Lewis Lung Cancer Research Articles

Effect of erlotinib combined with cisplatin on IL-6 and IL-12 in mice with Lewis lung cancer.

Effect of erlotinib combined with cisplatin on tumor growth, interleukin-6 (IL-6) and interleukin-12 (IL-12) in mice with Lewis lung cancer (LLC) was investigated. Forty-four pure inbred SPF C57BL/6J mice were modeled for LLC and randomized into groups A, B, C and D (n=11 each group). Mice in group A were given normal saline, group B was given erlotinib, group C was given cisplatin injection and group D erlotinib combined with cisplatin. Tumor growth of the mice was observed and the tumor mass was measured. Serum IL-6 and IL-12 levels were measured by enzyme-linked immunosorbent assay (ELISA) 40 days later. At different time-points after medication, tumor volume in group D was significantly lower than that in groups A, B and C (P<0.05), and that in groups B and C was significantly lower than that in group A (P<0.05), whereas there was no significant difference between groups B and C (P>0.05). Tumor mass in groups B, C and D was significantly lower than that in group A (P<0.05), and that in group D was significantly lower than that in groups B and C (P<0.05), whereas there was no significant difference between groups B and C (P>0.05). Compared with groups B and C, mice in group D had significantly lower IL-6 level (P<0.05), but significantly higher IL-12 level (P<0.05). There was no significant difference in IL-6 and IL-12 levels between groups B and C (P>0.05). In conclusion, erlotinib combined with cisplatin can inhibit the tumor growth of mice with LLC, and inhibition of IL-6 level and upregulation of IL-12 level may be one of its therapeutic mechanisms.

Antitumor immunity targeting fibroblast activation protein-α in a mouse Lewis lung carcinoma model.

The tumor stromal microenvironment is an integral part of the occurrence and development of tumor. Cancer-associated fibroblasts (CAFs) are a key component of most tumor stromal microenvironments. The present study aimed to investigate the use of CAFs-targeted immunotherapy to fibroblast activation protein-α (FAP-α) expressed in CAFs. Recombinant adenoviral vectors containing the mouse FAP-α cDNA (rAd-FAP-α) were constructed. C57BL/6 mice were immunized with rAd-FAP-α infected dendritic cells (DCs) against FAP-α, which is overexpress in CAFs. The results demonstrated that mice vaccinated with rAd-FAP-α DCs gave rise to potent FAP-α-specific cytotoxic T lymphocytes capable of lysing Lewis lung cancer (LLC) CAFs. Furthermore, mice vaccinated with rAd-FAP-α-transduced DCs induced an effective therapeutic or protective antitumor immunity to LLC in a subcutaneous model, and prolonged overall survival time compared with mice vaccinated with the control recombinant adenovirus-transduced DCs (rAd-c DCs) or DCs alone. The results of the present study suggested that FAP-α, which is preferentially expressed in CAFs, may be considered as a potential target for killing or destroying CAFs within the tumor stromal microenvironment, and may be exploited to develop immunogenic tumor vaccines.

Synergistic effect of a combination of granulocyte macrophage colony-stimulating factor and thymosin α1 on Lewis lung cancer transplanted tumor in mice

