Neutralization diversity of HIV-1 Indian subtype C envelopes obtained from cross sectional and followed up individuals against broadly neutralizing monoclonal antibodies having distinct gp120 specificities.

BACKGROUND: The potential use of the broadly neutralizing monoclonal antibodies (bnAbs) towards prophylaxis and treatment to HIV-1 is currently being explored. While a number of promising bnAbs have been discovered and a few of them have progressed towards clinical development, their extent of neutralization coverage with respect to global HIV-1 variants given the existence of genetically distinct subtypes and recombinants circulating globally is not clearly known. In the present study, we examined the variation in the neutralization susceptibility of pseudoviruses expressing 71 full length primary HIV-1 subtype C envs obtained from limited cross-sectional individuals over different time points against four bnAbs that target gp120 with distinct specificities: VRC01, CAP256-VRC26.25, PGDM1400 and PGT121. RESULTS: We found significant variations in the susceptibility of Indian clade C to these four bnAbs. These variations were found to be distinct to that observed in African subtype C based on the existing datasets and concordant with their sequence diversity. Trend analysis indicated an increasing neutralization resistance observed over time with CAP25-VRC26.25, PGDM1400 and PGT121 when tested on pseudoviruses expressing envs obtained from 1999 to 2016. However, inconsistent trend in neutralization susceptibility was observed, when pseudoviruses expressing envs obtained from three followed up individuals were examined. Finally, through predictive analysis of the 98 Indian subtype C including those assessed in the present study by employing additive model implemented in CombiNAber ( http://www.hiv.lanl.gov ), we observed two possibilities where combinations of three bnAbs (VRC01/CAP56-VRC26.25/PGT121 and PGDM1400/CAP256-VRC26.25/PGT121) could achieve near 100% neutralization coverage. CONCLUSIONS: Our findings not only indicate disparate intra-clade C genetic vis-à-vis neutralization diversities but also warrant the need for more comprehensive study using additional isolates towards comparing inter and intra-clade neutralization diversities which will be necessary for selecting the bnAb combinations suitable for optimal coverage of the region-specific HIV-1 circulating subtypes. Expanding these efforts is imperative for designing efficacious bnAb based intervention strategies for India as well as subtype C in general.

FcγRIIIa receptor polymorphism influences NK cell mediated ADCC activity against HIV.

BACKGROUND: HIV-specific Antibody Dependent Cell Cytotoxicity (ADCC) has shown to be important in HIV control and resistance. The ADCC is mediated primarily by natural killer cell activated through the binding of FcγRIIIa receptor to the Fc portion of antibody bound to the antigen expressed on the infected cells. However, no data is available on the influence of the polymorphism in FcγRIIIa receptor on HIV-specific ADCC response. METHODS: The Sanger's method of sequencing was used to sequence the exon of FcγRIIIa receptor while the ADCC activity was determined using NK cell activation assay. The polymorphism in FcγRIIIa receptor was assessed in HIV-infected Indian individuals with or without HIV-specific ADCC antibodies and its influence on the magnitude of HIV-specific ADCC responses was analyzed. RESULTS: Two polymorphisms: V176F (rs396991) and Y158H (rs396716) were observed. The Y158H polymorphism is reported for the first time in Indian population. Both, V176F (V/V genotype) (p = 0.004) and Y158H (Y/H genotype) (p = 0.032) were found to be significantly associated with higher magnitude of HIV-specific ADCC response. CONCLUSION: The study underscores the role of polymorphism in the FcγRIIIa receptor on HIV-specific ADCC response and suggests that the screening of the individuals for FcγRIIIa-V176F and Y158H polymorphisms could be useful for prediction of efficient treatment in monoclonal antibody-based therapies aimed at ADCC in HIV infection.

Higher expression of human telomerase reverse transcriptase in productively-infected CD4 cells possibly indicates a mechanism for persistence of the virus in HIV infection.

