Intradermal delivery of the third dose of the mRNA-1273 SARS-CoV-2 vaccine: safety and immunogenicity of a fractional booster dose.

OBJECTIVES: The aim of this study was to assess the safety and immunogenicity of a dose-sparing fractional intradermal (ID) booster strategy with the mRNA-1273 COVID-19 vaccine. METHODS: COVID-19 naive adults aged 18-30 years were recruited from a previous study on primary vaccination regimens that compared 20 µg ID vaccinations with 100 µg intramuscular (IM) vaccinations with mRNA-1273 as the primary vaccination series. Participants previously immunized with ID regimens were randomly assigned (1:1) to receive a fractional ID booster dose (20 µg) or the standard-of-care intramuscular (IM) booster dose (50 µg) of the mRNA-1273 vaccine, 6 months after completing their primary series (ID-ID and ID-IM group, respectively). Participants that had received a full dose IM regimen as the primary series, received the IM standard-of-care booster dose (IM-IM group). In addition, COVID-19 naive individuals aged 18-40 years who had received an IM mRNA vaccine as the primary series were recruited from the general population to receive a fractional ID booster dose (IM-ID group). Immunogenicity was assessed using IgG anti-spike antibody responses and neutralizing capacity against SARS-CoV-2. Cellular immune responses were measured in a sub-group. Safety and tolerability were monitored. RESULTS: In January 2022, 129 participants were included in the study. Fractional ID boosting was safe and well tolerated, with fewer systemic adverse events compared with IM boosting. At day 28 post-booster, anti-spike S1 IgG geometric mean concentrations were 9106 (95% CI, 7150-11 597) binding antibody units (BAU)/mL in the IM-IM group and 4357 (3003-6322) BAU/mL; 6629 (4913-8946) BAU/mL; and 5264 (4032-6873) BAU/mL in the ID-IM, ID-ID, and IM-ID groups, respectively. DISCUSSION: Intradermal boosting provides robust immune responses and is a viable dose-sparing strategy for mRNA COVID-19 vaccines. The favourable side-effect profile supports its potential to reduce vaccine hesitancy. Fractional dosing strategies should be considered early in the clinical development of future mRNA vaccines to enhance vaccine availability and pandemic preparedness.

Immunogenicity and reactogenicity of intradermal mRNA-1273 SARS-CoV-2 vaccination: a non-inferiority, randomized-controlled trial.

Fractional dosing can be a cost-effective vaccination strategy to accelerate individual and herd immunity in a pandemic. We assessed the immunogenicity and safety of primary intradermal (ID) vaccination, with a 1/5th dose compared with the standard intramuscular (IM) dose of mRNA-1273 in SARS-CoV-2 naïve persons. We conducted an open-label, non-inferiority, randomized controlled trial in the Netherlands between June and December 2021. One hundred and fifty healthy and SARS-CoV-2 naïve participants, aged 18-30 years, were randomized (1:1:1) to receive either two doses of 20 µg mRNA-1273 ID with a standard needle (SN) or the Bella-mu® needle (BM), or two doses of 100 µg IM, 28 days apart. The primary outcome was non-inferiority in seroconversion rates at day 43 (D43), defined as a neutralizing antibody concentration threshold of 465 IU/mL, the lowest response in the IM group. The non-inferiority margin was set at -15%. Neutralizing antibody concentrations at D43 were 1789 (95% CI: 1488-2150) in the IM and 1263 (951-1676) and 1295 (1020-1645) in the ID-SN and ID-BM groups, respectively. The absolute difference in seroconversion proportion between fractional and standard-dose groups was -13.95% (-24.31 to -3.60) for the ID-SN and -13.04% (-22.78 to -3.31) for the ID-BM group and exceeded the predefined non-inferiority margin. Although ID vaccination with 1/5th dose of mRNA-1273 did not meet the predefined non-inferior criteria, the neutralizing antibody concentrations in these groups are far above the proposed proxy for protection against severe disease (100 IU/mL), justifying this strategy in times of vaccine scarcity to accelerate mass protection against severe disease.

Transforming Parkinson's Care in Africa (TraPCAf): protocol for a multimethodology National Institute for Health and Care Research Global Health Research Group project.

