Antitumor effect of pharmacologic ascorbate in the B16 murine melanoma model

Because 5-year survival rates for patients with metastatic melanoma remain below 25%, there is continued need for new therapeutic approaches. For some tumors, pharmacologic ascorbate treatment may have a beneficial antitumor effect and may work synergistically with standard chemotherapeutics. To investigate this possibility in melanoma, we examined the effect of pharmacologic ascorbate on B16-F10 cells. Murine models were employed to compare tumor size following treatment with ascorbate, and the chemotherapeutic agents dacarbazine or valproic acid, alone or in combination with ascorbate. Results indicated that nearly all melanoma cell lines were susceptible to ascorbate-mediated cytotoxicity. Compared to saline controls, pharmacologic ascorbate decreased tumor size in both C57BL/6 (P<0.0001) and NOD-scid tumor bearing mice (P<0.0001). Pharmacologic ascorbate was superior or equivalent to dacarbazine as an antitumor agent. Synergy was not apparent when ascorbate was combined with either dacarbazine or valproic acid; the latter combination may have additional toxicities. Pharmacologic ascorbate induced DNA damage in melanoma cells, as evidenced by increased phosphorylation of the histone variant, H2A.X. Differences were not evident in tumor samples from C57BL/6 mice treated with pharmacologic ascorbate compared to tumors from saline-treated controls. Together, these results suggest that pharmacologic ascorbate has a cytotoxic effect against melanoma that is largely independent of lymphocytic immune functions and that continued investigation of pharmacologic ascorbate in cancer treatment is warranted.     

The effect of high intensity focused ultrasound treatment on metastases in a murine melanoma model

This study aims to assess the risk of high intensity focused ultrasound (HIFU) therapy on the incidence of distant metastases and to investigate its association with HIFU-elicited anti-tumor immunity in a murine melanoma (B16-F10) model. Tumor-bearing legs were amputated immediately after or 2 days following HIFU treatment to differentiate the contribution of the elicited anti-tumor immunity. In mice undergoing amputation immediately after mechanical, thermal, or no HIFU treatment, metastasis rates were comparable (18.8%, 13.3%, and 12.5%). In contrast, with a 2-day delay in amputation, the corresponding metastasis rates were 6.7%, 11.8%, and 40%, respectively. Animal survival rate was higher and CTL activity was enhanced in the HIFU treatment groups. Altogether, our results suggest that HIFU treatment does not increase the risk of distant metastasis. Instead, HIFU treatment can elicit an anti-tumor immune response that may be harnessed to improve the overall effectiveness and quality of cancer therapy.     

Tumour-stromal interactions in acid-mediated invasion: A mathematical model

It is well established that the tumour microenvironment can both promote and suppress tumour growth and invasion, however, most mathematical models of invasion view the normal tissue as inhibiting tumour progression via immune modulation or spatial constraint. In particular, the production of acid by tumour cells and the subsequent creation of a low extracellular pH environment has been explored in several ‘acid-mediated tumour invasion’ models where the acidic environment facilitates normal cell death and permits tumour invasion. In this paper, we extend the acid-invasion model developed by Gatenby and Gawlinski (1996) to include both the competitive and cooperative interactions between tumour and normal cells, by incorporating the influence of extracellular matrix and protease production at the tumour-stroma interface. Our model predicts an optimal level of tumour acidity which produces both cell death and matrix degradation. Additionally, very aggressive tumours prevent protease production and matrix degradation by excessive normal cell destruction, leading to an acellular (but matrix filled) gap between the tumour and normal tissue, a feature seen in encapsulated tumours. These results sugest, counterintuitively, that increasing tumour acidity may, in some cases, prevent tumour invasion.     

Antitumor effect of the streptococcal preparation OK-432 in a murine model of ovarian cancer

Summary The antitumor effects of the streptococcal preparation OK-432 were analyzed in a murine ovarian teratocarcinoma (MOT) model. Administration of OK-432 i.p. prevented tumor outgrowth in 75% of mice challenged with 10³ MOT cells i.p. 24 h previously. Treatment was less successful in mice challenged with 10⁴ or 10⁵ cells, preventing tumor growth in 25% of the former and only 5% of the latter group. Tumor-challenged mice cured by injections of OK-432 were not rendered resistant to a subsequent challenge with 10³ MOT cells 75 days after initial treatment. Only the i.p. route of administration was effective as i.v. OK-432 did not prolong survival of tumor-challenged mice. An antitumor response was detected as early as 24 h after i.p. treatment. This correlated temporally with an influx of neutrophils into the peritoneal cavity. Peritoneal cells obtained between 6 and 24 h after treatment were capable of lysing MOT targets in vitro. A single cell cytotoxicity assay demonstrated that peritoneal neutrophils, elicited by i.p. injection of OK-432, could bind to and lyse MOT targets. These data indicate that OK-432 is effective against small tumor cell inocula in this murine model of ovarian cancer and, furthermore, that the neutrophilic response into the peritoneal cavity plays a role in tumor rejection.     

