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1.
FEBS Open Bio ; 11(3): 911-920, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33455075

RESUMO

Multiple clinical trials have shown that monoclonal antibodies (mAbs) against programmed death-ligand 1 (PD-1/PD-L1) can benefit patients with lung cancer by increasing their progression-free survival and overall survival. However, a significant proportion of patients do not respond to anti-PD-1/PD-L1 mAbs. In the present study, we investigated whether galectin (Gal)-3 inhibitors can enhance the antitumor effect of PD-L1 blockade. Using the NSCLC-derived cell line A549, we examined the expression of Gal-3 in lung cancer cells under hypoxic conditions and investigated the regulatory effect of Gal-3 on PD-L1 expression, which is mediated by the STAT3 pathway. We also explored whether Gal-3 inhibition can facilitate the cytotoxic effect of T cells induced by PD-L1 blockade. The effects of combined use of a Gal-3 inhibitor and PD-L1 blockade on tumor growth and T-cell function were also investigated, and we found that hypoxia increased the expression and secretion of Gal-3 by lung cancer cells. Gal-3 increased PD-L1 expression via the upregulation of STAT3 phosphorylation, and administration of a Gal-3 inhibitor enhanced the effect of PD-L1 blockade on the cytotoxic activity of T cells against cancer cells in vitro. In a mouse xenograft model, the combination of a Gal-3 inhibitor and PD-L1 blockade synergistically suppressed tumor growth. Furthermore, the administration of a Gal-3 inhibitor enhanced T-cell infiltration and granzyme B release in tumors. Collectively, our results show that Gal-3 increases PD-L1 expression in lung cancer cells and that the administration of a Gal-3 inhibitor as an adjuvant enhanced the antitumor activity of PD-L1 blockade.

2.
Int J Mol Med ; 47(2): 708-718, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33416098

RESUMO

A large human natural single­chain fragment variable (scFv) phage library was constructed based on Cre­LoxP recombination, and used to successfully identify antibodies against proprotein convertase subtilisin/kexin type 9 (PCSK9). The library was derived from 400 blood samples, 30 bone marrow samples, and 10 cord blood samples from healthy donors. Lymphocytes were isolated from each sample and cDNA was synthesized using reverse transcription­quantitative PCR. Two­step overlap PCR was then used for scFv synthesis using a LoxP peptide as the linker. The scFv gene was inserted into the phagemid vector pDF by enzymatic digestion and ligation, and then transformed into Escherichia coli (E. coli) SS320 to establish a primary antibody library in the form of scFvs. A primary antibody library consisting of 5x107 peripheral blood and umbilical cord blood sources, as well as a primary antibody library of 5x107 bone marrow samples were obtained. By optimizing the recombination conditions, the primary phage library was used to infect E. coli BS1365 strain (which expresses the Cre enzyme), and a human scFv recombinant library with a size of 1x1011 was obtained through Cre­LoxP enzyme­mediated heavy and light chain replacement and recombination. This constructed recombinant library was employed to screen for antibodies against recombinant PCSK9. After four rounds of selection, a fully human antibody (3D2) was identified with a binding affinity of 1.96±1.56ⅹ10­10 M towards PCSK9. In vitro, the PCSK9/low­density lipoprotein receptor (LDLR) pathway of Hep­G2 cells was inhibited by 3D2 treatment, thereby increasing LDL uptake in these cells. In addition, combination treatment with 3D2 and statin was more effective at increasing LDLR levels than treatment with 3D2 or statin alone. Furthermore, 3D2 resulted in a 3­fold increase in hepatic LDLR levels, and lowered total serum cholesterol by up to 61.5% in vivo. Taken together, these results suggest that the constructed human Cre­LoxP scFv phage display library can be used to screen fully human scFv, and that 3D2 may serve as a candidate hypolipidemic therapy.