Purpose: To study the synergistic effect of a combination of granulocyte-macrophage colonystimulating factor and thymosin-α1 on the treatment of Lewis lung cancer transplanted tumor.Methods: C57BL/6 mice were used. A mouse model of Lewis lung cancer was established using Lewis lung cancer cell lines. The mice were randomly divided into blank control group, polyene taxol (DTX) group, DTX thymosin α1 (Tα-1) group, and DTX granulocyte-macrophage colony-stimulating factor (GM-CSF) group, with 8 mice per group. The degree of tumor inhibition, thymus mass, thymus index, spleen mass, spleen index, IL-6, TNF-1, IFN-1, CD4+, CD8+ T cells and the ratio of CD4+/CD8+ were determined by ELISA and flow cytometry.Results: Body mass, thymus mass, thymus index, spleen mass, spleen index, IL-6, TNF-1, IFN-1, CD4+, CD8+ T cells and the ratio of CD4+/CD8+ in DTX + Tα-1 group, DTX + GM-CSF group and DTX + Tα-1 + GM-CSF group were significantly elevated (p &lt; 0.05), relative to the corresponding levels in DTXmice (p &lt; 0.05). Body mass, degree of tumor inhibition, thymus mass, thymus index, spleen mass, spleen index, IL-6, TNF-1, IFN-1, CD4, CD8 T cells and CD4+/CD8+ ratio in DTX + Tα-1 + GM-CSF mice were significantly elevated, relative to the DTX + Tα-1 and DTX + GM-CSF groups (p &lt; 0.05). Thestate of the tumor was significantly improved in the DTX + Tα-1 and DTX + GM-CSF mice.Conclusion: A combination treatment of GM-CSF, Tα-1 and DEX effectively enhances the resistance of mice and suppresses chemotherapy-induced decrease in body weight. This finding may be of clinical significance.&#x0D; Keywords: Granulocyte macrophage, Colony-stimulating factor, Thymosin, Docetaxel, Lewis lung cancer, Transplanted tumor

Insulin-like growth factor-1 receptor induces immunosuppression in lung cancer by upregulating B7-H4 expression through the MEK/ERK signaling pathway.

The Insulin-like growth factor-1/Insulin-like growth factor-1 receptor (IGF1/IGF1R) axis contributes to immunosuppression during tumor progression; however, the underlying mechanism remains unclear. In the present study, we found that IGF1 stimulation or IGF1R overexpression (IGF1R-OE) could upregulate the expression of B7-H4, while IGF1R inhibition downregulated B7-H4 in both A549 and SPC-A-1 lung cancer cell lines. IGF1R-OE conferred the inhibition of CD8+ T cells by cancer cells in vitro, and induction of B7-H4 expression was mediated by the activation of the MEK/ERK1/2 signaling pathway. The in vitro findings were further confirmed in vivo using a Lewis lung cancer mouse model. IGF1R-OE promoted tumor growth and inhibited tumor infiltration by CD8+ T cells in the mouse model. However, this effect was suppressed when B7-H4 was knocked down in IGF1R-OE cells. Our findings suggest that IGF1R could induce immunosuppression in lung cancer by upregulating the expression of B7-H4 through the MEK/ERK pathway. B7-H4 may therefore be a potential therapeutic target for lung cancer immunotherapy.

INKT cell modulation of tumor microenvironment (TME) and antitumor immunity: the role of antitumorigenic and protumorigenic iNKT cells

Abstract Preclinical and clinical data indicate that iNKT cells play important roles in antitumor immunity. High number of iNKT cells in the PBL of cancer patients usually correlates with a favorable prognosis. We found that compared to wild-type (WT) mice, depletion of iNKT cells did not affect tumor progression or metastasis in mice bearing melanoma or Lewis lung cancer; restoring iNKT cells by transferring thymocytes into iNKT KO mice dramatically enhanced antitumor immunity. The underlying mechanism is no known. Herein, we report progress on the mechanistic study. Thymocyte transfer only reconstituted iNKT cells in liver, spleen and blood, but not in LN of iNKT KO mice. This suggests that two functionally opposite iNKT subsets exist in the immune system, and thymocyte transfer only restored antitumorigenic iNKT, but not protumorigenic iNKT cells. Transferring IFNγ KO thymocytes diminished iNKT reconstitution-induced antitumor immunity; transferring IL-17A KO thymocytes enhanced iNKT reconstitution-enhanced antitumor immunity. Compared to WT mice or iNKT KO mice, iNKT reconstitution in iNKT KO mice up regulated the expression of XCL1, FLT3L and CCL5, down regulated IL-6, TGFβ1, TGFβ2, COX2 expression, increased IRF8+ cDC1 cells and Siglec F− DC, and decreased αSMA+ FAP+ cancer-associated fibroblasts (CAF) and MDSC in the tumor. Taken together, these results support that antitumorigenic iNKT cells produce IFNγ dominant cytokine and facilitate NK/cDC1 infiltration and activation in the tumor to induce antitumor immunity; protumorigenic iNKT cells induce CAF to produce TGFβ, which induces a myeloid cell-mediated inhibitory TME. The balance between antitumorigenic and protumorigenic iNKT cells shapes the TME and antitumor immunity.

