TRAIL/Apo2L ligand selectively induces apoptosis and overcomes drug resistance in multiple myeloma: therapeutic applications

Blood ◽  
2001 ◽  
Vol 98 (3) ◽  
pp. 795-804 ◽  
Author(s):  
Constantine S. Mitsiades ◽  
Steven P. Treon ◽  
Nicholas Mitsiades ◽  
Yoshihito Shima ◽  
Paul Richardson ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Drug Resistance ◽  
Mononuclear Cells ◽  
Nuclear Translocation ◽  
Death Receptor 5 ◽  
Therapeutic Applications ◽  
Trail Receptors ◽  
Induced Apoptosis

Abstract Multiple myeloma (MM) remains incurable and novel treatments are urgently needed. Preclinical in vitro and in vivo evaluations were performed to assess the potential therapeutic applications of human recombinant tumor necrosis factor (TNF)–related apoptosis-inducing ligand/Apo2 ligand (TRAIL/Apo2L) in MM. TRAIL/Apo2L potently induced apoptosis of MM cells from patients and the majority of MM cell lines, including cells sensitive or resistant to dexamethasone (Dex), doxorubicin (Dox), melphalan, and mitoxantrone. TRAIL/Apo2L also overcame the survival effect of interleukin 6 on MM cells and did not affect the survival of peripheral blood and bone marrow mononuclear cells and purified B cells from healthy donors. The status of the TRAIL receptors (assessed by immunoblotting and flow cytometry) could not predict TRAIL sensitivity of MM cells. The anti-MM activity of TRAIL/Apo2L was confirmed in nu/xid/bg mice xenografted with human MM cells; TRAIL (500 μg intraperitoneally daily for 14 days) was well tolerated and significantly suppressed the growth of plasmacytomas. Dox up-regulated the expression of the TRAIL receptor death receptor 5 (DR5) and synergistically enhanced the effect of TRAIL not only against MM cells sensitive to, but also against those resistant to, Dex- or Dox-induced apoptosis. Nuclear factor (NF)-κB inhibitors, such as SN50 (a cell-permeable inhibitor of the nuclear translocation and transcriptional activity of NF-κB) or the proteasome inhibitor PS-341, enhanced the proapoptotic activity of TRAIL/Apo2L against TRAIL-sensitive MM cells, whereas SN50 reversed the TRAIL resistance of ARH-77 and IM-9 MM cells. Importantly, normal B lymphocytes were not sensitized to TRAIL by either Dox, SN50, or PS-341. These preclinical studies suggest that TRAIL/Apo2L can overcome conventional drug resistance and provide the basis for clinical trials of TRAIL-based treatment regimens to improve outcome in patients with MM.

scholarly journals Perifosine, an oral bioactive novel alkylphospholipid, inhibits Akt and induces in vitro and in vivo cytotoxicity in human multiple myeloma cells

Blood ◽  
2006 ◽  
Vol 107 (10) ◽  
pp. 4053-4062 ◽  
Author(s):  
Teru Hideshima ◽  
Laurence Catley ◽  
Hiroshi Yasui ◽  
Kenji Ishitsuka ◽  
Noopur Raje ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Mononuclear Cells ◽  
Antitumor Agents ◽  
Mek Inhibitor ◽  
Peripheral Blood Mononuclear ◽  
Akt Phosphorylation ◽  
Human Multiple Myeloma ◽  
Induced Apoptosis

Perifosine is a synthetic novel alkylphospholipid, a new class of antitumor agents which targets cell membranes and inhibits Akt activation. Here we show that baseline phosphorylation of Akt in multiple myeloma (MM) cells is completely inhibited by perifosine [octadecyl-(1,1-dimethyl-piperidinio-4-yl)-phosphate] in a time- and dose-dependent fashion, without inhibiting phosphoinositide-dependent protein kinase 1 phosphorylation. Perifosine induces significant cytotoxicity in both MM cell lines and patient MM cells resistant to conventional therapeutic agents. Perifosine does not induce cytotoxicity in peripheral blood mononuclear cells. Neither exogenous interleukin-6 (IL-6) nor insulinlike growth factor 1 (IGF-1) overcomes Perifosine-induced cytotoxicity. Importantly, Perifosine induces apoptosis even of MM cells adherent to bone marrow stromal cells. Perifosine triggers c-Jun N-terminal kinase (JNK) activation, followed by caspase-8/9 and poly (ADP)-ribose polymerase cleavage. Inhibition of JNK abrogates perifosine-induced cytotoxicity, suggesting that JNK plays an essential role in perifosine-induced apoptosis. Interestingly, phosphorylation of extracellular signal–related kinase (ERK) is increased by perifosine; conversely, MEK inhibitor synergistically enhances Perifosine-induced cytotoxicity in MM cells. Furthermore, perifosine augments dexamethasone, doxorubicin, melphalan, and bortezomib-induced MM cell cytotoxicity. Finally, perifosine demonstrates significant antitumor activity in a human plasmacytoma mouse model, associated with down-regulation of Akt phosphorylation in tumor cells. Taken together, our data provide the rationale for clinical trials of perifosine to improve patient outcome in MM.