3.
Mol Med Rep ; 23(2): 1, 2021 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-33355363

RESUMO

The vital functions of long non­coding (lnc)RNAs have been verified in gastric carcinoma (GC). However, as a novel cancer­related lncRNA, the influence of leukemia inhibitory factor receptor antisense RNA 1 (LIFR­AS1) in GC cell biological behaviors remains unreported. The present study explored the biological effects of lncRNA LIFR­AS1 on GC progression. Reverse transcription­quantitative PCR was performed to examine lncRNA LIFR­AS1 expression in GC tissues and cells. Cell Counting Kit­8, 5­ethynyl­2'­deoxyuridine incorporation, cell wound healing and Transwell invasion assays were used to assess the functions of lncRNA LIFR­AS1 in GC cell proliferation, migration and invasion. Additionally, associations among lncRNA LIFR­AS1, microRNA (miR)­4698 and microtubule­associated tumor suppressor 1 (MTUS1) were investigated via bioinformatics software and a luciferase reporter system. In addition, western blotting was used to examine the expression of MEK and ERK. Decreased lncRNA LIFR­AS1 expression was observed in GC tissues and cells. Upregulated lncRNA LIFR­AS1 inhibited GC cell proliferation, migration and invasion. Upregulated miR­4698 and downregulated MTUS1 were identified in GC tissues and cells. The inhibitory interaction between lncRNA LIFR­AS1 and miR­4698 was confirmed. Additionally, MTUS1 was predicted as a target gene of miR­4698 positively regulated by lncRNA LIFR­AS1. The MEK/ERK pathway was inhibited by lncRNA LIFR­AS1 via regulating MTUS1. These findings revealed the inhibitory functions of lncRNA LIFR­AS1 in GC cell proliferation, migration and invasion. The process was mediated via miR­4698, MTUS1 and the MEK/ERK pathway.

4.
Biomed Res Int ; 2020: 7586521, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32904490

RESUMO

cir-ITCH, a well-known tumor-suppressive circular RNA, plays a critical role in different cancers. However, its expression and functional role in prostate cancer (PCa) are unclear. Herein, we explored the potential mechanism and tumor-inhibiting role of cir-ITCH in PCa. Using reverse transcriptase polymerase chain reaction assay, we analyzed the expression of cir-ITCH in PCa and paired adjacent nontumor tissue samples resected during surgical operation, as well as in two cell lines of human PCa (LNCaP and PC-3) and the immortalized normal prostate epithelial cell line (RWPE-1). Cell viability and migration of PCa cell lines were evaluated using CCK-8 and wound-healing assays. Expression of key proteins of the Wnt/ß-catenin and PI3K/AKT/mTOR pathways was detected using western blotting. We found that cir-ITCH expression was typically downregulated in the tissues and cell lines of PCa compared to that in the peritumoral tissue and in RWPE-1 cells, respectively. The results showed that cir-ITCH overexpression significantly inhibits the proliferation, migration, and invasion of human PCa cells and that reciprocal inhibition of expression occurred between cir-ITCH and miR-17. Proteins in the Wnt/ß-catenin and PI3K/AKT/mTOR pathways were downregulated by overexpression of cir-ITCH both in androgen receptor-positive LNCaP cells and androgen receptor-negative PC-3 cells. Taken together, these data demonstrated that cir-ITCH plays a tumor-suppressive role in human PCa cells, partly through the Wnt/ß-catenin and PI3K/AKT/mTOR pathways. Thus, cir-ITCH may serve as a novel therapeutic target for the treatment of PCa, especially castration-resistant prostate cancer.

5.
Onco Targets Ther ; 13: 7229-7241, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32801752

RESUMO

Tremendous progress has been achieved in the field of immune checkpoint inhibitors (ICIs) therapy in lung cancer in recent years. To generate robust, long-lasting anti-tumor immune responses in lung cancer patients, combinational ICI therapies have been explored deeply. Conventionally, chemotherapy was considered as immunosuppressive. It is now recognized that chemotherapy could also reinstate cancer cell immune-surveillance and enable the perception of cancer cells as dangerous. That is to say that chemotherapeutic drugs are not only a source of direct cytotoxic effects but also an adjuvant for anti-tumor immunity. Recently, multiple clinical studies of ICIs combined with chemotherapeutic drugs have been explored and proved effective. However, there are still crucial questions that are not well addressed, such as the optimal dose and schedule for a given combination may differ across disease indications, and the appropriate strategy of selecting patient population that can benefit from ICIs remains unclear. To facilitate more rational lung cancer ICIs therapy development, this review summarizes the immune-regulatory effects and related mechanisms of chemotherapeutic drugs and the clinical progress of ICIs and their combination with chemotherapies in lung cancer treatment.