M1 macrophage-derived exosome-encapsulated cisplatin can enhance its anti-lung cancer effect.

The aim of this study was to study the efficacy of cisplatin-loaded M1 macrophage secreted-exosomes (DDP-M1-Exos) in enhancing DDP antitumor effect. M1-Exos were first extracted using density gradient centrifugation, and the DDP-M1-Exos system was established via electroporation. Then, the morphology, particle size and maker proteins of the DDP-M1-Exos system were assessed using transmission electron microscope, DLS and Western blotting (WB), respectively. The uptake of Exos by mouse Lewis lung cancer (LLC) cells was observed under a confocal laser scanning microscope, and the influence of the DDP-M1-Exos system on the viability of Lewis cells was determined using methyl thiazolyl tetrazolium assay and 4',6-diamidino-2-phenylindole staining. Besides, its impacts on the messenger ribonucleic acid (mRNA) levels and protein expression levels of B-cell lymphoma 2 (Bcl-2), Bcl-2 associated X protein (Bax) and Caspase-3 in LLC cells were examined using quantitative real-time PCR (qRT-PCR) and WB, respectively. Finally, with the LLC-bearing mouse model as the object of study, the efficacy was evaluated using terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) and hematoxylin-eosin staining staining in vivo. Saucer-shaped Exos with a bilayer membrane, uniform particle size and highly expressed CD81, Alix and HSP70 were successfully isolated from M1 macrophages via density gradient centrifugation, and they were absorbed by LLC cells. Meanwhile, the DDP-M1-Exos drug delivery system was successfully prepared using electroporation technique. Compared with Control group, DDP and DDP-M1-Exos groups exhibited a decrease in the proliferation rate of mouse LLC cells and an increase in their apoptosis rate (*P<0.05), and the apoptosis rate in DDP-M1-Exos group was substantially higher than that in DDP group (#P<0.01). According to the qRT-PCR and WB results, DDP-M1-Exos up-regulated Bax and Caspase-3 in the apoptosis signaling pathway to induce tumor cell apoptosis, thereby resisting tumors. Moreover, through in-vivo experiments, it was found that M1-Exos alone had a potential to suppress tumor growth, and that the carrier of M1-Exos could not only kill tumor cells, but also encapsulate DDP to enhance its anti-lung cancer effect. The DDP-M1-Exos drug delivery system is prepared using Exos extracted via density gradient centrifugation by electroporation, and the drug-loaded Exos are able to effectively inhibit the proliferation of tumor cells and induce their apoptosis, exerting an antitumor effect.

Effect of Fei-Liu-Ping ointment combined with cyclophosphamide on lung cancer cell proliferation and acidic microenvironment

