Kyle Crassini

Immunoglobulin G subclass deficiency and infection risk in 150 patients with chronic lymphocytic leukemia

Hypogammaglobulinemia is a common complication of chronic lymphocytic leukemia (CLL), but the significance of immunoglobulin G (IgG) subclass deficiency is unknown. We analyzed the prevalence of immunoglobulins G, A and M, IgG subclass deficiency and infection in 150 patients with CLL. Low IgG, IgA and IgM levels were observed in 27.3%, 30.7% and 56.7% of patients, respectively. IgG subclass deficiency was frequent, with reduced IgG1, IgG2, IgG3 and IgG4 in 28%, 19.3%, 52% and 22.7% of patients, respectively. IgG subclass deficiency (total 64.6%) and hypogammaglobulinemia (27.3%) were more prevalent than clinically significant infection (16%). Recurrent or significant infections were seen in 24 patients (16%), of whom 50% had hypogammaglobulinemia but 100% had at least one IgG subclass deficiency, indicating that half the patients with infection had IgG subclass deficiency but normal total IgG level. Deficiencies of IgG3 and IgG4 were statistically associated with infection risk. Normal immunoglobulin and IgG subclass levels were seen in 26 patients (17%) and none had infections. IgG subclass deficiency is commonly observed in patients with CLL with both normal and reduced total IgG levels, and is associated with infection. Screening patients with CLL for IgG subclass deficiency may be a useful adjunct in stratifying their infection risk.

The MEK1/2 inhibitor, MEKi-1, induces cell death in chronic lymphocytic leukemia cells under conditions that mimic the tumor microenvironment and is synergistic with fludarabine.

The Raf-1/MEK/ERK1/2 pathway has become a focus for novel cancer therapies. This study sought to investigate whether targeting MEK1/2 may represent a therapeutic option for chronic lymphocytic leukemia (CLL). The MEK1/2 inhibitor, MEKi-1, induced apoptosis of CLL cells and was synergistic with fludarabine under conditions that mimic the tumor microenvironment, irrespective of poor-risk characteristics. MEKi-1 down-regulated the activities of AKT and ERK1/2 and was synergistic with fludarabine through a mechanism that involved potentiation of DNA damage and attenuation of the activity of ERK1/2 and expression of Mcl-1. This study highlights the significant role of the mitogen-activated protein kinase (MAPK)–ERK1/2 pathway in mediating the effects of the CLL tumor microenvironment and suggests that targeting MEK1/2 in CLL cells may impact upon the activity of both ERK1/2 and AKT. Inhibitors of MEK1/2 as single agents or in combination with DNA-damaging agents may represent a novel therapeutic strategy for CLL.

Targeting chronic lymphocytic leukemia cells in the tumor microenviroment: A review of the in vitro and clinical trials to date.

Chronic lymphocytic leukemia (CLL) is the most common leukemia in the western world. Despite significant advances in therapy over the last decade CLL remains incurable. Current front-line therapy often consists of chemoimmunotherapy-based regimens, most commonly the fludarabine, cyclophosphamide plus rituximab combination, but rates of relapse and refractory disease are high among these patients. Several key signaling pathways are now known to mediate the survival and proliferation of CLL cells in vivo, the most notable of which are the pathways mediated by the B-cell receptor (BCR) and cytokine receptors. A better understanding of the pathogenesis of the disease, the underlying biology of the CLL-cell and the roles of the tumour microenvironment has provided the rationale for trials of a range of novel, more targeted therapeutic agents. In particular, clinical trials of ibrutinib and idelalisib, which target the Brutons tyrosine kinase and the delta isoform of phosphoinositol-3 kinase components of the BCR signaling pathway respectively, have shown extremely promising results. Here we review the current literature on the key signaling pathways and interactions of CLL cells that mediate the survival and proliferation of the leukemic cells. For each we describe the results of the recent clinical trials and in vitro studies of novel therapeutic agents.