Inhibition of Notch signaling induces apoptosis of myeloma cells and enhances sensitivity to chemotherapy

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 2220-2229 ◽  
Author(s):  
Yulia Nefedova ◽  
Daniel M. Sullivan ◽  
Sophia C. Bolick ◽  
William S. Dalton ◽  
Dmitry I. Gabrilovich
Keyword(s):  
Multiple Myeloma ◽  
Drug Resistance ◽  
Notch Signaling ◽  
Mononuclear Cells ◽  
De Novo ◽  
Drug Induced ◽  
Peripheral Blood Mononuclear ◽  
Myeloma Cells ◽  
Induced Apoptosis

Drug resistance remains a critical problem in the treatment of patients with multiple myeloma. Recent studies have de-termined that Notch signaling plays a major role in bone marrow (BM) stroma-mediated protection of myeloma cells from de novo drug-induced apoptosis. Here, we investigated whether pharmacologic inhibition of Notch signaling could affect the viability of myeloma cells and their sensitivity to chemotherapy. Treatment with a γ-secretase inhibitor (GSI) alone induced apoptosis of myeloma cells via specific inhibition of Notch signaling. At concentrations toxic for myeloma cell lines and primary myeloma cells, GSI did not affect normal BM or peripheral blood mononuclear cells. Treatment with GSI prevented BM stroma-mediated protection of myeloma cells from drug-induced apoptosis. The cytotoxic effect of GSI was mediated via Hes-1 and up-regulation of the proapoptotic protein Noxa. In vivo experiments using xenograft and SCID-hu models of multiple myeloma demonstrated substantial antitumor effect of GSI. In addition, GSI significantly improved the cytotoxicity of the chemotherapeutic drugs doxorubicin and melphalan. Thus, this study demonstrates that inhibition of Notch signaling prevents BM-mediated drug resistance and sensitizes myeloma cells to chemotherapy. This may represent a promising approach for therapeutic intervention in multiple myeloma.

scholarly journals Targeting NSD2-mediated SRC-3 liquid–liquid phase separation sensitizes bortezomib treatment in multiple myeloma

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Jing Liu ◽  
Ying Xie ◽  
Jing Guo ◽  
Xin Li ◽  
Jingjing Wang ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Drug Resistance ◽  
Phase Separation ◽  
Liquid Phase ◽  
Liquid Phase Separation ◽  
Acquired Drug Resistance ◽  
Bortezomib Treatment ◽  
Liquid Liquid Phase Separation

AbstractDevelopment of chemoresistance is the main reason for failure of clinical management of multiple myeloma (MM), but the genetic and epigenetic aberrations that interact to confer such chemoresistance remains unknown. In the present study, we find that high steroid receptor coactivator-3 (SRC-3) expression is correlated with relapse/refractory and poor outcomes in MM patients treated with bortezomib (BTZ)-based regimens. Furthermore, in immortalized cell lines, high SRC-3 enhances resistance to proteasome inhibitor (PI)-induced apoptosis. Overexpressed histone methyltransferase NSD2 in patients bearing a t(4;14) translocation or in BTZ-resistant MM cells coordinates elevated SRC-3 by enhancing its liquid–liquid phase separation to supranormally modify histone H3 lysine 36 dimethylation (H3K36me2) modifications on promoters of anti-apoptotic genes. Targeting SRC-3 or interference of its interactions with NSD2 using a newly developed inhibitor, SI-2, sensitizes BTZ treatment and overcomes drug resistance both in vitro and in vivo. Taken together, our findings elucidate a previously unrecognized orchestration of SRC-3 and NSD2 in acquired drug resistance of MM and suggest that SI-2 may be efficacious for overcoming drug resistance in MM patients.

