Decoupling of normal CD40/interleukin-4 immunoglobulin heavy chain switch signal leads to genomic instability in SGH-MM5 and RPMI 8226 multiple myeloma cell lines

Hwang, William Ying Khee; PhD, Charles A. Gullo; Shen, Jie; Poh, Chee Kok; Tham, Sim-Mun; Cow, Geraline; Au, Melvin; Chan, Emily; Teoh, Gerrard

doi:10.1038/sj.leu.2404099

Cited by 5 publications

(9 citation statements)

References 34 publications

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“…However, CD40 is also known to exert proliferative effects as well. For example, soluble CD40 ligand is sufficient to induce proliferation of MM cells whereas normal B cells require both CD40 ligand and IL-4 for full activation [25], suggesting that CD40 ligand act differently on different cells, and that MM cells exist at a stage of differentiation or activation that does not require a second signal such as IL-4. Finally, CD40 stimulation has been shown to induce apoptosis or proliferation in different cell types despite the activation of the NF-κB pathway in both instances [46].…”

Section: Discussionmentioning

confidence: 99%

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Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling

et al. 2010

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BackgroundMultiple myeloma (MM) is a malignancy of terminally-differentiated plasma cells, and the second most prevalent blood cancer. At present there is no cure for MM, and the average prognosis is only three to five years. Current treatments such as chemotherapy are able to prolong a patient's life but rarely prevent relapse of the disease. Even hematopoietic stem cell transplants and novel drug combinations are often not curative, underscoring the need for a continued search for novel therapeutics. CD137 and its ligand are members of the Tumor Necrosis Factor (TNF) receptor and TNF superfamilies, respectively. Since CD137 ligand cross-linking enhances proliferation and survival of healthy B cells we hypothesized that it would also act as a growth stimulus for B cell cancers.Methodology/Principal FindingsProliferation and survival of B cell lymphoma cell lines were not affected or slightly enhanced by CD137 ligand agonists in vitro. But surprisingly, they had the opposite effects on MM cells, where CD137 ligand signals inhibited proliferation and induced cell death by apoptosis. Furthermore, secretion of the pro-inflammatory cytokines, IL-6 and IL-8 were also enhanced in MM but not in non-MM cell lines in response to CD137 ligand agonists. The secretion of these cytokines in response to CD137 ligand signaling was consistent with the observed activation of the classical NF-κB pathway. We hypothesize that the induction of this pathway results in activation-induced cell death, and that this is the underlying mechanism of CD137-induced MM cell death and growth arrest.Conclusions/SignificanceThese data point to a hitherto unrecognized role of CD137 and CD137 ligand in MM cell biology. The selective inhibition of proliferation and induction of cell death in MM cells by CD137 ligand agonists may also warrant a closer evaluation of their therapeutic potential.

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Section: Discussionmentioning

confidence: 99%

“…DOHH-2 and SUDHL-4 were obtained from the Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (DSMZ, Germany). The SGH-MM5 and SHG-MM6 human MM cell lines were developed in our laboratory from a patient with MM using a modified Dexter-type long-term tissue culture system, as described previously [25].…”

Section: Methodsmentioning

confidence: 99%

Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling

et al. 2010

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“…Mutations in the CD40 receptor in MM cell lines and patient specimens have been described previously by us and others (3,49). This consideration as well as the propensity of irreversible genomic instability through CD40 binding (50) are also potential explanations for CD40 agonism and antagonism converging to a common apoptotic outcome in myeloma cells. Clearly, further molecular studies that are based on our gene expression array findings may help resolve this conundrum.…”

Section: Discussionmentioning

confidence: 99%

Growth Inhibition of Human Multiple Myeloma Cells by an Oncolytic Adenovirus Carrying the CD40 Ligand Transgene