BACKGROUND: Parkinson's disease (PD) is the second most common neurodegenerative disorder and, according to the Global Burden of Disease estimates in 2015, was the fastest growing neurological disorder globally with respect to associated prevalence, disability, and deaths. Information regarding the awareness, diagnosis, phenotypic characteristics, epidemiology, prevalence, risk factors, treatment, economic impact and lived experiences of people with PD from the African perspective is relatively sparse in contrast to the developed world, and much remains to be learned from, and about, the continent. METHODS: Transforming Parkinson's Care in Africa (TraPCAf) is a multi-faceted, mixed-methods, multi-national research grant. The study design includes multiple sub-studies, combining observational (qualitative and quantitative) approaches for the epidemiological, clinical, risk factor and lived experience components, as appropriate, and interventional methods (clinical trial component). The aim of TraPCAf is to describe and gain a better understanding of the current situation of PD in Africa. The countries included in this National Institute for Health and Care Research (NIHR) Global Health Research Group (Egypt, Ethiopia, Ghana, Kenya, Nigeria, South Africa and Tanzania) represent diverse African geographies and genetic profiles, with differing resources, healthcare systems, health and social protection schemes, and policies. The research team is composed of experts in the field with vast experience in PD, jointly led by a UK-based and Africa-based investigator. DISCUSSION: Despite the increasing prevalence of PD globally, robust data on the disease from Africa are lacking. Existing data point towards the poor awareness of PD and other neurological disorders on the continent and subsequent challenges with stigma, and limited access to affordable services and medication. This multi-site study will be the first of its kind in Africa. The data collected across the proposed sub-studies will provide novel and conclusive insights into the situation of PD. The selected country sites will allow for useful comparisons and make results relevant to other low- and middle-income countries. This grant is timely, as global recognition of PD and the public health challenge it poses builds. The work will contribute to broader initiatives, including the World Health Organization's Intersectoral global action plan on epilepsy and other neurological disorders. TRIAL REGISTRATION: https://doi.org/10.1186/ISRCTN77014546 .

Gender differences in microRNA expression in levodopa-naive PD patients.

Gender is an important factor influencing epidemiological and clinical features of Parkinson's disease (PD). We aimed to evaluate gender differences in the expression of a panel of miRNAs (miR-34a-5p, miR-146a, miR-155, miR-29a, miR-106a) possibly involved in the pathophysiology or progression of disease. Serum samples were obtained from 104 PD patients (58 men and 46 women) never treated with levodopa. We measured levels of miRNAs using quantitative PCR. Correlations between miRNA expression and clinical data were assessed using the Spearman's correlation test. We used STRING to evaluate co-expression relationship among target genes. MiR-34a-5p was significantly upregulated in PD male patients compared to PD female patients (fc: 1.62; p < 0.0001). No correlation was found with age, BMI, and disease severity, assessed by UPDRS III scale, in male and female patients. MiR-146a-5p was significantly upregulated in female as compared to male patients (fc: 3.44; p < 0.0001) and a significant correlation was also observed between disease duration and mir-146a-5p. No differences were found in the expression of miR-29a, miR-106a-5p and miR-155 between genders. Predicted target genes for miR-34a-5p and miR-146-5p and protein interactions in biological processes were reported. Our study supports the hypothesis that there are gender-specific differences in serum miRNAs expression in PD patients. Follow-up of this cohort is needed to understand if these differences may affect disease progression and response to treatment.

SARS-CoV-2-specific CD4+ and CD8+ T cell responses can originate from cross-reactive CMV-specific T cells.

Detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific CD4+ and CD8+ T cells in SARS-CoV-2-unexposed donors has been explained by the presence of T cells primed by other coronaviruses. However, based on the relatively high frequency and prevalence of cross-reactive T cells, we hypothesized cytomegalovirus (CMV) may induce these cross-reactive T cells. Stimulation of pre-pandemic cryo-preserved peripheral blood mononuclear cells (PBMCs) with SARS-CoV-2 peptides revealed that frequencies of SARS-CoV-2-specific T cells were higher in CMV-seropositive donors. Characterization of these T cells demonstrated that membrane-specific CD4+ and spike-specific CD8+ T cells originate from cross-reactive CMV-specific T cells. Spike-specific CD8+ T cells recognize SARS-CoV-2 spike peptide FVSNGTHWF (FVS) and dissimilar CMV pp65 peptide IPSINVHHY (IPS) presented by HLA-B*35:01. These dual IPS/FVS-reactive CD8+ T cells were found in multiple donors as well as severe COVID-19 patients and shared a common T cell receptor (TCR), illustrating that IPS/FVS-cross-reactivity is caused by a public TCR. In conclusion, CMV-specific T cells cross-react with SARS-CoV-2, despite low sequence homology between the two viruses, and may contribute to the pre-existing immunity against SARS-CoV-2.

Human T cells recognize HLA-DP-bound peptides in two orientations.

Human leukocyte antigen (HLA) molecules present small peptide antigens to T cells, thereby allowing them to recognize pathogen-infected and cancer cells. A central dogma over the last 50+ y is that peptide binding to HLA molecules is mediated by the docking of side chains of particular amino acids in the peptide into pockets in the HLA molecules in a conserved N- to C-terminal orientation. Whether peptides can be presented in a reversed C- to N-terminal orientation remains unclear. Here, we performed large-scale identification of peptides bound to HLA-DP molecules and observed that in addition to peptide binding in an N- to C-terminal orientation, in 9 out of 14 HLA-DP allotypes, reverse motifs are found, compatible with C- to N-terminal peptide binding. Moreover, we isolated high-avidity human cytomegalovirus (CMV)-specific HLA-DP-restricted CD4+ T cells from the memory repertoire of healthy donors and demonstrate that such T cells recognized CMV-derived peptides bound to HLA-DPB1*01:01 or *05:01 in a reverse C- to N-terminal manner. Finally, we obtained a high-resolution HLA-DPB1*01:01-CMVpp65(142-158) peptide crystal structure, which is the molecular basis for C- to N-terminal peptide binding to HLA-DP. Our results point to unique features of HLA-DP molecules that substantially broaden the HLA class II bound peptide repertoire to combat pathogens and eliminate cancer cells.

Immune Determinants of Viral Clearance in Hospitalised COVID-19 Patients: Reduced Circulating Naïve CD4+ T Cell Counts Correspond with Delayed Viral Clearance.

Virus-specific cellular and humoral responses are major determinants for protection from critical illness after SARS-CoV-2 infection. However, the magnitude of the contribution of each of the components to viral clearance remains unclear. Here, we studied the timing of viral clearance in relation to 122 immune parameters in 102 hospitalised patients with moderate and severe COVID-19 in a longitudinal design. Delayed viral clearance was associated with more severe disease and was associated with higher levels of SARS-CoV-2-specific (neutralising) antibodies over time, increased numbers of neutrophils, monocytes, basophils, and a range of pro-inflammatory cyto-/chemokines illustrating ongoing, partially Th2 dominating, immune activation. In contrast, early viral clearance and less critical illness correlated with the peak of neutralising antibodies, higher levels of CD4 T cells, and in particular naïve CD4+ T cells, suggesting their role in early control of SARS-CoV-2 possibly by proving appropriate B cell help. Higher counts of naïve CD4+ T cells also correlated with lower levels of MIF, IL-9, and TNF-beta, suggesting an indirect role in averting prolonged virus-induced tissue damage. Collectively, our data show that naïve CD4+ T cell play a critical role in rapid viral T cell control, obviating aberrant antibody and cytokine profiles and disease deterioration. These data may help in guiding risk stratification for severe COVID-19.

