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Aims Trastuzumab, the first humanized monoclonal antibody that targets human epidermal growth factor receptor 2 (ERBB2/HER2), is currently used as a first-line treatment for HER2 (+) tumours. However, trastuzumab increases the risk of cardiac complications without affecting myocardial structure, suggesting a distinct mechanism of cardiotoxicity. Methods and results We used medium from trastuzumab-treated human umbilical vein endothelial cells (HUVECs) to treat CCC-HEH-2 cells, the human embryonic cardiac tissue-derived cell lines, and human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) to assess the crosstalk between vascular endothelial cells (VECs) and cardiomyocytes. Protein mass spectrometry analysis was used to identify the key factors from VECs that regulate the function of cardiomyocytes. We applied RNA-sequencing to clarify the mechanism, by which PTX3 causes cardiac dysfunction. We used an anti-human/rat HER2 (neu) monoclonal antibody to generate a rat model that was used to evaluate the effects of trastuzumab on cardiac structure and function and the rescue effects of lapatinib on trastuzumab-induced cardiac side effects. Medium from trastuzumab-treated HUVECs apparently impaired the contractility of CCC-HEH-2 cells and iPSC-CMs. PTX3 from VECs caused defective cardiomyocyte contractility and cardiac dysfunction in mice, phenocopying trastuzumab treatment. PTX3 affected calcium homoeostasis in cardiomyocytes, which led to defective contractile properties. EGFR/STAT3 signalling in VECs contributed to the increased expression and release of PTX3. Notably, lapatinib, a dual inhibitor of EGFR/HER2, could rescue the cardiac complications caused by trastuzumab by blocking the release of PTX3. Conclusion We identified a distinct mode of cardiotoxicity, wherein the activation of EGFR/STAT3 signalling by trastuzumab in VECs promotes PTX3 excretion, which contributes to the impaired contractility of cardiomyocytes by inhibiting cellular calcium signalling. We confirmed that lapatinib could be a feasible preventive agent against trastuzumab-induced cardiac complications and provided the rationale for the combined application of lapatinib and trastuzumab in cancer therapy.

PDF file - 2686K, The effect of bortezomib (BTZ) and ATRA on RARα expression and NBT reduction ability in HL60-vector and HL60-shRNA-STAT1 cells.

PDF file - 186K, The effect of bortezomib (BTZ) and ATRA on PML-RARα expression in NB4 cells.

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PDF file - 1344K, The effect of bortezomib (BTZ) on NF-κB pathway activity in HL60 cells.

PDF file - 1900K, Bortezomib (BTZ) synergized with ATRA to inhibit AML cell proliferation.

All-trans-retinoic acid (ATRA) has held great promise for differentiation-based therapy but reportedly downregulates retinoic acid receptor-α (RARα) in a proteasome-dependent manner, which leads to decreased acute myeloid leukemia (AML) cell differentiation efficiency. Therefore, research strategies that seek to further sensitize cells to retinoids and extend the range of retinoid-affected myeloid malignancies beyond acute promyelocytic leukemia (APL) are key investigative avenues. Here, we show that bortezomib, the first proteasome inhibitor approved for newly diagnosed and relapsed multiple myeloma, exhibited strong synergism with ATRA to promote HL60 and NB4 AML cell differentiation. We observed that bortezomib sensitized AML cells to ATRA-induced morphologic, biochemical, and functional changes, indicative of myeloid differentiation without cell death. In addition, treatment of human leukemia HL60 xenografts with bortezomib and ATRA together did not increase bortezomib-induced progressive weight loss but resulted in significant tumor growth inhibition in addition to increased differentiation (P < 0.05). These enhanced differentiation effects were accompanied by RARα stabilization and STAT1 activation. Taken together, our study was the first to evaluate bortezomib and ATRA synergy in AML cell differentiation and to assess new opportunities for bortezomib and ATRA combination as a promising approach for future differentiation therapy. Mol Cancer Ther; 12(2); 195–206. ©2012 AACR.

Supp Figure 1. The treatment of CT-707 reduced the protein level of CTGF under hypoxia. Supp Figure 2. CT-707 elicited apoptotic cell death but not cell cycle arrest under hypoxia. Supp Figure 3. The protein level of HIF-1α under hypoxia was not influenced by YAP depletion by siRNA. Supp Figure 4. FAK inhibition was not required for the hypoxic activity of CT-707. Supp Figure 5. The semi-quantitative analyses using density measurements on the protein levels displayed in Figure 5. Supp Figure 6. The correlation of FAK, IGF-1R and YAP target genes expression with the overall survival days of liver cancer patients. Supp Figure 7. The in vivo activities of CT-707 on HepG2-, Bel-7402-xenograft and H22-allograft models. Supp Figure 8. PF-562271 treatment caused death of HepG2-xenografted nude mice but imposed less activity than CT-707.

