Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of non-Hodgkin's lymphoma, with the combination of rituximab and chemotherapy being the standard treatment for it. Although rituximab monotherapy has a remarkable response rate, drug resistance with unclear mechanisms and lack of effective second-line therapy limit the survival benefits of patients with lymphoma. Here, we report that MORTALIN is highly expressed and correlates with resistance to rituximab-based therapy and poor survival in patients with DLBCL. Mechanistically, gain- and loss-of-function experiments revealed that the voltage-dependent anion channel 1-binding protein, MORTALIN, regulated Ca2+ release from the endoplasmic reticulum through mitochondria-associated membrane, facilitating AP1-mediated cell proliferation and YY-1-mediated downregulation of FAS in DLBCL cells. These dual mechanisms contribute to rituximab resistance. In mouse models, genetic depletion of MORTALIN markedly increased the antitumor activity of rituximab. We shed mechanistic light on MORTALIN-Ca2+-CaMKII-AP1-mediated proliferation and MORTALIN-Ca2+-CaMKII-inhibited death receptor in DLBCL, leading to rituximab resistance, and propose MORTALIN as a novel target for the treatment of DLBCL.

OTUD7B is a deubiquitinase and has been reported as a prognostic factor in various solid tumors. However, its prognostic value in lymphoma patients remains unclear. We detected OTUD7B expression levels in 160 diffuse large B-cell lymphoma (DLBCL) tissue samples by immunohistochemistry, and analyzed correlations between its expression and clinic-pathologic parameters as well as clinical outcomes. We also investigated association between OTUD7B expression and chemotherapeutic drugs anti-tumor activity in vitro. We found that OTUD7B overexpressed in 129 (80.6%) cases, and patients with overexpression of OTUD7B experienced better overall survival comparing to those with OTUD7B low expression (P=0.021). Multivariate Cox regression analysis illustrated that OTUD7B was an independent prognostic indicator. In DLBCL cell lines, we found that Chidamide could up-regulate OTUD7B in several DLBCL cell lines, and also had synergistic effect with doxorubicin at low concentration. Our data illustrated that OTUD7B deficiency is a negative predictor of clinical outcome, and might be a potential therapeutic target in the treatment of diffuse large B-cell lymphoma.

Previous studies show that mortalin, a HSP70 family member, contributes to the development and progression of ovarian cancer. However, details of the transcriptional regulation of mortalin remain unknown. We aimed to determine whether NF-kappa B p65 participates in the regulation of mortalin expression in ovarian cancer cells and to elucidate the underlying mechanism. Chromatin immunoprecipitation and luciferase reporter assay were used to identify mortalin gene sequences, to which NF-kappa B p65 binds. Results indicated that NF-kappa B p65 binds to the mortalin promoter at a site with the sequence 'CGGGGTTTCA'. Using lentiviral pLVX-NF-kappa B-puro and Lentivirus-delivered NF-kappa B short hairpin RNA (shRNA), we created ovarian cancer cell lines in which NE-kappa B p65 was stably up-regulated and down-regulated. Using these cells, we found that downregulation of NF-kappa B p65 inhibits the growth and migration of ovarian cancer cells. Further experimental evidence indicated that downregulation of NF-kappa B p65 reduced mortalin, and upregulation of mortalin rescued the proliferation and migration of ovarian cancer cells reduced by NF-kappa B p65 knockdown. In conclusion, NF-kappa B p65 binds to the mortalin promoter and promotes ovarian cancer cells proliferation and migration via regulating mortalin.

p38 mitogen-activated protein kinases are signaling molecules with major involvement in cancer. A detailed mechanistic understanding of how p38 MAPK family members function is urgently warranted for cancer targeted therapy. The conformational dynamics of the most common member of p38 MAPK family, p38α, are crucial for its function but poorly understood. Here we found that, unlike in other cancer types, p38α is significantly activated in pancreatic adenocarcinoma samples, suggesting its potential for anti-pancreatic cancer therapy. Using a state of the art supercomputer, Anton, long-timescale (39 μs) unbiased molecular dynamics simulations of p38α show that apo p38α has high structural flexibility in six regions, and reveal potential catalysis mechanism involving a “butterfly” motion. Moreover, in vitro studies show the low-selectivity of the current p38α inhibitors in both human and mouse pancreatic cancer cell lines, while computational solvent mapping identified 17 novel pockets for drug design. Taken together, our study reveals the conformational dynamics and potentially druggable pockets of p38α, which may potentiate p38α-targeting drug development and benefit pancreatic cancer patients.