6.
Onco Targets Ther ; 13: 3703-3716, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32440140

RESUMO

Purpose: Based on the continuous exploration of solid tumor immunotherapy, we focused on hepatocellular carcinoma with a high level of morbidity and mortality. We confirm the stability of mcDNA-based CAR T cell generating platform, and investigate the antitumor activity of CD44-CAR T cells against hepatocellular carcinoma both in vitro and in vivo. Materials and Methods: We fused anti-CD44 scFv structure with transmembrane domain and intracellular domain. Using a non-viral mcDNA vector to load CD44-CAR gene, then transfected the mcDNA-CD44-CAR into human T cells by electroporation. We exhibited the transfection efficacy of CAR T cells and the CD44 expression of tumor cell lines by flow cytometry. The antitumor efficacy of CD44-CAR T cells in vitro and in vivo was detected through CCK-8 and ELISA assays, and xenograft mouse models, respectively. Results: We obtained mcDNA-CD44-CAR with a high level of density after repeated extraction and purification. The expression efficacy of CD44-CAR in T cells was more than 50% after seven days electroporation and the phenotype of CD44-CAR T cells was no difference compared with normal T cells. For CD44-positive hepatocellular carcinoma xenograft mice, CD44-CAR T cells had stronger tumor growth suppression compared to normal T and mock T cells. The same results occurred on the in vitro experiments including cytokine secretion and cytotoxicity assays. H&E staining graphs revealed that CD44-CAR T cells did not induce side effects in xenograft mice. Conclusion: The strategy for generating CAR T cells targeting cancer stem cell antigens was efficient and concise. The mcDNA had superior transgene ability without virus-related adverse effects. CD44-CAR T cells had strong suppression capacity against hepatocellular carcinoma.

7.
Onco Targets Ther ; 13: 559-571, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32021298

RESUMO

Purpose: This study aimed to investigate the regulatory effects and mechanisms of long non-coding RNA (LncRNA) metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) on gastric cancer (GC) cells. Methods: The expression of MALAT1 was detected in GC tissues and two GC cell lines (SGC-7901 and BGC-823). MALAT1 was overexpressed and silenced in GC cells by the transfection of pcDNA-MALAT1 and siRNA-MALAT1, respectively. The proliferation and apoptosis of transfected cells, as well as the tumor volume and weight in mice injected with transfected cells were determined. After identifying the interaction between microRNA-22-3p (miR-22-3p) and MALAT1/epidermal growth factor receptor 3 (ErbB3), the effects of miR-22-3p/ErbB3 silencing on the proliferation and apoptosis of GC cells were evaluated. Results: MALAT1 was significantly upregulated in GC tissues and cells and negatively associated with the survival of GC patients. Overexpression of MALAT1 significantly promoted the proliferation and inhibited the apoptosis of SGC-7901 cells, while silencing of MALAT1 exerts contrary effects on BGC-823 cells. Silencing of MALAT1 also significantly inhibited the tumor growth in mice. In addition, MALAT1 negatively regulated its target miR-22-3p. Silencing of miR-22-3p reversed the anti-tumor effects of MALAT1 silencing on GC cells. MiR-22-3p negatively regulated its target ErbB3. Silencing of ErbB3 reversed the tumor-promoting effects of miR-22-3p silencing on GC cells. Conclusion: Silencing of MALAT1 inhibited the proliferation and promoted the apoptosis of GC cells through upregulating miR-22-3p and downregulating ErbB3.