The chick embryo chorioallantoic membrane as a model for tumor biology

Among the in vivo models, the chick embryo chorioallantoic membrane (CAM) has been used to implant several tumor types as well as malignant cell lines to study their growth rate, angiogenic potential and metastatic capability. This review article is focused on the major compelling literature data on the use of the CAM to investigate tumor growth and the metastatic process.     

Apatinib-loaded nanoparticles inhibit tumor growth and angiogenesis in a model of melanoma

Anti-angiogenic drugs are an effective therapeutic method for the treatment of melanomas. Apatinib is a small-molecule tyrosine kinase inhibitor, which has potent inhibitory activity on tumor angiogenesis. Due to the low water solubility and stability of Apatinib, we aimed to design and develop poly (lactic-co-glycolic acid) (PLGA) and Poloxamer 407 nanoparticles to encapsulate Apatinib (Apa/p NPs) to improve the efficacy of application in melanoma treatment. The size and morphology of the nanoparticles were characterized by dynamic light scattering (DLS) and transmission electron microscopy (TEM). In vitro proliferation assays were used to assess the capacity of Apa/p NPs to suppress the growth of B16 cells. Furthermore, we constructed melanoma models using C57BL/6 mice, and preliminary evaluation of the effect and mechanism of Apa/p NPs on tumor inhibition was performed in vivo. The results showed that the size of Apa/p NPs averaged 136 ± 0.27 nm and the nanoparticles were evenly dispersed. Moreover, Apa/p NPs significantly inhibited the growth of B16 cells and melanoma tumors, compared with the naked drug treatment and control groups. The protein levels of VEGFR-2, phosphorylated (p)-VEGFR-2 and p-ERK1/2 in tumor tissues were inhibited by Apa/p NP treatment, as detected by Western blot. The results of this study suggested that Apa/p NPs could inhibit the growth of melanoma tumors by inhibiting the phosphorylation and expression of VEGFR-2 and downstream ERK1/2, providing a theoretical basis for the clinical application of Apatinib in the treatment of melanoma.     

Galectin-9 in tumor biology: A jack of multiple trades

Galectin family members have been shown to exert multiple roles in the context of tumor biology. Several recent findings support a similar multi-faceted role for galectin-9. Galectin-9 expression is frequently altered in cancer as compared to normal tissues. In addition, an increasing amount of evidence suggests that galectin-9 is involved in several aspects of tumor progression, including tumor cell adhesion and survival, immune escape and angiogenesis. Also, galectin-9 shows potential as a prognostic marker and a therapeutic target for several malignancies. In this review we summarize both the established and the emerging roles of galectin-9 in tumor biology and discuss the potential application of galectin-9 in anti-cancer therapy.     

Accumulation of nano-sized particles in a murine model of angiogenesis

Purpose

To evaluate the ability of nm-scaled iron oxide particles conjugated with Azure A, a classic histological dye, to accumulate in areas of angiogenesis in a recently developed murine angiogenesis model.

Materials and methods

We characterised the Azure A particles with regard to their hydrodynamic size, zeta potential, and blood circulation half-life. The particles were then investigated by Magnetic Resonance Imaging (MRI) in a recently developed murine angiogenesis model along with reference particles (Ferumoxtran-10) and saline injections.

Results

The Azure A particles had a mean hydrodynamic diameter of 51.8 ± 43.2 nm, a zeta potential of −17.2 ± 2.8 mV, and a blood circulation half-life of 127.8 ± 74.7 min. Comparison of MR images taken pre- and 24-h post-injection revealed a significant increase in R₂^* relaxation rates for both Azure A and Ferumoxtran-10 particles. No significant difference was found for the saline injections. The relative increase was calculated for the three groups, and showed a significant difference between the saline group and the Azure A group, and between the saline group and the Ferumoxtran-10 group. However, no significant difference was found between the two particle groups.

