Establishment of Patient-Derived Xenograft (PDX) Zebrafish Model of Multiple Myeloma and Its Application in Drug Screening / 中国实验血液学杂志

OBJECTIVE@#To establish a MM patient-derived tumor xenograft model (MM-PDX) in zebrafish, and to evaluate the anti-myeloma activity of indirubin-3'-monoxime(I3MO) using this model.@*METHODS@#Zebrafish embryos 2 days after fertilization were transplanted with fluorescence labeled myeloma primary tumor cells, the survival of primary tumor cells in zebrafish was observed at 0,16 and 24 hours after cell injection. The zebrafish embryos after tumor cell transplantation were randomly divided into control group, BTZ treatment and I3MO treatment group. Before and 24 hours after treatment with BTZ and I3MO, the positive area with calcein or Dil in zebrafish were observed under fluorescence microscope to reflect the survival of tumor cells, and it was verified.@*RESULTS@#MM patient derived tumor cells survived in zebrafish. The construction of MM-PDX was successful. Compared with control group, the fluo- rescence area of the BTZ and I3MO treatment groups in zebrafish were significantly decreased(P<0.05), and BTZ and I3MO significantly inhibited the survival of MM cells in zebrafish.@*CONCLUSION@#MM-PDX model was successfully established. Zebrafish model derived from tumor cells of MM patients can be used as a tool for drug screening of MM.

Efficacy and cardiotoxicity observation of different doses of capecitabine in patient-derived tumor xenograft of mice with colorectal cancer / 肿瘤研究与临床

Objective:To explore the correlation of the dose of capecitabine with the efficacy and cardiotoxicity in patient-derived tumor xenograft (PDX) model of mice with colorectal cancer.Methods:The fresh cancer tissues of 1 colorectal cancer patient were transplanted into the bilateral axillary subcutaneous of immunodeficient NOG mice to establish PDX model and passage stably. And then the morphology of tumor cells in primary generation and the second-generation tumor tissues was observed by using HE staining. The expression of tumor markers was detected by using immunohistochemistry method, and the model was evaluated. Mice were intragastrically infused with 200, 300 and 400 mg/kg capecitabine once a day, which were treated as low, middle and high dose groups respectively, 5 rats in each group; in the control group, 0.9% NaCl solution was perfused into the stomach; 14 d in total, use stop for 7 d, consecutively administered in this way. The body weight was measured every day and the tumor volume was measured every 3 days. After 100 days of observation, the mice were killed, and the tumor tissue was taken to measure the tumor weight and then the tumor volume, tumor volume inhibition rate and tumor inhibition rate were calculated. The morphology of tumor tissues was observed by using HE staining. The protein levels of anti-tumor effect indexes like rasP21, cyclooxygenase 2 (COX2), prostaglandin E2 (PGE2), cardiac troponin Ⅰ (cTn-Ⅰ) and brain natriuretic peptide (BNP) in serum of mice were detected by using enzyme linked immunosorbent assay (ELISA).Results:PDX model of mice with colorectal cancer was successfully constructed, and the histological characteristics of the primary tumor in the model were well preserved. During administration, 1 mouse died in the capecitabine high dose group; a slow down in tumor volume growth could be found with the increased dose of capecitabine. There was no statistically significant difference in body weight among 4 groups until all mice were killed ( P > 0.05). The tumor volume and tumor weight in the low, middle and high dose groups were lower than those in the control group (all P < 0.05), and the tumor volume and tumor weight showed an obvious decrease with the increase in dose. The tumor volume inhibition rates of low, middle and high dose groups were 42.61%, 67.61% and 77.27%, respectively, and the tumor inhibition rates were 35.53%, 67.77% and 75.09%, respectively. The serum anti-tumor effect indexes rasP21, COX2 and PGE2 in the middle and high dose groups were decreased compared with those in the control group (all P < 0.05), while cTn-Ⅰ and BNP levels were increased compared with those in the control group (all P < 0.05). Conclusions:The established PDX model of mice with colorectal cancer can better retain the histological characteristics of the original tumor. After treatment of middle and high dose of capecitabine, the tumor inhibition effect is obvious, but the risk of myocardial damage should be noticed.