8.
SLAS Discov ; 25(3): 310-319, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31560248

RESUMO

D-dimer is an essential diagnostic index of thrombotic diseases. Since the existing anti-D-dimer antibodies vary in quality and specificity, a search for alternative anti-D-dimer antibodies is required. The present study aimed to screen a novel monoclonal antibody (mAb) against D-dimer using a light-initiated chemiluminescence assay (LiCA). In this work, mice were immunized with antigen prepared from human plasma by enzyme hydrolysis. After screening, a novel mAb, DD 2G11, was obtained. The results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis indicated that DD 2G11 could be used as a standard marker for D-dimer. The isotype of DD 2G11 was IgG1, the Ka value was 0.646 nM-1, and the Kd value was 50 nM, indicating that the binding affinity to D-dimer was very high. Furthermore, no cross-reactivity between DD 2G11 and other fibrinogen degradation products (FgDPs) was found. Finally, the correlation between DD 2G11 and the reference antibody (commercial antibody) was investigated by analyzing 56 clinical samples using a latex-enhanced turbidimetric immunoassay (LTIA). The R2 value of the linear regression was 0.94538, indicating that DD 2G11 met clinical requirements. In conclusion, the present study provides a more expeditious protocol to screen mAbs and provides a clinically usable mAb against D-dimer.

9.
Mol Cancer Ther ; 19(1): 178-186, 2020 01.
Artigo em Inglês | MEDLINE | ID: mdl-31582530

RESUMO

Viral-based chimeric antigen receptor-engineered T (CAR T)-cell manufacturing has potential safety risks and relatively high costs. The nonviral minicircle DNA (mcDNA) is safer for patients, cheaper to produce, and may be a more suitable technique to generate CAR T cells. In this study, we produced mcDNA-based CAR T cells specifically targeting prostate stem cell antigen (PSCA; mcDNA-PSCA-CAR T cells). Our results showed that mcDNA-PSCA-CAR T cells persisted in mouse peripheral blood as long as 28 days and demonstrated more CAR T-cell infiltration, higher cytokine secretion levels, and better antitumor effects. Together, our results suggest that mcDNA-CAR can be a safe and cost-effective platform to produce CAR T cells.

10.
Am J Transl Res ; 11(9): 5740-5751, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31632544

RESUMO

The down-regulation of long non-coding RNA (lncRNA) MEG3 has been observed in various cancers; nonetheless, underlying mechanisms are still unclear. The current research work aims at exploring the roles of MEG3 in the pathogenesis of CRC and the associated mechanism. We observed that MEG3 was significantly down-regulated in both CRC tumor tissue and cell lines; also, the transient over-expression of MEG3 in CRC cell line SW480 and LoVo inhibited the proliferation and the migration and clone formation capability of cells; on the other hand, the knockdown of MEG3 has revealed opposite effects. Eventually, we figured it out that target miR-376 directly targeted both MEG3 and PRDK1 in SW480 and LoVo cells. To conclude, as our findings proved, MEG3 is likely to act as a tumor suppressor in the pathogenesis of CRC by means of the regulation of the miR-376/PRDK1 signal axis, suggesting that MEG3 has the potential to become a novel therapeutic target for the treatment of CRC.

11.
Am J Cancer Res ; 9(5): 945-958, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31218103

RESUMO

Colorectal cancer is one of the most common malignancies worldwide, as it is often diagnosed at an advanced stage. Chimeric antigen receptor (CAR) T cell therapy has demonstrated remarkable success and emerged as one of the most promising therapeutic strategies in multiple malignancies. The purpose of this study was to investigate the anti-tumor activity of NKG2D CAR-T cells against human colorectal cancer cells. A non-viral third-generation NKG2D CAR was constructed, and subsequently transduced into T cells to obtain the NKG2D CAR-T cells. In vitro, NKG2D CAR-T cells showed cytotoxicity against human colorectal cancer cells in a dose-dependent manner compared with untransduced T cells. In addition, IL-2 and IFN-γ secreted by these cells were significantly higher than those by untransduced T cells. In vivo, NKG2D CAR-T cells significantly suppressed tumor growth, reduced tumor sizes and extended overall survival of mice in a xenograft model of HCT-116 cells. Furthermore, human NKG2D-positive lymphocytes infiltration could be found in the tumor sections of NKG2D CAR-T cells-treated mice. There were no severe pathological changes found in vital organs in any of the treatment groups. NKG2D CAR-T cells showed excellent killing effect and represented a promising immunotherapeutic strategy against human colorectal cancer.

