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Tumor-associated macrophages (TAMs) are correlated with the prognosis of different types of solid tumors and lymphoma, according to many clinical studies. In vitro experiments have demonstrated the roles of these cells in myeloma cell survival, angiogenesis, immunomodulation, drug resistance, and the interaction between malignant myeloma cells and the microenvironment. Here, we investigated the prognostic significance of TAMs in patients with multiple myeloma (MM). We evaluated the polarized functional status of bone marrow infiltrated by TAMs by immunohistochemical staining of CD68, iNOS, and CD163 in 240 patients with MM from January 2009 to December 2014. The overall response rates to chemotherapy were lower in patients with high CD68+ or CD163+ TAM densities than in those with low densities. Kaplan-Meier analysis showed that the progression-free survival (PFS, p = 0.001) and overall survival (OS, p < 0.001) of patients with low CD163+ TAM density were significantly higher than those of patients with high CD163+ TAM density. Furthermore, combined analysis of iNOS+ and CD163+ TAMs (iNOS/CD163 signature) exhibited greater power in predicting patient outcomes for both PFS (p < 0.001) and OS (p < 0.001). Moreover, Cox regression analysis identified iNOS+ and CD163+ TAMs as independent prognostic factors (p = 0.007, p < 0.001, respectively). These factors could be combined with the international staging system (ISS) to generate a predictive nomogram for patient outcomes. Our findings suggest that the mosaic of diametrically polarized TAMs is a novel independent prognostic factor that could be integrated into the evaluation of and therapy for MM.

Epigenetic abnormalities play important roles in the pathogenesis of Hodgkin lymphoma (HL). Highly expressed class I histone acetyltransferase (HDAC) and hyper-methylation of the promoter region of tumor suppressor genes have been demonstrated in Hodgkin lymphoma. In this paper, we investigated the synergistic effects of combination treatment of chidamide, a selective HDAC inhibitor, and decitabine, a demethylation agent, for HL cell lines and explored a new strategy for treating HL. The apoptosis rates, cell cycle, mitochondrial transmembrane potentials, epigenetic changes and gene expression of HL cell lines treated with chidamide as a single agent and in combination with decitabine were tested. We found that chidamide inhibited the proliferation of HL cells through increased apoptosis. Interestingly, after combined with decitabine, the inhibition rate and apoptotic death in HL cells were significantly increased. Further studies demonstrated that when combined with decitabine the expression of acetylated histone H3 and H3K9 induced by chidamide in HL cells increased, and also the expression of tumor suppressor genes PU.1 and KLF4, which exert inhibition through apoptosis pathway. Therefore, we could come to a conclusion that chidamide and decitabine can synergistically induce apoptosis of Hodgkin lymphoma cells by up-regulating the expression of PU.1 and KLF4. These results provide a new sight in combining two different epigenetic regulatory agents for treating HL.

PIG7 localizes to lysosomal membrane in leukemia cells. Our previous work has shown that transduction of pig7 into a series of leukemia cell lines did not result in either apoptosis or differentiation of most tested cell lines. Interestingly, it did significantly sensitize these cell lines to chemotherapeutic drugs. Here, we further investigated the mechanism underlying pig7-induced improved sensitivity of acute leukemia cells to chemotherapy. Our results demonstrated that the sensitization effect driven by exogenous pig7 was more effective in drug-resistant leukemia cell lines which had lower endogenous pig7 expression. Overexpression of pig7 did not directly activate the caspase apoptotic pathway, but decreased the lysosomal stability. The expression of pig7 resulted in lysosomal membrane permeabilization (LMP) and lysosomal protease (e.g. cathepsin B, D, L) release. Moreover, we also observed increased reactive oxygen species (ROS) and decreased mitochondrial membrane potential (ΔΨm) induced by pig7. Some autophagy markers such as LC3I/II, ATG5 and Beclin-1, and necroptosis maker MLKL were also stimulated. However, intrinsic antagonism such as serine/cysteine protease inhibitors Spi2A and Cystatin C prevented downstream effectors from triggering leukemia cells, which were only on the "verge of apoptosis". When combined with chemotherapy, LMP increased and more proteases were released. Once this process was beyond the limit of intrinsic antagonism, it induced programmed cell death cooperatively via caspase-independent and caspase-dependent pathways.