A short-term chick embryo in vivo xenograft model to study retinoblastoma cancer stem cells

Purpose: Cancer stem cells (CSCs) reported in various tumors play a crucial role in tumorigenesis and metastasis of retinoblastoma (Rb). Following the efforts to reduce, replace, and refine the use of mammalian models, we aimed to establish a short?term xenograft for Rb to evaluate the CSC properties of CD133? Rb Y79 cells, using the well?established chick embryo chorioallantoic membrane (CE?CAM) assay. Methods: Y79 cells were cultured, labeled with two different dyes (CM?Dil Y79 and enhanced green fluorescent protein (eGFP)) and sorted for CD133? and CD133 + subsets. Two million cells from each of the labeled groups were transplanted onto the abraded CAM on embryonic day 7 (E7). On E14, the tumor nodule formation on CAM and spontaneous metastasis to the embryos were evaluated by confocal microscopy, in vivo imaging, and histology. Results: Y79 cells formed pink–white raised perivascular nodules with feeder vessels on the CAM with both the types of labeled CD133? cells. CD133? cells, when compared to CD133 + cells, demonstrated significantly larger tumor volume (40.45 ± 7.744 mm3 vs 3.478 ± 0.69 mm3, P = 0.0014) and higher fluorescence intensity (CM?Dil: AUF = 6.37 × 107 ± 7.7 × 106 vs 1.08 × 107 ± 1.6 × 106; P < 0.0001; eGFP: AUF = 13.94 × 104 ± 2.54 × 104 vs AUF = 1.39 × 104 ± 0.4 × 104; P = 0.0003). The metastatic potential of CD133? cells was also observed to be higher as noted by in vivo imaging and histopathology. Conclusion: This study highlights that CE?CAM is a feasible alternative nonmammalian model for evaluating tumorigenicity and metastatic potential of Y79 CSCs. Increased tumorigenicity and metastatic potential of CD133? subset of tumor cells substantiate their CSC properties

A novel second-generation platinum derivative and evaluation of its anti-cancer potential

Abstract Cisplatin is the primary anti-cancer agent for the treatment of most solid tumors. However, platinum-based anti-cancer chemotherapy produces severe side effects due to its poor specificity. There are a broad interest and literature base for a novel mechanism of action on platinum derivatives. Additionally, combining cisplatin with histone deacetylase inhibitors (HDACi) such as 4-hydroxybenzoic acid derivatives showed promising results in treating solid tumors. Here we aimed to conjugate 4-hydroxybenzoic acid with platinum to obtain a novel platinum derivative that can overcome cisplatin resistance. Cis-4-hydroxyphenylplatinum(II)diamine compound was synthesized under mild conditions and characterized. Cytotoxicity assay was performed on SKOV3-Luc and A549-Luc cells. Hemocompatibility and serum protein binding analysis were performed. Treatment potential was evaluated in xenograft tumor models. Biodistribution was tested on tumor-bearing mice via Pt analysis in organs with ICP-MS, ex vivo. In this study, cis-4-hydroxyphenylplatinum (II) diamine was synthesized with a yield of 62%. The MTT assay on A549-Luc and SKOV3-Luc cell lines resulted in IC50 values of 17.82 and 7.81 µM, respectively. While tumor growth was continued in the control group, the tumor volume decreased in the treatment group. All results point to the conclusion that the new compound has the potential to treat solid tumors

Application and expectation of patient-derived tumor xenograft models in the treatment of pancreatic cancer / 国际外科学杂志

Pancreatic cancer has a poor prognosis, systemic comprehensive therapy including chemotherapeutic and molecular targeted therapy is the key to improve the postoperative prognosis of pancreatic cancer, as well as to prolong the survival time of advanced pancreatic cancer. However, due to the drug resistance and heterogeneity of pancreatic cancer cells, systemic comprehensive treatment still does not reach the ideal effect, and personalized therapy will be an important approach to solve this problem. The personalized therapy for pancreatic cancer will become possible with the application of patient-derived tumor xenograft (PDX) models. Here, this article will review the progress of the application of PDX models in comprehensive therapy of pancreatic cancer based on reviewing the methods of establishing pancreatic cancer PDX models, aiming to provide new ideas for personalized therapy of pancreatic cancer.