scholarly journals PIWIL1 Drives Chemoresistance in Multiple Myeloma by Modulating Mitophagy and the Myeloma Stem Cell Population

2022 ◽  
Vol 11 ◽  
Author(s):  
Yajun Wang ◽  
Lan Yao ◽  
Yao Teng ◽  
Hua Yin ◽  
Qiuling Wu
Keyword(s):  
Multiple Myeloma ◽  
Drug Resistance ◽  
Stem Cell ◽  
Cell Population ◽  
Side Population ◽  
Chemotherapeutic Agents ◽  
Stem Cell Population ◽  
Exact Function

As an important member of the Argonaute protein family, PIWI-like protein 1 (PIWIL1) plays a key role in tumor cell viability. However, the exact function of PIWIL1 in multiple myeloma (MM) and the underlying mechanism remain unclear. Here, we revealed that PIWIL1 was highly expressed in myeloma cell lines and newly diagnosed MM patients, and that its expression was notably higher in refractory/relapsed MM patients. PIWIL1 promoted the proliferation of MM cells and conferred resistance to chemotherapeutic agents both in vitro and in vivo. More importantly, PIWIL1 enhanced the formation of autophagosomes, especially mitophagosomes, by disrupting mitochondrial calcium signaling and modulating mitophagy-related canonical PINK1/Parkin pathway protein components. Mitophagy/autophagy inhibitors overcome PIWIL1-induced chemoresistance. In addition, PIWIL1 overexpression increased the proportion of side population (SP) cells and upregulated the expression of the stem cell-associated genes Nanog, OCT4, and SOX2, while its inhibition resulted in opposite effects. Taken together, our findings demonstrated that PIWIL1 induced drug resistance by activating mitophagy and regulating the MM stem cell population. PIWIL1 depletion significantly overcame drug resistance and could be used as a novel therapeutic target for reversing resistance in MM patients.

scholarly journals HNRNPA2B1 promotes multiple myeloma progression by increasing AKT3 expression via m6A-dependent stabilization of ILF3 mRNA

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Fengjie Jiang ◽  
Xiaozhu Tang ◽  
Chao Tang ◽  
Zhen Hua ◽  
Mengying Ke ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Cell Proliferation ◽  
Cellular Proliferation ◽  
Aberrant Expression ◽  
The Stability ◽  
Induced Apoptosis ◽  
Proliferation In Vitro

AbstractN6-methyladenosine (m6A) modification is the most prevalent modification in eukaryotic RNAs while accumulating studies suggest that m6A aberrant expression plays an important role in cancer. HNRNPA2B1 is a m6A reader which binds to nascent RNA and thus affects a perplexing array of RNA metabolism exquisitely. Despite unveiled facets that HNRNPA2B1 is deregulated in several tumors and facilitates tumor growth, a clear role of HNRNPA2B1 in multiple myeloma (MM) remains elusive. Herein, we analyzed the function and the regulatory mechanism of HNRNPA2B1 in MM. We found that HNRNPA2B1 was elevated in MM patients and negatively correlated with favorable prognosis. The depletion of HNRNPA2B1 in MM cells inhibited cell proliferation and induced apoptosis. On the contrary, the overexpression of HNRNPA2B1 promoted cell proliferation in vitro and in vivo. Mechanistic studies revealed that HNRNPA2B1 recognized the m6A sites of ILF3 and enhanced the stability of ILF3 mRNA transcripts, while AKT3 downregulation by siRNA abrogated the cellular proliferation induced by HNRNPA2B1 overexpression. Additionally, the expression of HNRNPA2B1, ILF3 and AKT3 was positively associated with each other in MM tissues tested by immunohistochemistry. In summary, our study highlights that HNRNPA2B1 potentially acts as a therapeutic target of MM through regulating AKT3 expression mediated by ILF3-dependent pattern.