// Yi Li 1,2,* , Yuhuan Zheng 2,3,* , Tianshu Li 2,* , Qiang Wang 2 , Jianfei Qian 2 , Yong Lu 2 , Mingjun Zhang 2 , Enguang Bi 2 , Maojie Yang 2 , Frederic Reu 4 , Qing Yi 2,** and Zhen Cai 1,** 1 Bone Marrow Transplantation Center, Department of Hematology, Zhejiang University, Hangzhou, Zhejiang, China 2 Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, OH, USA 3 Department of Hematology, Sichuan University, West China School of Medicine, Chengdu, Sichuan, China 4 Taussig Cancer Institute, Cleveland Clinic, Cleveland, OH, USA * These authors have contributed equally to this manuscript ** These authors have contributed equally as senior authors to this manuscript Correspondence to: Zhen Cai, email: // Qing Yi, email: // Keywords : multiple myeloma, chemokine, macrophage, bone marrow Received : April 24, 2015 Accepted : June 05, 2015 Published : June 19, 2015 Abstract We previously showed that macrophages (MΦs) infiltrate the bone marrow (BM) of patients with myeloma and may play a role in drug resistance. This study analyzed chemokines expressed by myeloma BM that are responsible for recruiting monocytes to the tumor bed. We found that chemokines CCL3, CCL14, and CCL2 were highly expressed by myeloma and BM cells, and the levels of CCL14 and CCL3 in myeloma BM positively correlated with the percentage of BM-infiltrating MΦs. In vitro, these chemokines were responsible for chemoattracting human monocytes to tumor sites and in vivo for MΦ infiltration into myeloma-bearing BM in the 5TGM1 mouse model. Surprisingly, we also found that these chemokines stimulated MΦ in vitro proliferation induced by myeloma cells and in vivo in a human myeloma xenograft SCID mouse model. The chemokines also activated normal MΦ polarization and differentiation into myeloma-associated MΦs. Western blot analysis revealed that these chemokines promoted growth and survival signaling in MΦs via activating the PI3K/Akt and ERK MAPK pathways and c-myc expression. Thus, this study provides novel insight into the mechanism of MΦ infiltration of BM and also potential targets for improving the efficacy of chemotherapy in myeloma.

// Mingjun Zhang 1, 2 , Jin He 1 , Zhiqiang Liu 1 , Yong Lu 1, 2 , Yuhuan Zheng 1, 2 , Haiyan Li 1, 2 , Jingda Xu 1 , Huan Liu 1 , Jianfei Qian 1, 2 , Robert Z. Orlowski 1 , Larry W. Kwak 1 , Qing Yi 1, 2 , Jing Yang 1, 3 1 Department of Lymphoma/Myeloma, Division of Cancer Medicine, The University of Texas MD Anderson Cancer Center, Houston, Texas, USA 2 Department of Cancer Biology, Lerner Research Institute, Cleveland Clinic, Cleveland, Ohio, USA 3 Cancer Research Institute and Cancer Hospital, Guangzhou Medical University, Guangzhou, China Correspondence to: Jing Yang, e-mail: jiyang@mdanderson.org Qing Yi, e-mail: yiq@ccf.org Keywords: multiple myeloma, anti-β 2 M monoclonal antibody, bortezomib, autophagy, NF-κ p65 Received: November 17, 2014 Accepted: January 29, 2015 Published: March 31, 2015 ABSTRACT Our previous studies showed that anti-β 2 M monoclonal antibodies (mAbs) have strong and direct apoptotic effects on multiple myeloma (MM) cells, suggesting that anti-β 2 M mAbs might be developed as a novel therapeutic agent. In this study, we investigated the anti-MM effects of combination treatment with anti-β 2 M mAbs and bortezomib (BTZ). Our results showed that anti-β 2 M mAbs enhanced BTZ-induced apoptosis of MM cell lines and primary MM cells. Combination treatment could also induce apoptosis of BTZ-resistant MM cells, and the enhanced effect depended on the surface expression of β 2 M on MM cells. BTZ up-regulated the expression of autophagy proteins, whereas combination with anti-β 2 M mAbs inhibited autophagy. Sequence analysis of the promoter region of beclin 1 identified 3 putative NF-κB-binding sites from –615 to –789 bp. BTZ treatment increased, whereas combination with anti-β 2 M mAbs reduced, NF-κB transcription activities in MM cells, and combination treatment inhibited NF-κB p65 binding to the beclin 1 promoter. Furthermore, anti-β 2 M mAbs and BTZ combination treatment had anti-MM activities in an established MM mouse model. Thus, our studies provide new insight and support for the clinical development of an anti-β 2 M mAb and BTZ combination treatment to overcome BTZ drug resistance and improve MM patient survival.