To observe the effects of Fei-Liu-Ping ointment and chemotherapy on mice with lung cancer, and to explore the inherent mechanism of action from the point of acidic microenvironment and apoptosis. First of all, the Lewis lung cancer transplanted mouse model was established. Therefore, they were treated by Fei-Liu-Ping ointment, cyclophosphamide, Fei-Liu-Ping ointment + cyclophosphamide and the saline as control. All the groups' tumor size, tumor growth rate and food consumption were recorded. The mice were sacrificed and the tumors were took out after 15 days' interventions. Then lactate relative concentrations were detected with lactate kits and the protein expressions of glucose transporter 4 (GLUT4), hexokinase 1 (HK1), glucose-regulated protein 78 (GRP78), carbonic anhydrase-IX (CA-IX) were detected through immunohistochemical staining. Flow cytometry was adopted to detect the percentage of apoptotic tumor cells and regulatory T cells (Treg), and the expression of hypoxia-inducible factor-1α (HIF-1α), Bcl-2, Bax, Caspase-3, interleukin-2 (IL-2) were tested through western blot. The strongest inhibition effect and the lowest tumor growth rate was found in Fei-Liu-Ping ointment + cyclophosphamide group. There were significant differences between Fei-Liu-Ping ointment + cyclophosphamide group and saline group(P<0.05). And the highest food consumption was found in Fei-Liu-Ping ointment + cyclophosphamide group while there were no significant differences between Fei-Liu-Ping ointment + cyclophosphamide group and saline group (P>0.05). Further molecular biological detections found that the lowest lactate level and regulatory T cells ratio were found in Fei-Liu-Ping ointment + cyclophosphamide group and these expressions of GLUT4, HK1, GRP78, CA-IX were suppressed. There were significant differences between Fei-Liu-Ping ointment+cyclophosphamide group and saline group (P<0.05). In addition, the Fei-Liu-Ping ointment + cyclophosphamide group's cell apoptosis increased significantly compared with saline group and there were significant differences on expressions of HIF-1α, Bcl-2, Bax, Caspase-3, IL-2 for this group compared with saline group. Chemotherapy and Fei-Liu-Ping ointment had the synergistic effect on inhibiting tumor growth and improving the general conditions of tumor-bearing mice. The effect was partly owed to the improvement on tissue hypoxia, the inhibition of HIF-1α expression and the regulations on its downstream proteins, such as GLUT4, HK1, GRP78, and CA-IX. And then all these alterations led to the modulation tumor acidic microenvironment, the induced tumor cells apoptosis and suppression of T cells to regulatory T cells differentiation.

Comprehensive mutanome analysis of Lewis lung cancer reveals immunogenic neoantigens for therapeutic vaccines

Personalized neoantigen vaccines are capable of eliciting vigorous T-cell responses and have been demonstrated to achieve striking therapeutic effects against cancer. Here we performed comprehensive mutanome analysis of the mouse Lewis lung cancer cells to identify tumor neoantigens followed by prediction of their MHC affinity and immunogenicity. We adopted a strategy that enables us to select neoantigens that were predicted to have high affinity to both MHC I and MHC II. Ten neoantigens were selected to synthesize peptide vaccines and tested in vivo for immunogenicity. Four neoantigen peptide vaccines were found to elicit robust immune reactivity and were further examined for tumor inhibition in mice with xenografted LLC tumors. Two neoantigen peptide vaccines showed significant inhibition on tumor growth and prolonged the survival of tumor-bearing mice. Our studies explored the neoantigen peptide vaccines to treat lung cancer and provide rationale for the optimization of tumor neoantigen selection for therapeutic vaccines.

Folate receptor-targeted RNAi nanoparticles for silencing STAT3 in tumor-associated macrophages and tumor cells

We developed a STAT3 silencing siRNA to both tumor cells and M2 macrophages. The dual-targeting system prepared by electronic self-assembly was composed of folic acid-conjugated carboxymethyl chitosan for targeting and cationic chitosan derivatives for siRNA package. The effects of siRNA delivery was investigated in M2 macrophages and Lewis lung cancer cells (LLC). Due to the enhanced delivery efficiency, the dual-targeting delivery system exhibited a higher efficacy compared with non-targeting nanoparticles, resulting in a dramatically reduction of STAT3 expression in both cells, and a successful shift from M2 phenotypes (pro-tumor) to M1 phenotypes (anti-tumor) for macrophages. Additionally, the influence of the nanoparticles on LLC cells co-cultured with M2 macrophages was also investigated. The increased apoptosis and inhibition of proliferation of LLC cells were observed. In vivo therapeutic effect was also evaluated in s.c. tumor models, tumor growth was effectively inhibited and the level of M2 macrophages in tumor tissues was dramatically reduced.