Given that Yes-associated protein (YAP) signaling acts as a critical survival input for hypoxic cancer cells in hepatocellular carcinoma (HCC), disruption of YAP function and the maintenance of hypoxia is an attractive way to treat HCC. Utilizing a cell-based YAP-TEAD luciferase reporter assay and functional analyses, we identified CT-707, a China-FDA approved multi-kinase inhibitor under clinical trial with remarkable inhibitory activity against YAP function. CT-707 exhibited prominent cytotoxicity under hypoxia on HCC cells, which was attributable to the inhibition of YAP signaling. CT-707 arrested tumor growth in HepG2, Bel-7402, and HCC patient-derived xenografts. Mechanistically, the inhibitory activity of CT-707 on YAP signaling was due to the interruption of hypoxia-activated IGF1R. Overall, these findings not only identify CT-707 as a promising hypoxia-targeting agent against HCC, but they also unveil IGF1R as a new modulator specifically regulating hypoxia-activated YAP signaling.Significance: CT-707 may represent a novel clinical approach for patients with HCC suffering poor drug response due to intratumor hypoxia. Cancer Res; 78(14); 3995–4006. ©2018 AACR.

Materials and Methods for Supplemental Figures

Supplemental Table 1. The primer sequences for qRT-PCR assay Supplemental Table 2. The targeting sequences for siRNA Supplemental Table 3. The list of compounds for YAP-TEADs luciferase screening

Hepatotoxicity of regorafenib is one of the most noteworthy concerns for patients, however the mechanism has poorly understood. Hence, lack of effective intervention strategy. In this study, we found that regorafenib-induced liver injury mainly derived from its non-therapeutic target EPH receptor A2 (EphA2) by comparing the target with sorafenib. EphA2 deficiency attenuated liver damage and cell apoptosis under regorafenib treatment. Overexpression of EphA2-S898A could directly cause hepatocyte apoptosis, while recovering EphA2 Ser898 phosphorylation greatly alleviated regorafenib-induced liver injury. Mechanistically, regorafenib inhibits EphA2 Ser897 phosphorylation and reduced ubiquitination of p53 by altering the intracellular localization of mouse double minute 2 (MDM2) through affecting the extracellular signal-regulated kinase (ERK)/MDM2 axis. Meanwhile, we found Schisandrin C which could upregulate the phosphorylation of EphA2 at Ser897 also had protective effect against the toxicity in vivo. Collectively, our findings identify the inhibition of EphA2 Ser897 phosphorylation as a key cause of regorafenib-induced hepatotoxicity, and chemical activation on EphA2 Ser897 represents a potential therapeutic strategy to prevent regorafenib-induced hepatotoxicity.

Liver dysfunction is an outstanding dose-limiting toxicity of gefitinib, an EGFR (epidermal growth factor receptor)-tyrosine kinase inhibitor (TKI), in the treatment of EGFR mutation-positive non-small cell lung cancer (NSCLC). We aimed to elucidate the mechanisms underlying gefitinib-induced hepatotoxicity, and provide potentially effective intervention strategy. We discovered that gefitinib could sequentially activate macroautophagy/autophagy and apoptosis in hepatocytes. The inhibition of autophagy alleviated gefitinib-induced apoptosis, whereas the suppression of apoptosis failed to lessen gefitinib-induced autophagy. Moreover, liver-specific Atg7+/− heterozygous mice showed less severe liver injury than vehicle, suggesting that autophagy is involved in the gefitinib-promoted hepatotoxicity. Mechanistically, gefitinib selectively degrades the important anti-apoptosis factor COX6A1 (cytochrome c oxidase subunit 6A1) in the autophagy-lysosome pathway. The gefitinib-induced COX6A1 reduction impairs mitochondrial respiratory chain complex IV (RCC IV) function, which in turn activates apoptosis, hence causing liver injury. Notably, this autophagy-promoted apoptosis is dependent on PLK1 (polo like kinase 1). Both AAV8-mediated Plk1 knockdown and PLK1 inhibitor BI-2536 could mitigate the gefitinib-induced hepatotoxicity in vivo by abrogating the autophagic degradation of the COX6A1 protein. In addition, PLK1 inhibition could not compromise the anti-cancer activity of gefitinib. In conclusion, our findings reveal the gefitinib-hepatotoxicity pathway, wherein autophagy promotes apoptosis through COX6A1 degradation, and highlight pharmacological inhibition of PLK1 as an attractive therapeutic approach toward improving the safety of gefitinib-based cancer therapy. Abbreviations: 3-MA: 3-methyladenine; AAV8: adeno-associated virus serotype 8; ATG5: autophagy related 5; ATG7: autophagy related 7; B2M: beta-2-microglobulin; CCCP: carbonyl cyanide m-chlorophenylhydrazone; CHX: cycloheximide; COX6A1: cytochrome c oxidase subunit 6A1; c-PARP: cleaved poly(ADP-ribose) polymerase; CQ: chloroquine; GOT1/AST: glutamic-oxaloacetic transaminase 1, soluble; GPT/ALT: glutamic pyruvic transaminase, soluble; HBSS: Hanks´ balanced salt solution; H&E: hematoxylin and eosin; MAP1LC3/LC3: microtubule associated proteins 1 light chain 3; PLK1: polo like kinase 1; RCC IV: respiratory chain complex IV; ROS: reactive oxygen species; TUBB8: tubulin beta 8 class VIII

