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Background: Impaired generation of antibody responses define 'predominantly antibody immuno-deficiencies' (PAD) with reduced IgG and impaired vaccination responses. However, the antibody repertoire defects underpinning PAD are unknown. Here, we examine the antibody repertoire using mass spectrometry-based proteomics (MS-proteomics) in PAD and healthy controls (HC). Method(s): Following SARS-CoV-2 vaccination, anti -S1 ELISA, and live-virus neutralisation assays were assessed. Purified anti-S1 IgG and IgM was sequenced by MS-Proteomics to define immunoglobulin heavy chain variable region subfamily (IGHVsf) usage and somatic hypermutation (SHM). Result(s): 12 vaccine responsive PAD subjects were included, matched to 11 HC. Neutralisation and anti-S1 titres were reduced in PAD. Strikingly, all PAD subjects demonstrated restricted IgG IGHVsf utilisation, [median 3, (range 2-4), vs 6 (5-11) in HC, p<0.001], irrespective neutralisation or total antibody response. IgG SHM and IgM repertoire did not differ but IgG IGHV 3-7 utilisation was less frequent in PAD. Conclusion(s): MS proteomics uncovers stereotyped anti-S1 IgG IGHVsf restriction in PAD subjects following vaccination. Our results suggest that a relatively pauci-clonal antibody repertoire can produce a functional immune response, otherwise masked by traditional serology measures. Further studies to uncover the determinants of antibody repertoire breadth and elaborate on this novel approach to assessing serological responses are required. Copyright © 2022
ABSTRACT
Background: Adults and childrenwith cancer are susceptible to severe SARS-CoV-2 disease. Vaccination is protective;data beyond initial response and regarding effect of booster doses are lacking in cancer patients. Method(s): The SerOzNET study assesses SARS-CoV-2 vaccine response in haematological and solid cancer patients aged 5 and older. Patients are recruited pre dose 1 and receive standard BNT162b2 (Pfizer) or ChadOx1-S (AstraZeneca) vaccine. Blood is taken at baseline and after each dose. Neutralizing antibody (NAb) titre, absolute antibody titre (Abbott), T cell response (IFN-gamma) and epigenetics are analysed. Clinical data are collected. Patients are followed for up to 3 months beyond dose 5. Result(s): 105 children (64% haem, 36% solid cancers) and 399 adults (35% haem, 65% solid cancers) were enrolled. In adults, NAb response rate increased after dose 3 (Post 2: 40% haem, 87%solid;Post 3:70%haem,97%solid). Post dose 2, predictors of nonresponse were ChadOx1-S vaccine (OR 3 p = .02), haem cancer (OR 14 p < .001), ECOG >=1 (OR 2.6 p = .01) and steroids (OR 5 p = .01). Post dose 3, only haem cancer predicted non-response (OR 16). IFN-gamma response is available for a subset, detectable in 41/90 (46%) postdose 1, 78/96 (81%) post-dose 2 and 35/42 (83%) post-dose 3;without significant difference between haem and solid cancer. In children, NAb response post dose 2 is available for 50 patients. Response rate between haem (19/31, 61%) and solid patients (13/19, 68%) was similar. IFN-gamma response post dose 2 was also similar: (14/22, 63%) vs solid patients (12/14, 85%) (p = .25). Analysis is ongoing. Conclusion(s): Response to two doses of SARS-CoV-2 vaccine is suboptimal in patients with cancer. The third priming dose is integral, with significantly higher response rates observed. 36% of children did not develop neutralizing antibodies post dose 2;subsequent doses are likely to be important for young patients.