Conclusion

Ultrahigh-field MRI revealed localisation of both types of iron oxide particles to areas of neovasculature. However, the Azure A particles did not show any enhanced accumulation relative to Ferumoxtran-10, suggesting the accumulation in both cases to be passive.     

Antitumor effects of combined therapy of recombinant heat shock protein 70 and hyperthermia using magnetic nanoparticles in an experimental subcutaneous murine melanoma

Heat shock proteins (HSPs) are recognized as significant participants in cancer immunity. We previously reported that HSP70 expression following hyperthermia using magnetic nanoparticles induces antitumor immunity. In the present study, we examine whether the antitumor immunity induced by hyperthermia is enhanced by administration of recombinant HSP70 protein into the tumor in situ. Hyperthermia was conducted using our original magnetite cationic liposomes (MCLs), which have a positive surface charge and generate heat in an alternating magnetic field (AMF) due to hysteresis loss. MCLs and recombinant mouse HSP70 (rmHSP70) were injected into melanoma nodules in C57BL/6 mice, which were subjected to AMF for 30 min. Temperature within the tumor reached 43°C and was maintained by controlling the magnetic field intensity. The combined treatment strongly inhibited tumor growth over a 30-day period and complete regression of tumors was observed in 20% (2/10) of mice. It was also found that systemic antitumor immunity was induced in the cured mice. This study suggests that novel combined therapy using exogenous HSP70 and hyperthermia has great potential in cancer treatment.     

Analysis of HLA class I expression in progressing and regressing metastatic melanoma lesions after immunotherapy

Despite the potential efficacy of cancer immunotherapy in preclinical studies, it did not show yet significant positive clinical results in humans with only a small number of cancer patients demonstrating objective tumor regression. This poor clinical outcome can be explained by the generation of sophisticated tumor immune escape mechanism, in particular, abnormalities in the expression of HLA class I antigens. We have studied the expression of HLA class I antigens in ten metastatic lesions obtained from a melanoma patient undergoing immunotherapy. Five lesions were obtained after Interferon-alpha-2b treatment and five after autologous vaccination plus BCG (M-VAX). Eight metastases were regressing after immunotherapy while two were progressing. The eight regressing metastases showed high level of HLA class I expression, whereas the two progressing lesions had low levels as measured by real time PCR and immunohistological techniques. These results indicate a strong association between HLA class I expression and progression or regression of the metastatic lesions. Our data support the hypothesis that the level of HLA class I expression is an important parameter of tumor immune escape that needs to be monitored.     

Antitumor effect of intratumoral administration of dendritic cell combination with vincristine chemotherapy in a murine fibrosarcoma model

A new antitumor therapeutic strategy utilizing the combined effect of chemotherapy and DC (dendritic cell)-based immunotherapy was designed, and the effect of intratumoral injections of unpulsed, immature DCs was evaluated after in vivo pretreatment of vincristine on tumor growth in a murine fibrosarcoma tumor model. Vincristine exerted a much more potent apoptosis/necrosis-inducing effect on MCA-102 tumor cells than on DCs both in vitro and in vivo. Moreover, CD11c, CD40, CD80 and CD86 molecules on DCs were not downregulated after treatment with vincristine either in vitro or in vivo. The growth of tumor significantly regressed in the group which received the combined vincristine chemotherapy with intratumoral administration of DCs in contrast to the untreated group, the group treated with DCs alone, and the group treated with vincristine alone. In particular, an upregulated expression of CD40, CD80 and CD86 molecules on DCs was found in the combination treatment group. Furthermore, the number of CD4+ and CD8+ T cells and the staining intensity of their CD4 and CD8 surface molecules also increased after the combination treatment. Therefore, our results indicate the feasibility of this combination therapy with vincristine chemotherapy and DC-based immunotherapy as an efficient antitumor strategy for the treatment of fibrosarcoma.     