Discovery of novel hydroxyamidine derivatives as indoleamine 2,3-dioxygenase 1 inhibitors with in vivo anti-tumor efficacy

Indoleamine 2,3-dioxygenase 1 (IDO1) is closely associated with immune escape in many tumor tissues, and is considered to be a valuable therapeutic target in cancer immunotherapy. In this study, the modification of amino sidechain was performed with the hydroxyamidine core kept intact to optimize lead compound Epacadostat. 19 new compounds with hydrazide, thietane or sulfonamide moiety as polar capping group in sidechain were prepared and their IDO1 inhibitory activities were evaluated. Sulfonamide 3a showed potent IDO1 inhibition in both enzymatic and cellular assays with the IC50 value of 71 nM and EC50 value of 11 nM, respectively. Furthermore, in vivo Lewis lung cancer (LLC) allograft studies of 3a indicated that it handicapped the tumor growth with similar efficacy to Epacadostat. Molecular docking demonstrated that the change of polar capping group affords influence on the orientation of amino ethylene side chain and forms new hydrogen bonding.

Low Dose Cyclophosphamide Modulates Tumor Microenvironment by TGF-β Signaling Pathway.

The tumor microenvironment has been recently recognized as a critical contributor to cancer progression and anticancer therapy-resistance. Cyclophosphamide (CTX) is a cytotoxic agent commonly used in clinics for the treatment of cancer. Previous reports demonstrated that CTX given at low continuous doses, known as metronomic schedule, mainly targets endothelial cells and circulating Tregs with unknown mechanisms. Here, we investigated the antitumor activity of two different metronomic schedules of CTX along with their corresponding MTD regimen and further explored their effect on immune function and tumor microenvironment. Toxicity evaluation was monitored by overall survival rate, weight loss, and histopathological analysis. A nude mouse model of Lewis lung cancer was established to assess the anti-metastatic effects of CTX in vivo. CD4+, CD8+, and CD4+CD25+FoxP3 T cells were selected by flow cytometry analysis. Low and continuous administration of CTX was able to restore immune function via increase of CD4+/CD8+ T cells and depletion of T regulatory cells, not only in circulatory and splenic compartments, but also at the tumor site. Low-dose CTX also reduced myofibroblasts, accompanied with an increased level of E-cadherin and low N-cadherin, both in the primary tumor and lung through the TGF-β pathway by the downregulated expression of TGF-β receptor 2. Our data may indicate that several other molecular mechanisms of CTX for tumor may be involved in metronomic chemotherapy, besides targeting angiogenesis and regulatory T cells.

EGFR E746-A750 deletion in lung cancer represses antitumor immunity through the exosome-mediated inhibition of dendritic cells.

EGFR-mutant lung cancer (LC) patients display a poor response to PD-1/PD-L1 blockade. In the absence of independent genetic validation, whether EGFR mutation distorts host antitumor immunity is unknown. Here, we showed that in the clinic, LC with the E746-A750 deletion mutation (EGFR-19del) displayed a temporal association with the loss of intratumoral CD8+ T cells. In a xenograft model, EGFR-19del-expressing Lewis lung cancer (LLC) tumors had a low T cell density at the early stage of tumor development, along with dendritic cells (DCs) exhibiting variant phenotypes in the tumors and draining lymph nodes (LNs). Importantly, EGFR-19del DCs were observed in the LNs of tumor-bearing mice and LC patients. The proliferative activity of T cells within the LN was significantly dampened. In vitro experiments indicated that the function of DCs was repressed by EGFR-19del LLC cells through exosome uptake in which exosomes derived from the EGFR-19del LLC cells could efficiently transfer active EGFR-19del to the surface of the DCs. Injection of EGFR-19del tumor-derived exosomes promoted LLC tumor progression and induced immunosuppression. The combination of gefitinib and GM-CSF treatment recovered tumor T cell infiltration in EGFR-19del tumors by rescuing the function of DCs and increasing the efficacy of anti-PD-L1 treatment. Together, these results indicated that LC with the EGFR E746-A750 deletion mutation induced anergic DCs to repress antitumor immunity through exosomes.