// Xi Yang 1, * , Tianyou Yan 1, * , Yuping Gong 1 , Xuehua Liu 1 , Huaqin Sun 2 , Wenming Xu 2 , Chunsen Wang 3 , Duolan Naren 1 , Yuhuan Zheng 1 1 Department of Hematology, West China Hospital, Sichuan University, Chengdu, China 2 Sichuan University-The Chinese University of Hong Kong Joint Laboratory for Reproductive Medicine, West China Second University Hospital, Chengdu, China 3 Department of Hematology, Sichuan Provincial People’s Hospital, Affiliated Hospital of University of Electronic Science and Technology of China, Chengdu, China * These authors contributed equally to this work Correspondence to: Yuping Gong, email: gongyuping2010@aliyun.com Keywords: CFTR, Ph + acute leukemia, PP2A Received: May 30, 2016 Accepted: February 06, 2017 Published: February 19, 2017 ABSTRACT Cystic fibrosis transmembrane conductance regulator (CFTR) is classified as an anion channel transporter of Cl − and HCO3 − . Through interactions with its PDZ domain, CFTR is capable of regulating other proteins, such as protein phosphatase 2A (PP2A). The aberrant expression and mutation of CFTR have been observed in several tumor, but not in philadelphia chromosome–positive(Ph + ) acute leukemia, including Ph + B cell acute lymphoblastic leukemia(Ph + B-ALL) and chronic myelogenous leukemia blast crisis phases (CML-BC). In this study, we demonstrated the mean expression level of CFTR in Ph + acute leukemia cells was markedly higher than that in Ph- B-ALL and CML-chronic phase cells. CFTRinh-172, a classic CFTR inhibitor, down-regulated the expression of CFTR, p-BCR-ABL and classical Wnt/β-catenin signaling in Ph + acute leukemia cells, while imatinib had no effect on CFTR. Importantly, reduced efficacy of CFTRinh-172 was closely associated with elevated PP2A phosphatase activity. Furthermore, we confirmed an interaction between CFTR and the PP2AA subunit in K562 cells. In addition, we demonstrated CFTR and PP2AA interact in the cytosol, resulting in PP2 A complex inactivation and increased degradation of PP2 A substrates via the lysosomal/proteasome pathway. In conclusion, our results showed CFTR was highly expressed in Ph + acute leukemia, which protected and maintained the continuous activation of BCR-ABL and the canonical Wnt/β-catenin signaling pathway by decreasing PP2A phosphatase activity. According to this working model of the CFTR-PP2A-BCR-ABL axis, targeting the CFTR protein will activate PP2A and may offer a new treatment strategy for Ph + acute leukemia, especially for patients exhibiting high levels of CFTR expression.

Epigenetic abnormalities play important roles in the pathogenesis of Hodgkin lymphoma (HL). Highly expressed class I histone acetyltransferase (HDAC) and hyper-methylation of the promoter region of tumor suppressor genes have been demonstrated in Hodgkin lymphoma. In this paper, we investigated the synergistic effects of combination treatment of chidamide, a selective HDAC inhibitor, and decitabine, a demethylation agent, for HL cell lines and explored a new strategy for treating HL. The apoptosis rates, cell cycle, mitochondrial transmembrane potentials, epigenetic changes and gene expression of HL cell lines treated with chidamide as a single agent and in combination with decitabine were tested. We found that chidamide inhibited the proliferation of HL cells through increased apoptosis. Interestingly, after combined with decitabine, the inhibition rate and apoptotic death in HL cells were significantly increased. Further studies demonstrated that when combined with decitabine the expression of acetylated histone H3 and H3K9 induced by chidamide in HL cells increased, and also the expression of tumor suppressor genes PU.1 and KLF4, which exert inhibition through apoptosis pathway. Therefore, we could come to a conclusion that chidamide and decitabine can synergistically induce apoptosis of Hodgkin lymphoma cells by up-regulating the expression of PU.1 and KLF4. These results provide a new sight in combining two different epigenetic regulatory agents for treating HL.