3.1. Methotrexate
Methotrexate (MTX) is widely used in AIIRD. It inhibits dihydrofolate reductase, thereby preventing the reduction of dihydrobiopterin (BH2) to tetrahydrobiopterin (BH4). This leads to nitric oxide synthase uncoupling and increased sensitivity of T cells to apoptosis [
19]. It also increases adenosine release which then activates adenosine receptor A2a. These mechanisms inhibit the activation of nuclear factor-κB (NF-κB) and serve to diminish immune responses [
19].
Various studies have reported reduced humoral response to mRNA and inactivated virus vaccines in patients with AIIRD receiving MTX [
8,
20,
21,
22,
23,
24]. Furer et al. found that the seropositivity rate measured 2–6 weeks after the second vaccine dose (Pfizer—BioNTech BNT162b2 or Moderna mRNA-1273) was 92% in patients on MTX monotherapy compared with 100% in controls (
p = 0.02) [
8]. Similarly, Moyon et al. found that MTX use in SLE patients was associated with lower anti-spike antibody production measured 14 days after the second dose (β = −122, 95% CI: −184 to −61,
p p 20]. MTX in combination with other immunosuppressants also significantly reduced seroconversion rates compared to that in controls (84% vs. 100%,
p 8]. MTX when used in combination with tumor necrosis factor inhibitors (TNFi) reduced the rate of seropositivity to 93% (
p = 0.04) [
8]. A cohort study suggested that although the early immunosuppressive of MTX was attenuated in patients with concomitant TNFi use, at 6 months this combination had the lowest adequate humoral response rate of any medication even when compared to all MTX use (alone or in combination with other medications) [
25].
There were a few studies on the impact of MTX on the cellular immune response in AIIRD patients receiving the COVID-19 vaccine. In a study by Moyon et al., the T cell responses measured by IGRA following SARS-CoV2 spike protein stimulation was 57% in patients with detectable neutralising activity, and only 10% in those without detectable neutralising activity [
20]. Another observational study demonstrated reduced CD4 T cell response (interleukin(IL)-2 and interferon-gamma (IFN-γ) production) and CD8 T cell response (S-induced Granzyme A/B detection) in MTX compared to TNFi use, leflunomide use, and healthy controls (CD4 T cell response: 80% vs. 100%, 100% and 100%, respectively; CD8 T cell response: 76% vs. 100%, 100% and 92%, respectively) [
26]. Compared with healthy adults and AIIRD patients not on MTX, the production of activated CD8 T cells (expressing Ki67 and CD38) and the granzyme B-producing subset of these activated CD8 T cells were not induced in AIIRD patients on MTX [
23].
Various treatment modification strategies have been proposed to improve the immunogenicity of the SARS-CoV-2 vaccine in AIIRD patients on MTX. While the American College of Rheumatology (ACR) Guidance recommended withholding MTX for 1–2 weeks after each of the two mRNA vaccine doses, Moutsopoulos et al. recommended withholding anti-metabolites 10 days before and 10 days after each vaccination dose [
27,
28]. Tzioufas et al. found that AIIRD responders either on MTX monotherapy or combined MTX-based regimes underwent more frequently extended treatment modifications as Moutsopoulos recommended than non-responders (21.49 vs. 4.28%, OR = 6.115, 95% CI: 2.033–18.91,
p = 0.0001) [
9]. These patients developed antibody titres that were comparable with patients without immunomodulatory therapies and significantly higher than patients on immunomodulatory therapies that were not suspended [
9]. The VROOM study was a randomised open-label superiority trial which showed that a 2-week interruption of MTX in AIIRD patients improved S1-receptor binding domain (RBD) antibody response. However, the authors cautioned about the high risk of bias due to the exclusion of participants after randomisation for previous SARS-CoV-2 infections and disease flare-ups, a dropout rate of 33%, and different dropout rates in the suspended MTX group compared to the continued treatment group [
29]. A single-centre open-labeled, assessor-masked, randomised controlled trial in India demonstrated that the final antibody titres were higher among patients with either psoriatic arthritis or rheumatoid arthritis who withhold MTX for 2 weeks after both doses or only after the second dose of ChAdOx1 nCov-19 (Oxford—AstraZeneca) vaccine compared to those who did not withhold MTX. Of interest, the humoral response was similar between patients withholding MTX for 2 weeks after both doses or only after the second dose of ChAdOx1 nCov-19 (Oxford—AstraZeneca) vaccine, and fewer flare-ups of arthritis in the latter group [
30].
3.4. Azathioprine
Azathioprine, a purine analogue, inhibits lymphocyte proliferation by blocking purine synthesis, thereby dampening inflammation [
32]. Santos et al. observed a seroconversion rate of 57% in AIIRD patients on azathioprine following two vaccine doses (mRNA SARS-CoV-2 vaccine). Azathioprine was shown to impair humoral responses with reduced antibody titres compared to healthy controls (178.5 ± 142 vs. 526.3 ± 2078 BAU/mL,
p = 0.01) [
26]. Additionally, only 42% of these patients exhibited a complete cellular response, significantly lower than the 96% average in healthy controls, although this finding is based on a small sample size of seven [
26]. In another study, Carruthers et al. reported a seroconversion rate of 87.5% in patients taking azathioprine for ANCA-associated vasculitis [
33].
3.5. Cyclosporin/Tacrolimus
Calcineurin inhibitors, such as cyclosporin and tacrolimus, are known to inhibit T cell proliferation [
32]. A prospective observational study by Zecca et al. identified calcineurin inhibitors as an independent predictor of impaired humoral responses and vaccination failure in patients, with an odds ratio of 17.5 (95% CI: 3.1–99.0,
p = 0.0012) for vaccination failure, although the risks may be confounded by concomitant MMF treatment [
34]. Prendecki et al. reported a 67.7% seroconversion rate among patients with autoimmune rheumatic and glomerular diseases treated with tacrolimus [
35]. While the proportion of cellular responses in tacrolimus patients did not significantly differ from non-tacrolimus patients, the magnitude of this response was significantly lower in the former group [
35].
3.6. Mycophenolate Mofetil
MMF, a prodrug of mycophenolic acid, blocks de novo purine synthesis, thereby reducing cell proliferation [
32]. MMF treatment has been shown to significantly reduce seroconversion rates (69% in MMF group vs. 100% in healthy controls,
p 36]. A similar dose-dependent impact on humoral immune response was observed in another study [
37]. Sieiro et al. showed that 70% of patients on MMF showed a complete cellular response after a two-dose mRNA COVID-19 vaccine [
26]. Additionally, Wieske et al. found that a third vaccine dose could enhance seroconversion rates in MMF-treated patients, increasing from 52.6% after the second dose to 89.5% after the third [
38].