Increased levels of myeloid-derived suppressor cells in esophageal cancer patients is associated with the complication of sepsis

BackgroundTo evaluate whether the level of myeloid-derived suppressor cells is related to the complication of sepsis after esophageal cancer surgery and whether changing the myeloid-derived suppressor cells levels can improve the prognosis of patients cancer-related sepsis. MethodsA total of 178 esophageal cancer patients from Harbin Medical University Cancer Hospital were included in this study. Blood samples were taken from the patients for the analysis of the levels of G-MDSCs and M-MDSCs by flow cytometry. The conditions of the patients was recorded. Male C57BL/6 mice were implanted with Lewis lung cancer cells (2 × 106/mice) by subcutaneous injection into the iliac fossa. Three weeks later, we performed CLP in the mice. All-trans-retinoic acid (ATRA) was intraperitoneally injected at 20 mg/kg, and the control group was injected with 0.9 % NS. We observed the mortality of the mice with cancer-related sepsis. ResultsIn all, 95 % of the esophageal cancer patients had a high level of G-MDSCs (>50 %). A high level of G-MDSCs (>82.5 %) can lead to high morbidity from sepsis after surgery. The increase in M-MDSCs was suggestive of a poor prognosis in patients with cancer-related sepsis. ATRA can improve the survival of patients with cancer-related sepsis. ConclusionsA high level of G-MDSCs can be used to determine the incidence of sepsis in preoperative esophageal cancer patients, M-MDSCs might be effective prognostic indicators for cancer-sepsis patients, and changing the MDSC levels can improve the mortality of patients with cancer-related sepsis.

Molecular hybrid design, synthesis, in vitro and in vivo anticancer evaluation, and mechanism of action of N-acylhydrazone linked, heterobivalent β-carbolines.

A series of N-acylhydrazone-linked, heterobivalent β-carboline derivatives was designed and synthesized from l-tryptophan in a nine-step reaction sequence. The effort resulted in the heterobivalent β-carbolines 10a-t in good yields. The target compounds were characterized by 1H NMR, 13C NMR and high-resolution mass spectrometry (HRMS). The in vitro cytotoxic activity of the synthesized compounds was evaluated against normal EA.HY926 cells and five cancer cell lines: LLC (Lewis lung carcinoma), BGC-823 (gastric carcinoma), CT-26 (murine colon carcinoma), Bel-7402 (liver carcinoma), and MCF-7 (breast carcinoma). Compound 10e, with an IC50 value of 2.41μM against EA.HY926 cells, was the most potent inhibitor. It showed cytotoxicity against all five cancer cell lines of different origin - murine and human, with IC50 values ranging from 4.2±0.7 to 18.5±3.1μM. A study of structure-activity relationships indicated that the influence on cytotoxic activities of the substituent in the R9'-position followed the tendency, 2,3,4,5,6-perfluorophenylmethyl>4-fluorobenzyl>3-phenylpropyl group. The antitumor efficacies of the selected compounds were also evaluated in mice. Compound 10e exhibited potent antitumor activity, with tumor inhibition of more than 40% for Sarcoma 180 and 36.7% for Lewis lung cancer. Furthermore, the pharmacological mechanisms showed that compound 10e has a certain impairment in the motility of LLC cells, which suggests the anti-metastatic potential. And compound 10e inhibited angiogenesis in chicken chorioallantoic membrane assay, and the anti-angiogenetic potency was more potent than the reference drug combretastatin A4-phosphate (CA4P) at a concentration 50μM.

Preparation of Cholesteryl-Modified Aminated Pullulan Nanoparticles to Evaluate Nanoparticle of Hydrophobic Degree on Drug Release and Cytotoxicity

The formation of nanoparticles (NPs) and surface properties such as size and charge are affected by the amphiphilic property of polymer, which is vital for evaluating their function. Here, we synthesized cholesteryl-modified aminated pullulan polymers (CHPNs) with different amounts of cholesterol succinate (CHS). We characterized the three hydrophobically modified polymers (CHPN1, CHPN2, and CHPN3) (CHS: Pu‐NH2=1/5,2/5,3/5) by Fourier transform infrared spectrometry. Dynamic light scattering (DLS) was used to measure particle size and zeta potential of CHPN NPs. The particle sizes of the three NPs CHPN1, CHPN2, and CHPN3 were 178.0, 144.4, and 97.8 nm, respectively. The particle size was related to the cholesteryl substitution of polymers to a certain extent: the stronger the hydrophobicity, the smaller the particle size. In 48 h, the drug release for CHPN3 and CHPN1 NPs was 57.8% and 72.7%. Thus, the NPs showed good sustained drug release: the greater the degree of hydrophobic substitution, the better the sustained release. The cytotoxicity findings were reversed: CHPN1 NPs, with low hydrophobic substitution, showed the best inhibition of Lewis lung cancer cells.