Mechanisms involved in survival of productively-infected memory CD4+cells after initial antigenic stimulation and their subsequent reversion to the resting state are critical for the development of a predominant replication-competent HIV reservoir. These mechanisms may also counter their elimination after HIV reactivation through latency-reversing agents (LRA). Thus, their evaluation is critical when using an appropriate HIV latency model that recapitulates the predominant replication-competent HIV reservoir to develop strategies for HIV eradication. The model for evaluating the possible survival mechanisms after T cell receptor (TCR) stimulation was developed by infecting memory CD4+cells with an HIV-1C primary isolate and cytokine secretion and gene expression patterns determined. Infected cells showed compromised functionality as evident from 6.8-fold lower secretion of IL-2 than from uninfected control cells. After TCR stimulation, the infected cells showed significantly higher fold increases in CD27 and CCR5 and smaller increases in CD5 mRNA over baseline values. Because CD27 expression may influence telomerase activity through AKT phosphorylation, CD27, human telomerase reverse transcriptase (hTERT) and pAKT expression in productively-infected cells from HIV-infected patients was evaluated by flow cytometry. HIV harbored in memory CD4+ cells was reactivated by HIV-1 envelope peptides, which have been shown to act as effective LRA. P24+CD4+cell showed significantly higher expression of CD27, hTERT and pAKT than P24-CD4+cells. These findings indicate compromised functionality of HIV-infected cells after TCR stimulation, which may interfere with their elimination by the immune system. They also indicate that pAKT and hTERT induction are possible survival mechanisms of productively-infected CD4+cells.

The effects of antiretroviral treatment initiation on cognition in HIV-infected individuals with advanced disease in Pune, India.

There has been a reduction in the most severe cases of HIV-associated neurocognitive disorders (HAND) with advances in antiretroviral treatment (ART). But the prevalence of milder forms of HAND still remains high. Data from systematically conducted studies on the effects of ART on cognition are scanty in India, where HIV-1 clade C is prevalent. The purpose of the present study was to assess the effect of antiretroviral therapy in HIV-seropositive (HIV+) individuals (n = 92) with CD4 cell counts <200 cells/mm(3). The overall and domain-specific levels of cognitive functioning were determined using a locally recruited normative sample, and a change in neurocognitive functioning at the 1-year follow-up visit was analyzed. Results revealed cognitive impairment in 44.6 % of the HIV+ group at baseline. At the 1-year follow-up, the group showed significant improvement in the Learning domain (p < 0.05). HIV+ individuals showing improvement in the global cognitive scores had a significantly lower baseline CD4 cell count compared to others. Overall, the degree of improvement associated with the magnitude of rise in CD4 suggests the possibility that early, mild subclinical deficits may also benefit from treatment.

Duplicate analysis method: a cheaper alternative to commercial IQC materials in limited resource settings for monitoring CD4 testing.

BACKGROUND: India has a large number of HIV infected patients being followed up at anti-retroviral therapy (ART) centers. The patients are regularly offered CD4 count estimation for deciding their eligibility for ART initiation as well as for monitoring response to ART, making CD4 count estimation a very critical test. Hence, quality control of CD4 testing is utmost important for ultimate success of ART program. As the commercial controls are very expensive, internal quality control (IQC), at present, is being done by duplicate analysis method using previous day samples in most of the laboratories. Hence the study was undertaken to review performance of duplicate analysis method for monitoring daily IQC. METHODS: Quality control (QC) data from 11 Indian laboratories using duplicate analysis and/or commercial controls for IQC of CD4 testing was collected for reviewing information on QC parameters such as precision, accuracy and trend monitoring. Precision was determined by r(2) values and mean % variation for duplicate analysis and coefficient of variation (% CV) for commercial controls. Accuracy was monitored by rate of QC failures for both the types of control and trend monitoring was done by plotting LJ charts for commercial controls and by plotting daily % variation for duplicate analysis. RESULTS: The laboratories using duplicate analysis for IQC showed good precision with mean % variation ranging from 0.5 to 7.2. There was good match between r(2) values and % CV of the laboratories performing both the types of QC methods. Rates of QC failures were 2.3 for duplicate analysis and 3 per laboratory-year for IMMUNO-TROL controls. Daily trend monitoring showed fluctuation of daily counts around mean in LJ charts and of percent variation around 0% in duplicate analysis method. Commercially available controls showed limitations such as altered specimen quality leading to difficulties in manual gating and issues with the establishment of laboratory range. CONCLUSION: Duplicate analysis can serve as a cheaper alternative to commercially available controls for IQC of CD4 testing especially when supplemented with other QC measures for controlling variations caused by reagent, equipment, staff and environment in addition to the successful participation in External Quality Assurance programme.