Effects of silencing AEG-1 gene by shRNA on vasculogenic mimicry of a glioma xenograft model / 西安交通大学学报(医学版)

【Objective】 To investigate the effects of silencing AEG-1 gene by shRNA on vasculogenic mimicry (VM) of a glioma xenograft model. 【Methods】 U87 glioma cells were infected with AEG-1 shRNA lentivirals. Real-time PCR and Western blotting were used to detect the mRNA and protein expressions of AEG-1 in U87 cells after infected by the AEG-1 shRNA lentivirals. A glioma xenograft model was generated and CD 34/PAS double-staining was performed to detect the VM channels in vivo. The immunohistochemical assay was performed to evaluate the expressions of MMP-2, MMP-9, VE-cadherin, and VEGF in glioma xenograft models. 【Results】 AEG-1 shRNA lentivirals could significantly inhibit the AEG-1 expression in glioma cells (P<0.01). Meanwhile, they also decreased the number of VM in the glioma xenograft model (P<0.01). Furthermore, the expressions of MMP-2, MMP-9, VE-cadherin, and VEGF in glioma significantly decreased in vivo (P<0.01). 【Conclusion】 These results suggest that silencing AEG-1 gene by shRNA can significantly inhibit VM of glioma in vivo, the mechanism of which may partly be through regulating MMP-2, MMP-9, VEGF, and VE-cadherin expressions.

Establishment of a simple and efficient platform for car-t cell generation and expansion: from lentiviral production to in vivo studies

ABSTRACT Introduction: Adoptive transfer of T cells expressing a CD19-specific chimeric antigen receptor (CAR) has shown impressive response rates for the treatment of CD19 + B-cell malignancies in numerous clinical trials. The CAR molecule, which recognizes cell-surface tumor-associated antigen independently of human leukocyte antigen (HLA), is composed by one or more signaling molecules to activate genetically modified T cells for killing, proliferation, and cytokine production. Objectives: In order to make this treatment available for a larger number of patients, we developed a simple and efficient platform to generate and expand CAR-T cells. Methods: Our approach is based on a lentiviral vector composed by a second-generation CAR that signals through a 41BB and CD3-ζ endodomain. Conclusions: In this work, we show a high-level production of the lentiviral vector, which was successfully used to generate CAR-T cells. The CAR-T cells produced were highly cytotoxic and specific against CD19+ cells in vitro and in vivo, being able to fully control disease progression in a xenograft B-cell lymphoma mouse model. Our work demonstrates the feasibility of producing CAR-T cells in an academic context and can serve as a paradigm for similar institutions. Nevertheless, the results presented may contribute favoring the translation of the research to the clinical practice.

Establishment of a lung cancer mouse model with humanized immunity and its role in efficacy evaluation of programmed death-1 inhibitors / 上海交通大学学报（医学版）

Objective:To establish a lung cancer mouse model with humanized peripheral blood mononuclear cells (PBMC) expressing programmed death-ligand 1 (PD-L1), and study the role of the model in evaluating the efficacy of programmed death-1 (PD-1) inhibitors. Methods:Fresh biopsy tissue samples or tumor cells in malignant pleural effusion from the patients with advanced non-small cell lung cancer were inoculated subcutaneously in CB17-SCID mice to establish patient-derived xenograft (PDX) models. The expression of PD-L1 in PDX models was detected by immunohistochemistry. The mature human PBMC and PDX model tumor cells were mixed and then inoculated into NCG mice to establish a PDX model of lung cancer with humanized immunity, on which the efficacy of PD-1 inhibitor was verified. Results:Among the PDX models established by 16 clinical samples, 2 were strongly positive for PD-L1, 4 were positive, and the rest were negative. In the PDX model with strongly positive PD-L1, the tumor growth inhibition rate of cindilimab, an inhibitor of PD-1, was 82.6%, 21 days after the initial administration; in the PDX model with negative PD-L1, the inhibitor of PD-1 showed no antitumor activity. Conclusion:A PD-L1-expressing lung cancer mouse model with humanized immunity is successfully established and the efficacy of PD-1 inhibitor can be evaluated on the model.

Comparison of B-NDG and BALB/c mouse models bearing patient-derived xenografts of esophageal squamous cell carcinoma / 南方医科大学学报