Human umbilical cord mesenchymal stem cells suppress lung cancer via TLR4/NF-κB signalling pathway

The present study investigated the inhibitory effect of human umbilical cord mesenchymal stem cells (hucMSCs) on the growth of lung cancer in mice and its relationship with the TLR4/NF-κB signalling pathway. A mouse model of Lewis lung cancer was constructed by inoculating mice with Lewis lung cancer cell suspension under the axilla. Twenty mice were randomly and evenly divided into experimental group and control group. The experimental group received a caudal vein injection of 1 × 106 hucMSCs each week for two consecutive weeks, and the control group, an equal amount of saline at the same time. Two weeks after treatment with hucMSCs, the mice were sacrificed and the tumour volume and tumour weight were recorded. Enzyme-linked immunosorbent assay was used to determine the IL-1, IL-6 and TNF-α contents in lung cancer tissues; real-time polymerase chain reaction, to measure the level of mRNA; and Western blotting, to determine the protein expression. The results showed that injection of hucMSCs inhibited the growth of Lewis lung cancer tissues, reduced Lewis lung cancer tissue weight, enhanced thymus index, and suppressed inflammatory responses. hucMSCs inhibited the expression of inflammatory factors IL-1, IL-6 and TNF-α by activating TLR4/NF-κB signalling pathway, and facilitated the expression of Bax. Moreover, huMSCs induced the expression of apoptogenic factor Bax by activating TLR4/NF-κB signalling pathway, and inhibited the expression of antiapoptotic factor bcl-2. The present study demonstrates that hucMSCs reduce inflammatory responses by mediating TLR4/NF-κB signalling pathway, promote apoptosis of tumour cells, and inhibit tumour proliferation in mice with Lewis lung cancer.

Ginsenoside 3β-O-Glc-DM (C3DM) enhances the antitumor activity of Taxol on Lewis lung cancer by targeting the interleukin-6/Jak2/STAT3 and interleukin-6/AKT signaling pathways

Objective: Nonsmall-cell lung cancer (NSCLC) is an aggressive, highly chemoresistant disease. Taxol is an effective chemotherapeutic drug widely used for the treatment of NSCLC. However, the clinical use of Taxol is limited due to the occurrence of adverse side effects under high therapeutic doses. Therefore, it is desirable to explore combination therapy to reduce the dose of chemotherapeutic drugs and achieve excellent outcomes. A biosynthetic ginsenoside, 3-O-β-D-glucopyranosyl-dammar-24-ene-3β, 20S-diol (3β-O-Glc-DM, C3DM) is obtained from microbial fermentation by metabolic engineering. Based on previous study findings, we aimed to explore the mechanism of combination therapy with C3DM and Taxol and its increasing antitumor effect on Lewis lung cancer (LLC) in this study. Materials and Methods: A thiazolyl blue tetrazolium bromide (MTT) assay was performed to evaluate cell viability; the apoptotic effect was studied using cell apoptosis assay. The Lewis tumor xenograft experiment was performed to determine the effects of C3DM combined with Taxol on tumor growth in vivo, and western blotting was performed to analyze protein expressions. Results: C3DM effectively inhibited the proliferation of NSCLC cells. Moreover, C3DM increased the antiproliferative activity of Taxol and significantly enhanced cell apoptosis induced by Taxol in Lewis lung cancer cells. C3DM alone also suppressed Lewis tumor growth and enhanced the antitumor activity of Taxol in vivo. Western blot analysis revealed that the effects of the antiproliferation and apoptosis induction of C3DM treatment alone or in combination with Taxol on Lewis lung cancer were mediated by inhibiting the interleukin-6 (IL-6)/Jak2/STAT3 and IL-6/AKT signaling pathways. Conclusions: The results showed that C3DM has the potential to be used in combination therapy with Taxol against NSCLC.