Background and Purpose Lapatinib, a widely-used dual inhibitor of EGFR/ERBB1 and HER2/ERBB2 effectively targeting HER2 positive breast cancer, has been seriously limited due to cutaneous toxicity. However, the specific mechanism of lapatinib-induced cutaneous toxicity has not been clarified, leading to a lack of effective strategy targets to improve clinical safety. Here, we aimed to identify molecular mechanism occurs in this process and strive for effective intervention strategies against lapatinib-induced cutaneous side effects. Experimental and Approach C57BL/6 mice were subjected to lapatinib via intragastric administration, serum was used for ELISA assay, skin tissue was collected and performed with histopathological analysis. Apoptotic assay was analyzed in HaCaT and NHEK cell lines, comet assay was conducted to measure the damage of DNA, real-time PCR was used to assess the level of HMGB and inflammatory factors. Key Results We found that lapatinib could induce mitochondrial dysfunction, lead to DNA damage and finally cause apoptosis of keratinocytes. In addition, we found that lapatinib could induce aberrant immune response and promote the release of inflammatory factors in vitro and in vivo. Mechanistically, downregulated expression of HMGB1 played a critical role in these toxic reaction processes. Delightfully, we found that saikosaponin A could significantly rescue the reduced HMGB1 transcription, which could alleviate lapatinib-induced DNA damage, inhibit keratinocyte apoptosis and further prevent toxicity of lapatinib in mice. Conclusion and Implications Our study provided molecular mechanism of lapatinib-induced cutaneous toxicity, and shed new light on the prevention of cutaneous adverse drug reactions induced by EGFR inhibitors.

PD-1/PD-L1 inhibitors are a group of immune checkpoint inhibitors as front-line treatment of multiple types of cancer. However, the serious immune-related adverse reactions limited the clinical application of PD-1/PD-L1 monoclonal antibodies, despite the promising curative effects. Therefore, it is urgent to develop novel inhibitors, such as small molecules, peptides or macrocycles, targeting the PD-1/PD-L1 axis to meet the increasing clinical demands. Our review discussed the mechanism of action of PD-1/PD-L1 inhibitors and presented clinical trials of currently approved PD-1/PD-L1 targeted drugs and the incidence of related adverse reactions, helping clinicians pay more attention to them, better formulate their intervention and resolution strategies. At last, some new inhibitors whose patent have been published are listed, which provide development ideas and judgment basis for the efficacy and safety of novel PD-1/PD-L1 inhibitors.

High incidence of cutaneous toxicity ranging from 29.2% to 71.2% has been reported during clinical use of vandetanib, which is a multi-target kinase inhibitor indicated for the treatment of unresectable medullary thyroid carcinoma. The cutaneous toxicity of vandetanib has limited its clinical benefits, but the underlying mechanisms and protective strategies are not well studied. Hence, we firstly established an in vivo model by continuously administrating vandetanib at 55 mg/kg/day to C57BL/6 for 21 days and verified that vandetanib could induce skin rash in vivo, which was consistent with the clinical study. We further cultured HaCaT and NHEK cells, the immortalized or primary human keratinocyte line, and investigated vandetanib (0–10 μM, 0–24 h)-caused alteration in cellular survival and death processes. The western blot showed that the expression level of apoptotic-related protein, c-PARP, c-Caspase 3 and Bax were increased, while the anti-apoptotic protein Bcl2 and MCL1 level were decreased. Meanwhile, vandetanib downregulated mitochondrial membrane potential which in turn caused the release of Cytochrome C, excessive production of reactive oxygen species and DNA damage. Furthermore, we found that 5 μM bisdemethoxycurcumin partially rescued vandetanib-induced mitochondria pathway-dependent keratinocyte apoptosis via activation of autophagy in vivo and in vitro, thereby ameliorated cutaneous toxicity. Conclusively, our study revealed the mechanisms of vandetanib-induced apoptosis in keratinocytes during the occurrence of cutaneous toxicity, and suggested bisdemethoxycurcumin as a potential protective drug. This work provided a potentially promising therapeutic strategy for the treatment of vandetanib-induced cutaneous toxicity.

Fibroblast growth factor receptor 4 (FGFR4) together with co-receptors modulate the activation of downstream proteins that regulate fundamental processes, and elevated FGFR4 activity is associated with Hepatocellular Carcinoma (HCC). Hence, FGFR4 is a promising therapeutic target for HCC. Based on BLU9931, we designed and synthesized a series of phenylquinazoline derivatives as novel inhibitors of FGFR4 through the covalent reversible strategy. Among them, a novel compound (C3) showed FGFR4 and cell proliferation inhibitory activity. Cellular mechanism studies demonstrated that compound C3 induced apoptosis via the FGFR4 signaling pathway blockage. Further mechanism study showed that C3 has the reversible covalent binding capacity, could be used as a reference for the development of novel FGFR4 covalent reversible inhibitors.

Renal fibrosis is a failed wound-healing process of the kidney tissue after chronic, sustained injury, which is a common pathway and pathological marker of virtually every type of chronic kidney disease (CKD), regardless of cause. However, there is a lack of effective treatment specifically targeting against renal fibrosis per se to date. The main pathological feature of renal fibrosis is the massive activation and proliferation of renal fibroblasts and the excessive synthesis and secretion of extracellular matrix (ECM) deposited in the renal interstitium, leading to structural damage, impairment of renal function, and eventually end-stage renal disease. In this review, we summarize recent advancements regarding the participation and interaction of many types of kidney residents and infiltrated cells during renal fibrosis, attempt to comprehensively discuss the mechanism of renal fibrosis from the cellular level and conclude by highlighting novel therapeutic targets and approaches for development of new treatments for patients with renal fibrosis.