OBJECTIVE@#To investigate the difference of tumor formation in different mouse strains bearing patient-derived xenograft of esophageal squamous cell carcinoma(ESCC) and establish a better animal model for preclinical study of individualized treatment of ESCC.@*METHODS@#The tumor tissues collected from 22 ESCC patients were used to establish tumor-bearing mouse models in B-NDG (NSG) mice and BALB/c nude mice. The tumor formation rate and tumor formation time were compared between the two mouse models, and HE staining, immunohistochemistry and genome sequencing were carried out to assess the consistency between transplanted tumor tissues in the models and patient-derived tumor tissues.@*RESULTS@#The tumor-bearing models were established successfully in both NSG mice (50%, 11/22) and BALB/c nude mice (18.18%, 4/22). The average tumor formation time was significantly shorter in NSG mice than in BALB/c nude mice (75.95 91.67 days, < 0.001). In both of the mouse models, the transplanted tumors maintained morphological characteristics identical to those of patient-derived ESCC tumors. Genetic analysis showed that the xenografts in NSG mice had a greater genetic similarity to the patients' tumors than those in BALB/c nude mice ( < 0.0001).@*CONCLUSIONS@#Mouse models bearing xenografts of patient-derived ESCC can be successfully established in both NSG mice and BALB/c nude mice, but the models in the former mouse strain can be more reliable.

Advances in the mouse models of myeloid leukemia / 生物医学工程学杂志

Mouse animal models are the most commonly used experimental tools in scientific research, which have been widely favored by researchers. The animal model of mouse leukemia appeared in the 1930s. During the past 90 years, researchers have developed various types of mouse leukemia models to simulate the development and treatment of human leukemia in order to promote effectively the elucidation of the molecular mechanism of leukemia' development and progression, as well as the development of targeted drugs for the treatment of leukemia. Considering that to myeloid leukemia, especially acute myeloid leukemia, there currently is no good clinical treatment, it is urgent to clarify its new molecular mechanism and develop new therapeutic targets. This review focuses on the various types of mouse models about myeloid leukemia used commonly in recent years, including mouse strains, myeloid leukemia cell types, and modeling methods, which are expected to provide a reference for relevant researchers to select animal models during myeloid leukemia research.

Establishment of patient derived xenograft model of high-grade mucinous carcinoma peritonei accompanied with signet ring cells and identification of biological characteristics / 中华肿瘤杂志

Objective@#To establish the patient derived xenograft (PDX) model of pseudomyxoma peritonei (PMP), and identify the key characteristics of tumor biology of this model, in order to provide a reliable model for studying the pathological mechanisms and new therapeutic strategies of PMP.@*Methods@#PMP tumor tissue was obtained from surgery and cut into pieces after washing. Then tumor pieces were implanted subcutaneously in BAL B/c-nu mice for 6 stable passages. In the 7th passage, tumor tissue was implanted orthotopically into abdomen. Subcutaneous tumor and orthotopic tumor were then homogenized to make tumor cell suspension, implanted into abdomen of 10 BAL B/c-nu mice through midline laparotomy, 100 μl for each. The key experimental parameters including body weight changes in the observation period, experimental peritoneal cancer index (ePCI) score at the autopsy, histopathological and immunohistochemical characteristics, and gene expression profiles by high-throughput whole-genome exon sequencing were detected and recorded.@*Results@#The successful rate of established orthotopic PDX model of human PMP was 100% (10/10). The animals showed smooth body weight increases after tumor inoculation until day 27, then the body weight began to decrease steadily. Widespread tumor dissemination of PMP tumor through the whole abdomen was found by autopsy, including the diaphragm, liver, spleen, stomach, kidney, parietal peritoneum, bowel and mesenterium. Gelatinous ascites was also observed in abdominopelvic cavity. The ePCI score ranged from 5 to 9, with a 8 of median ePCI. Histopathological studies showed peritoneal mucinous carcinomatosis accompanied with signet ring cells (PMCA-S), obvious tumor cell atypia and parenchymal invasion.Immunohistochemistry showed the expressions of MUC1, MUC2, MUC5AC, CEA, CA199, CK20, CDX-2 and Ki-67 were positive, MUC6, CK7 and p53 were negative. Whole-exome sequencing identified that the most significant genetic alteration is the exon10 missense mutation c. 1621A>C of KIT gene, the mutation abundance was 89.7%.@*Conclusion@#PDX model of PMCA-S is successfully established, which displays the characters of high-degree malignancy, high proliferation and strong aggressiveness.