HLA class I expression in metastatic melanoma correlates with tumor development during autologous vaccination

Our knowledge of the mechanisms underlying tumor-specific immune response and tumor escape has considerably increased. HLA class I antigen defects remain an important tumor escape mechanism since they influence the interactions between tumor cells and specific T and NK cells in the course of malignant disease. We have studied here HLA class I expression in six subcutaneous metastases obtained from a melanoma patient immunized with an autologous melanoma cell vaccine (M-VAX). We report in this paper that HLA class I antigen expression on these metastatic lesions strongly correlated with the course of the disease. The three metastases that were partially regressing at the time of their excision showed a strong HLA class I expression, whereas the progressing ones showed a very weak or negative staining with most of the anti-HLA class I mAbs used. Real-time quantitative PCR of the samples obtained from microdissected tumor tissue revealed a significant difference in the mRNA levels of HLA-ABC heavy chain and beta2m between the two types of metastases, i.e., lower levels in progressing metastases and high levels in regressing ones, confirming the immunohistological findings. This is, to our knowledge, the first report where the clinical outcome of different HLA class I positive and negative melanoma metastases can be clearly correlated with the regression and progression of the disease, respectively.     

The dynamic evolution of the power exponent in a universal growth model of tumors

We have previously reported that a universal growth law, as proposed by West and collaborators for all living organisms, appears to be able to describe also the growth of tumors in vivo after an initial exponential growth phase. In contrast to the assumption of a fixed power exponent p (assumed by West et al. to be equal to 3/4), we propose in this paper a dynamic evolution of p, using experimental data from the cancer literature. In analogy with results obtained by applying scaling laws to the study of fragmentation of solids, the dynamic behaviour of p is related to the evolution of the fractal topology of neoplastic vascular systems. Our model might be applied for diagnostic purposes to mark the emergence of an efficient neo-angiogenetic structure if the results of our in silico experiments are confirmed by clinical observations.     

The multifaceted role of CD146/MCAM in the promotion of melanoma progression

Human malignant melanoma is a common primary malignant cutaneous tumour derived from transformed epidermal melanocytes. Patients with melanoma have a high rate of mortality due to resistance to chemotherapeutic drugs, a major obstacle to a successful treatment. Several reports have suggested that CD146 plays an important role as a signalling molecule in human melanoma. This role includes CD146 as a participant in inflammation, differentiation, adhesion, tumourigenicity, metastasis, invasion and angiogenesis among other processes, which suggests that this molecule promotes the progression of human melanoma as a multifaceted regulator. In this article, we explore the effects and corresponding mechanisms with respect to the role of CD146/MUC18 in the promotion of human melanoma progression. Collectively, the studies indicated that targeting CD146, because it is a suitable marker of poor patient outcome, might be useful in the design of future strategies for the prevention and treatment of human melanoma.     

Knockdown of Atg7 suppresses Tumorigenesis in a murine model of liver cancer

Hepatocellular Carcinoma (HCC) is the most common type of primary liver cancer in adults and a leading cause of cancer-related deaths worldwide. Studies have shown that autophagy is significantly involved in carcinogenesis, in particular, driven by activated RAS signaling. Autophagy related 7 (Atg7) is a critical component for the formation of autophagosome and required for autophagy processes. We investigated the role of autophagy in RAS-driven tumorigenesis in the liver, via the knockdown of Atg7 in the model. Transposon vectors encoding short hairpin RNAs targeting Atg7 (Atg7 shRNA) were constructed. Inhibition of autophagy via Atg7 knockdown was tested in Hep3B cells cultured in nutrient-starved medium. Formation of autophagosome was suppressed in nutrient-starved Hep3B cells expressing Atg7 shRNA, demonstrating that it efficiently inhibited autophagy in HCC cells. Transposons encoding Atg7 shRNA were mixed with those expressing HRAS^G12V and p53 shRNA, and subsequently used for hydrodynamic injection to 5-week-old C57BL/6 mice. Tumorigenesis in livers induced by HRAS^G12V and p53 shRNA was significantly suppressed by Atg7 knockdown. The inhibition of autophagy led to a decreased proliferation of cancer cells, as determined by Ki-67 staining. Our data indicate that knockdown of Atg7 led to a significant decrease in tumorigenesis in a murine HCC model induced by activated RAS. Inhibition of autophagosome formation is expected to be a therapeutic option for liver cancer.     

Tumor invasion and metastasis in Drosophila: a bold past, a bright future

Invasion and metastasis are the most deadly hallmarks of cancer.Once a cancer has acquired the ability to colonize new sites in the body it becomes dramatically more difficult to treat.This has made it a focus of much of cancer research.The humble fruit fly,Drosophila melanogaster,has despite its relative simplicity,made significant contributions to the understanding of tumor progression.In this review we outline and highlight those with an emphasis on modeling the genetic and epigenetic changes required for invasion and metastasis.We will revisit the early years of cancer modeling in Drosophila where the first parallels were drawn between Drosophila and vertebrate neoplasms and highlight recent advances using genetic screens and interactions with the epithelial microenvironment and innate immune system.We focus on the power and limitations of current fly models of metastasis.     