Fernandes

Gomes

Butcher

et al. 2009

Clinical Cancer Research

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Purpose: The growth-inhibitory activity of recombinant CD40 ligand (CD40L) is well documented in human multiple myeloma (MM). We examined MM-targeted delivery of CD40L by a conditional replicative oncolytic adenovirus, AdEHCD40L. Experimental Design: The growth-regulatory activity of AdEHCD40L was determined in vitro and in vivo. Differential analysis with AdEHCD40L and parental virus (AdEHNull)-infected cultures allowed the identification of cellular and molecular pathways modulated by the CD40L transgene. Results: Conditional expression of viral E1A and CD40L transgene was shown in human MM lines RPMI 8226 [interleukin (IL)-6 independent] and Kas-6/1 (IL-6 dependent) under hypoxic conditions commonly found in MM in situ. AdEHCD40L inhibited MM cell growth more effectively than AdEHNull. This enhanced growth-inhibitory activity was abrogated by cotreatment with a CD40L antibody. Chemoresistant MM lines (MR20 and LR5) were similarly susceptible to AdEHCD40L treatment. AdEHCD40L induced apoptosis and S-phase cell cycle blockade while uniquely up-regulating the previously described proapoptotic elements tumor necrosis factor-related apoptosis-inducing ligand, Fas, and IL-8. Intratumoral injections of AdEHCD40L reduced the growth of severe combined immunodeficient/hu RPMI 8226 xenografts by >50% compared with 28% reduction by AdEHNull. Adenoviral hexon and CD40L were detected in AdEHCD40L-treated tumors at day 35 after infection primarily in necrotic areas, suggesting viral replicative activity. Conclusions: These findings show that CD40L acts in concert with viral oncolysis to produce MM growth inhibition through activation of cellular apoptosis. The direct growth-inhibitory activity of AdEHCD40L, together with the well-known immunepotentiating features of CD40L, may be clinically applicable for the experimental treatment of MM or plasma cell leukemia. Multiple myeloma (MM) is a B-cell malignancy characterizedby the clonal expansion and accumulation of malignant plasma cells in the bone marrow (1). Despite recent advances in induction therapy, the long-term outlook for patients with this disease is poor. Promising new agents, such as thalidomide-like immunomodulatory drugs (lenalidomide) and proteasome inhibitors (bortezomib), have improved prognosis and survival (2), but MM is still incurable.CD40 belongs to the tumor necrosis factor (TNF) receptor superfamily member and is commonly expressed on human B cells. The CD40 pathway is best known for its costimulatory role in B-cell and T-cell proliferation and differentiation. Interestingly, CD40 is commonly expressed on human MM cells (3) but not normal, terminally differentiated plasma cells (4). This differential pattern of expression presents an opportunity for exploiting the CD40 growth-regulatory pathway for growth inhibition of MM. We and others have found that binding of the CD40 receptor by its natural ligand (CD40L, CD154) produced

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“…RPMI 8226 MM, CESS Epstein-Barr virus (EBV)-transformed normal B cells, K562 chronic myeloid leukemia (CML), and HL-60 APL human cell lines were all purchased from the American Type Culture Collection (ATCC, Rockville, MD). The EBV-negative SGH-MM5 human MM cell line (CD10+ CD19- CD20- CD38+ CD40+ CD45+ CD56+ CD138+) was developed in our laboratory under the Singapore General Hospital (SGH) Institutional Review Board (IRB) good research practice guidelines, from a patient with MM using a modified Dexter-type long-term tissue culture system, which was previously described [ 23 ]. All cell lines were cultured in RPMI 1640 medium (Invitrogen, Gibco, Grand Island, NY) supplemented with 10% fetal calf serum (FCS) at 37°C and 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Several recent studies have suggested that the Ku86 variant seen in lymphocytes may be due to cleavage by proteases induced during biochemical isolation [ 20 - 22 ]. Considering the increased amount of genomic instability seen in MM, the disregulated CD40/interleukin-4 (IL-4) pathway in MM cells [ 23 ], and the role of Ku in DNA DSBR and non-homologous end-joining (NHEJ), we investigated the presence of Ku86 and its variants in MM cells, and compared them to T lymphocytes and other cell lines. We found that unlike human T lymphocytes, the detection of 69-kDa Ku86 (Ku86v-N) variant is not likely due to in vitro generated protease cleavage.…”

Section: Introductionmentioning

confidence: 99%

Ku86 exists as both a full-length and a protease-sensitive natural variant in multiple myeloma cells

PhD

Cow

et al. 2008

Cancer Cell International

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BackgroundTruncated variants of Ku86 protein have previously been detected in 86% to 100% of freshly isolated patient multiple myeloma (MM) cells. Since, the Ku70/Ku86 heterodimer functions as the regulatory subunit of the DNA repair enzyme, DNA-dependent protein kinase, we have been interested in the altered expression and function of Ku86 variant (Ku86v) proteins in genome maintenance of MM.ResultsAlthough, a number of studies have suggested that truncated forms of Ku proteins could be artificially generated by proteolytic degradation in vitro in human lymphocytes, we now show using whole cell immunoblotting that the RPMI-8226 and SGH-MM5 human MM cell lines consistently express full-length Ku86 as well as a 69-kDa Ku86v; a C-terminus truncated 69-kDa variant Ku86 protein. In contrast, Ku86v proteins were not detected in the freshly isolated lymphocytes as was previously reported. Data also indicates that the Ku86v was not generated as a result of carbohydrate modification but that serine proteases may act on the full-length form of the protein.ConclusionThese data confirm that MM cells contain bona fide Ku86v proteins that were generated intracellularly by a post-transcriptional mechanism, which required proteolytic processing.

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Decoupling of normal CD40/interleukin-4 immunoglobulin heavy chain switch signal leads to genomic instability in SGH-MM5 and RPMI 8226 multiple myeloma cell lines

Cited by 5 publications

References 34 publications

Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling

Inhibition of Proliferation and Induction of Apoptosis in Multiple Myeloma Cell Lines by CD137 Ligand Signaling

Growth Inhibition of Human Multiple Myeloma Cells by an Oncolytic Adenovirus Carrying the CD40 Ligand Transgene

Ku86 exists as both a full-length and a protease-sensitive natural variant in multiple myeloma cells

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