Recent efforts to develop tumor-targeted photodynamic therapy (PDT) photosensitizers (PSs) have greatly advanced the potential of PDT in cancer therapy, although complete eradication of tumor cells by PDT alone remains challenging. As a way to improve PDT efficacy, we report a new combinatory PDT therapy technique that specifically targets multilayers of cells. Simply mixing different PDT PSs, even those that target distinct receptors (this may still lead to similar cell-killing pathways), may not achieve ideal therapeutic outcomes. Instead, significantly improved outcomes likely require synergistic therapies that target various cellular pathways. In this study, we target two proteins upregulated in cancers: the cannabinoid CB2 receptor (CB2R, a G-protein coupled receptor) and translocator protein (TSPO, a mitochondria membrane receptor). We found that the CB2R-targeted PS, IR700DX-mbc94, triggered necrotic cell death upon light irradiation, whereas PDT with the TSPO-targeted IR700DX-6T agent led to apoptotic cell death. Both PSs significantly inhibited tumor growth in vivo in a target-specific manner. As expected, the combined CB2R- and TSPO-PDT resulted in enhanced cell killing efficacy and tumor inhibition with lower drug dose. The median survival time of animals with multilayer PDT treatment was extended by as much as 2.8-fold over single PDT treatment. Overall, multilayer PDT provides new opportunities to treat cancers with high efficacy and low side effects.Statement of SignificancePhotodynamic therapy (PDT) is increasingly used as a minimally invasive, controllable and effective therapeutic procedure for cancer treatment. However, complete eradication of tumor cells by PDT alone remains challenging. In this study, we investigate the potential of multilayer PDT in cancer treatment with high efficacy and low side effects. Through PDT targeting two cancer biomarkers located at distinct subcellular localizations, remarkable synergistic effects in cancer cell killing and tumor inhibition were observed in both in vitro and in vivo experiments. This strategy may be widely applied to treat various cancer types by using strategically designed PDT photosensitizers that target corresponding upregulated receptors at tactical subcellular localization. (C) 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Salvianolic acid B is an antioxidative ingredient derived from Radix Salviae miltiorrhizae that has been widely used to treat liver diseases. However, the therapeutic mechanism underlying Salvianolic acid B has remained largely unknown. Our studies verified that Salvianolic acid B efficiently blocked mitochondrial deformation and dysfunction induced by H2O2 in the human hepatocyte cell line HL7702. Mortalin, a mitochondrial molecular chaperone, maintains mitochondrial morphology stabilization and function integrity. Previous results showed that mortalin overexpression has been observed in hematoma carcinoma cells and that mortalin maintains mitochondrial homeostasis and antagonizes oxidative stress damage. We found that Salvianolic acid B significantly up-regulated mortalin protein expression levels. In addition, Salvianolic acid B lost the function of preventing mitochondrial deformation and dysfunction induced by oxidative stress under mortalin knockdown conditions. We further found that mortalin overexpression increases the mRNA expression of mitofusin-related factor Mfn1 and mitofission-related factor hFis1. In conclusion, Salvianolic acid B maintains the mitochondrial structure stabilization and functional integrity by up-regulating mortalin, which may be associated with increased mitofusin factor Mfn1 and reduced mitofission factor hFis1.

Mortalin is frequently overexpressed in human malignancies. Previous studies have suggested that mortalin contributes to ovarian cancer development and progression, but further investigation is warranted. The aim of this study is to elucidate the mechanism of mortalin in ovarian cancer development and progression. In this study, lentivirus‐delivered mortalin short hairpin RNA (shRNA) was used to knockdown mortalin expression in A2780 and A2780/cis ovarian cancer cell lines, and lentiviral mortalin‐pLVX‐AcGFP was used to generate mortalin‐overexpressing cell lines. The results demonstrated that decreased mortalin expression reduced ovarian cancer cell proliferation, colony formation, migration and invasion by Cell Counting Kit‐8 assay, colony formation assay, wounding healing assay and Transwell cell invasion assay, respectively. Flow cytometry results suggested that mortalin promotes the G1 transition, leading to faster restoration of a normal cell‐cycle distribution. Cell‐cycle proteins, including C‐myc and Cyclin‐D1, significantly increased, and Cyclin‐B1 remarkably decreased upon mortalin down‐regulation. Western blot analysis showed that mortalin knockdown significantly decreased p‐c‐Raf and phospho‐extracellular–regulated protein kinases (p‐ERK1/2) pathways but not the Jun N‐terminal kinase pathway, whereas mortalin overexpression had the opposite effect. Taken together, these results indicate that mortalin is an oncogenic factor, and mitogen‐activated protein kinase‐ERK signalling pathway activation by mortalin may contribute to ovarian cancer development and progression.