The CXCL8-CXCR1/2 pathways in cancer

Persistent infection or chronic inflammation contributes significantly to tumourigenesis and tumour progression. C-X-C motif ligand 8 (CXCL8) is a chemokine that acts as an important multifunctional cytokine to modulate tumour proliferation, invasion and migration in an autocrine or paracrine manner. Studies have suggested that CXCL8 and its cognate receptors, C-X-C chemokine receptor 1 (CXCR1) and C-X-C chemokine receptor 2 (CXCR2), mediate the initiation and development of various cancers including breast cancer, prostate cancer, lung cancer, colorectal carcinoma and melanoma. CXCL8 also integrates with multiple intracellular signalling pathways to produce coordinated effects. Neovascularisation, which provides a basis for fostering tumour growth and metastasis, is now recognised as a critical function of CXCL8 in the tumour microenvironment. In this review, we summarize the biological functions and clinical significance of the CXCL8 signalling axis in cancer. We also propose that CXCL8 may be a potential therapeutic target for cancer treatment.     

Antitumor effect of malaria parasite infection in a murine Lewis lung cancer model through induction of innate and adaptive immunity

Background

Lung cancer is the most common malignancy in humans and its high fatality means that no effective treatment is available. Developing new therapeutic strategies for lung cancer is urgently needed. Malaria has been reported to stimulate host immune responses, which are believed to be efficacious for combating some clinical cancers. This study is aimed to provide evidence that malaria parasite infection is therapeutic for lung cancer.

Methodology/Principal Findings

Antitumor effect of malaria infection was examined in both subcutaneously and intravenously implanted murine Lewis lung cancer (LLC) model. The results showed that malaria infection inhibited LLC growth and metastasis and prolonged the survival of tumor-bearing mice. Histological analysis of tumors from mice infected with malaria revealed that angiogenesis was inhibited, which correlated with increased terminal deoxynucleotidyl transferase-mediated (TUNEL) staining and decreased Ki-67 expression in tumors. Through natural killer (NK) cell cytotoxicity activity, cytokine assays, enzyme-linked immunospot assay, lymphocyte proliferation, and flow cytometry, we demonstrated that malaria infection provided anti-tumor effects by inducing both a potent anti-tumor innate immune response, including the secretion of IFN-γ and TNF-α and the activation of NK cells as well as adaptive anti-tumor immunity with increasing tumor-specific T-cell proliferation and cytolytic activity of CD8⁺ T cells. Notably, tumor-bearing mice infected with the parasite developed long-lasting and effective tumor-specific immunity. Consequently, we found that malaria parasite infection could enhance the immune response of lung cancer DNA vaccine pcDNA3.1-hMUC1 and the combination produced a synergistic antitumor effect.

Conclusions/Significance

Malaria infection significantly suppresses LLC growth via induction of innate and adaptive antitumor responses in a mouse model. These data suggest that the malaria parasite may provide a novel strategy or therapeutic vaccine vector for anti-lung cancer immune-based therapy.     

Tumor specific phage particles promote tumor regression in a mouse melanoma model

Within cancer research, phage display libraries have been widely used for the identification of tumor targeting peptides and antibodies. Additionally, phages are known to be highly immunogenic; therefore we evaluated the immunotherapeutic potential of tumor specific phages to treat established solid tumors in a mouse model of melanoma. We developed two tumor specific phages, one derived from a peptide phage display library and one Fab expressing phage with known specificity, for the treatment of mice bearing palpable B16-F10 or B16/A2K^b tumors. Therapy in B16-F10 tumor bearing mice with tumor specific phages was superior to treatment with non-tumor specific phages and lead to delayed tumor growth and increased survival. In B16/A2K^b tumor bearing mice, therapy with tumor specific phages resulted in complete tumor regression and long-term survival in 50% of the mice. Histological analysis of tumors undergoing treatment with tumor specific phages revealed that phage administration induced a massive infiltration of polymorphonuclear neutrophils. Furthermore, phages induced secretion of IL-12 (p70) and IFN-γ as measured in mouse splenocyte culture supernatants. These results demonstrate a novel, immunotherapeutic cancer treatment showing that tumor specific phages can promote regression of established tumors by recruitment of inflammatory cells and induction of Th1 cytokines.