12.
FEBS Lett ; 593(18): 2566-2573, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31254364

RESUMO

Aquaporin 1 (AQP1) plays an important role in endothelial functions and is regulated by MEF2C. However, how AQP1 level is stabilized to maintain endothelial water homeostasis is still not clear. Here, we show that AQP1 expression is significantly upregulated by MEF2C transcriptionally and inhibited by miR-133a-3p.1 post-transcriptionally. Meanwhile, MEF2C activates the expression of miR-133a1. Simultaneous overexpression of MEF2C and miR-133a-3p.1 suppresses the aptitude of changes in AQP1 expression caused by either MEF2C or miR-133a-3p.1. Accordingly, the changes in migration and tube formation of human umbilical vein endothelial cells (HUVECs) caused by MEF2C or miR133a-3p.1 are blunted by coexpression of both of them. These data demonstrate that the homeostasis and physiological function of AQP1 in endothelial health are maintained by the MEF2C and miR-133a-3p.1 regulatory circuit.


Assuntos
Aquaporina 1/genética , Regulação da Expressão Gênica/genética , Homeostase/genética , Células Endoteliais da Veia Umbilical Humana/metabolismo , MicroRNAs/genética , Água/metabolismo , Sequência de Bases , Movimento Celular/genética , Células Endoteliais da Veia Umbilical Humana/citologia , Humanos , Fatores de Transcrição MEF2/metabolismo , Transcrição Genética
13.
FEBS Open Bio ; 9(7): 1305-1314, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31141316

RESUMO

Aberrant expression of microRNAs (miRNAs) may contribute to the initiation and development of multiple types of human cancer. Several miRNAs have been found to be strongly correlated with the diagnosis, progression, and prognosis of colorectal carcinoma (CRC), but the role of miR-125a in CRC remains unclear. In the present study, the function of miR-125a on the expression of Smad ubiquitin regulatory factor 1 (Smurf1) was investigated in vitro and in vivo. We verified that Smurf1 is a downstream target gene of miR-125a and is involved in miR-125a-mediated regulation of CT26 cell (colon cancer cell) proliferation and migration. Overexpression of miR-125a suppresses CT26 cell growth by inhibiting cell proliferation. Additionally, wound healing assays were performed to show that overexpression of miR-125a significantly reduced CT26 cell migration, which was reversed by overexpression of Smurf1. In vivo, miR-125a overexpression downregulated the expression of Ki67 and Smurf1, thus leading to a marked reduction in tumor growth. These results revealed that miR-125a plays a critical role in CRC by directly targeting Smurf1, a finding that may facilitate the development of improved diagnostic and therapeutic techniques for CRC.


Assuntos
Neoplasias Colorretais/genética , MicroRNAs/genética , Ubiquitina-Proteína Ligases/metabolismo , Regiões 3' não Traduzidas/genética , Animais , Carcinogênese/genética , Carcinogênese/metabolismo , Ciclo Celular , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Camundongos , Camundongos Nus , MicroRNAs/metabolismo , Invasividade Neoplásica/genética , Transdução de Sinais , Ubiquitina-Proteína Ligases/genética , Ensaios Antitumorais Modelo de Xenoenxerto/métodos
14.
Immunotherapy ; 11(7): 599-616, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30943862

RESUMO

AIM: To produce dendritic cells (DCs) from CD34+ stem cells from cord blood and explore their prophylactic and curative effect against tumors by vaccinating humanized NSG mice. MATERIALS & METHODS: Separated CD34+ stem cells from cord blood were cultured for 30 days, and the resultant DCs (CD34-DCs) were collected. The basic function of the CD34-DCs and the cytotoxicity of CD34-cytotoxic-T lymphocytes (CTLs) were tested in vitro, and tumor inhibition in a humanized NSG mouse tumor model was observed. RESULTS: The number of CD34-DCs reached approximately 9 log. These cells performed functions similar to those of DCs derived from monocytes from peripheral blood (PBMC-DCs). The CTLs of the CD34-DCs (CD34-CTLs) presented a better antitumor effect in vitro. The obvious prophylactic and therapeutic antitumor effects of the CD34-DC vaccine were observed in the humanized NSG mouse models. CONCLUSION: CD34-DCs from cord blood were sufficient in quantity and quality as a vaccine agent against tumors in vitro and in vivo.