Photodynamic therapy (PDT) has been proven to be a minimally invasive and effective therapeutic strategy for cancer treatment. It can be used alone or as a complement to conventional cancer treatments, such as surgical debulking and chemotherapy. The mitochondrion is an attractive target for developing novel PDT agents, as it produces energy for cells and regulates apoptosis. Current strategy of mitochondria targeting is mainly focused on utilizing cationic photosensitizers that bind to the negatively charged mitochondria membrane. However, such an approach is lack of selectivity of tumor cells. To minimize the damage on healthy tissues and improve therapeutic efficacy, an alternative targeting strategy with high tumor specificity is in critical need. Herein, we report a tumor mitochondria-specific PDT agent, IR700DX-6T, which targets the 18 kDa mitochondrial translocator protein (TSPO). IR700DX-6T induced apoptotic cell death in TSPO-positive breast cancer cells (MDA-MB-231) but not TSPO-negative breast cancer cells (MCF-7). In vivo PDT study suggested that IR700DX-6T-mediated PDT significantly inhibited the growth of MDA-MB-231 tumors in a target-specific manner. These combined data suggest that this new TSPO-targeted photosensitizer has great potential in cancer treatment.Statement of SignificancePhotodynamic therapy (PDT) is an effective and minimally invasive therapeutic technique for treating cancers. Mitochondrion is an attractive target for developing novel PDT agents, as it produces energy to cells and regulates apoptosis. Current mitochondriatargeted photosensitizers (PSs) are based on cationic molecules, which interact with the negatively charged mitochondria membrane. However, such PSs are not specific for cancerous cells, which may result in unwanted side effects. In this study, we developed a tumor mitochondria-targeted PS, IR700DX-6T, which binds to translocator protein (TSPO). This agent effectively induced apoptosis in TSPO-positive cancer cells and significantly inhibited tumor growth in TSPO-positive tumor-bearing mice. These combined data suggest that IR700DX-6T could become a powerful tool in the treatment of multiple cancers that upregulate TSPO. (C) 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The heat shock protein mortalin is frequently overexpressed in human malignancies. In this study, an assessment of mortalin expression using ovarian cancer tissue microarrays suggested that mortalin overexpression might be related to drug resistance. Using a short hairpin (sh) RNA approach, we evaluated the effect of reducing mortalin expression in vitro and in vivo. The results of our study show that elevated levels of mortalin expression increase cancer cell resistance to cisplatin-induced cytotoxicity and identify mortalin as a potential therapeutic target for improved treatment of drug-resistant ovarian cancer.

During glucose deprivation (GD)-induced cellular stress, the molecular chaperone glucose-regulated protein 75 (Grp75)/Mortalin/PBP74/mtHSP70 (hereafter termed "Grp75") plays an important role in the suppression of apoptosis by inhibiting the Bax conformational change that delays the release of cytochrome c. The molecular pathways by which it carries out these functions are still unclear. We hypothesize that the anti-apoptotic effect by the overexpression of Grp75 was through the signal of AKT activated by classic phosphoinositide 3-kinase (PI3K) and also involved PI3K-independent pathways. Using the PC12 cell GD model, we demonstrated a novel mechanism of Grp75 activating AKT, which may be PI3K independent and associated with Raf/MEK (mitogen-activated protein kinase/ERK kinase)/ERK signaling. The PI3K inhibitor LY294002 did not influence the activation of AKT by the Grp75 overexpression under GD; however, the MEK inhibitor U0126 dramatically inhibited AKT phosphorylation in the same assay. In addition to the PI3K/AKT signal pathway, Grp75 overexpression also inhibited the Bax conformational change through the Raf/MEK/ERK signal pathway. In conclusion, Grp75 overexpression in activating AKT can be PI3K independent and associated with Raf/MEK/ERK signaling under GD. At the same time, PI3K may also crosstalk with Raf-1, in which the prosurvival signal of PI3K maintains the expression of Raf-1. The activated AKT and extracellular signal-regulated protein kinases 1 and 2 by Grp75 inhibited the Bax conformational change and subsequent apoptosis.