The microRNA miR-29a is associated with human immunodeficiency virus latency.

BACKGROUND: Latent reservoirs of HIV-1 provide a major challenge to its cure. There are increasing reports of interplay between HIV-1 replication and host miRNAs. Several host miRNAs, which potentially target the nef-3'LTR region of HIV-1 RNA, including miR-29a, are proposed to promote latency. FINDINGS: We used two established cellular models of HIV-1 latency - the U1 monocytic and J1.1 CD4+ T cell lines to show an inverse relationship between HIV-1 replication and miR-29a levels, which was mediated by the HIV-1 Nef protein. Using a miR-29a responsive luciferase reporter plasmid, an expression plasmid and an anti-miR29a LNA, we further demonstrate increased miR-29a levels during latency and reduced levels following active HIV replication. Finally, we show that miR-29a levels in the PBMCs and plasma of HIV infected persons also correlate inversely with latency and active viral replication. CONCLUSIONS: The levels of miR-29a correlate inversely with active HIV-1 replication in cell culture models and in HIV infected persons. This links miR-29a to viral latency and suggests another approach to activate and destroy latent HIV-1 reservoirs.

Clinical profile of HIV infected patients attending a HIV referral clinic in Pune, India.

BACKGROUND & OBJECTIVES: Human immunodeficiency virus (HIV) has infected several million individuals in India. Various interventions have been implemented for early detection and prevention of transmission of HIV infection. This has progressively changed the clinical profile of HIV infected individuals and this study documents the clinical presentation of individuals positive for HIV in 2010, in Pune, Maharashtra, India. METHODS: This cross-sectional study included subjects who had come to the HIV referral clinic for HIV testing from January to December 2010. Children as well as individuals with indeterminate HIV result were excluded from the study, and data for 1546 subjects were finally analysed. RESULTS: The HIV positivity rate among all referred cases for the year 2010 was 35 per cent (male 55% and females 45%). The median age (Q1, Q3) was 31 (25.75, 39) yr. The median CD4 cell count for all HIV infected individuals (whose CD4 count was available n=345) was 241 cells/µl and for asymptomatic HIV infected individuals was 319 cells/µl. There were 673 (43.5%) symptomatic and 873 (56.5%) asymptomatic participants. Fever, breathlessness, cough with expectoration, weight loss, loss of appetite, generalized weakness, pallor and lymphadenopathy (axillary and cervical) were found to be associated (P<0.001) with HIV positivity. On multivariate analysis, history of Herpes zoster [AOR 11.314 (6.111-20.949)] and TB [AOR 11.214 (6.111-20.949)] was associated with HIV positivity. INTERPRETATION & CONCLUSIONS: Signs and symptoms associated with HIV positivity observed in this study can be used by health care providers to detect HIV infection early. Moreover, similar to HIV testing in patients with tuberculosis, strategies can be developed for considering Herpes zoster as a predictor of HIV infection.

Seropersistence of SII-ChAdOx1 nCoV-19 (COVID-19 vaccine): 6-month follow-up of a randomized, controlled, observer-blind, phase 2/3 immuno-bridging study in Indian adults.

