Objectives The mRNA-based COVID-19 vaccines are now employed globally and have shown high efficacy in preventing SARS-CoV-2 infection. However, less is known about the vaccine efficacy in immune suppressed individuals. This study sought to explore whether humoral immunity to the COVID-19 vaccine BNT162b2 is altered in rheumatoid arthritis patients treated with Janus kinase inhibitors by analyzing antibodies titer, neutralization activity and B cell responses. Methods We collected plasma samples from 12 rheumatoid arthritis patients who were treated with Janus kinase inhibitors and received two doses of the BNT162b2 vaccine, as well as 26 healthy individuals who were vaccinated with the same vaccine. We analyzed the quantity of the anti-spike IgG and IgA antibodies that were elicited following the BNT162b2 vaccination, the plasma neutralization capacity and the responsiveness of the B-lymphocytes. We used ELISA to quantify antibody titers, and plasma neutralization assay was used to determine virus neutralization capacity. Alteration in expression of genes that are associated with B cell activation and germinal center response were analyzed by qPCR. Results Reduced levels of anti-spike IgG antibodies and neutralization capacity were seen in the rheumatoid arthritis patients who were treated with JAK inhibitors in comparison with healthy individuals. Furthermore, B cell responsiveness to the SARS-CoV-2 spike protein were reduced in the rheumatoid arthritis patients. Conclusion Rheumatoid arthritis patients who are treated with JAK inhibitors show suppressed humoral response following BNT162b2 vaccination, as revealed by the quantity and quality of the anti-spike antibodies.