SARS-CoV-2-specific CD4+ and CD8+ T cell responses can originate from cross-reactive CMV-specific T cells

Detection of SARS-coronavirus-2 (SARS-CoV-2) specific CD4+ and CD8+ T cells in SARS-CoV-2-unexposed donors has been explained by the presence of T cells primed by other coronaviruses. However, based on the relative high frequency and prevalence of cross-reactive T cells, we hypothesized CMV may induce these cross-reactive T cells. Stimulation of pre-pandemic cryo-preserved PBMCs with SARS-CoV-2 peptides revealed that frequencies of SARS-CoV-2-specific T cells were higher in CMV-seropositive donors. Characterization of these T cells demonstrated that membrane-specific CD4+ and spike-specific CD8+ T cells originate from cross-reactive CMV-specific T cells. Spike-specific CD8+ T cells recognize SARS-CoV-2 spike peptide FVSNGTHWF (FVS) and dissimilar CMV pp65 peptide IPSINVHHY (IPS) presented by HLA-B*35:01. These dual IPS/FVS-reactive CD8+ T cells were found in multiple donors as well as severe COVID-19 patients and shared a common T cell receptor (TCR), illustrating that IPS/FVS-cross-reactivity is caused by a public TCR. In conclusion, CMV-specific T cells cross-react with SARS-CoV-2, despite low sequence homology between the two viruses, and may contribute to the pre-existing immunity against SARS-CoV-2.

Prolonged activation of nasal immune cell populations and development of tissue-resident SARS-CoV-2-specific CD8+ T cell responses following COVID-19.

Systemic immune cell dynamics during coronavirus disease 2019 (COVID-19) are extensively documented, but these are less well studied in the (upper) respiratory tract, where severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) replicates1-6. Here, we characterized nasal and systemic immune cells in individuals with COVID-19 who were hospitalized or convalescent and compared the immune cells to those seen in healthy donors. We observed increased nasal granulocytes, monocytes, CD11c+ natural killer (NK) cells and CD4+ T effector cells during acute COVID-19. The mucosal proinflammatory populations positively associated with peripheral blood human leukocyte antigen (HLA)-DRlow monocytes, CD38+PD1+CD4+ T effector (Teff) cells and plasmablasts. However, there was no general lymphopenia in nasal mucosa, unlike in peripheral blood. Moreover, nasal neutrophils negatively associated with oxygen saturation levels in blood. Following convalescence, nasal immune cells mostly normalized, except for CD127+ granulocytes and CD38+CD8+ tissue-resident memory T cells (TRM). SARS-CoV-2-specific CD8+ T cells persisted at least 2 months after viral clearance in the nasal mucosa, indicating that COVID-19 has both transient and long-term effects on upper respiratory tract immune responses.

Convalescent Plasma in a Patient with Protracted COVID-19 and Secondary Hypogammaglobulinemia Due to Chronic Lymphocytic Leukemia: Buying Time to Develop Immunity?

It is not exactly clear yet which type of immune response prevails to accomplish viral clearance in coronavirus disease 2019 (COVID-19). Studying a patient with chronic lymphocytic leukemia and hypogammaglobulinemia who suffered from COVID-19 provided insight in the immunological responses after treatment with COVID-19 convalescent plasma (CCP). Treatment consisted of oxygen, repeated glucocorticosteroids and multiple dosages of CCP guided by antibody levels. Retrospectively performed humoral and cellular immunity analysis made clear that not every serological test for COVID-19 is appropriate for follow-up of sufficient neutralizing antibodies after CCP. In retrospect, we think that CCP merely bought time for this patient to develop an adequate cellular immune response which led to viral clearance and ultimately clinical recovery.

Prolonged activation of nasal immune cell populations and development of tissue-resident SARS-CoV-2 specific CD8 T cell responses following COVID-19

The immune system plays a major role in Coronavirus Disease 2019 (COVID-19) pathogenesis, viral clearance and protection against re-infection. Immune cell dynamics during COVID-19 have been extensively documented in peripheral blood, but remain elusive in the respiratory tract. We performed minimally-invasive nasal curettage and mass cytometry to characterize nasal immune cells of COVID-19 patients during and 5-6 weeks after hospitalization. Contrary to observations in blood, no general T cell depletion at the nasal mucosa could be detected. Instead, we observed increased numbers of nasal granulocytes, monocytes, CD11c+ NK cells and exhausted CD4+ T effector memory cells during acute COVID-19 compared to age-matched healthy controls. These pro-inflammatory responses were found associated with viral load, while neutrophils also negatively correlated with oxygen saturation levels. Cell numbers mostly normalized following convalescence, except for persisting CD127+ granulocytes and activated T cells, including CD38+ CD8+ tissue-resident memory T cells. Moreover, we identified SARS-CoV-2 specific CD8+ T cells in the nasal mucosa in convalescent patients. Thus, COVID-19 has both transient and long-term effects on the immune system in the upper airway.