AZD1222 (ChAdOx1 nCoV-19) is a replication-deficient adenoviral vectored coronavirus disease-19 (COVID-19) vaccine that is manufactured as SII-ChAdOx1 nCoV-19 by the Serum Institute of India Pvt Ltd following technology transfer from Oxford University/AstraZeneca. The non-inferiority of SII-ChAdOx1 nCoV-19 with AZD1222 was previously demonstrated in an observer-blind, phase 2/3 immuno-bridging study (trial registration: CTRI/2020/08/027170). In this analysis of immunogenicity and safety data 6 months post first vaccination (Day 180), 1,601 participants were randomized 3:1 to SII-ChAdOx1 nCoV-19 or AZD1222 (immunogenicity/reactogenicity cohort n = 401) and 3:1 to SII-ChAdOx1 nCoV-19 or placebo (safety cohort n = 1,200). Immunogenicity was measured by anti-severe acute respiratory syndrome coronavirus 2 spike (anti-S) binding immunoglobulin G and neutralizing antibody (nAb) titers. A decline in anti-S titers was observed in both vaccine groups, albeit with a greater decline in SII-ChAdOx1 nCoV-19 vaccinees (geometric mean titer [GMT] ratio [95% confidence interval (CI) of SII-ChAdOx1 nCoV-19 to AZD1222]: 0.60 [0.41-0.87]). Consistent similar decreases in nAb titers were observed between vaccine groups (GMT ratio [95% CI]: 0.88 [0.44-1.73]). No cases of severe COVID-19 were reported following vaccination, while one case was observed in the placebo group. No causally related serious adverse events were reported through 180 days. No thromboembolic or autoimmune adverse events of special interest were reported. Collectively, these data illustrate that SII-ChAdOx1 nCoV-19 maintained a high level of immunogenicity 6 months post-vaccination. SII-ChAdOx1 nCoV-19 was safe and well tolerated.

Heparin and EDTA anticoagulants differentially affect the plasma cytokine levels in humans.

Cytokines and chemokines are the cell signaling proteins which are considered as important biomarkers of inflammation and immunity. However the fragile nature of these markers results in the concentration variation due to various external factors. We assessed the influence of commonly used anticoagulants (EDTA and heparin) on various cytokine levels from 32 paired plasma samples using highly sensitive multiple cytokine estimation assay. Out of 17 cytokines estimated, 15 were detectable in more than 80% of the samples from both the groups. TNF-α, IFN-γ, IL-4, IL-5, and G-CSF levels were significantly higher (p values < 0.05 for all) in plasma with EDTA, whereas the levels of IL-6, IL-8, IL-10, IL-17, MIP-1ß, GM-CSF and MCP-1 were found to be significantly higher (p values < 0.05 for all) in plasma with heparin. There was no significant difference in the levels of IL-7, IL-12 (P70) and IL-13 in both the groups. The study showed that the anticoagulants significantly affect the measurement of certain cytokines. Hence, it is important to choose an appropriate anticoagulant before the estimation of cytokines for reliable use of plasma cytokines as biomarkers in patient management.

CD4 estimating reagents in dry format are compatible with conventional flow cytometer; FACSCalibur for estimation of absolute CD4 count & percentages.

BACKGROUND & OBJECTIVES: Reliable CD4 counts are important for successful implementation of antiretroviral treatment (ART). Availability of dry CD4 reagents can eliminate cold chain requirement reducing shipment and storage cost. An attempt was made in this study to validate the ReaPan and Rea T Count dry reagents developed by ReaMetrix against the original BD Biosciences liquid reagents. METHOD: Absolute counts and percentages of CD4, CD8 and CD3 + T cells obtained in 100 HIV infected individuals using the test and reference reagents were analyzed for correlation and agreement using Pearson's correlation and Bland Altman bias analysis . The stability of the reagents and of the stained samples was analyzed at ambient temperature and at 37 °C. RESULTS: The absolute CD4 + T cell count and percentages obtained using test and reference reagents showed correlation coefficients ranging from 833 to 981. A mean bias between dry and reference reagents ranged from 0.8 to 26.4. The ReaPan and Rea T Count reagents were stable up to one month at 37 °C also. The samples stained with ReaPan reagents were stable at ambient temperature till day 7 whereas the samples stained with Rea T Count reagents were stable at ambient temperature and at 37° C for 10 days. INTERPRETATION & CONCLUSIONS: The ReaPan dry reagents can be used on existing FACSCalibur machines with additional training on Cell Quest Pro software without incurring any additional equipment cost and this can eliminate the requirement of cold chain during transport and on site storage. The stability of the stained samples has great clinical significance preventing redrawing of the blood samples from the patients.

Use of first line antiretroviral therapy from a free ART programme clinic in Pune, India - a preliminary report.