Assuntos
Vacinas Anticâncer/imunologia , Neoplasias Colorretais/terapia , Células Dendríticas/imunologia , Imunoterapia Adotiva/métodos , Animais , Antígenos CD34/metabolismo , Diferenciação Celular , Células Cultivadas , Neoplasias Colorretais/imunologia , Células Dendríticas/transplante , Modelos Animais de Doenças , Feminino , Sangue Fetal/citologia , Xenoenxertos , Humanos , Camundongos , Camundongos Endogâmicos , Camundongos SCID , Transplante de Neoplasias
15.
Cancer Chemother Pharmacol ; 83(5): 911-920, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30848330

RESUMO

Activation of programmed death-1 (PD-1) and cytotoxic T-lymphocyte antigen-4 (CTLA-4) on T cells leads to T cell exhaustion and ultimately facilitates tumor progression. Recent success of using immune cell checkpoint inhibitors offers a great promise to treat various cancers, including bladder cancer. However, the expression pattern and therapeutic value of PD-1 and CTLA-4 in peripheral blood T cells remain largely unexplored. In this study, we presume that disruption of the potential dysregulated checkpoint molecules in peripheral blood T cells may improve the anti-tumor efficacy of cytotoxic T cells in bladder cancer. We showed that both PD-1 and CTLA-4 expression were specifically elevated on CD8 + T cells but not CD4 + T cells in peripheral blood of patients with bladder cancer compared with that in healthy donors. Notably, CTLA-4 expression was significantly higher in muscle-invasive bladder cancer (MIBC) and correlated with tumor size. By blocking CTLA-4 with anti-CTLA-4 antibody and CRISPR-Cas9-mediated CTLA-4 disruption, we revealed that CTLA-4-disrupted CTLs had enhanced cellular immune response and superior cytotoxicity to the CD80/CD86-positive bladder cancer cells in vitro. Moreover, the CTLA-4-disrupted CTLs exhibited a pronounced anti-tumor effect in vivo as demonstrated by prophylactic assay and therapeutic assay in the subcutaneous xenograft model. Collectively, our findings confirm improved therapeutic efficacy of CTLA-4-disrupted CTLs and provides the potential strategy for targeting immune checkpoints to enhance the promising immunotherapy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Antígeno CTLA-4/genética , Receptor de Morte Celular Programada 1/genética , Neoplasias da Bexiga Urinária/patologia , Animais , Estudos de Casos e Controles , Linhagem Celular Tumoral , Feminino , Humanos , Imunoterapia/métodos , Masculino , Camundongos , Camundongos SCID , Pessoa de Meia-Idade , Neoplasias da Bexiga Urinária/imunologia , Ensaios Antitumorais Modelo de Xenoenxerto
16.
Biomed Pharmacother ; 114: 108708, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-30913493

RESUMO

PURPOSE: Dynamic remodeling of the extracellular matrix (ECM) around tumor cells is crucial for the tumor progressions. However, the mechanism is not well defined. Here, we aimed to reveal the underlying mechanism of ECM induced metastasis and provide innovative strategy to suppress the distant metastasis induced by ECM. MATERIALS AND METHODS: IHC was used to detect the expression of target proteins. H&E staining was used to evaluate the growth of tumor in vivo. Using wound healing and transwell assay, we examined the ability of cell to metastasis. We employed IF and Western blot to detect the expression of target proteins. And qRT-PCR was used to examine the target genes in mRNA level. We also applied flow cytometry to examine the percent of CD133+ cell population. RESULTS: Herein, we observed elevated expression of type I collagen in colorectal cancer tissues from patients with high metastasis. Additionally, colorectal cancer cells cultured on 2D collagen reveal obviously enhanced capability of metastasis and tumorigenesis both in vitro and in vivo. We demonstrated that the activation of PI3K/AKT signal induced by integrin α2ß1 resulted in the enhanced metastatic capability and stemness of colorectal cancer cells. Moreover, we found that Snail worked as the downstream of PI3K/AKT signaling, resulting in the intensive invasion and metastasis of colorectal cancer. Blocking the pathway by applying E7820 successfully reversed the type I collagen induced distant metastasis in colorectal cancer. CONCLUSION: Combining E7820 and chemotherapeutic agents to block the integrin α2ß1/PI3K/AKT/Snail signaling pathway revealed dramatic enhanced tumor suppression and provided an innovative approach for clinical colorectal cancer treatment.