Modeling the chronic lymphocytic leukemia microenvironment in vitro

Abstract Microenvironments within the lymph node and bone marrow promote proliferation and drug resistance in chronic lymphocytic leukemia (CLL). Successful treatment of CLL must therefore target the leukemic cells within these compartments. A better understanding of the interaction between CLL cells and the tumor microenvironment has led to the development of in vitro models that mimic the mechanisms that support leukemic cell survival and proliferation in vivo. Employing these models as part of the pre-clinical evaluation of novel therapeutic agents enables a better approximation of their potential clinical efficacy. In this review we summarize the current literature describing how different aspects of the tumor microenvironment have been modeled in vitro and detail how these models have been employed to study the biology of the disease and potential efficacy of novel therapeutic agents.

MEK1/2 inhibition by binimetinib is effective as a single agent and potentiates the actions of Venetoclax and ABT-737 under conditions that mimic the chronic lymphocytic leukaemia (CLL) tumour microenvironment

The survival and proliferation of chronic lymphocytic leukaemia (CLL) cells is driven by multiple signalling pathways, including those mediated by the B cell, Toll‐like and chemokine receptors. Many of these pathways converge on the same signalling molecules, including those involved in the Raf‐1/MEK/Erk1/2‐MAPK pathway. We investigated the effects of the MEK1/2 (also termed MAP2K1/2) inhibitor, binimetinib, against CLL cells cultured under conditions that mimic aspects of the tumour microenvironment. Binimetinib blocked CLL cell survival induced by stroma‐conditioned media and phorbol myristylate (PMA). Binimetinib was also significantly more toxic towards CLL cells cultured in the presence of either anti‐IgM antibody or stroma‐derived factor‐1α (SDF‐1α) and reduced CLL cell cycle progression and proliferation. Furthermore, binimetinib significantly increased the sensitivity of CLL cells co‐cultured with CD40 ligand (CD40L)‐expressing fibroblasts to the BH3‐mimetics ABT‐737 and Venetoclax (ABT‐199) via a mechanism involving down‐regulation of Mcl‐1 (MCL1) activity and Bim (BCL2L11) and Bcl‐xL (BCL2L1) expression. Collectively, these data suggest that binimetinib may have both cytotoxic and cytostatic effects on CLL cells by blocking microenvironment‐derived signals known to drive survival and proliferation. The combination of binimetinib with a BH3 mimetic may be an effective treatment strategy for CLL, particularly against the proliferative fraction of the disease within the tumour microenvironment.

Humoral immune failure defined by immunoglobulin class and immunoglobulin G subclass deficiency is associated with shorter treatment‐free and overall survival in Chronic Lymphocytic Leukaemia

Immune dysfunction attributed to hypogammaglobulinaemia is common in chronic lymphocytic leukaemia (CLL) and infection is a major contributor to morbidity and mortality. A higher incidence of multiple immunoglobulin and immunoglobulin G (IgG) subclass deficiency was associated with more advanced disease (P < 0·001 and P < 0·001, respectively) in a cohort of 147 CLL patients. Multiple immunoglobulin and IgG subclass deficiency were significantly associated with shorter treatment‐free survival (TFS) (P < 0·001 and P = 0·006, respectively). The association between disease stage and immune dysfunction demonstrated by these data suggest aspects of immune deficiency correlate with disease severity and may be associated with shorter TFS in CLL.

The clinical significance of hypogammaglobulinaemia and serum immunoglobulin G subclass deficiency in patients with chronic lymphocytic leukaemia (CLL).

