gossypol-acetic-acid and Leukemia--Lymphocytic--Chronic--B-Cell

The failure of apoptosis (programmed cell death) underpins the development of many tumors and often renders them resistant to cytotoxic therapies. In hematologic malignancies, this impairment of apoptosis is often caused by overexpression of the pro-survival protein BCL2. Because abnormally high levels of BCL2 sustain these tumors, there has been much interest in targeting BCL2 as a novel approach to treating various hematologic malignancies. One such approach is the development of BH3 mimetic compounds, small molecules that mimic the action of the BH3-only proteins, natural antagonists of BCL2 and its pro-survival relatives. These compounds act by restoring the ability of a cell to undergo apoptotic cell death. Some of them have shown very encouraging results in early-phase clinical trials that are currently underway, particularly in patients with chronic lymphocytic leukemia and some non-Hodgkin lymphomas, diseases marked by BCL2 overexpression. In this review, we discuss the rationale behind targeting BCL2, highlight the recent findings from clinical trials, and pinpoint the next steps in the clinical development of this interesting and promising class of targeted agents, particularly for the treatment of lymphoid malignancies.

Topics: Aniline Compounds; Antineoplastic Agents; Bridged Bicyclo Compounds, Heterocyclic; Gossypol; Hematologic Neoplasms; Humans; Indoles; Leukemia, Lymphocytic, Chronic, B-Cell; Peptide Fragments; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Pyrroles; Sulfonamides

Over-expression of anti-apoptotic BCL2 has been reported in chronic lymphocytic leukaemia (CLL), but targeting BCL2 alone did not yield appreciable clinical results. However, it was demonstrated that BCL2 inhibitors enhanced the clinical efficacy of chemo and immunotherapeutics. Lenalidomide, an immunomodulator, is clinically effective in CLL and can enhance the anti-CLL effects of CD20 targeting monoclonal antibody, rituximab. Here, we investigated the mechanism of immune-directed killing of lenalidomide in CLL and evaluated if concurrent targeting of CD20 and BCL2 can enhance this effect. In vitro treatment with lenalidomide enhanced the antibody-mediated cellular cytotoxicity (ADCC) directed by rituximab in autologous leukaemic cells. Furthermore, peripheral blood mononuclear cells obtained from patients after treatment with lenalidomide and rituximab showed increased ADCC in vitro versus control (pre-treatment sample). This effect was further enhanced with pre-treatment of tumour cells with AT-101 (a BH3 mimetic that functions as BCL2 antagonist). Our data suggest that AT-101 in combination with lenalidomide can potentially be an effective therapeutic regimen for CLL.

Topics: Antibodies, Monoclonal, Murine-Derived; Antineoplastic Agents, Phytogenic; Antineoplastic Combined Chemotherapy Protocols; Down-Regulation; Female; Gene Expression Regulation, Leukemic; Gossypol; Humans; Immunologic Factors; Lenalidomide; Leukemia, Lymphocytic, Chronic, B-Cell; Male; Proto-Oncogene Proteins c-bcl-2; Rituximab; Thalidomide; Tumor Cells, Cultured

Chronic lymphocytic leukaemia (CLL) is the most common haematological malignancy in the U.S. The course of the disease has been shown to be negatively impacted by increased levels of BCL2. Strategies to downregulate BCL2 and shift the balance towards cellular demise are actively being explored. Therefore, we examined whether the investigational agent MLN2238 could inhibit the proteasomal machinery and induce CLL cell death while also downregulating BCL2. MLN2238-induced cell death was studied in peripheral blood mononuclear cells from 28 CLL patients. MLN2238 produced a dose-dependent reduction in BCL2 and CLL cell viability with maximum cell death observed at a 50 nmol/l concentration by 48 h. Annexin-V staining, PARP1 and caspase-3 cleavage along with an increase in mitochondrial membrane permeability were noted after cells were treated with MLN2238; however, apoptosis was only partially blocked by the pan-caspase inhibitor z-VAD.fmk. Furthermore, we observed enhanced anti-CLL effects in tumour cells treated with either a combination of MLN2238 and the BH3 mimetic AT-101 or MLN2238 and fludarabine. Together, our data suggest the potential for proteasome inhibitor based therapy in CLL and the rationale design of drug combination strategies based on CLL biology.