Deacetylation Induced Nuclear Condensation of HP1γ Promotes Multiple Myeloma Drug Resistance

2021 ◽  
Author(s):  
Zhiqiang Liu ◽  
Xin Li ◽  
Sheng Wang ◽  
Ying Xie ◽  
Hongmei Jiang ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Dna Damage ◽  
Drug Resistance ◽  
Heterochromatin Protein 1 ◽  
Nuclear Condensation ◽  
Repair Capacity ◽  
Acquired Drug Resistance ◽  
Histone Deacetylase 1

Abstract Acquired chemoresistance to proteasome inhibitors (PIs) is a major obstacle that results in failure to manage patients with multiple myeloma (MM) in the clinic; however, the key regulators and underlying mechanisms are still unclear. In this study, we found that high levels of a chromosomal modifier, heterochromatin protein 1 gamma (HP1γ), are accompanied by a low acetylation level in bortezomib-resistant (BR) MM cells, and aberrant DNA repair capacity is correlated with HP1γ overexpression. Mechanistically, the deacetylation of HP1γ at lysine 5 by histone deacetylase 1 (HDAC1) alleviates HP1γ ubiquitination, and the stabilized HP1γ recruits the mediator of DNA damage checkpoint 1 (MDC1) to induce DNA damage repair. Simultaneously, deacetylation modification and MDC1 recruitment enhance the nuclear condensate of HP1γ, which facilitates the chromatin accessibility of genes governing sensitivity to PIs, such as FOS, JUN and CD40. Thus, targeting HP1γ stability using the HDAC1/2 inhibitor, romidepsin, sensitizes PIs treatment and overcomes drug resistance both in vitro and in vivo. Our findings elucidate a previously unrecognized role of HP1γ in the acquired drug resistance of MM and suggest that targeting HP1γ may be efficacious for overcoming drug resistance in MM patients.

scholarly journals Anti-CD38-blocked ricin: an immunotoxin for the treatment of multiple myeloma [see comments]

Blood ◽  
1994 ◽  
Vol 84 (9) ◽  
pp. 3017-3025 ◽  
Author(s):  
VS Goldmacher ◽  
LA Bourret ◽  
BA Levine ◽  
RA Rasmussen ◽  
M Pourshadi ◽  
...  
Keyword(s):  
Multiple Myeloma ◽  
Tumor Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Tumor Cell Line ◽  
Covalent Attachment ◽  
Myeloma Cells ◽  
Colony Forming Units

Abstract We report the development of a potent anti-CD38 immunotoxin capable of killing human myeloma and lymphoma cell lines. The immunotoxin is composed of an anti-CD38 antibody HB7 conjugated to a chemically modified ricin molecule wherein the binding sites of the B chain have been blocked by covalent attachment of affinity ligands (blocked ricin). Conjugation of blocked ricin to the HB7 antibody has minimal effect on the apparent affinity of the antibody and no effect on the ribosome-inactivating activity of the ricin A-chain moiety. Four to six logs of CD38+ tumor cell line kill was achieved at concentrations of HB7-blocked ricin in the range of 0.1 to 3 nmol/L. Low level of toxicity for normal bone marrow (BM) granulocyte-macrophage colony- forming units (CFU-GM), burst-forming units-erythroid (BFU-E), colony- forming units-granulocyte/erythroid/monocyte/macrophage (CFU-GEMM) cells was observed. Greater than two logs of CD38+ multiple myeloma cells were depleted from a 10-fold excess of normal BM mononuclear cells (BMMCs) after an exposure to HB7-blocked ricin under conditions (0.3 nmol/L) that were not very toxic for the normal BM precursors. HB7- blocked ricin was tested for its ability to inhibit protein synthesis in fresh patients' multiple myeloma cells and in normal BMMCs isolated from two healthy volunteers; tumor cells from four of five patients were 100-fold to 500-fold more sensitive to the inhibitory effect of HB7-blocked ricin than the normal BM cells. HB7 antibody does not activate normal resting peripheral blood lymphocytes, and HB7-blocked ricin is not cytotoxic toward these cells at concentrations of up to 1 nmol/L. The potent killing of antigen-bearing tumor cells coupled with a lack of effects on peripheral blood T cells or on hematopoietic progenitor cells suggests that HB7-blocked ricin may have clinical utility for the in vivo or in vitro purging of human multiple myeloma cells.

scholarly journals Jianpi Yangwei decoction promotes apoptosis and suppresses proliferation of 5-fluorouracil resistant gastric cancer cells in vitro and in vivo