Assuntos
Colágeno/metabolismo , Neoplasias Colorretais/metabolismo , Integrinas/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição da Família Snail/metabolismo , Animais , Carcinogênese/metabolismo , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Transição Epitelial-Mesenquimal/fisiologia , Feminino , Células HCT116 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus
17.
Eur J Pharm Sci ; 130: 78-90, 2019 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-30684657

RESUMO

Many strategies have been employed to improve oral drug delivery. One such approach involves the use of supersaturable delivery systems such as amorphous self-micellizing solid dispersions (SmSDs). SmSDs have attracted more attention recently, but little is known regarding the impact of production methods on profiles and internal mechanisms of final SmSDs in spite of its importance. In this study, amorphous SmSDs containing self-micellizing Soluplus® and BCS II drug (either indomethacin (IND) or fenofibrate (FEN)) were generated using various methods: solvent evaporation (SOL), freeze-drying (FD), microwave radiation-quench cooling (MQC), and hot melt extrusion (HME). Microscopic morphology, amorphous state, thermal behavior, dissolution/solubility, and "spring-parachute" data were used to assemble physicochemical profiles for SmSD systems prepared using each method. Analysis of intermolecular interactions, solubilization, and crystallization inhibition further uncovered internal mechanisms explaining observed physicochemical properties. Generally, SmSD/IND and SmSD/FEN systems generated using HME exhibited superior dissolution, solubility, and spring-parachute profiles. The superior advantages of HME-generated SmSD/IND systems were attributed to relatively stronger intermolecular interactions than observed in SmSD/IND systems fabricated using other methods. Moreover, self-micellizing Soluplus® carrier was able to solubilize IND or FEN and suppress drug crystallization from a supersaturated state, which seemed to be an important mechanism for the properties enhancement caused by SmSD/FENHME. This knowledge should be useful for guiding further development of self-micellizing solid dispersions and for gaining deeper understanding of how HME technology can improve supersaturable drug delivery based on SmSDs strategy.


Assuntos
Química Farmacêutica/métodos , Fenofibrato/química , Temperatura Alta , Indometacina/química , Micelas , Polietilenoglicóis/química , Polivinil/química , Anti-Inflamatórios não Esteroides/química , Anti-Inflamatórios não Esteroides/farmacocinética , Relação Dose-Resposta a Droga , Fenofibrato/farmacocinética , Hipolipemiantes/química , Hipolipemiantes/farmacocinética , Indometacina/farmacocinética , Polietilenoglicóis/farmacocinética , Polivinil/farmacocinética , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier/métodos
18.
Cancer Res Treat ; 51(1): 252-266, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-29690747

RESUMO

PURPOSE: We investigated the role of tumor-associated macrophages (TAMs) on the epithelial to mesenchymal transition (EMT) of colorectal cancer cells and determined the potential mechanism involved in the metastatic process. Materials and Methods: In this study, flow cytometry was used to detect the expression of target proteins. We used transwell assay to evaluate the migration of cancer cells under specific conditions. Using real-time polymerase chain reaction, we examined the expressions of cytokines and EMT-related markers in mRNA level. Animal assay was performed for analysis in vivo and hematoxylin and eosin was used to visualize the effect of TAMs on tumor metastasis. We also used immunohistochemistry and Western blotting to detect the expression of target proteins. RESULTS: Here, we observed enrichment of TAMs in colorectal tumor tissues, resulting in high metastasis in clinical therapy. Moreover, those TAMs could facilitate the EMT progression of colorectal cancer cells, which is induced by the transforming growth factor-ß (TGF-ß) derived from TAMs, leading to the invasion and migration of cancer cells. CONCLUSION: Our results demonstrated that TAMs contributed the EMT progression through a TGF-ß/Smad2,3-4/Snail signaling pathway, and disrupting this pathway with TGF-ß receptor inhibitor could suppress metastasis, readjusting our focus to the connection of TAMs and cancer metastasis.