The outbreak of COVID-19 caused by SARS-CoV-2 infection has become a serious hazard to global health. Apart from attacking respiratory system, it can induce multiorgan dysfunction, including cardiovascular system, liver, kidney, gastrointestinal, nervous system, and immune system. However, there are few reviews focusing on summary and comparison of diagnostic methods and complications induced by SARS-CoV-2 infection, which places a significant limit on the effective management.This review is a blend of evidence obtained by literature retrieval from PubMed, clinical experience, and the authors' opinions. We searched PubMed using the terms 'COVID-19diagnosis' and 'COVID-19complications' and selected the most relevant articles. Here we summarize the diagnostic methods that are available in clinic and discuss their different characters. Furthermore, the review offers an insight into the symptoms, incidence, and clinical strategies of complications associated with SARS-CoV-2 infection.COVID-19 has been a global pandemic, which requires rapid response. The comparison between different characters of the diagnostic methods and the summary of the symptoms, incidence, and clinical strategies of complications given in this review are not only significant for the optimal use of diagnostic methods, but also beneficial for the prevention and management of complications.

Numerous protein kinases encoded in the genome have become attractive targets for the treatment of different types of cancer. As of January 2020, a total of 52 small-molecule kinase inhibitors (SMKIs) have been approved by the FDA. With the numerous clinical trials and a heavy focus on drug safety, SMKI-induced cardiotoxicity, which is a life-threatening risk, has greatly attracted the attention of researchers. In this review, the SMKIs with cardiotoxicity incidence were described exhaustively. The data were collected from 42 clinical trials, 25 FDA-published documents, seven meta-analysis/systematic reviews, three case reports and more than 50 other types of articles. To date, 73% (38 of 52) of SMKIs have reported treatment-related cardiotoxicity. Among the 38 SMKIs with known cardiotoxicity, the rates of incidence of cardiac adverse events were QT prolongation: 47% (18 of 38), hypertension: 40% (15 of 38), left ventricular dysfunction: 34% (13 of 38), arrhythmia: 34% (13 of 38), heart failure: 26% (10 of 38) and ischemia or myocardial infarction: 29% (11 of 38). In the development process of novel SMKIs, more attention should be paid to balancing the treatment efficacy and the risk of cardiotoxicity. In preclinical drug studies, producing an accurate and reliable cardiotoxicity evaluation model is of key importance. To avoid the clinical potential cardiotoxicity risk and discontinuation of a highly effective drug, patients treated with SMKIs should be proactively monitored on the basis of a global standard. Moreover, the underlying mechanisms of SMKI-induced cardiotoxicity need to be further studied to develop new therapies for SMKI-induced cardiotoxicity.

Dasatinib shows remarkable activity against imatinib-refractory chronic myelogenous leukemia (CML) and Philadelphia chromosome positive acute lymphoblastic leukemia (Ph+ALL). However, severe cardiovascular toxicity limits the clinical applications of dasatinib. Since the underlying mechanism of dasatinib-induced cardiotoxicity is still elusive, we aim to clarify this. Recent studies have shown that necroptosis and apoptosis participate in multiple toxicity development. Here, we first report that dasatinib could directly induce cardiomyocytes death, as analyzed by the Sulforhodamine B (SRB) assay. This type of cardiomyocytes death was mediated by the necrosis pathway rather than apoptosis, as determined by using flow cytometry to characterize the mode of dasatinib-induced cell death. Inhibition of receptor-interacting protein kinase 1 (RIP1)activity and knockdown of receptor-interacting protein kinase 3 (RIP3)expression can block dasatinib-evoked cardiotoxicity, which further confirmed the involvement of necroptosis. We next found that the classic substrates of RIP3, mixed lineage kinase domain-like protein (MLKL) and Ca2+-calmodulin-dependent protein kinase II (CaMKII) were not involved in dasatinib-induced cardiomyocytes necroptosis. What's more, unlike the inflammation-associated necroptosis, dasatinib-triggered necroptosis was dependent on intracellular instead of secreted High-mobility group box 1 (HMGB1) protein. Collectively, our study revealed that dasatinib-induced cardiotoxicity acted via leading cardiomyocytes to HMGB1-mediated necroptosis, indicating a viable strategy for prevention of dasatinib-induced cardiotoxicity.

Adriamycin (ADR) is one of the most widely used and effective anthracycline antitumor agents in the treatment of certain hematological malignancies and solid tumors. However, the severe cardiotoxicity of ADR limits its clinical application. So far, the mechanism of the cardiotoxicity of ADR has not been completely clarified. In our research, cardiomyocyte autophagy and cardiac damage were observed in accompany ADR treatment, and autophagy appeared earlier than cardiac damage. Inhibition of autophagy by silencing ATG7 improved the survival rate of cardiomyocytes treated with ADR. The release of HMGB1 increased after ADR treatment, and silencing HMGB1 could reverse cardiomyocyte damage by attenuating autophagy. In addition, the expression of YAP was decreased, and overexpressing YAP down-regulated HMGB1 and alleviated cardiomyocyte damage. These results indicated that autophagy was the leading cause of ADR-induced cardiotoxicity, and HMGB1 played a vital role in the process of up-regulating autophagy.

