protocol (0·02-0·41; p=0·031) and 0·22 mITT (0·01-0·39; p=0·041). INTERPRETATION: A three-dose regimen of PfSPZ Vaccine was safe, well tolerated, and conferred 51% vaccine efficacy against intense natural P falciparum transmission, similar to 52% vaccine efficacy reported for a five-dose regimen in a previous trial. FUNDING: US National Institute of Allergy and Infectious Diseases, National Institutes of Health, Sanaria. TRANSLATION: For the French translation of the abstract see Supplementary Materials section.

Severe Hepatocellular Liver Injury After COVID-19 Vaccination Without Autoimmune Hepatitis Features: A Case Series

Various vaccines were developed and administered to end the pandemic caused by the novel coronavirus diseases 2019 (COVID-19). These vaccines caused rare side effects not known during clinical trials. We report 2 cases of idiosyncratic drug-induced liver injury where the Pfizer BioNTech COVID-19 mRNA vaccine was administered. Both cases showed hepatocellular liver injury with negative autoimmune features on serology and histology and progressed to a protracted course of severe jaundice and pruritus before resolving without any immunosuppressants. We hope to raise awareness of possible rare side effects of the COVID-19 vaccine so that physicians can be vigilant.

Identification of warning signs in Malaysian patients having COVID-19 infection who progress to severe form of the illness

INTRODUCTION: Coronavirus disease 2019 (COVID-19) is an infectious disease caused by a novel coronavirus, now widely known as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), which has caused 3 major pandemic waves in Malaysia. We aimed to identify the warning signs as indicators that predict the progression of disease. MATERIALS AND METHODS: This is a retrospective cohort study of adult patients more than 12 years of age presenting with laboratory-confirmed COVID-19 admitted in three separate hospitals around the country. RESULTS: Of the 228 patients initially admitted with mild illness, 47 had progressed requiring oxygen. The median time from admission to deterioration was 3 days (IQR 2 - 5). Age more than ≥50years old (median age = 42.5, IQR = 28.8 - 57.0), higher temperature (mean = 37.3, IQR 36.8 - 38.0), MEWS score >3 (9, 19.1%), Neutrophil-to-lymphocyte ratio (NLR) >3.13 , (18, 38.3%) C-reactive protein (CRP) >5. (12, 27.3%), multiple zonal involvement on the chest radiography on admission (2, IQR 1-3) were more common in the deteriorated group on admission. On multivariate analysis, multiple comorbidities (HR = 7.40, 95 percent CI 2.58-21.2, p0.001), presence of persistent fever (HR = 2.88, 95 percent CI 1.15 - 7.2, p = 0.024), MEWS scoring >3 (HR of 6.72 ;95 percent CI 2.81-16.0, p0.001) were associated with progression to severe illness. CONCLUSION: In our cohort, we found that several factors were associated with the severity of COVID19. Early detection of these factors could correctly identify patients who need more intensive monitoring, and early referral for ICU care.

Transient knockdown of Anopheles stephensi LRIM1 using RNAi increases Plasmodium falciparum sporozoite salivary gland infections.

BACKGROUND: Plasmodium falciparum (Pf) sporozoites (PfSPZ) can be administered as a highly protective vaccine conferring the highest protection seen to date. Sanaria® PfSPZ vaccines are produced using aseptically reared Anopheles stephensi mosquitoes. The bionomics of sporogonic development of P. falciparum in A. stephensi to fully mature salivary gland PfSPZ is thought to be modulated by several components of the mosquito innate immune system. In order to increase salivary gland PfSPZ infections in A. stephensi and thereby increase vaccine production efficiency, a gene knock down approach was used to investigate the activity of the immune deficiency (IMD) signaling pathway downstream effector leucine-rich repeat immune molecule 1 (LRIM1), an antagonist to Plasmodium development. METHODS: Expression of LRIM1 in A. stephensi was reduced following injection of double stranded (ds) RNA into mosquitoes. By combining the Gal4/UAS bipartite system with in vivo expression of short hairpin (sh) RNA coding for LRIM1 reduced expression of LRIM1 was targeted in the midgut, fat body, and salivary glands. RT-qPCR was used to demonstrate fold-changes in gene expression in three transgenic crosses and the effects on P. falciparum infections determined in mosquitoes showing the greatest reduction in LRIM1 expression. RESULTS: LRIM1 expression could be reduced, but not completely silenced, by expression of LRIM1 dsRNA. Infections of P. falciparum oocysts and PfSPZ were consistently and significantly higher in transgenic mosquitoes than wild type controls, with increases in PfSPZ ranging from 2.5- to tenfold. CONCLUSIONS: Plasmodium falciparum infections in A. stephensi can be increased following reduced expression of LRIM1. These data provide the springboard for more precise knockout of LRIM1 for the eventual incorporation of immune-compromised A. stephensi into manufacturing of Sanaria's PfSPZ products.

Stabilization of vaccines

The purpose of vaccines is to prevent and/or reduce morbidity and mortality in individuals and interrupt transmission of infectious agents within populations. Vaccines have saved many millions of lives and are forefront in the struggle to contain new emerging diseases including Ebola and Covid-19. Vaccines to ∼25 human viral and bacterial diseases are licensed in the US. On the horizon is the first licensed human vaccine against a eukaryotic pathogen, Plasmodium falciparum, responsible for >98% of deaths from malaria. Vaccine stabilization methods are designed to minimize potency loss during manufacture and losses during storage, distribution, and in-clinic storage/use. Methods vary according to the immunogen, whether an adjuvant is incorporated and anticipated shelf life. Liquid or lyophilized formulations with storage and distribution at 2-8 °C utilize refrigerators that are particularly unreliable, and vaccine efficacy is often compromised. The Ebola vaccine, ERBEVO, is stored/distributed at -60 to -80 °C in specialized freezers or on dry ice, which is even more difficult than 2 to 8 °C. Alternative stabilization methods that maintain adequate vaccine potency at ambient temperatures – increasingly foam drying and spray drying – are used experimentally, but not yet for any licensed vaccines. Eukaryotic whole cell vaccines including PfSPZ Vaccine and several veterinary vaccines (as well as CAR-T and cellular therapies), require cryopreservation with storage and distribution below -150 °C. Little to no loss in potency occurs below Tg, compared to ∼1 log loss over 12 months typical of lyophilized vaccines at 2 to 8 °C. Cryogenic storage is favored for some RNA- and DNA-based vaccines particularly in anticipation of future epidemics requiring long term stockpiling. Cryogenic storage in standard vaccine vials is problematic: these tolerate temperatures down to -80 °C but for CAR-T products and PfSPZ Vaccine new custom cryovials that incorporate a septum for needle access and maintain CCIT at liquid nitrogen temperatures have been developed. And the -150 °C cold chain is more practical than one based on -80 °C or dry ice. Funding: NIH/NIAID 5SB1AI077262-07 DoD W81XWH18C0326 Conflict of Interest: None to disclose