Intratumoral injection of anlotinib hydrogel enhances antitumor effects and reduces toxicity in mouse model of lung cancer

This study was conducted to determine the antitumor effects and ability of an anlotinib (AL) hydrogel (AL–HA–Tyr) to reduce toxicity in a mouse model of Lewis lung cancer (LLC). We constructed a drug carrier system for AL, verified its effectiveness and systemic safety, and provided a preliminary experimental foundation for clinical carrier transformation. AL–HA–Tyr was prepared by encapsulating AL with hyaluronic acid–tyramine (HA–Tyr) conjugates. Colony and tube formation assays showed that AL–HA–Tyr restrained the proliferation of human umbilical vein endothelial cells (HUVECs) and LLC cells, respectively, in vitro, and that AL exerted significant anti-angiogenesis and anti-tumor effects. The invasion and migration of HUVECs and LLC cells were efficiently suppressed by AL according to transwell assays. HUVEC and LLC cell-cycle and apoptosis analysis clarified the direct anti-tumor effects of AL–HA–Tyr. Mice engrafted with LLC cells in vivo were administered oral saline, oral AL, or an intratumoral injection of HA–Tyr or AL–HA–Tyr. The results showed that AL–HA–Tyr obviously reduced visceral toxicity and decreased Ki67 and VEGF-A expression in tumor cells compared with AL. Furthermore, AL–HA–Tyr significantly prolonged the survival of tumor-bearing mice. Overall, AL–HA–Tyr enhanced antitumor effects and reduced toxicity in the LLC model. It provided a foundation for the clinical transformation of drug carrier systems.

养阴扶正汤调节Lewis肺癌荷瘤鼠T细胞耗竭的实验研究

养阴扶正汤调节Lewis肺癌荷瘤鼠T细胞耗竭的实验研究

The Combinatorial Effect of Cisplatin and Moxibustion on Tumor Growth Inhibition with Special Reference to Modulation of the Immune Microenvironment in Lewis Lung Cancer Mice.

Objective As a first-line treatment for non-small cell lung cancer (NSCLC), the efficacy of chemotherapy is still unsatisfactory. Moxibustion has been shown to improve the side effects of radiotherapy and chemotherapy and regulate immune function. This study aimed to explore the antitumor effects and potential mechanisms of combinatorial cisplatin and moxibustion treatment for NSCLC by targeting the tumor microenvironment. Methods Lewis lung cancer (LLC)-bearing mice were induced and treated with cisplatin or/and moxibustion at ST36 (Zusanli), and the growth, weight, and area of the tumor were evaluated. The numbers of various T cell subsets and myeloid cells in the tumor were assessed by flow cytometry, and the gene expression of related markers and cytokines was detected with real-time quantitative polymerase chain reaction (RT-qPCR). In addition, the tumor vascular structure was investigated using CD31 and α-smooth muscle actin (α-SMA) immunofluorescence staining. The expression of the vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) was detected by immunohistochemical staining. Results Both cisplatin and moxibustion inhibited LLC tumor growth and reduced both the tumor area and weight, with the combinatorial therapy showing superior outcomes. Moxibustion upregulated the infiltration of CD4+ T cells and Th1 cells in the tumor, and the combinatorial therapy increased the proportion of CD8+ cytotoxic T cells (CTLs), CD4+T cells, Th1, Th9 cells, and M1 macrophages, as well as the expression of Cd69, Ifng, and Cd86 mRNA. The combinatorial therapy improved vascular normalization by increasing both the microvessel density (MVD) and pericyte coverage (α-SMA area density) and inhibiting the expression of the VEGF. Conclusions Combinatorial cisplatin and moxibustion treatment inhibited the LLC tumor growth by mechanisms related to the improvement of the tumor immune microenvironment and vascular normalization, providing an effective combinatorial therapy beneficial for patients with NSCLC.