Given that Yes-associated protein (YAP) signaling acts as a critical survival input for hypoxic cancer cells in hepatocellular carcinoma (HCC), disruption of YAP function and the maintenance of hypoxia is an attractive way to treat HCC. Utilizing a cell-based YAP-TEAD luciferase reporter assay and functional analyses, we identified CT-707, a China-FDA approved multi-kinase inhibitor under clinical trial with remarkable inhibitory activity against YAP function. CT-707 exhibited prominent cytotoxicity under hypoxia on HCC cells, which was attributable to the inhibition of YAP signaling. CT-707 arrested tumor growth in HepG2, Bel-7402, and HCC patient-derived xenografts. Mechanistically, the inhibitory activity of CT-707 on YAP signaling was due to the interruption of hypoxia-activated IGF1R. Overall, these findings not only identify CT-707 as a promising hypoxia-targeting agent against HCC, but they also unveil IGF1R as a new modulator specifically regulating hypoxia-activated YAP signaling. Significance: CT-707 may represent a novel clinical approach for patients with HCC suffering poor drug response due to intratumor hypoxia. Cancer Res; 78(14); 3995–4006. ©2018 AACR.

Background: Hepatocellular carcinoma (HCC) is one of the most common types of malignant tumour and has poor prognosis. Currently, systematic chemotherapy is the only approach to prolong survival. Thus the development of new treatment regimens is urgently needed to improve the therapeutic efficacy. Our study intended to assess the combination of dasatinib and irinotecan against HCC and made an effort to develop a potential medical choice for advanced HCC patients. Methods: We used SRB colorimetric assay and clonogenic assay to assess antitumour effect in vitro and HCC xenograft model to assess antitumour effect in vivo. We applied flow cytometry and western blotting to explore the mechanism of the combined therapy. Knockdown and overexpression of PLK1 are also applied for validation. Results: We confirmed that dasatinib has synergistic effect with irinotecan (or SN38) on HCC both in vitro and in vivo. The effect is due to arisen apoptosis rate of HCC cells that is accompanied by mitochondria dysfunction. The enhanced antitumour efficacy of SN38 could be explained by additional inhibition of PLK1, which is triggered by dasatinib. Unlike existed PLK1 inhibitors, dasatinib does not inhibit PLK1 activity in a direct way. Instead, we found that dasatinib reduces PLK1 level by interfering with its protein synthesis progress. We validated that this kind of downregulation of PLK1 level has a key role in the synergistic effect of the two agents. Conclusions: Dasatinib is able to reinforce the anti-HCC efficacy of irinotecan/SN38 by downregulation of PLK1 synthesis. The combination of the two agents might be a potential medical choice for HCC therapy.

Capecitabine, as the first-line treatment for multiple tumor types, has a serious drawback of hand-foot-syndrome (HFS) that limits its clinical use. However, the pathophysiology and mechanism of capecitabine-induced HFS is rarely known. Here we built the experimental mouse model of HFS induced by capecitabine at first and it was shown that 3 of 6 mice appeared HFS in the 5th day and 5 mice occurred HFS in the 30th day. The corneous layer was reduced in capecitabine-induced HFS in vivo. Moreover, we found that capecitabine could significantly induce keratinocytes cells death in vitro through activated apoptosis pathway and decreased mitochondrial membrane potential. In conclusion, these results suggested that HFS of capecitabine may be developed from reduction of corneous layer through stimulation of intracellular mitochondrial dysfunction following activation of caspase-dependent apoptosis pathway.

Hand–foot skin reaction (HFSR), among the most significant adverse effects of sorafenib, has been limiting the clinical benefits of this frontline drug in treating various malignant tumors. The mechanism underlying such toxicity remains poorly understood, hence the absence of effective intervention strategies. In the present study, we show that vascular endothelial cells are the primary cellular target of sorafenib-induced HFSR wherein soluble heparin-binding epidermal growth factor (s-HBEGF) mediates the crosstalk between vascular endothelial cells and keratinocytes. Mechanistically, s-HBEGF released from vascular endothelial cells activates the epidermal growth factor receptor (EGFR) on keratinocytes and promotes the phosphorylation of c-Jun N-terminal kinase 2 (JNK2), which stabilizes sirtuin 1 (SIRT1), an essential keratinization inducer, and ultimately gives rise to HFSR. The administration of s-HBEGF in vivo could sufficiently induce hyper-keratinization without sorafenib treatment. Furthermore, we report that HBEGF neutralization antibody, Sirt1 knockdown, and a classic SIRT1 inhibitor nicotinamide could all significantly reduce the sorafenib-induced HFSR in the mouse model. It is noteworthy that nicotinic acid, a prodrug of nicotinamide, could substantially reverse the sorafenib-induced HFSR in ten patients in a preliminary clinical study. Collectively, our findings reveal the mechanism of vascular endothelial cell-promoted keratinization in keratinocytes and provide a potentially promising therapeutic strategy for the treatment of sorafenib-induced HFSR.

Sunitinib, a multikinase inhibitor approved for a number of cancer indications has a low response rate. Identifying mechanisms of resistance could lead to rational combination regimens that could improve clinical outcomes. Here we report that resistance to sunitinib therapy was driven by autophagic degradation of TP53/p53. Deletion of ATG7 or ATG5 suppressed TP53 degradation, as did knockdown of SQSTM1/p62. Mechanistically, the transport of TP53 from the nucleus to the cytoplasm was essential for the sunitinib-induced autophagic degradation of TP53 and did not require TP53 nuclear export signals (NESs). Moreover, TP53 degradation was achieved by the transport of its nuclear binding target, HMGB1, which shifted TP53 from the nucleus to the cytoplasm. The inhibition of HMGB1 sensitized cancer cells to sunitinib. Importantly, sunitinib induced the degradation of all TP53 proteins, except for TP53 proteins with mutations in the interaction domain of TP53 with HMGB1 (amino acids 313 to 352). In conclusion, our data identify an alternative HMGB1-mediated TP53 protein turnover mechanism that participates in the resistance of sunitinib and suggest HMGB1 as a potential therapeutic target for improving clinical outcomes of sunitinib.