Chinese Herbal Medicine for Osteosarcoma in the Mouse: A Systematic Review and Meta-Analysis / 中国结合医学杂志

OBJECTIVE@#To summarize and critically assess the inhibitory effects of Chinese herbal medicine (CHM) on tumor volume and tumor weight for the treatment of osteosarcoma (OS) in mouse models.@*METHODS@#PubMed, Embase, Web of Science, China Knowledge Resource Integrated Database (CNKI), Wanfang Database, VIP Database, and Chinese BioMedical (CBM) were searched since their inception dates to March 10, 2016. Two reviewers independently selected the controlled studies estimating effects of CHM on mouse OS by administration in vivo. A pair-wise meta-analysis was performed. Twenty-five studies with adequate randomization were included in the systematic review.@*RESULTS@#CHM may significantly inhibit OS growth in mice, as assessed using the tumor weight [20 studies, n=443; 290 for CHM and 153 for the control: pooled mean difference (MD)=-2.90; 95% confidence interval (Cl): -3.50 to -2.31: P<0.01], tumor volume (16 studies, n=382; 257 for CHM and 125 for the control; pooled MD =-2.57; 95% Cl: -3.33 to -1.80; P<0.01) and tumor growth inhibition rate.@*CONCLUSION@#CHM could significantly inhibit the growth of OS in mouse models, which might be supportive for the design of preclinical and clinical trials in future.

Bortezomib and obatoclax for dual blockade of protein degradation pathways show synergistic anti-tumor effect in human acute T lymphoblastic leukemia cells / 南方医科大学学报

OBJECTIVE@#To explore whether bortezomib and a Bcl-2 inhibitor exhibit synergistic anti-tumor effect in human acute T lymphoblastic leukemia cells.@*METHODS@#MTT assay was used to determine the cytotoxicity of bortezomib in the absence or presence of Bcl-2 inhibitors (obatoclax, AT-101 and ABT-199) in Jurkat cells. The effects of drug treatment on the expression of Bcl-2 family proteins, LC3B, p62, ubiquitin, BiP/Grp78, p-JNK, p-p38 and CHOP proteins were examined by Western blotting. Flow cytometry was used to determine the effects of bortezomib and Bcl-2 inhibitors (obatoclax, AT-101 and ABT-199) on cell apoptosis. Quantitative real-time PCR was used to measure the mRNA expression levels of the key regulatory factors of unfolded protein reaction (UPR). A zebrafish xenograft model was used to study the anti-tumor effect of bortezomib, obatoclax and their combination in vivo.@*RESULTS@#Bortezomib or Bcl-2 inhibitors alone inhibited the cell viability of Jurkat cells, but only obatoclax and bortezomib showed synergistic cytotoxicity and pro-apoptotic effect. Obatoclax, rather than AT-101 and ABT- 199, blocked autophagic flux in the cells evidenced by concomitant accumulation of LC3B-Ⅱ and p62. Both bortezomib and obatoclax alone caused accumulation of polyubiquinated proteins, and their combination showed a synergistic effect, which was consistent with their synergistic cytotoxicity. The dual blockade of proteasome and autophagy by the combination of bortezomib and obatoclax triggered unfolded protein response followed by cell apoptosis. Preventing UPS dysfunction by tauroursodeoxycholic acid (TUDCA) significantly attenuated the cytotoxicity and pro-apoptotic effect of bortezomib in combination with obatoclax. In zebrafish xenograft models, bortezomib combined with obatoclax significantly decreased tumor foci formation.@*CONCLUSIONS@#Bortezomib and obatoclax for dual blockade of protein degradation pathways show synergistic anti-tumor effect in human acute T lymphoblastic leukemia cells.

Glioma stem cell clones and molecular genetics characteristics of primary and recurrent gliomas / 中华神经医学杂志