Simultaneous Deletion of Endogenous TCRαß for TCR Gene Therapy Creates an Improved and Safe Cellular Therapeutic.

Generation of an optimal T cell therapeutic expressing high frequencies of transgenic T cell receptor (tgTCR) is essential for improving TCR gene therapy. Upon TCR gene transfer, presence of endogenous TCRαß reduces expression of tgTCR due to TCR mixed-dimer formation and competition for binding CD3. Knockout (KO) of endogenous TCRαß was recently achieved using CRISPR/Cas9 editing of the TRAC or TRBC loci, resulting in increased expression and function of tgTCR. Here, we adopt this approach into current protocols for generating T cell populations expressing tgTCR to validate this strategy in the context of four clinically relevant TCRs. First, simultaneous editing of TRAC and TRBC loci was reproducible and resulted in high double KO efficiencies in bulk CD8 T cells. Next, tgTCR expression was significantly higher in double TRAC/BC KO conditions for all TCRs tested, including those that contained structural modifications to encourage preferential pairing. Finally, increased expression of tgTCR in edited T cell populations allowed for increased recognition of antigen expressing tumor targets and prolonged control of tumor outgrowth in a preclinical model of multiple myeloma. In conclusion, CRISPR/Cas9-mediated KO of both endogenous TCRαß chains can be incorporated in current T cell production protocols and is preferential to ensure an improved and safe clinical therapeutic.

Multiparametric MR imaging of Parkinsonisms at 3 tesla: Its role in the differentiation of idiopathic Parkinson's disease versus atypical Parkinsonian disorders.

OBJECTIVES: The Nigrosome-1 and putaminal hypointensity depicted on susceptibility-weighted imaging (SWI), and midbrain atrophy assessed on T1-weighted are some of the most common radiological parameters to diagnose Parkinsonism at Magnetic Resonance (MR) imaging. Our aim is to assess the feasibility of these signs in the differentiation of Idiopathic Parkinson's disease (IPD) patients versus disease (DC) and healthy controls (HC) and in the assessment of the Atypical Progressive Parkinsonisms (APPs). METHODS: Presence or loss of the Nigrosome-1 was assessed retrospectively on multiple-echo SWI obtained on a 3 T scan by two neuroradiologists. Results were compared with the 123I-FP-CIT SPECT images. Morphologic diagnostic features suggestive of APPs such as midbrain atrophy and putaminal hypointensity were evaluated by qualitative scores. The midbrain and putaminal scores were summed (combined score) and then added to the Nigrosome-1 score (global score). RESULTS: The study included 126 patients with IPD (n = 56), APPs patients (n = 30; 18 PSP, 3 MSA-C, 9 MSA-P), 16 DC and 24 HC. Sensitivity and specificity of the Nigrosome-1 in discriminating IPD from controls were 96,43% and 85.00%, APPs from controls were 100% and 85%, IPD from APPs were 96,43% and 0% respectively. Combined score for midbrain atrophy and putaminal hypointensity resulted in the most accurate for distinguishing APPs from IPD with a value of ≥ 2 (AUC = 0.98). CONCLUSION: Nigrosome-1 is a valid tool to differentiate IPD-APPs from controls. The combined score of midbrain atrophy and putaminal hypointensity represents a valid diagnostic pointer in the differential diagnosis of APPs from IPD.

The relationship between cerebral vascular disease and parkinsonism: The VADO study.