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Huijuan Tang ◽  
Wenjie Huang ◽  
Qiang Yang ◽  
Ying Lin ◽  
Yihui Chen ◽  
...  
Keyword(s):  
Gastric Cancer ◽  
Drug Resistance ◽  
Cell Line ◽  
Real Time ◽  
Mtt Assay ◽  
Xenograft Tumor ◽  
Rt Pcr ◽  
Induced Apoptosis

Abstract Background The exploration of new therapeutic agents targeting 5-Fu resistance may open a new opportunity to gastric cancer treatment. The objective is to establish a 5-Fu resistant gastric cancer cell line and observe the effect of Jianpi Yangwei decoction (JPYW) on its apoptosis and drug-resistance related proteins. Methods MTT assay was used to measure the effect of JPYW on the BGC823 cells proliferation, and the apoptosis was observed by flow cytometry and Hoechst fluorescence staining. The BGC823 xenograft tumor nude mice models were established, the apoptosis was detected by Tunel method. BGC-823/5-Fu was established by repeated low-dose 5-Fu shocks, the drug resistance index and proliferation were detected by the MTT assay; MDR1 mRNA was detected by real-time RT-PCR; Western blot was used to detect the ratio of p-AKT to AKT; The BGC823/5-Fu xenograft tumor nude mice models were established and apoptosis was measured. The expressions of MRP1, MDR1, ABCG2, AKT, p-AKT, caspase-3 and bcl-2 were detected by immunohistochemistry and the AKT mRNA expression was detected by real-time RT-PCR. Results JPYW induced apoptosis in BGC823 cells; Drug-resistant cell line BGC-823/5-Fu was sucessfully established; JPYW induced apoptosis of BGC823/5-Fu cells, down-regulated the expression of MRP1, MDR1 and ABCG2 in vitro and in vivo, and further decreased MDR1 expression when combined with pathway inhibitor LY294002 (P < 0.05); JPYW down-regulated the ratio of p-AKT to AKT in vitro in a dose-dependent manner, the same as after the combination with LY294002 (P < 0.05). Conclusion JPYW can induce apoptosis of BGC823 and BGC823/5-Fu cells, and down-regulate the expression of MDR1, MRP1, ABCG2 in vitro and in vivo. Its in vitro effect is related to the PI3K/AKT signaling pathway.

Azacytidine Suppressors Autocrine IL-6 Secretion and Demonstrates In Vitro and In Vivo Activity Against Multiple Myeloma.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1566-1566
Author(s):  
Tiffany Khong ◽  
Janelle Sharkey ◽  
Andrew Spencer
Keyword(s):  
Multiple Myeloma ◽  
Murine Model ◽  
Dna Methyltransferase ◽  
Myeloma Cell ◽  
Apoptotic Pathway ◽  
P16 Gene ◽  
Induced Apoptosis ◽  
The Impact

Abstract Azacytidine (AZA), a DNA methyltransferase inhibitor, has been shown to inhibit cell growth and induce apoptosis in some cancer cells. We determined the impact of AZA on a panel of human myeloma cell lines (HMCL); KMS 12PE, KMS 18, LP-1, NCI-H929, OPM-2, RPMI-8226 and U266 and in an in vivo murine model of multiple myeloma (5T33 model). Dose responsiveness to AZA was determined via MTS assays with a range of AZA doses (1–10mM) for 72 hours. FACS and cell cycle analysis were used to evaluate the profile of the cells after exposure to AZA for 72 hours. MTS assays demonstrated a dose and time dependent AZA-induced inhibition of HMCL viability with effective concentrations of AZA ranging from 1–10 mM. This was associated with accumulation of cells in the Go/G1 phase with decreasing number of cells in the S and G2/M phases. Western Blot analysis using antibodies against caspases 3,8,10, PARP, phospho-ERK, ERK, Stat3 and phospho -Stat3 were performed to help characterize the mechanism(s) of cell killing. Cleavage of caspases 3,8,10 and PARP within 24 hours of AZA treatment confirmed early AZA-induced HMCL apoptosis. phospho-ERK which was absent in untreated U266 appeared after 48 hours exposure to 5mM AZA. Similarly inhibitors of caspases 3,8 and 9 were used to determine which apoptotic pathway was being preferentially activated by AZA. Inhibitors of both caspase 3 and 9 effectively abrogated AZA-induced apoptosis in U266 and NCI-H929. In contrast caspase 8 inhibitor was less effective which is consistent with AZA acting via the mitochondrial apoptotic pathway. Reactivation of p16 gene by AZA-induced hypomethylation was assessed with methylation specific PCR. MSP-PCR of the p16 gene indicated a loss of methylation and up-regulated transcription after 48 hours treatment with 5 mM AZA. The level of IL-6 in conditioned media from U266 cells treated with AZA was determined by ELISA assay and demonstrated a rapid fall in autocrine IL-6 production. RT-PCR demonstrated rapid AZA-induced cessation of IL-6 transcription temporarily associated with the disappearance of upstream phospho -Stat3. Addition of exogenous IL-6 did not rescue U266 from AZA-induced apoptosis. AZA was also administered to a 5T33 murine model of multiple myeloma at increasing concentrations (1, 3, 10 mg/kg). At 10 mg/kg the median survival of vehicle versus AZA treated mice was 28 days versus 30+ days (p=0.003). These findings justify further evaluation of AZA as a potential therapeutic agent for multiple myeloma.