ABSTRACT
Background: Patients (pts) with indolent lymphomas are at increased risk of severe COVID-19 infection. We have shown limited seroconversion and live viral neutralisation (VN), but preserved COVID-specific T cell responses after 2 doses of mRNA COVID-19 vaccination in such pts. (Beaton, B ASH 2021, 149348). A 3rd vaccine dose to complete primary vaccination has since been recommended. Aims: To assess humoral & cellular immune responses to a 3rd COVID-19 (mRNA) vaccination in pts with follicular lymphoma (FL) & Waldenström Macroglobulinemia (WM), including assessment of response after pausing BTKi therapy in WM pts. Methods: Patients with WM, FL & healthy controls (HC) were enrolled in a prospective observational study to measure immune responses 21-28 days after a 3rd mRNA COVID-19 vaccine. Immune response was measured by mean fluorescence intensity (MFI) of anti-SARS-CoV-2 spike antibodies (ASAb) obtained using a high-sensitivity live cell assay, live VN to a panel of SARS-CoV-2 variants of concern, and CD4+ & CD8+ antigen-specific T cell responses. The associated TRIBECA (TReatment Interruption of BTKi to Enhance COVID-19 Antibody response) study sought to determine if a superior immune response could be gained by pause of BTKi therapy prior to and up to 4 weeks after 3rd vaccine dose. Patients were closely monitored during the BTKi pause with weekly clinical, full blood count and IgM assessments. WM pts receiving a 3rd dose while continuing on BTKi served as a control. Statistical analysis of medians between cohorts were compared by the non-parametric Mann-Whitney (Graphpad Prism). Comparison of medians between paired grouped data was assessed by 2-way ANOVA. Results: To date, 56 of 125 pts had their ASAb measured following 3rd vaccine dose administered between October 2021 and January 2022: 28 WM pts (including 6/9 WM pts on the BTKi pause sub-study), 24 FL pts and 4 HC. Median age was 68 years with 21 females and 35 males. Median follow up from 2nd dose was 140 days (range: 79- 170 days). In antibody responders, median MFI fell from 163042 (IQR 82663-249934) 28 days post 2nd dose to 52117 (IQR 19942-60973) (p<0.0001) immediately before the 3rd dose. Median MFI in all FL & WM pts pre- 3rd dose vaccine was 17111 (IQR 0-52650), rising significantly post 3rd dose to a median of 86730 (IQR 0-221937). Only 4/20 pts without measurable ASAb prior to the 3rd dose (2 WM, 2 FL) developed measurable ASAb following the 3rd dose: 16/20 patients (8 WM, 8FL, all treated) had no ASAb response. The median MFI in pts who underwent a BTKi treatment pause rose from 9151 (IQR 1671-21232) pre-3rd dose to 87720 (IQR 2785-152195) post 3rd dose, significantly higher than the median MFI in WM pts who did not pause their BTKi , which rose from 16769 (IQR 218-22447) pre- to 20252 (IQR 168-114262) post 3rd dose, (p = 0.016). Of the 5/125 with COVID infection in the study to date, only one patient (without measurable ASAb) in this 3-dose cohort had COVID, requiring intensive care support. Summary/Conclusion: Most WM & FL pts who responded to a 2nd dose COVID vaccine showed a decline in ASAb titre over time which increased following a 3rd mRNA vaccine. Only 20% of pts without detectable ASAb pre- 3rd dose showed improvement post 3rd dose, highlighting the importance of other COVID protection strategies in these pts. Although initial numbers are small, there may be a higher increment in ASAb when BTKi therapy is paused around the time of vaccination. Comprehensive immune analysis, including VN and T-cell response on the entire FL & WM cohort will be presented at the EHA congress.