Hypoxia is associated with resistance to chemotherapy in a number of human cancer types; particularly in hepatocellular carcinoma (HCC), which is a highly vascularized tumor. To develop a potential combination therapy strategy that is capable of overcoming the hypoxia‑induced insensitivity to chemotherapy, the HCC cell SMMC‑7721 was employed to investigate the hypoxia‑induced chemoresistance to etoposide. Increased levels of hypoxia‑inducible factor‑1α (HIF‑1α) and vascular endothelial growth factor (VEGF) were observed when SMMC‑7721 cells were exposed to hypoxia, and exposure of tumor cells to hypoxia impaired etoposide‑induced DNA damage, as indicated by the failure of upregulation of γHA2X. Etoposide‑induced apoptosis and cell cycle arrest of SMMC‑7721 was also impaired in hypoxia. However, co‑treatment with anti‑VEGF significantly restored etoposide‑induced cell apoptosis and cell cycle arrest, as indicated by the elimination of B‑cell lymphoma 2 (Bcl‑2), procaspase 3, cyclin B1 and Cdc2. Furthermore, anti‑VEGF eliminated phosphorylation of AKT, ERK and IκB‑α resulting from hypoxia, suggesting the involvement of VEGF in the activation of the survival pathways. In conclusion, the present study suggests a significant role of VEGF in the chemoresistance of etoposide in hypoxia. A rational chemotherapy should be developed based on a combination of etoposide and anti‑VEGF.

A series of novel indene-derived retinoic acid receptor α (RARα) agonists have been designed and synthesized. The use of receptor binding, cell proliferation and cell differentiation assays demonstrated that most of these compounds exhibited moderate RARα binding activity and potent antiproliferative activity. In particular, 4-((3-isopropoxy-2,3-dihydro-1H-inden-5-yl)-carbamoyl)benzoic acid (36d), which showed a moderate binding affinity, exhibited a great potential to induce the differentiation of NB4 cells (68.88% at 5 μM). Importantly, our work established indene as a promising skeleton for the development of novel RARα agonists.

Hepatocellular carcinoma is among the leading causes of cancer-related deaths worldwide, and the development of new treatment regimens is urgently needed to improve therapeutic approach. In our study, we found that the combination of a Met inhibitor, cabozantinib, and a novel FAK inhibitor, CT-707, exerted synergistic antitumor effects against hepatocellular carcinoma in vitro and in vivo. Interestingly, further studies showed that therapeutic concentrations of cabozantinib increased the phosphorylation of FAK, which might attenuate the antitumor activity of cabozantinib. The simultaneous exposure to CT-707 effectively inhibited the activation of FAK that was induced by cabozantinib, which contributes to the synergistic effect of the combination. Furthermore, cabozantinib increased the mRNA and protein levels of integrin α5, which is a canonical upstream of FAK, and the introduction of cilengitide to block integrin function could abrogate FAK activation by cabozantinib, indicating that cabozantinib upregulated the phosphorylation of FAK in an integrin-dependent manner. Similar synergy was also observed on PHA-665752, another selective MET inhibitor, indicating that this observation might be a common characteristic of MET-targeting strategies. Our findings not only favor the development of the novel FAK inhibitor CT-707 as a therapeutic agent against hepatocellular carcinoma but also provide a new strategy of combining MET and FAK inhibitors to potentiate the anticancer activities of these two types of agents for treating hepatocellular carcinoma patients. Mol Cancer Ther; 15(12); 2916–25. ©2016 AACR.

Background Chemotherapy is the only therapy option for the majority of AML patients, however, there are several limitations for this treatment. Our aim was to find a new chemotherapy strategy that is more effective and less toxic. Methods MTT assays and a xenograft mouse model were employed to evaluate the synergistic activity of all-trans retinoic acid (ATRA) combined with topotecan (TPT). Drug-induced DNA damage and apoptosis were determined by flow cytometry analysis with PI and DAPI staining, the comet assay and Western blots. Short hairpin RNA (shRNA) and a RARα plasmid were used to determine whether RARα expression influenced DNA damage and apoptosis. Results We found that ATRA exhibited synergistic activity in combination with Topotecan in AML cells, and the enhanced apoptosis induced by Topotecan plus ATRA resulted from caspase pathway activation. Mechanistically, ATRA dramatically down regulated RARα protein levels and led to more DNA damage and ultimately resulted in the synergism of these two agents. In addition, the increased antitumor efficacy of Topotecan combined with ATRA was further validated in the HL60 xenograft mouse model. Conclusions Our data demonstrated, for the first time, that the combination of TPT and ATRA showed potential benefits in AML, providing a novel insight into clinical treatment strategies. Electronic supplementary material The online version of this article (doi:10.1186/s12885-015-2010-6) contains supplementary material, which is available to authorized users.