Topics: Adult; Aged; Aged, 80 and over; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Boron Compounds; Caspase 3; Caspase 9; Cell Death; Cell Survival; Dexamethasone; Drug Synergism; Enzyme Activation; Glycine; Gossypol; Humans; Intracellular Membranes; Leukemia, Lymphocytic, Chronic, B-Cell; Middle Aged; Mitochondria; Neoplasm Staging; Permeability; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Proto-Oncogene Proteins c-bcl-2; Tumor Cells, Cultured; Vidarabine

Despite real advances made in chemoimmunotherapy, chronic lymphocytic leukemia (CLL) is still an incurable disease. New therapeutic strategies based on the restoration of the cell death program seemed relevant. Some members of the Bcl-2 family are critical players in the defective apoptotic program in CLL cells and/or targets of apoptosis inducers in vitro. The concept of BH3 mimetics has led to the characterization of small molecules mimicking proapoptotic BH3-only members of the Bcl-2 family by their ability to bind and antagonize the prosurvival members. Some putative or actual BH3 mimetics are already being tested in clinical trials with somewhat promising results. However, none of them has a high enough interaction affinity with Mcl-1, a crucial antiapoptotic factor in CLL. It has been suggested that resistance to BH3 mimetics can be overcome by using inhibitors of Mcl-1 expression. An alternative and more direct strategy is to design mimetics of the Noxa BH3 domain, which is a specific antagonistic Mcl-1 ligand. The development of such Noxa-like BH3 mimetics, capable of directly interacting with Mcl-1 and efficiently neutralizing its antiapoptotic activity, is extremely important to evaluate their impact on the clinical outcome of patients with CLL.

Topics: Aniline Compounds; Apoptosis; Biomimetic Materials; Gossypol; Humans; Indoles; Leukemia, Lymphocytic, Chronic, B-Cell; Myeloid Cell Leukemia Sequence 1 Protein; Peptide Fragments; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Pyrroles; Sulfonamides

To investigate the effects of gossypol acetate on apoptosis in primary cultured cells from patients with acute lymphoblastic leukemia (ALL) and chronic lymphocytic leukemia (CLL) and its synergistic effect with dexamethasone. The apoptosis-inducing effect of gossypol acetate on primary cultured leukemia cells was analyzed by flow cytometry (FCM). The effect of gossypol acetate on survival rates of Raji cells and mononuclear cells (MNC) from normal bone marrow were evaluated by MTT assay. After co-treatment with gossypol acetate and dexamethasone, the apoptosis rate of Raji cells was detected by FCM. The results showed that gossypol acetate was able to induce apoptosis in primary cultured ALL cells at concentrations of ≥ 5 µmol/L. The effect was concentration and time dependent. Apoptosis-inducing concentration in CLL cells was higher than that in ALL cells. After exposing to 50 µmol/L gossypol acetate for 48 h, the apoptosis rate of ALL and CLL cells were (90.4 ± 6.2)% and (51.7 ± 10.3)% separately. No major growth inhibitory effect was observed in MNC from normal bone marrow when they were exposed to gossypol acetate at concentrations lower than 10 µmol/L. After exposing for 48 and 72 h, the IC(50) of gossypol acetate for MNC from normal bone marrow was 7.1 and 9.1 times as much as the IC(50) of Raji cells. Co-treatment with 10 µmol/L gossypol acetate and dexamethasone remarkably increased the apoptosis rate of Raji cells. It is concluded that the gossypol acetate has apoptosis-inducing activity in primary cultured leukemia cells from patients diagnosed as ALL and CLL in vitro. The inhibitory effect of gossypol acetate on MNC from normal bone marrow is less prominent than that on Raji cells. Co-treatment with gossypol acetate and dexamethasone notably amplified the pro-apoptosis activity of the latter in Raji cells.

Topics: Apoptosis; Cell Line; Dexamethasone; Gossypol; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Tumor Cells, Cultured

Resistance to apoptosis in CLL B cells is associated with overexpression of Bcl-2 family antiapoptotic proteins. Their expression is endogenous, but is also induced by signals from the microenvironment resulting in intrinsic and extrinsic drug resistance. Because AT-101 binds to the BH3 motif of all Bcl-2-family antiapoptotic proteins, we hypothesized that this molecule could overcome resistance. AT-101 treatment (20 microM for 24 hours) resulted in a median 72% apoptosis in CLL cells (patients; n = 32, P < .001). Stromal cells protected CLL B cells from spontaneous and fludarabine-induced apoptosis (P = .003) by increasing the Mcl-1 protein levels. However, AT-101 induced similar extent of down-regulation of Mcl-1 and apoptosis in CLL lymphocytes cultured in suspension or on stroma (P = .999). Stromal cells expressed undetectable levels of antiapoptotic but high levels of activated ERK and AKT proteins and had low or no apoptosis with AT-101. Collectively, these data demonstrate that AT-101 induces apoptosis in CLL B cells and overcomes microenvironment-mediated resistance while sparing normal stromal cells.

Topics: Antineoplastic Agents; Antineoplastic Agents, Phytogenic; Apoptosis; B-Lymphocytes; Cell Communication; Dose-Response Relationship, Drug; Down-Regulation; Drug Resistance, Neoplasm; Gossypol; Humans; Leukemia, Lymphocytic, Chronic, B-Cell; Myeloid Cell Leukemia Sequence 1 Protein; Proto-Oncogene Proteins c-bcl-2; Stromal Cells; Tumor Cells, Cultured; Vidarabine