We read with interest the paper by Svensson et al. [1] regarding IgG subclass defi ciency in chronic lymphocytic leukaemia (CLL). In contrast to our study [2], Svensson et al. failed to fi nd an association between infection with either IgG subclass defi ciency or hypogammaglobulinaemia (reduced total IgG level), while our study found an association with both. There are likely to be several reasons for this, including the fact that Svensson ’ s cohort comprised almost entirely stage A patients and included some patients already on immunoglobulin replacement therapy. Differences in assays and reference ranges, lack of IgG4 level analysis, differences in the defi nition of infection, and the fact that Svensson et al. focused on a single calendar year, may also have contributed to the contrast with our fi ndings. Despite the focus on mainly early stage A disease, they found a remarkably high incidence of infection in their patients. Therefore, we believe that both studies show that infection is an important cause of morbidity, and mortality, in patients with CLL, with relatively similar rates of severe infection (12.6% vs 16%). Importantly, Svensson et al. failed to fi nd an association between infection and hypogammaglobulinaemia, an association that has been well documented for many years [3], including by our recent study [2]. The primary purpose of this letter is to express our concerns that these fi ndings may be interpreted by some that reduced immunoglobulin levels in CLL are of no consequence. We believe it is very important to emphasize to clinicians and patients that hypogammaglobulinaemia is an important factor contributing to infection risk in CLL and should be regularly monitored. Furthermore, patients with CLL together with hypogammaglobulinaemia and recurrent episodes of infection, especially severe and potentially life-threatening infections, may benefi t substantially from immunoglobulin replacement therapy and this should be considered an essential component of disease management in patients with CLL.

Inhibition of the Raf-1 kinase inhibitory protein (RKIP) by locostatin induces cell death and reduces the CXCR4-mediated migration of chronic lymphocytic leukemia cells

Abstract The Raf-1 kinase inhibitory protein (RKIP) is an important regulatory element in multiple signaling pathways, including MAPK-ERK1/2. We investigated whether targeted disruption of RKIP is a therapeutic option for chronic lymphocytic leukemia (CLL). The RKIP inhibitor locostatin-induced apoptosis of CLL cells, irrespective of poor prognostic indications or treatment history. Locostatin down-regulated MAPK-ERK1/2 and AKT phosphorylation, decreased expression of the chemokine receptor CXCR4 (pu2009=u2009.04) and reduced the migratory capacity of CLL cells toward stroma-derived factor 1α (SDF-1α, pu2009=u2009.02). Immuno-blotting and immuno-precipitation showed that RKIP is constitutively phosphorylated and highly expressed in CLL cells and that the actions of locostatin may be mediated by binding of G-protein receptor kinase-2 (GRK2) to MEK1 and AKT. Collectively, our data suggest that inhibition of RKIP may be effective against CLL, reducing the survival and migratory capacity of the leukemic cells through down-regulation of MAPK-ERK1/2 and AKT-mediated signaling.

Immune failure, infection and survival in chronic lymphocytic leukemia

Chronic lymphocytic leukemia (CLL) commonly leads to immune dysfunction especially hypogammaglobulinemia, increasing risk of infection. Infection remains a major cause of mortality and morbidity in CLL with studies showing this accounts for a third to a half of all deaths.[1][1]–[3][2]

The dual inhibitor of the phosphoinositol-3 and PIM kinases, IBL-202, is effective against chronic lymphocytic leukaemia cells under conditions that mimic the hypoxic tumour microenvironment

Despite significant advances in treatment, chronic lymphocytic leukaemia (CLL) remains an incurable disease. Ibrutinib and idelalisib, which inhibit Bruton Tyrosine kinase (BTK) and phosphoinositol‐3 (PI3) kinase‐δ respectively, have become important treatment options for the disease and demonstrate the potential of targeting components of the B‐cell receptor‐signalling pathway. IBL‐202 is a dual inhibitor of the PIM and PI3 kinases. Synergy between the pan‐PIM inhibitor, pPIMi, and idelalisib against a range of haematological cell lines and primary CLL cells supports the rationale for preclinical studies of IBL‐202 in CLL. Importantly, IBL‐202, but not idelalisib, was cytotoxic against CLL cells under in vitro conditions that mimic the hypoxic tumour microenvironment. The significant effects of IBL‐202 on CD49d and CXCR4 expression and migration, cycle and proliferation of CLL cells suggest the drug may also interfere with the migratory and proliferative capacity of the leukaemic cells. Collectively, these data demonstrate that dual inhibition of the PIM and PI3 kinases by IBL‐202 may be an effective strategy for targeting CLL cells, particularly within the environmental niches known to confer drug‐resistance.