All-trans retinoic acid (ATRA) has been successfully used in differentiation therapy for acute promyelocytic leukemia (APL) in the clinic. ATRA-induced differentiation of leukemia cells is accompanied by a G0/G1 arrest, yet how ATRA couples cell cycle arrest to differentiation remains largely unknown. Here we observed that the ubiquitin-proteasome pathway (UPP) was activated upon ATRA treatment in the human acute myeloid leukemia cell lines, NB4 and HL-60, as represented by the accumulation of ubiquitinated proteins, the up-regulation of ubiquitin mRNA and increased 20S proteasome activity. Interestingly, we found that complete inhibition of proteasome activity suppressed ATRA-induced proliferation/differentiation (P/D) transition in both cell lines. Furthermore, we demonstrate that the exact protein contributing to this phenomenon is different in these two cell lines. Cyclin-dependent kinase 2 (CDK2) and Cyclin E were degraded by the UPP; they accumulated significantly after complete inhibition of the proteasome in ATRA-treated NB4 and HL-60 cells, respectively. These findings suggested that the UPP might be indispensable in the ATRA-induced G0/G1 arrest and differentiation of leukemia cells. The exact protein degraded by the UPP to promote the myeloid maturation program set in motion by the retinoid may be cell type dependent.

Hypoxia is one of the inevitable circumstances of various tumors. It controls various levels of regulation in tumor progression and results in tumor resistance to radiotherapy and chemotherapy. Here we investigated a synthetic TPZ derivative, N-ethoxymethyl-3-amino-1,2,4-benzotriazine-1,4-dioxide (XQ2), a novel compound that induced anti-cancer effects both in normoxia and in hypoxia, cell proliferation assay found that XQ2 exhibited a potent inhibitory effect on the tested cancer cell lines both in normoxia and in hypoxia. Flow cytometry and western blot studies indicated that XQ2 induces G2/M arrest and a caspase-dependent apoptosis in A549 cells. Additionally, intracellular reactive oxygen species (ROS) appear to play a key role in the anticancer effect of XQ2 in hypoxia. Taken together, our data suggest that XQ2 exerted anticancer action by suppressing the ROS level and triggering cell-cycle arrest and the caspase-dependent pathway, which is associated with apoptosis.

P53 is under exquisitely fine regulation and acts as a transcription factor that regulates the expression of thousands of genes that control apoptosis, necroptosis, ferrotosis, cell cycle arrest, senescence, metabolism and fertility. Regulation of p53 turnover is essential not only for the response to chemotherapeutic drugs or radiation in cancer cell but also to keep p53 activity under control in a normal cell. Moreover, .p53 is found to be overactivated in various diseases including idiopathic pulmonary fibrosis (IPF), alcoholic liver disease (ALD) and non-alcoholic fatty liver disease (NAFLD). Promoting hyperactivated-p53 degradation can combat these diseases. However, the strategies that based on known proteasome-dependent degradation only can stabilize the expression level of p53 such as nutlins (inhibit the interaction between mdm2 and tumor suppressor p53). Autophagy-lysosomal degradation is the other pathway for controlling cellular protein stability. More and more researches have established that numerous proteins can be degraded through both proteasome and autophagy-lysosome pathways. However, very little is known to date about p53 turnover through autophagy-lysosome pathway. Here we discover that the activation of p53 autophagic degradation during an oral small molecule receptor tyrosine kinase inhibitor sunitinib in p53 wt cancer cells and normal cells. Mechanistically, the nucleus-to-cytoplasm shift is essential for the autophagic degradation of p53 induced by sunitinib, however, does not require p53 nuclear export signals. Indeed, p53 degradation is achieved by the nucleus-to-cytoplasm transport of its nuclear binding target HMGB1, shifting the distribution of p53 from the nucleus to the cytoplasm. Then cytoplasmic p53 directly interacts with the autophagy cargo receptor p62 to promote degradation. Importantly, inhibition of HMGB1 sensitizes cancer cell to sunitinib. Taken together, our study identified an alternative p53 protein turnover mechanism induced by sunitinib, thus not only elucidating the underlying mechanisms that limit sunitinib efficacy in cancer therapy but also opening an avenue for expanding the clinical indications of sunitinib. Note: This abstract was not presented at the meeting. Citation Format: Qiaojun He, Peihua Luo, Xiaochun Yang, Qinjie Weng, Ling Ding, Bo Yang. HMGB1 governs p53 to autophagic degradation via its nucleus-to-cytoplasm transport [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 2070. doi:10.1158/1538-7445.AM2017-2070