Objective To explore the molecular genetic characteristics of primary and recurrent glioblastomas (GBMs) from the same patient in vivo, primary glioma stem cells cultured in vitro, and patient-derived xenograft (PDX). Methods (1) The primary and recurrent GBM specimens from one patient during surgical resection were collected; and the expressions of glial fibrillary acidic protein (GFAP), nestin and Ki-67 were detected by immunohistochemical staining; the methylation of O6-methylguanine DNA methyltransferase (MGMT) gene, mutation of isocitrate dehydrogenase (IDH) gene and amplification of epidermal growth factor receptor (EGFR) gene were analyzed. (2) The primary and recurrent GBM stem cells were cultured in vitro and named as SU5-1 and SU5-2 cells, respectively; the expressions of nestin and CD133 were detected by immunohistochemical staining; GFAP expression was detected by immunohistochemical staining after induced differentiation, and the growth curve was detected by CCK-8 assay; Transwell invasion assay was used to detect the invasion ability; cell resistance to temozolomide (TMZ), carboplatin (CBP), cisplatin (DDP) and adriamycin (ADM) was detected by CCK-8 assay; the protein expression of programmed death receptor-ligand 1 (PD-L1) was detected by Western blotting. The rate of PD-L1 positive cells was detected by flow cytometry; genetic testing analysis was as above. (3) The primary and recurrent in situ PDX models in nude mice were established, and the expressions of nestin, GFAP and Ki-67 were detected by immunohistochemical staining. Results (1) As compared with the primary GBM, the recurrent GBM had significantly higher percentages of Ki-67 and nestin positive cells, while statistically lower percentage of GFAP positive cells (P<0.05); genetic analysis showed that there was no mutation in IDH gene in the primary GBM tissues and recurrent GBM tissues; the MGMT gene in the primary GBM tissues was methylated and EGFR gene was not amplified, while the MGMT gene in recurrent GBM tissues was demethylated and EGFR gene amplification was positive. (2) Both SU5-1 and SU5-2 cells expressed nestin and CD133, and GFAP was expressed after induced differentiation; the growth curve showed that the proliferation of SU5-2 cells started earlier than that of SU5-1 cells, the two were equal on the 3rd, 4th, and 5th d, and the proliferation of SU5-1 cells was faster than that of SU5-2 cells from the 6th d; the invasion ability of SU5-2 cells was statistically stronger than that of SU5-1 cells (P<0.05); the inhibition rates of SU5-2 cells treated with 5, 10, and 15 mmol/L CBP, 0.3125, 1.25, and 5 mmol/L DDP, 0.5 and 2 mmol/L ADM, and 125 and 500 mmol/L TMZ were significantly lower than those of SU5-1 cells treated with the same concentrations and same drugs (P<0.05); the protein expression of PD-L1 in SU5-2 cells was higher than that in SU5-1 cells; the positive rate of PD-L1 in SU5-2 cells was statistically higher than that in SU5-1 cells (P<0.05); the results of genetic analysis were consistent with those of the primary and recurrent GBM samples. (3) As compared with those in the primary PDX model, the nestin and Ki-67 expressions were significantly higher and GFAP expression was significantly lower in the recurrent PDX model (P<0.05); the results of genetic analysis were consistent with those of the primary and recurrent GBM samples. Conclusions Genetic differences are detected between primary and recurrent GBMs; recurrent GBM has stronger invasive capacity and multi-drug resistance. The primary stem cells derived from surgical specimens and corresponding PDX models could replicate the molecular genetic characteristics of original tumors, which provide a reliable experimental platform for both tumor translation researches and screening of molecular therapeutic targets.

Modeling guidelines and latest applications of patient-derived xenograft models / 国际外科学杂志

Tumor models are required to keep the most original characteristics of the primary tumor in precision treatment.Patient-derived xenograft models are established when cancerous cells or tissues directly from patients'primary tumors are transplanted into immunodeficient mice to mimic human tumor biology in vivo,which have been widely used in cancer research.In this review,we initially summarize the methodology and its progress to create patient-derived xenograft models from three aspects including grafts,hosts and grafting regions,and then go over recent applications of patient-derived xenograft models in basic cancer research on the areas of tumorigenesis,metastasis and drug resistance and in translational medical research of tumor,such as exploring cancer biomarkers,screening anti-cancer drugs and personalized therapy for neoplasm.Finally,we propose the problems of patientderived xenograft,which must be solved urgently.

Total Saponins of Rubus Parvifolius L. Exhibited Anti-Leukemia Effect in vivo through STAT3 and eIF4E Signaling Pathways / 中国结合医学杂志