Assuntos
Neoplasias Colorretais/metabolismo , Neoplasias Pulmonares/metabolismo , Macrófagos/metabolismo , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismo , Animais , Neoplasias Colorretais/genética , Neoplasias Colorretais/patologia , Transição Epitelial-Mesenquimal , Feminino , Células HCT116 , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Neoplasias Pulmonares/secundário , Macrófagos/patologia , Masculino , Camundongos , Invasividade Neoplásica , Transplante de Neoplasias , Proteína Smad2/genética , Proteína Smad2/metabolismo , Proteína Smad3/genética , Proteína Smad3/metabolismo , Proteína Smad4/genética , Proteína Smad4/metabolismo
19.
Front Pediatr ; 6: 293, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30356669

RESUMO

Purpose: To assess the outcomes of a novel laparoscopic assisted transcrotal orchidopexy (LATO) combined with percutaneous extraperitoneal closure (PEC) for palpable inguinal canalicular cryptorchidism accompany with indirect inguinal hernia, and evaluate its safety and efficiency. Materials and Methods: A retrospective cohort study for single-port LATO-PEC and traditional inguinal orchidopexy (TIO) was performed between 2011 and 2014. Totally 53 children with both palpable inguinal canalicular testes and indirect inguinal hernia were included. Median patient age was 15month (range, 6 months to 4 years). Of them, 35 patients underwent LATO-PEC procedure, utilizing an umbilical trocar for laparoscope, transcrotal dissection for orchidopexy, and an inner two-hooked cannula for ligation of the patent processus at the level of the internal ring. Three of them were bilateral, 12 on the left side and 20 on the right. Eighteen patients received TIO, seven of them on the left side and 11 on the right. Patient demographics, surgical technique, complications, and clinical outcomes were reviewed. Follow-up visits were performed to reassess position and size of the testes. Results: All 56 undescended testes were delivered into the scrotum successfully. In the LATO-PEC group, nine contralateral herniorrhaphy were accomplished simultaneously. Fifteen contralateral patent processus vaginalis (PPVs) in 32 unilateral undescended testis (UDT) were newly confirmed during the laparoscopy, while 6 of them received percutaneous extra-peritoneal herniorrhaphy for visible inguinal bubble in pneumoperitoneum condition. No additional port placement or conversion to open procedure was needed. Mean operative time for unilateral and bilateral LATO-PEC in this study was (37.81 ± 5.23) min and (53.33 ± 2.98) min, respectively. In TIO group, mean operative time was (41.11 ± 8.67) min. There was no statistical difference in operative time between the two approaches for unilateral UDTs (p = 0.098). Median follow-up interval was 24 months (range, 12-84 months). No operative complications were found in either group to date. Conclusions: Singe-port LATO-PEC is a safe, effective, and cosmetic choice for inguinal canalicular cryptorchidism accompany with indirect inguinal hernia, minimizing injuries to the vas deferens and testicular vessels. Laparoscopy can provide a diagnostic and therapeutic solution of contralateral PPV.

20.
Sci Rep ; 8(1): 2561, 2018 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-29416099

RESUMO

A murine monoclonal antibody (MAb-1) specific for GM3 has been generated by immunizing ß3Gn-T5 knockout mice with purified GM3 ganglioside. The binding specificity of MAb-1 (IgG3 subclass) was established by an enzyme-linked immunosorbent assay (ELISA) and FACS and the antibody showed high binding specificity with GM3. Cell viability assay showed that MAb-1 significantly suppressed cell growth. Immunohistochemistry analysis revealed that MAb-1 was strongly expressed in human ovarian cancer tissues, whereas it was hardly expressed in normal tissues. Finally, antibody-dependent cellular cytotoxicity (ADCC) activities were determined by measuring lactate dehydrogenase (LDH) releasing assay and the results showed high ADCC activities in two representative ovarian cancer cell lines (OVHM and ID8). All of these data indicate that MAb-1 may be potentially used as a therapeutic antibody against ovarian cancers in clinical trials.


Assuntos
Anticorpos Monoclonais Murinos/imunologia , Gangliosídeo G(M3)/análogos & derivados , Imunoglobulina G/imunologia , Neoplasias Ovarianas/imunologia , Animais , Anticorpos Monoclonais Murinos/genética , Especificidade de Anticorpos , Linhagem Celular Tumoral , Testes Imunológicos de Citotoxicidade , Feminino , Gangliosídeo G(M3)/imunologia , Humanos , Imunoglobulina G/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
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