// Xiaochun Yang 1 , Jincheng Wang 1 , Jiabin Dai 1 , Jinjin Shao 1 , Jian Ma 2 , Chao Chen 2 , Shenglin Ma 3 , Qiaojun He 1 , Peihua Luo 1 and Bo Yang 1 1 Institute of Pharmacology & Toxicology, College of Pharmaceutical Sciences, Zhejiang University, Hangzhou, China 2 Center for Drug Safety Evaluation and Research of Zhejiang University, Hangzhou, China 3 Nanjing Medical University, Affiliated Hangzhou Hospital, Hangzhou First People’s Hospital, Hangzhou, China Correspondence: Peihua Luo, email: // Bo Yang, email: // Keywords : Autophagy, p38 signaling, Hepatotoxicity, Dasatinib Received : October 14, 2014 Accepted : January 14, 2015 Published : January 31, 2015 Abstract Liver dysfunction is a common side effect associated with the treatment of dasatinib and its mechanism is poorly understood. Autophagy has been thought to be a potent survival or death factor for liver dysfunction, which may shed the light on a novel strategy for the intervention of hepatotoxicity caused by dasatinib. In this study, we show for the first time that autophagy is induced, which is consistent with the formation of liver damage. Autophagy inhibition exacerbated dasatinib-induced liver failure, suggesting that autophagy acted as a self-defense mechanism to promote survival. Oxidative stress has been shown to be an important stimulus for autophagy and hepatotoxicity. Interestingly, dasatinib increased the activity of p38, which is a critical modulator of the oxidative stress related to liver injury and autophagy. p38 silencing significantly blocked LC3-II induction and p62 reduction by dasatinib, which was accompanied by increased caspase-3 and PARP cleavage, indicating that autophagy alleviated dasatinib-induced hepatotoxicity via p38 signaling. Finally, the p38 agonist isoproterenol hydrochloride (ISO) alleviated dasatinib-induced liver failure by enhancing autophagy without affecting the anticancer activity of dasatinib. Thus, this study revealed that p38-activated autophagy promoted survival during liver injury, which may provide novel approaches for managing the clinical applications of dasatinib.

Adriamycin, a widely used anthracycline antibiotic in multiple chemotherapy regimens, has been challenged by the cardiotoxicity leading to fatal congestive heart failure in the worst condition. The present study demonstrated that Dihydromyricetin, a natural product extracted from ampelopsis grossedentat, exerted cardioprotective effect against the injury in Adriamycin-administrated ICR mice. Dihydromyricetin decreased ALT, LDH and CKMB levels in mice serum, causing a significant reduction in the toxic death triggered by Adriamycin. The protective effects were also indicated by the alleviation of abnormal electrocardiographic changes, the abrogation of proliferation arrest and apoptotic cell death in primary myocardial cells. Further study revealed that Dihydromyricetin-rescued loss of anti-apoptosis protein ARC provoked by Adriamycin was involved in the cardioprotection. Intriguingly, the anticancer activity of Adriamycin was not compromised upon the combination with Dihydromyricetin, as demonstrated by the enhanced anticancer effect achieved by Adriamycin plus Dihydromyricetin in human leukemia U937 cells and xenograft models, in a p53-dependent manner. These results collectively promised the potential value of Dihydromyricetin as a rational cardioprotective agent of Adriamycin, by protecting myocardial cells from apoptosis, while potentiating anticancer activities of Adriamycin, thus further increasing the therapeutic window of the latter one.

Dasatinib (BMS-354825) is a FDA-approved multitargeted kinase inhibitor of BCR/ABL and Src kinases. It is now used in the treatment of chronic myelogenous leukemia (CML) with resistance or intolerance to prior therapies, including imatinib. Here we report a novel effect of dasatinib on inducing the differentiation of acute myeloid leukemia (AML) cells through MEK/ERK-dependent activation of signal transducer and activator of transcription 1 (STAT1). We found that dasatinib could induce the differentiation of AML cells as demonstrated by the expression of differentiation marker CD11b, G0/G1 phase arrest and decreased ratio of nucleus to cytoplasm. Of note, dasatinib induced robust phosphorylation of STAT1 both at Tyr701 and Ser727 as well as the redistribution of STAT1 from the cytoplasm to the nucleus, thus leading to the transcription of STAT1-targeted genes. Knocking down STAT1 expression by shRNA significantly attenuated dasatinib-induced differentiation, indicating an important role of STAT1 in myeloid maturation. We further found that dasatinib-induced activation of STAT1 was regulated by the MEK/ERK kinases. The phosporylation of MEK and ERK occurred rapidly upon dasatinib treatment and increased progressively as differentiation was induced. MEK inhibitors PD98059 and U0216 not only inhibited the phosphorylation of STAT1, but also abrogated dasatinib-induced myeloid differentiation, suggesting that MEK/ERK dependent phosphorylation of STAT1 might be indispensable for the differentiating effect of dasatinib in AML cells. Taken together, our study suggests that STAT1 is an important mediator in dasatinib-induced differentiation of AML cells, whose activation requires the activation of MEK/ERK cascades.

All-trans-retinoic acid (ATRA) has held great promise for differentiation-based therapy but reportedly downregulates retinoic acid receptor-α (RARα) in a proteasome-dependent manner, which leads to decreased acute myeloid leukemia (AML) cell differentiation efficiency. Therefore, research strategies that seek to further sensitize cells to retinoids and extend the range of retinoid-affected myeloid malignancies beyond acute promyelocytic leukemia (APL) are key investigative avenues. Here, we show that bortezomib, the first proteasome inhibitor approved for newly diagnosed and relapsed multiple myeloma, exhibited strong synergism with ATRA to promote HL60 and NB4 AML cell differentiation. We observed that bortezomib sensitized AML cells to ATRA-induced morphologic, biochemical, and functional changes, indicative of myeloid differentiation without cell death. In addition, treatment of human leukemia HL60 xenografts with bortezomib and ATRA together did not increase bortezomib-induced progressive weight loss but resulted in significant tumor growth inhibition in addition to increased differentiation (P < 0.05). These enhanced differentiation effects were accompanied by RARα stabilization and STAT1 activation. Taken together, our study was the first to evaluate bortezomib and ATRA synergy in AML cell differentiation and to assess new opportunities for bortezomib and ATRA combination as a promising approach for future differentiation therapy. Mol Cancer Ther; 12(2); 195–206. ©2012 AACR.