OBJECTIVE@#To investigate the anti-leukemia effect of total saponins of Rubus parvifolius L. (TSRP) on K562 cell xenografts in nude mice and the mechanisms of action.@*METHODS@#The K562 cell xenografts in nude mice were established, and then randomly divided into 5 groups, the control group, the cytosine arabinoside group(Ara-c) and 3 TSRP groups (20, 40 and 100 mg/kg). The tumor volume and mass of each group of nude mice were measured and the anti-tumor rates of TSRP were calculated subsequently. The apoptosis status of tumor cells was detected by hematoxylin-eosin (HE) and terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) staining analysis. Finally, the activities of apoptosis related signaling of signal transducer and activator of transcription 3 (STAT3), eukaryotic initiation factor 4E (eIF4E) and B-cell lymphoma-2 (bcl-2) were determined with immunohistochemistry tests.@*RESULTS@#Subcutaneous injection of K562 cells induced tumor formation in nude mice, and the TSRP treated group showed a signifificant inhibitory effect on tumor formation. The nude mice treated with TSRP showed a signifificant decrease in tumor growth rate and tumor weight in comparison to the control group (all P<0.05). The HE staining and TUNEL assay showed that TSRP induced cell death by apoptosis. The immunohistochemical assay showed down-regulation of the bcl-2 gene in the TSRP treated cells. The phosphorylation levels of eIF4E and STAT3 were decreased obviously after the treatment of TSRP.@*CONCLUSION@#TSRP had an excellent tumor-suppressing effect on K562 cells in the nude mice xenograft model, suggesting that TSPR can be developed as a promising anti-chronic myeloide leukemia drug.

Demethylation effect of emodin combined with 5AzA-cdR on tumor suppressor gene p16, RASSF1A and ppENK in subcutaneous xenografts of pancreatic cancer nude mice model / 中华胰腺病杂志

Objective To investigate whether emodin combined with 5AzA-cdR can enhance the demethylation of tumor suppressor genes p16,RASSF1A and ppENK in nude mice with subcutaneously transplanted pancreatic cancer.Methods Pancreatic cancer cells Panc1 burdened subcutaneous xenograft nude mice model was established,which were randomly divided into control group,emodin group,5AzA-cdR group and emodin combined 5AzA-cdR group (combined group).The growth of transplanted tumors wasobserved in each group.Methylation specific PCR (MSP) was used to detect the methylation levels of p16,RASSF1A and ppENK in the xenograft tumor tissue among three groups.The mRNA and protein expression of three tumor suppressor genes were detected by FQ-PCR and Western blotting,respectively.Results The weight of xenografts in the control group,emodin group,5AzA-cdR group,and combination group were (0.28 ±0.01),(0.17 ± 0.01),(0.12 ± 0.02),(0.08 ± 0.01)g,respectively.The tumor volume was (517 ±0.02),(382 ± 0.01),(232 ± 0.03),(169 ± 0.01) mm3.The methylation levels of p16 were 1.00 ± 0.00,0.89 ± 0.02,0.63 ± 0.02,and 0.19 ± 0.01;the methylation levels of RASSF1A were 1.00 ± 0.00,0.88 ± 0.02,0.51 ± 0.01,and 0.32 ± 0.01;the methylation degree of ppENK was 1.00 ± 0.00,0.92 ± 0.02,0.77 ± 0.02 and 0.31 ± 0.01,respectively.The expression of p16 mRNA was 1.00 ± 0.00,1.71 ±0.02,2.67 ± 0.02,3.81 ± 0.01.The expression of RASSF1A mRNA was 1.00 ± 0.00,1.92 ±0.02,2.73 ± 0.03,3.77 ± 0.01.The expression of ppENK mRNA was 1.00 ± 0.00,1.69 ± 0.03,2.17 ± 0.02 and 4.28 ± 0.01.The expression of p16 protein was 1.00 ± 0.00,1.71 ± 0.02,2.67 ± 0.02,3.81 ± 0.01;the expression of RASSF1A protein was 1.00 ± 0.00,1.92 ± 0.02,2.73 ± 0.03.3.77 ± 0.01;ppENK protein expression levels were 1.00 ±0.00,1.69 ±0.03,2.17 ±0.02,4.28 ±0.01.The weight and volume of xenografts in the three treatment groups were significantly smaller than those in the control group.The methylation of three tumor suppressor genes was lower than that of the control group,and the expression of tumor suppressor mRNA and protein was all significantly higher than the control group,which the combination drug group was also significantly stronger than that in emodin group and 5AzA-cdR group,and the differences were statistically significant (P ＜ 0.05 or ＜ 0.01).Conclusions The combination of emodin and 5AzA-cdR can enhance the demethylation effect of 5A6A-cdR on the tumor suppressor genes p16,RASSF1A and ppENK in the tumor tissue of pancreatic cancer xenograft model.

