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LETTERS TO THE EDITOR
Immunologic characterization of COVID-19 patients
with hematological cancer
The rapid spread of the coronavirus disease 2019 (COVID-19) pandemic is having a profound impact in oncologic care,1 with recent analyses suggesting that can- cer patients may have an increased risk of severe compli- cations, including hospitalization, respiratory failure and death.2 Severe events from initial onset of COVID-19 appear to be more frequent in individuals with blood malignancies versus other cancer types.2,3 Furthermore, an increased risk of death in patients with hematological cancer infected by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been identified in some studies,4,5 though unconfirmed in others.6,7 Here, we ana- lyzed outcomes and immune response to SARS-CoV-2 in a large series of 513 patients with COVID-19 that includ- ed ten cases with blood cancer. This study confirms that the later have a greater risk of fatal COVID-19 and shows for the first time that hematological patients display spe- cific immune alterations that could compromise a response to the infection.
The immune system reacts to viral infection with cel- lular and humoral responses. Thus, myelo- and lympho- suppression caused by cancer itself as well as cytotoxic treatment may pose a challenge to COVID-19 patients with hematological tumors.1 Indeed, lymphopenia has been associated with worse prognosis in the general pop- ulation8 and preliminary data showed lower neutrophil and lymphocyte counts in COVID-19 patients bearing hematological cancer.9 However, there are conflicting results supporting that both worsening of lymphopenia during COVID-19 and prior to SARS-CoV-2 infection and during COVID-19 had a beneficial impact on survival.9 Accordingly, the role of some antineoplastic drugs (e.g., inhibitors of PD-1, BTK, JAK1/2, XPO-1 and tyrosine kinases, as well as thalidomide) in mitigating the harmful immune response associated with severe COVID-19 is being investigated.10 At the same time, it was found that checkpoint inhibitor immunotherapy is a risk factor for severe outcomes in COVID-19 patients with cancer.3 Thus, greater knowledge on the immune status of hema- tological patients may be useful to optimize prevention, risk stratification and treatment strategies.
From March to May 2020, 513 consecutive patients with a SARS-CoV-2 positive result had peripheral blood (PB) samples taken at presentation for immune profiling using multidimensional flow cytometry (MFC). Additional PB samples were collected during follow-up in 167 of the 513 cases. Data was analyzed with a semi- automated pipeline that performs batch-analyses of MFC data to avoid variability intrinsic to manual analysis, and unveils full cellular diversity based on unbiased cluster- ing. In PB samples from 14 COVID-19 patients, higher- resolution MFC was performed to characterize antigen- dependent differentiation of B and T cells. Furthermore, various myeloid subsets and antigen-presenting cells were isolated by fluorescence-activated cell sorting (FACS) for transcriptome analysis using RNA sequencing (RNAseq) (all methods are described in the Online Supplementary Appendix). The Clínica Universidad de Navarra Ethics Committee approved the protocol and informed consent forms, required prior to patient enroll- ment. The study was conducted according to the ethical principles of the Declaration of Helsinki.
Of the 513 COVID-19 patients included in this study, 10 had a hematological tumor (Online Supplementary Table S1). These patients showed a similar frequency of hospi-
Table 1. Outcome of patients diagnosed with COVID-19 (n=513) regarding hospitalization, need of intensive care and survival.
Outcome No tumor (N=503)
Hospitalization,N(%) 385(77%)
Hematological tumor (N=10)
8(80%)
5 (50%)
3 (30%)
P
0.324
<0.001
<0.001
Intensive care, N (%)
Death, N (%)
27 (5%)
20 (4%)
talization compared to those without an active tumor (80% vs. 77%). By contrast, the frequency of hematolog- ical cases requiring intensive care (50%) and dying from COVID-19 (30%) was significantly higher than that observed in patients without hematological cancer (5% and 4%, respectively) (Table 1). These outcomes are con- sistent with previous results observed in the general pop- ulation and some series of hematological patients infect- ed by SARS-CoV-2.2,4,5
Studying immunological biomarkers has become noto- riously important in COVID-19, since immunopathology was suggested as a primary driver of morbidity and mor- tality in these patients.11 However, there are no studies analyzing the immune response to SARS-CoV-2 at pres- entation and during follow-up in hematological cases. Here, we performed a holistic and unbiased analysis of MFC data that enabled the systematic quantification of 17 cell types (including five myeloid and 12 lymphoid subsets), in all 680 PB samples corresponding to 513 COVID-19 patients taken at presentation and during fol- low-up (Figure 1A). Those patients with a hematological tumor showed significantly decreased percentages of classical monocytes, immunoregulatory natural killer (NK) cells, double-positive T cells, and B cells, when compared to COVID-19 patients without hematological cancer (Figure 1B). Similarly, hematological patients tend- ed to have also reduced absolute cell counts of various cell types, reaching statistical significance in double-pos- itive T cells (Online Supplementary Figure S1). While the aspects of cancer and its treatment conferring risk of severe COVID-19 have remained largely unknown,3 this study exposes for the first time that hematological patients show significant alterations in the relative distri- bution of specific innate and adaptive cell types, which could compromise an initial response to the infection.
Effective viral clearance requires CD8 effector T-cell- mediated killing of virally infected cells as well as CD4 T-cell-dependent enhancement of CD8 and B-cell responses. Interestingly, deep phenotypic characteriza- tion of T- and B-cell compartments in PB of COVID-19 patients with (n=4) or without (n=10) hematological can- cer showed that the relative distribution of antigen- dependent maturation stages within the T-cell compart- ment was generally similar between both groups (Online Supplementary Figure S2A). By contrast, hematological cases displayed alterations in several B-cell subsets that reached statistical significance in memory B cells express- ing immunoglobulin (Ig) G and IgA subclasses (Online Supplemental Figure S2B). These findings are important because the humoral immune response is critical for the clearance of SARS-CoV-2 and could be a major part of the memory response that prevents reinfection.11
We next compared the immune kinetics from presenta- tion to the last follow-up in COVID-19 patients with and without a hematological tumor (n=6 and n=161, respec- tively), depicting those with favorable or fatal outcome (Figure 1C). There was a profound variation from the first to the latest PB sample in the relative distribution of all
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