BACKGROUND & OBJECTIVES: The treatment outcomes under national antiretroviral therapy (ART) programme are being evaluated in some ART centres in the country. We carried out this study to analyze the impact of first line antiretroviral therapy in HIV infected patients attending a free ART roll out national programme clinic in Pune, India. METHODS: Antiretroviral naive HIV infected patients attending the clinic between December 2005 and April 2008 and followed up till March 31, 2011 were included in the analysis. The enrolment and follow up of these patients were done as per the national guidelines. Viral load estimations were done in a subset of patients. results: One hundred and forty two patients with median CD4 count of 109 cells/µl (IQR: 60-160) were initiated on treatment. The median follow up was 44 months (IQR: 37-53.3 months). Survival analysis showed that the probability of being alive at the end of 5 years was 85 per cent. Overall increase in the median CD4 count was statistically significant (P<0.001). It was significant in patients with >95 per cent adherence (P<0.001). In 14 per cent patients, the absolute CD4 count did not increase by 100 or more cells/µl at the end of 12 months. Viral load estimation in a subset of 68 patients showed undetectable levels in 61 (89.7%) patients after a median duration of 46 months (IQR: 38.3-54.8). INTERPRETATION & CONCLUSIONS: The first line treatment was effective in patients attending the programme clinic. The adherence level influenced immunological and virological outcomes of patients.

Differential modulation of phenotypic composition of HIV-infected and -uninfected PBMCs during cryopreservation.

This article was designed to determine variations in phenotypic composition of fresh and frozen PBMCs for assessing utility of cryopreserved PBMCs for phenotypic assays. Relative percentages of effector memory cells increased significantly as against percentages of naïve cells which showed significant decrease after cryopreservation in HIV-uninfected samples. These differences were not significant in HIV-infected individuals. There was no significant difference in the expression of activation markers in fresh and frozen PBMCs except the HLA DR expression on CD8 cells in HIV-infected individuals, which was significantly decreased in frozen PBMCs. Thus, cryopreservation resulted in differential effect on phenotypic composition of PBMCs in HIV-infected and -uninfected individuals.

HIV-1 exploits Hes-1 expression during pre-existing HPV-16 infection for cancer progression.

High Risk Human Papilloma Viruses (HR-HPV) persistently infect women with Human Immunodeficiency Virus-1 (HIV-1). HPV-16 escapes immune surveillance in HIV-1 positive women receiving combined antiretroviral therapy (cART). HIV-1 Tat and HPV E6/E7 proteins exploit Notch signaling. Notch-1, a developmentally conserved protein, influences cell fate from birth to death. Notch-1 and its downstream targets, Hes-1 and Hey-1 contribute to invasive and aggressive cancers. Cervical cancer cells utilize Notch-1 and hyper-express CXCR4, a co-receptor of HIV-1. Accumulating evidence shows that HIV-1 affects cell cycle progression in pre-existing HPV infection. Additionally, Tat binds Notch-1 receptor for activation and influences cell proliferation. Oncogenic viruses may interfere or converge together to favor tumor growth. The molecular dialogue during HIV-1/HPV-16+ co-infections in the context of Notch-1 signaling has not been explored thus far. This in vitro study was designed with cell lines (HPV-ve C33A and HPV-16+ CaSki) which were transfected with plasmids (pLEGFPN1 encoding HIV-1 Tat and pNL4-3 encoding HIV-1 [full HIV-1 genome]). HIV-1 Tat and HIV-1 inhibited Notch-1expression, with differential effects on EGFR. Notch-1 inhibition nullified Cyclin D expression with p21 induction and increased G2-M cell population in CaSki cells. On the contrary, HIV-1 infection shuts down p21 expression through interaction of Notch-1 downstream genes Hes-1-EGFR and Cyclin D for G2-M arrest, DDR response and cancer progression. This work lays foundations for future research and interventions, and therefore is necessary. Our results describe for the first time how HIV-1 Tat cancers have an aggressive nature due to the interplay between Notch-1 and EGFR signaling. Notch-1 inhibitor, DAPT used in organ cancer treatment may help rescue HIV-1 induced cancers. Graphical abstract: The illustration shows how HIV interacts with HPV-16 to induce Notch 1 suppression for cancer progression (Created with BioRender.com). Supplementary Information: The online version contains supplementary material available at 10.1007/s13337-023-00809-y.

Higher proinflammatory responses possibly contributing to suppressed cytotoxicity in patients with COVID-19 associated mucormycosis.