Patient-Derived Xenograft Models of Epithelial Ovarian Cancer for Preclinical Studies / Journal of the Korean Cancer Association, 대한암학회지

PURPOSE: Patient-derived tumor xenografts (PDXs) can provide more reliable information about tumor biology than cell line models. We developed PDXs for epithelial ovarian cancer (EOC) that have histopathologic and genetic similarities to the primary patient tissues and evaluated their potential for use as a platform for translational EOC research. MATERIALS AND METHODS: We successfully established PDXs by subrenal capsule implantation of primary EOC tissues into female BALB/C-nude mice. The rate of successful PDX engraftment was 48.8% (22/45 cases). Hematoxylin and eosin staining and short tandem repeat analysis showed histopathological and genetic similarity between the PDX and primary patient tissues. RESULTS: Patients whose tumors were successfully engrafted in mice had significantly inferior overall survival when compared with those whose tumors failed to engraft (p=0.040). In preclinical tests of this model, we found that paclitaxel-carboplatin combination chemotherapy significantly deceased tumor weight in PDXs compared with the control treatment (p=0.013). Moreover, erlotinib treatment significantly decreased tumor weight in epidermal growth factor receptor–overexpressing PDX with clear cell histology (p=0.023). CONCLUSION: PDXs for EOC with histopathological and genetic stability can be efficiently developed by subrenal capsule implantation and have the potential to provide a promising platform for future translational research and precision medicine for EOC.

Establishing a colorectal cancer liver metastasis patient-derived tumor xenograft model for the evaluation of personalized chemotherapy

PURPOSE: In order to suggest optimal anticancer drugs for patient-tailored chemotherapy, we developed a colorectal cancer (CRC)-liver metastasis patient-derived tumor xenograft (PDTX) model. METHODS: Tissue obtained from a patient with CRC-liver metastasis (F0) was transplanted in a nonobese female mouse with diabetic/severe combined immune deficiency (F1) and the tumor tissue was retransplanted into nude mice (F2). When tumor volumes reached ~500 mm³, the F2 mice were randomly divided into 4 groups (n = 4/group) of doxorubicin, cisplatin, docetaxel, and nontreated control groups. The tumor tissues were investigated using H&E staining, terminal deoxynucleotidyl transferase dUTP nick end labeling assays, and immunohistochemistry. To determine where the mutant allele frequencies varied across the different passages, we isolated genomic DNA from the primary tumor, liver metastasis, and PDTX models (F1/F2). RESULTS: The physiological properties of the tumor were in accord with those of the patient's tumors. Anticancer drugs delayed tumor growth, inhibited proliferation, and caused apoptosis. Histological assessments revealed no observable heterogeneity among the intragenerational PDTX models. Target exon sequencing analysis without high-quality filter conditions revealed some genetic variations in the 83 cancer-related genes across the generations. However, when de novo mutations were defined as a total count of zero in F0 and ≥5 in F2, exactly prognostic impact of clone cancer profiling (EGFR, KRAS, BRAF, PIK3CA, NRAS, APC and TP53) were detected in the paired. CONCLUSION: A CRC liver metastasis PDTX model was established for the evaluation of chemotherapeutic efficacy. This model retained the physiological characters of the patient tumors and potentially provides a powerful means of assessing chemotherapeutic efficacy.

Inhibitory effect of 2,3',4,5'-tetramethoxystilbene on growth of endometrial carcinoma xenografts / 肿瘤

Objective: To investigate the effect of 2,3',4,5'-tetramethoxystilbene (TMS) on the growth of xenograft tumors of endometrial carcinoma Ishikawa cells, to explore its molecular mechanism, and to evaluate the adverse reaction of TMS. Methods: Human xenograft models were established in nude mice by inoculation with human endometrial carcinoma Ishikawa cells, and randomly divided into TMS group and control group by subcutaneously injection with 10 mgkg-1d-1 TMS and equal volume of olive oil, respectively. The volume and weight of xenograft tumors were calculated. The body weight of mice and the wet weight of tumor, heart, liver, spleen, lung, kidney, uterus and ovary in mice were measured. The solid organ damage was detected by HE staining. The expressions of cytochrome P4501B1 (CYP1B1), extracellular signal-regulated kinase (ERK) and phospho-ERK (p-ERK) proteins in xenograft tumors were detected by SP immunohistochemistry method. Results: After subcutaneous injection of TMS, the volume (P 0.05). No obvious lesion was found in various organs of nude mice in each group. The expressions of CYP1B1, ERK and p-ERK proteins were weakly positive in xenograft tumors. Conclusion: TMS can obviously inhibit the growth of human endometrial carcinoma xenografts in nude mice, and has no obvious toxic and side effects on tumor-bearing nude mice.