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Background: COVID-19 infection has poor outcomes for patients (pts) with cancer. Understanding vaccine response as a correlate of protection from severe infection is essential to advise pts regarding protective behaviours and optimal vaccine schedule. This Australian cohort is unique due to low rates of COVID-19 exposure at study entry (July-November 2021). and use of a 3 dose schedule. Pts initially received 2 doses of either BNT162b2 (Pf) at a 3 week interval, or ChadOx1-S (AZ) at a 6 week interval, all then received a 3rd dose, either mRNA-1273 (Mod) or Pf after 2-4 months, and finally a 4th dose at an interval of a further 3 months, for a subset. Methods: SerOzNET (ACTRN12621001004853) has enrolled pts with solid and haematological (haem) cancers prior to initial vaccination. Serial blood samples were processed for serum, PBMC and PMN at timepoints: 0, then 3-4 weeks post dose 1 then 2 then 3 then 4 (where administered). We report here neutralizing antibodies (nAb) against wild type (wt) and delta and omicron variants of concern (VOC);quantitative S-protein IgG antibody (Abbott);Tcell correlates measured by levels of interferon-g (IFN g), tumour necrosis factor-a, interleukins (IL-) 2/ 4/5/13;and epigenetic profiling of T cells. Results: The cohort consists of 401 pts with median age 58 (range 18-85);59% female;128 (32%) haem cancers. 377 (94%) are on current or recent (< 12 months) systemic therapy: 162 (43%) chemotherapy, 62 (16%) immunotherapy, 40 (10%) combined chemo/immunotherapy, 113 (29%) hormonal or targeted therapy. 42 (10%) received anti-CD20 therapy < 12 months, 6 (1.4%) had allogeneic stem cell transplant. NAb levels against wt are available for 256 pts post dose 1, 245 pts post dose 2 and 159 pts post dose 3 (will be updated). Response rates post dose were respectively 27%, 77% and 88%. Pts with haem cancer were less likely to respond to vaccination at any time compared to pts with solid cancer (p < 0.001, chi-squared test). After 3 doses, 3.8% of pts with solid cancer and 27.8% with haem cancer lacked NAb. NAb results to VOC delta are available for 92 pts post dose 2: 25/92 (27%) were negative, compared with a non-response rate to wt of 15% at same time in same pts. IFN-γ-Spike response was detectable in 18/31 (58%) and 24/30 (80%) pts post dose 1 and 2 respectively. 101 pts to date have received a 4th dose;data will be available at the meeting, as will epigenetic profiles and detailed clinicopathological correlations. Conclusions: This interim analysis shows that a significant proportion of pts with haem cancers (27.8%) lack protective Sars-CoV-2 antibodies following 3 vaccinations, whereas only 3.8% of solid cancer pts lack detectable response. Results from other B and T cell parameters may also be important in identifying pts less well protected by vaccination. Follow up is ongoing, response rate post 4th dose will be presented at the meeting.
ABSTRACT
Background: Defining cancer and treatment-related factors which influence protection against COVID-19 following vaccination are important given the worse outcomes following infection in this group. Sophisticated and detailed studies which go beyond a single measure are required particularly with correlation to multiple disease and treatment factors. This study cohort is unique due to (a) very low prior COVID-19 infection at time of sampling (July-Nov 2021), (b) vaccines studied were BNT162b2 (Pf) given 3 weeks apart or ChAdOx1 (AZ) spaced 12 weeks (dose 1, 2) (c) most participants then received a third dose 2 months later (heterologous for AZ). Methods: SerOzNET (ACTRN12621001004853) enrols Australian blood and solid cancer patients prior to vaccination, with serial blood analyses and qualitative measures. We measured neutralizing antibodies (nAb) against SARS-CoV-2 wild type (wt) and variants of concern delta and omicron, quantitative S-protein IgG antibody level (Abbott), and T-cell correlates (interferon-g, tumour necrosis factor-a, interleukins 2/4/5/13) and epigenetic profiling at baseline and 3-4 weeks post dose 1, 2 +/- 3.Results: 379 participants were included, median age 58 years (IQR 47-66) and 60% female. 30% participants had hematological malignancies with the remainder solid organ cancers. 90% patients were on current systemic cancer treatment (most commonly chemotherapy in 41%, chemoimmunotherapy or immunotherapy in 20%). In 331 patients where treatment intent was recorded, 47% was palliative. Only one patient had known prior COVID-19 infection. Of the initial 94 participants who received Pf vaccination, median (IQR) neutralizing antibody titre 4 weeks following dose 2 was 80 (40-160) for SARS-CoV-2 wt and 40 (0-80) for delta variant. Conclusion: Neutralizing antibody titres in this Australian cancer population following Pf vaccination appear lower than those reported elsewhere such as CAPTURE study (Fendler et al, 2021), possibly related to shorter interdose interval. Preliminary data highlights low nAb titres as expected in haematology patients but also in some cases with treatment not traditionally associated with immunosuppression such as hormonal therapy. These results will be updated in February 2022 with third dose, AZ and omicron variant data.