INTRODUCTION: COVID-19 Associated Mucormycosis (CAM), an opportunistic fungal infection, surged during the second wave of SARS Cov-2 pandemic. Since immune responses play an important role in controlling this infection in immunocompetent hosts, it is required to understand immune perturbations associated with this condition for devising immunotherapeutic strategies for its control. We conducted a study to determine different immune parameters altered in CAM cases as compared to COVID-19 patients without CAM. METHODOLOGY: Cytokine levels in serum samples of CAM cases (n = 29) and COVID-19 patients without CAM (n = 20) were determined using luminex assay. Flow cytometric assays were carried out in 20 CAM cases and 10 controls for determination of frequency of NK cells, DCs, phagocytes, T cells and their functionalities. The cytokine levels were analyzed for their association with each other as well as with T cell functionality. The immune parameters were also analyzed with respect to the known risk factors such as diabetes mellitus and steroid treatment. RESULTS: Significant reduction in frequencies of total and CD56 + CD16 + NK cells (cytotoxic subset) was noted in CAM cases. Degranulation responses indicative of cytotoxicity of T cell were significantly hampered in CAM cases as compared to the controls. Conversely, phagocytic functions showed no difference in CAM cases versus their controls except for migratory potential which was found to be enhanced in CAM cases. Levels of proinflammatory cytokines such as IFN-γ, IL-2, TNF-α, IL-17, IL-1ß, IL-18 and MCP-1 were significantly elevated in cases as compared to the control with IFN-γ and IL-18 levels correlating negatively with CD4 T cell cytotoxicity. Steroid administration was associated with higher frequency of CD56 + CD16- NK cells (cytokine producing subset) and higher MCP-1 levels. Whereas diabetic participants had higher phagocytic and chemotactic potential and had higher levels of IL-6, IL-17 and MCP-1. CONCLUSION: CAM cases differed from the controls in terms of higher titers of proinflammatory cytokines, reduced frequency of total and cytotoxic CD56 + CD16 + NK cell. They also had reduced T cell cytotoxicity correlating inversely with IFN-γ and IL-18 levels, possibly indicating induction of negative feedback mechanisms while diabetes mellitus or steroid administration did not affect the responses negatively.

Safety and Immunogenicity of SARS-CoV-2 Recombinant Spike Protein Vaccine in Children and Adolescents in India: A Phase 2-3 Randomized Clinical Trial.

Importance: The recombinant COVID-19 vaccine NVX-CoV2373 has demonstrated efficacy of approximately 90% in adults; however, its safety and efficacy in children is unknown. Objective: To assess the noninferiority of SII-NVX-CoV2373 in children and adolescents compared to adults and to evaluate its safety in comparison with placebo. Design, Setting, and Participants: This phase 2-3 observer-blind randomized clinical trial was conducted in 2 cohorts, children (aged 2 to 11 years) and adolescents (aged 12 to 17 years) between August 2021 and August 2022. Participants were randomized 3:1 to SII-NVX-CoV2373 or placebo and monitored for 179 days. The participants, study team, and laboratory staff were blinded. This was a multicenter study conducted across 10 tertiary care hospitals in India. Exclusion criteria included previous COVID-19 infection or vaccination, immunocompromised condition, and immunosuppressive medications. Interventions: Two doses of 0.5-mL SII-NVX-CoV2373 or placebo were administered intramuscularly on days 1 and 22. Main Outcomes and Measures: Primary outcomes were geometric mean titer ratio of both anti-spike (anti-S) IgG and neutralizing antibodies (NAbs) between both pediatric age groups to that of adults on day 36. Noninferiority was concluded if the lower bound of 95% CI of this ratio was greater than 0.67 for each age group. Both the antibodies were assessed for the index strain and for selected variants at various time points. Solicited adverse events (AEs) were recorded for 7 days after each vaccination, unsolicited AEs were recorded for 35 days, and serious AEs and AEs of special interest were recorded for 179 days. Results: A total of 460 children in each age cohort were randomized to receive vaccine or placebo. The mean (SD) age was 6.7 (2.7) years in the child cohort and 14.3 (1.6) years in the adolescent cohort; 231 participants (50.2%) in the child cohort and 218 in the adolescent cohort (47.4%) were female. Both anti-S IgG and NAb titers were markedly higher in the SII-NVX-CoV2373 group than in the placebo group on both day 36 and day 180. The geometric mean titer ratios compared to those in adults were 1.20 (95% CI, 1.08-1.34) and 1.52 (95% CI, 1.38-1.67) for anti-S IgG in adolescents and children, respectively; while for NAbs, they were 1.33 (95% CI, 1.17-1.50) and 1.93 (95% CI, 1.70-2.18) in adolescents and children, respectively, indicating noninferiority. SII-NVX-CoV2373 also showed immune responses against variants studied. Injection site reactions, fever, headache, malaise, and fatigue were common solicited AEs. There were no AEs of special interest and no causally related serious AEs. Conclusions and Relevance: SII-NVX-CoV2373 was safe and well tolerated in children and adolescents in this study. The vaccine was highly immunogenic and may be used in pediatric vaccination against COVID-19. Trial Registration: Clinical Trials Registry of India Identifier: CTRI/2021/02/031554.