BACKGROUND: The observation of gait abnormalities, parkinsonism and vascular lesions in elderly patients is often reported as vascular parkinsonism (VP). However the status of striatal dopamine transporter (DAT) and the effects of brain vascular lesions on motor features and levodopa responsiveness are poorly defined. METHODS: We recorded clinical features, chronic response to levodopa and vascular risk factors in a cross-sectional cohort of consecutive elderly patients with possible Parkinson's disease (PD) or VP recruited in 20 centers in Italy. RESULTS: We included a total of 158 patients. Onset of motor symptoms was asymmetric in 93 (59%) and symmetric in 65 patients (41%). Symmetric motor onset was associated with greater disease severity. Chronic levodopa response was positive in 75 (47.8%) and negative in 82 patients (52.2%). A negative response to levodopa was associated with greater frequency of symmetric onset of motor symptoms, worst disease severity, absence of dyskinesia and greater number of vascular risk factors. Frontal lobe showed largest vascular load. Striatal DAT was normal in 48 (30.4%) and abnormal in 110 (69.6%) patients. Patients with normal DAT binding showed higher vascular load at MRI. Significant predictive factors of worst disease severity and negative response to levodopa were hypertension, vascular lesions in basal ganglia/periventricular regions, and normal DAT uptake. CONCLUSIONS: Multiple cerebral vascular lesions modify clinical presentation and severity in patients with parkinsonism and this is underlined by specific risk factors primarily hypertension. Striatal DAT assessment is helpful in identifying patients where therapy benefit is less likely.

Drug-induced deactivation of inhibitory networks predicts pathological gambling in PD.

OBJECTIVE: Some patients with Parkinson disease (PD) develop pathological gambling when treated with dopamine agonists (DAs). However, little is known about DA-induced changes in neuronal networks that may underpin this drug-induced change in behavior in vulnerable individuals. In this case-control study, we aimed to investigate DA-induced changes in brain activity that may differentiate patients with PD with DA-induced pathological gambling (gamblers) from patients with PD without such a history (controls). METHODS: Following overnight withdrawal of antiparkinsonian medication, patients were studied with H2(15)O PET before and after administration of DA (3 mg apomorphine) to measure changes in regional cerebral blood flow as an index of regional brain activity during a card selection game with probabilistic feedback. RESULTS: We observed that the direction of DA-related activity change in brain areas that are implicated in impulse control and response inhibition (lateral orbitofrontal cortex, rostral cingulate zone, amygdala, external pallidum) distinguished gamblers from controls. DA significantly increased activity in these areas in controls, while gamblers showed a significant DA-induced reduction of activity. CONCLUSIONS: We propose that in vulnerable patients with PD, DAs produce an abnormal neuronal pattern that resembles those found in nonparkinsonian pathological gambling and drug addiction. DA-induced disruption of inhibitory key functions--outcome monitoring (rostral cingulate zone), acquisition and retention of negative action-outcome associations (amygdala and lateral orbitofrontal cortex)--together with restricted access of those areas to executive control (external pallidum)--may well explain loss of impulse control and response inhibition in vulnerable patients with PD, thereby fostering the development of pathological gambling.

Cerebral activity modulation by extradural motor cortex stimulation in Parkinson's disease: a perfusion SPECT study.

Extradural motor cortex stimulation (EMCS) has been proposed as alternative to deep brain stimulation (DBS) in the treatment of Parkinson's disease (PD). Its mechanisms of action are still unclear. Neuroimaging evidenced motor cortical dysfunction in PD that can be reversed by therapy. We performed left hemisphere EMCS surgery in six advanced PD patients fulfilling CAPSIT criteria for DBS with the exception of age >70 years. After 6 months, we measured regional cerebral blood flow (rCBF) at rest with SPECT and Tc-99m cysteinate dimer bicisate off-medication with stimulator off and on. Clinical assessment included Unified Parkinson's Disease Rating Scale part II and III, Abnormal Involuntary Movement Scale and mean dopaminergic medication dosage. We used statistical parametric mapping for imaging data analysis. Clinically we observed no mean changes in motor scales, although blinded evaluation revealed some benefit in individual patients. We found significant rCBF decrements in the pre-central gyrus, pre-motor cortex and caudate nucleus bilaterally, left prefrontal areas and right thalamus. Perfusion increments were found in cerebellum bilaterally. EMCS determined significant modulation of neuronal activity within the cortico-basal ganglia-thalamo-cortical motor loop in our cohort of advanced PD patients. However, these effects were paralleled by mild and variable clinical efficacy.