ABSTRACT
Introduction Lymphoid malignancies are a risk factor for severe COVID-19. Vaccination with BNT162b2 protects the general population from severe disease, but recent studies have shown limited seroconversion after vaccination in patients with lymphoid malignancy. This reduced response is likely related to disease and treatment factors altering both humoral and cellular immunity. Assessing response in patients with the indolent lymphomas, Waldenström's Macroglobulinaemia (WM) and Follicular Lymphoma (FL), including cohorts on differing treatment regimens, may help elucidate some of these factors. Australia has had low prevalence of SARS-CoV-2 infection to date, affording a unique opportunity to assess efficacy to vaccination without the confounding impact of endemic infection. Methods Patients with WM and FL and controls were enrolled in a prospective study of immune response after two doses of BNT162b2 administered 21 days apart. The study had Human Research Ethics Committee approval and all patients gave informed consent prior to participation. Recruitment was targeted to obtain comparable proportions of controls to treatment cohorts. PBMC and sera were collected from participants immediately prior to the first dose (T1), at day 21 immediately prior to the second dose (T2), and day 49 (+/-7d) (T3). Immune response was measured by: flow cytometric detection of anti-SARS-CoV-2 spike antibodies (ASAb), performed using our recently validated flow cytometric live cell assay (Tea et.al. PLoS Medicine 2021) with increased sensitivity compared to currently available commercial ELISAs;live virus neutralisation to a panel of SARS-CoV-2 variants of concern;and CD4+and CD8+ antigenspecific T cell responses. Statistical analysis of medians between cohorts were compared by the Mann-Whitney non-parametric test using Graphpad Prism. Initial ASAb IgG data for T1 and T2 is presented here. Complete immune response data at all time points will be available for the ASH meeting. Results Eighty-five participants received their first dose of BNT162b2 from 18 May 2021 to 7 June 2021 with a second dose 21 days later: 72 lymphoma patients (WM and FL) and 13 age-matched healthy volunteers (controls). Of 37 with WM [19 (51.3%) female, median 71 years (IQR 63-74)] 9 were treatment naïve (WMN), 15 had received rituximab-chemotherapy (WMT), and 13 were currently treated with a Bruton Tyrosine Kinase inhibitor (BTKi): 5 ibrutinib, 8 zanubrutinib (WMB). Of 35 patients with FL [16 (45.7%) female, median 65 years (IQR 54-71)], 11 were treatment naïve (FLN), and 24 had received rituximab-chemotherapy (FLT). Of the 13 controls 8 (61.5%) were female, median age 72 years (IQR 57-74)]. No participants had detectable ASAb at T1, confirming no prior SARS-CoV-2 exposure. Figure 1 shows ASAb results at T2. The median mean fluorescence intensity (MFI) of healthy controls: 60802 (IQR 17565 -78443), is higher than all WM: 0 (IQR 0-15010) p<0.0001, and all FL patients: 1687 (IQR 0-25421) p=0.002, Fig 1A. The median MFI of controls was higher than WMN (p=0.036), but not higher than FLN (p=0.28). The median MFI of WMN: 20074 (IQR 5421-35695), is higher than WMB: 0 (IQR 0-4217) p=0.018, but not significantly higher than WMT: 0 (IQR 0-14356) p=0.13, Fig 1B. Median MFI of FLN: 31476 (IQR 19351-51317), is higher than FLT: 0 (IQR 0-32849) p=0.01, Fig 1B. Conclusion These early serological data show measurable ASAb in all healthy controls 21 days post first dose of BNT162b2 vaccination. Treatment naïve patients had a better response than treated patients, and this did not differ significantly to healthy controls in the FL cohort. In WM, patients on BTKi had a significantly reduced response compared to treatment-naïve patients. This same reduction was not observed in the chemotherapy-rituximab cohort, but the characteristics of early responders versus non-responders, including time since last therapy, is being analysed. FL patients treated with chemotherapy-rituximab had a significantly reduced response compared to the treatmentnaïve cohort. Time constraints before the submission deadline prevented reporting of all mature vaccination response data. Measurement of ASAb one month after second vaccination, live virus neutralisation to a panel of SARS-CoV-2 variants of concern, and CD4+ and CD8+ antigen-specific T cell responses at T1, T2 and T3, to fully characterise the immune response to BNT162b2, will be reported at the ASH meeting.