A Phase 3, randomized, non-inferiority study of a heterologous booster dose of SARS CoV-2 recombinant spike protein vaccine in adults.

Due to waning immunity following primary immunization with COVID-19 vaccines, booster doses may be required. The present study assessed a heterologous booster of SII-NVX-CoV2373 (spike protein vaccine) in adults primed with viral vector and inactivated vaccines. In this Phase 3, observer-blind, randomized, active controlled study, a total of 372 adults primed with two doses of ChAdOx1 nCoV-19 (n = 186) or BBV152 (n = 186) at least six months ago, were randomized to receive a booster of SII-NVX-CoV2373 or control vaccine (homologous booster of ChAdOx1 nCoV-19 or BBV152). Anti-S IgG and neutralizing antibodies (nAbs) were assessed at days 1, 29, and 181. Non-inferiority (NI) of SII-NVX-CoV2373 to the control vaccine was assessed based on the ratio of geometric mean ELISA units (GMEU) of anti-S IgG and geometric mean titers (GMT) of nAbs (NI margin > 0.67) as well as seroresponse (≥ 2 fold-rise in titers) (NI margin -10%) at day 29. Safety was assessed throughout the study period. In both the ChAdOx1 nCoV-19 prime and BBV152 prime cohorts, 186 participants each received the study vaccines. In the ChAdOx1 nCoV-19 prime cohort, the GMEU ratio was 2.05 (95% CI 1.73, 2.43) and the GMT ratio was 1.89 (95% CI 1.55, 2.32) whereas the difference in the proportion of seroresponse was 49.32% (95% CI 36.49, 60.45) for anti-S IgG and 15% (95% CI 5.65, 25.05) for nAbs on day 29. In the BBV152 prime cohort, the GMEU ratio was 5.12 (95% CI 4.20, 6.24) and the GMT ratio was 4.80 (95% CI 3.76, 6.12) whereas the difference in the proportion of seroresponse was 74.08% (95% CI 63.24, 82.17) for anti-S IgG and 24.71% (95% CI 16.26, 34.62) for nAbs on day 29. The non-inferiority of SII-NVX-CoV2373 booster to the control vaccine for each prime cohort was met. SII-NVX-CoV2373 booster showed significantly higher immune responses than BBV152 homologous booster. On day 181, seroresponse rates were ≥ 70% in all the groups for both nAbs and anti-S IgG. Solicited adverse events reported were transient and mostly mild in severity in all the groups. No causally related SAE was reported. SII-NVX-CoV2373 as a heterologous booster induced non-inferior immune responses as compared to homologous boosters in adults primed with ChAdOx1 nCoV-19 and BBV152. SII-NVX-CoV2373 showed a numerically higher boosting effect than homologous boosters. The vaccine was also safe and well tolerated.

Safety and immunogenicity of SII-NVX-CoV2373 (COVID-19 vaccine) in adults in a phase 2/3, observer-blind, randomised, controlled study.

