Effects of the BTN162b2 mRNA COVID-19 vaccine in humoral and cellular immunity in patients with chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL), the most common leukemia in the western countries, is characterized by immunosuppression due to disease itself and cytotoxic treatments. Since the beginning of COVID-19 pandemic, patients with CLL appear to be a vulnerable population. In addition, phase III mRNA vaccine trials did not provide information about the efficacy in immunocomprised population. In CLL, the antibody-mediated response to SARS-CoV-2 vaccine is impaired. The goal of this study was to evaluate the effects of SARS-CoV-2 vaccination on humoral immune response and on cellular immunity in CLL patients. Humoral immune response to BNT162b2 messenger RNA COVID-19 vaccine was evaluated in 44 CLL patients comprising 20 treatment-naïve, 14 under treatment with ibrutinib and 10 in follow-up after completion of therapy. A positive serological response to SARS-CoV-2 vaccination with IgG titers higher than 13 UA/ml was detected in 54.6% of CLL patients with a higher response in patients who obtained remission after treatment. Reduced antibody response was detected in patients under ibrutinib treatment. T-cell response to overlapping pool of peptides representing the spike region was assessed in paired CLL samples collected before and after 1 month from the second dose of COVID-19 vaccine in treatment-naïve and ibrutinib-treated CLL patients using cytokine secretion assay. Both CD3+ CD4+ and CD3+ CD8+ T cells are able to mount a cellular response to spike peptides with secretion of IFNγ and TNFα before and after vaccination in both treatment naïve and ibrutinib-treated patients and this cellular immune response is independent by COVID-19 vaccination. Collectively, T cell response to spike peptides appeared more blunted in CLL patients under treatment with ibrutinib compared to untreated ones. Our study supports the need for optimization of vaccination strategy to achieve an adequate immune response keeping strict preventive measures by CLL patients against COVID-19.

The dynamic functions of IRF4 in B cell malignancies.

The trajectory of B cell development goes through subsequent steps governed by complex genetic programs, strictly regulated by multiple transcription factors. Interferon regulatory factor 4 (IRF4) regulates key points from pre-B cell development and receptor editing to germinal center formation, class-switch recombination and plasma cell differentiation. The pleiotropic ability of IRF4 is mediated by its "kinetic control", allowing different IRF4 expression levels to activate distinct genetic programs due to modulation of IRF4 DNA-binding affinity. IRF4 is implicated in B cell malignancies, acting both as tumor suppressor and as tumor oncogene in different types of precursors and mature B cell neoplasia. Here, we summarize the complexity of IRF4 functions related to different DNA-binding affinity, multiple IRF4-specific target DNA motif, and interactions with transcriptional partners. Moreover, we describe the unique role of IRF4 in acute leukemias and B cell mature neoplasia, focusing on pathogenetic implications and possible therapeutic strategies in multiple myeloma and chronic lymphocytic leukemia.

Role of Notch2 pathway in mature B cell malignancies.

In recent decades, the Notch pathway has been characterized as a key regulatory signaling of cell-fate decisions evolutionarily conserved in many organisms and different tissues during lifespan. At the same time, many studies suggest a link between alterations of this signaling and tumor genesis or progression. In lymphopoiesis, the Notch pathway plays a fundamental role in the correct differentiation of T and B cells, but its deregulated activity leads to leukemic onset and evolution. Notch and its ligands Delta/Jagged exhibit a pivotal role in the crosstalk between leukemic cells and their environment. This review is focused in particular on Notch2 receptor activity. Members of Notch2 pathway have been reported to be mutated in Chronic Lymphocytic Leukemia (CLL), Splenic Marginal Zone Lymphoma (SMZL) and Nodal Marginal Zone Lymphoma (NMZL). CLL is a B cell malignancy in which leukemic clones establish supportive crosstalk with non-malignant cells of the tumor microenvironment to grow, survive, and resist even the new generation of drugs. SMZL and NMZL are indolent B cell neoplasms distinguished by a distinct pattern of dissemination. In SMZL leukemic cells affect mainly the spleen, bone marrow, and peripheral blood, while NMZL has a leading nodal distribution. Since Notch2 is involved in the commitment of leukemic cells to the marginal zone as a major regulator of B cell physiological differentiation, it is predominantly affected by the molecular lesions found in both SMZL and NMZL. In light of these findings, a better understanding of the Notch receptor family pathogenic role, in particular Notch2, is desirable because it is still incomplete, not only in the physiological development of B lymphocytes but also in leukemia progression and resistance. Several therapeutic strategies capable of interfering with Notch signaling, such as monoclonal antibodies, enzyme or complex inhibitors, are being analyzed. To avoid the unwanted multiple "on target" toxicity encountered during the systemic inhibition of Notch signaling, the study of an appropriate pharmaceutical formulation is a pressing need. This is why, to date, there are still no Notch-targeted therapies approved. An accurate analysis of the Notch pathway could be useful to drive the discovery of new therapeutic targets and the development of more effective therapies.