JS-K, a GST-Activated Nitric Oxide Generator, Induces Apoptosis and Overcomes In Vitro Drug Resistance in Multiple Myeloma Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 1593-1593
Author(s):  
Tanyel Kiziltepe ◽  
Kenji Ishitsuka ◽  
Teru Hideshima ◽  
Noopur Raje ◽  
Norihiko Shiraishi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Multiple Myeloma ◽  
Drug Resistance ◽  
Tumor Cells ◽  
Cell Lines ◽  
Biological Effects ◽  
Treatment Modalities ◽  
Cancer Drug ◽  
Induced Apoptosis

Abstract Multiple myeloma (MM) is currently an incurable hematological malignancy. A major reason for the failure of currently existing therapies is the chemotherapeutic resistance acquired by the MM cells upon treatment. Overexpression of glutathione S-transferases (GST) has been shown as one possible mechanism of anti-cancer drug resistance in a broad spectrum of tumor cells. JS-K (O2-(2,4-Dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate) belongs to a class of pro-drugs which are designed to release nitric oxide (NO) on reaction with GST. JS-K can possibly turn GST overexpression to the tumor’s disadvantage by (1) consuming intracellular GSH and preventing drug inactivation; and (2) by exposing tumor cells to high intracellular concentrations of NO. JS-K has potent in vitro and in vivo anti-leukemic activity. The purpose of the present study is to examine the biological effects of JS-K on human MM cells. We demonstrate that JS-K has significant in vitro cytotoxicity on MM cell lines, with an IC50 of 0.3-2 mM at 48 hours. JS-K also induces cytotoxicity on cell lines that are resistant to conventional chemotherapy (i.e., MM1R, RPMI-Dox40, RPMI-LR5, RPMI-MR20). Importantly, no cytotoxic effects of JS-K were detected on peripheral blood mononuclear cells (PBMNC) obtained from healthy volunteers at these doses. Moreover, JS-K could overcome the survival and growth advantages conferred by interleukin-6 (IL-6) and insulin-like growth factor-1 (IGF-1), or by adherence of MM cells to bone marrow stromal cells (BMSC). JS-K caused a transient G2/M arrest followed by apoptosis, as determined by flow cytometric analysis using PI, Annexin V and Apo2.7 staining. JS-K-induced apoptosis was associated with caspase 8, 7, 9 and 3 activation. Interestingly, Fas was upregulated by JS-K, suggesting the involvement of death receptor pathway in induction of apoptosis. JS-K also triggered Mcl-1 cleavage and Bcl-2 phosphorylation, suggesting the involvement of mitochondrial pathway. In addition, apoptosis inducing factor (AIF), endonuclease G (EndoG) and cytochrome c were released into the cytosol during apoptosis. Taken together, these findings suggest the involvement of both intrinsic and extrinsic apoptotic pathways in JS-K-induced apoptosis in MM cells. In summary, our studies demonstrate that JS-K induces apoptosis and overcomes in vitro drug resistance in MM cells. Therefore, JS-K is a novel compound which carries significant potential to be included in the repertoire of existing treatment modalities for MM. Ongoing studies are delineating the mechanism of action of JS-K to provide the preclinical rationale for combination therapies to overcome drug resistance and improve patient outcome.

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