Background: NVX-CoV2373, a Covid-19 vaccine was developed in the USA with ∼90% efficacy. The same vaccine is manufactured in India after technology transfer (called as SII-NVX-CoV2373), was evaluated in this phase 2/3 immuno-bridging study. Methods: This was an observer-blind, randomised, phase 2/3 study in 1600 adults. In phase 2, 200 participants were randomized 3:1 to SII-NVX-CoV2373 or placebo. In phase 3, 1400 participants were randomized 3:1 to SII-NVX-CoV2373 or NVX-CoV2373 (940 safety cohort and 460 immunogenicity cohort). Two doses of study products (SII-NVX-CoV2373, NVX-CoV2373 or placebo) were given 3 weeks apart. Primary objectives were to demonstrate non-inferiority of SII-NVX-CoV2373 to NVX-CoV2373 in terms of geometric mean ELISA units (GMEU) ratio of anti-S IgG antibodies 14 days after the second dose (day 36) and to determine the incidence of causally related serious adverse events (SAEs) through 180 days after the first dose. Anti-S IgG response was assessed using an Enzyme-Linked Immunosorbent Assay (ELISA) and neutralizing antibodies (nAb) were assessed by a microneutralization assay using wild type SARS CoV-2 in participants from the immunogenicity cohort at baseline, day 22, day 36 and day 180. Cell mediated immune (CMI) response was assessed in a subset of 28 participants from immunogenicity cohort by ELISpot assay at baseline, day 36 and day 180. The total follow-up was for 6 months. Trial registration: CTRI/2021/02/031554. Findings: Total 1596 participants (200 in Phase 2 and 1396 in Phase 3) received the first dose. SII-NVX-CoV2373 was found non-inferior to NVX-CoV2373 (anti-S IgG antibodies GMEU ratio 0.91; 95% CI: 0.79, 1.06). At day 36, there was more than 58-fold rise in anti-S IgG and nAb titers compared to baseline in both the groups. On day 180 visit, these antibody titers declined to levels slightly lower than those after the first dose (13-22 fold-rise above baseline). Incidence of unsolicited and solicited AEs was similar between the SII-NVX-CoV2373 and NVX-CoV2373 groups. No adverse event of special interest (AESI) was reported. No causally related SAE was reported. Interpretation: SII-NVX-CoV2373 induced a non-inferior immune response compared to NVX-CoV2373 and has acceptable safety profile. Funding: SIIPL, Indian Council of Medical Research, Novavax.

Utility of the point of care CD4 analyzer, PIMA, to enumerate CD4 counts in the field settings in India.

BACKGROUND: In resource limited settings non-availability of CD4 count facility at the site could adversely affect the ART roll out programme. Point of care CD4 enumerating equipments can make the CD4 count available at the site of care and improve the patients' management considerably. This study is aimed at determining the utility of a Point of Care PIMA CD4 analyzer (Alere, Germany) in the field settings in India. METHOD: The blood samples were collected from 1790 participants at 21 ART centers from different parts of the country and tested using PIMA and the reference methods (FACSCalibur, FACSCount and CyFlow SL3). The paired finger prick and venous blood samples from 175 participants were tested by the PIMA CD4 Analyzer and then by FACSCalibur. RESULT: The CD4 counts obtained by PIMA CD4 analyzer showed excellent correlation with the counts obtained by the reference methods; for venous blood the Pearson's r was 0.921, p < 0.001 and the relative bias was 0.2% (range: -42 to 42%) and for finger prick samples, the Pearson's r was 0.856 and the relative bias was -9.1% (range: -46% to 27%). For CD4 ranges; <250, 251-350, 351-500 and >500 cells/mm3, the differences in the median CD4 counts obtained by the reference method and the PIMA analyzer were not significant (P > 0.05) and the relative bias were low (-7 to 5.1%). The Intermachine comparison showed variation within the acceptable limit of%CV of 10%. CONCLUSION: In the field settings, the POC PIMA CD4 analyzer gave CD4 counts comparable to the reference methods for all CD4 ranges. The POC equipment could identify the patients eligible for ART in 91% cases. Adequate training is necessary for finger prick sample collection for optimum results. Decentralization of CD4 testing by making the CD4 counts available at primary health centers, especially in remote areas with minimum or no infrastructure would reduce the missed visits and improve adherence of the patients.