Indoleamine 2, 3-Dioxygenase 1 Mediates Survival Signals in Chronic Lymphocytic Leukemia via Kynurenine/Aryl Hydrocarbon Receptor-Mediated MCL1 Modulation.

The indoleamine 2,3-dioxygenase 1 (IDO1) metabolic circuitry, comprising the first tryptophan (Trp) catabolite L-kynurenine (Kyn) and the aryl hydrocarbon receptor (AHR), has emerged as a mechanism of cancer immune evasion. Here, we investigated the functional role of the IDO1/Kyn/AHR axis in chronic lymphocytic leukemia (CLL). Our data show that CLL cells expressed an active form of the IDO1 enzyme and microenvironmental stimuli can positively modulate its expression. Interferon (IFN)-γ induces IDO1 expression through the Jak/STAT1 pathway and mediates Kyn production concomitantly with Trp consumption in CLL-conditioned media, while INCB018424 (ruxolitinib), a JAK1/2 inhibitor, impaired both effects. To characterize the involvement of IDO1 in leukemic cell maintenance, we overexpressed IDO1 by vector transfection measuring enhanced resistance to spontaneous apoptosis. IDO1 pro-survival influence was confirmed by treating CLL cells with Kyn, which mediated the increase of induced myeloid leukemia cell differentiation protein (MCL1). Conversely, AHR silencing or its blockade via CH-223191 improved the apoptosis of leukemic clones and mitigated MCL1 expression. Moreover, Kyn-treated CLL cells are less affected by the pro-apoptotic effect of ABT-199 (venetoclax), while CH-223191 showed synergistic/additive cytotoxicity with this drug. Lastly, targeting directly MCL1 in CLL cells with AMG-176, we abrogate the pro-survival effect of Kyn. In conclusion, our data identify IDO1/Kyn/AHR signaling as a new therapeutic target for CLL, describing for the first time its role in CLL pathobiology.

Notch2 Increases the Resistance to Venetoclax-Induced Apoptosis in Chronic Lymphocytic Leukemia B Cells by Inducing Mcl-1.

Chronic lymphocytic leukemia (CLL) has experienced a clinical revolution-thanks to the discovery of crucial pathogenic mechanisms. CLL is still an incurable disease due to intrinsic or acquired resistance of the leukemic clone. Venetoclax is a Bcl-2 inhibitor with a marked activity in CLL, but emerging patterns of resistance are being described. We hypothesize that intrinsic features of CLL cells may contribute to drive mechanisms of resistance to venetoclax. We analyzed the expression of Interferon Regulatory Factor 4 (IRF4), Notch2, and Mcl-1 in a cohort of CLL patients. We evaluated CLL cell viability after genetic and pharmaceutical modulation of Notch2 expression in patients harboring trisomy 12. We tested venetoclax in trisomy 12 CLL cells either silenced or not for Notch2 expression or in combination with an inhibitor of Mcl-1, AMG-176. Trisomy 12 CLL cells were characterized by low expression of IRF4 associated with high levels of Notch2 and Mcl-1. Notch2 and Mcl-1 expression determined protection of CLL cells from spontaneous and drug-induced apoptosis. Considering the involvement of Mcl-1 in venetoclax resistance, our data demonstrated a contribution of high levels of Notch2 and Mcl-1 in a reduced response to venetoclax in CLL cells carrying trisomy 12. Furthermore, reduction of Mcl-1 expression by silencing Notch2 or by treatment with AMG-176 was able to restore the response of CLL cells to venetoclax. The expression of Notch2 identifies a subset of CLL patients, mainly harboring trisomy 12, characterized by high levels of Mcl-1. This biological mechanism may compromise an effective response to venetoclax.