mart-1-antigen and Disease-Models--Animal

Uveal melanoma (UM) and conjunctival melanoma (CM) are ocular malignancies that give rise to life-threatening metastases. Although local disease can often be treated successfully, it is often associated with significant vision impairment and treatments are often not effective against metastatic disease. Novel treatment modalities that preserve vision may enable elimination of small tumors and may prevent subsequent metastatic spread. Very few mouse models of metastatic CM and UM are available for research and for development of novel therapies. One of the challenges is to follow tumor growth in-vivo and to determine the right size for treatment, mainly of the posterior, choroidal melanoma. Hence, the purpose of this study was to establish a simple, noninvasive imaging tool that will simplify visualization and tumor follow-up in mouse models of CM and UM. Tumors were induced by inoculation of murine B16LS9 cells into the sub-conjunctival or the choroidal space of a C57BL/6 mouse eye under a surgical microscope. Five to ten days following injection, tumor size was assessed by Phoenix MicronIV™ image-guided Optical Coherence Tomography (OCT) imaging, which included a real-time camera view and OCT scan of the conjunctiva and the retina. In addition, tumor size was evaluated by ultrasound and histopathological examination of eye sections. Tumor growth was observed 5-9 days following sub-conjunctival or sub-retinal injection of seven-thousand or seventy-thousand cells, respectively. A clear tumor mass was detected at these regions using the MicronIV™ imaging system camera and OCT scans. Histology of eye sections confirmed the presence of tumor tissue. OCT allowed an accurate measurement of tumor size in the UM model and a qualitative assessment of tumor size in the CM model. Moreover, OCT enabled assessing the success rate of the choroidal tumor induction and importantly, predicted final tumor size already on the day of cell inoculation. In conclusion, by using a simple, non-invasive imaging tool, we were able to follow intraocular tumor growth of both CM and UM, and to define, already at the time of cell inoculation, a grading scale to evaluate tumor size. This tool may be utilized for evaluation of new mouse models for CM and UM, as well as for testing new therapies for these diseases.

Topics: Animals; Biomarkers, Tumor; Cell Line, Tumor; Conjunctival Neoplasms; Disease Models, Animal; Immunohistochemistry; MART-1 Antigen; Melanoma; Melanoma-Specific Antigens; Mice; Mice, Inbred C57BL; Monophenol Monooxygenase; Neoplasm Proteins; Tomography, Optical Coherence; Ultrasonography; Uveal Neoplasms

Adult wild-type mice are not supposed to be proper models for ultraviolet radiation (UVR)-induced melanoma since melanocytes are confined to hair follicles and cannot be sufficiently reached by UVR. On the other hand, in mutated mouse models used for melanoma research limitations, including an altered immune system and selection of affected pathways, lead to tumors phenotypically quite different from naturally occurring melanomas. We compared the distribution of epidermal melanocytes in UVR and not-UVR-exposed wild-type C57BL/6 mice. Starting at the age of 8 weeks, mice were exposed to physiologic doses of UVR three times weekly over 16 weeks. Back skin biopsies were taken 4, 8, 12 and 16 weeks after initiation of exposure, and stained for Melan-A, representing a highly selective marker for melanocytes. Surprisingly, after exposure to UVR, Melan-A positive cells were detected also in the interfollicular epidermis of C57BL/6 mice. We conclude that UVR is capable of inducing interfollicular epidermal melanocytes in wild-type mice.

Topics: Animals; Biomarkers; Biopsy; Disease Models, Animal; Epidermal Cells; Epidermis; Female; Hair Follicle; Humans; MART-1 Antigen; Melanocytes; Melanoma; Mice; Mice, Inbred C57BL; Ultraviolet Rays

To establish a mouse model with histologic characteristics of uveal melanoma for investigation of intraocular tumor biology of melanoma.. After injection of 1 × 10(5) of HCmel12 melanoma cells, a cutaneous melanoma cell line, into the vitreous of CX3CR1(+/GFP) or C57Bl/6 mice (n = 12), tumor growth patterns, clinicopathological features, angiogenesis and metastatic behavior were analyzed by histology (hematoxylin and eosin, periodic acid-Schiff without hematoxylin) and immunohistochemistry (HMB45/MART-1-Ab, F4/80-Ab, green fluorescent protein (GFP)-Ab and VE-cadherin-Ab).. HCmel12 cells formed intraocularly growing tumor masses, which showed histologic features of intraocular melanoma such as angiotropism, intratumoral endothelial-lined vasculature, vasculogenic mimicry including prognostic significant extravascular matrix patterns, and invasion by inflammatory cells, in particular macrophages. There was no difference in tumor growth characteristics between CX3CR1(+/GFP) and C57Bl/6 mice. Five of 10 mice proceeded to extrascleral tumor growth and three of these developed metastases.. Intraocularly injected HCmel12 cells developed tumor masses with histologic characteristics of aggressive melanoma similar to human uveal melanoma. Since hematogenous dissemination to the liver was not observed, intravitreally injected HCmel12 cells do not qualify as a model for metastasizing intraocular melanoma. However, since the eye represents a semi-closed compartment with access to constant blood supply, these intraocular tumors represent a model for studies of isolated parameters in general tumor biology of intraocular melanoma.

Topics: Animals; Antigens, CD; Antigens, Differentiation; Biomarkers, Tumor; Cadherins; Cell Line, Tumor; Disease Models, Animal; Female; gp100 Melanoma Antigen; Green Fluorescent Proteins; Humans; Intravitreal Injections; Male; MART-1 Antigen; Melanoma; Melanoma-Specific Antigens; Mice; Mice, Inbred C57BL; Neovascularization, Pathologic; Skin Neoplasms; Transplantation, Heterologous; Uveal Neoplasms

Rhododendrol, 4-(4-hydroxyphenyl)-2-butanol, Rhododenol(®) (RD), a naturally occurring phenolic compound, was developed as a tyrosinase inhibitor for skin-lightening/whitening cosmetics. In 2013, skin depigmentation was reported in consumers using RD-containing skin-brightening cosmetics; this condition is called RD-induced leukoderma.. The etiology of RD-induced leukoderma is still largely unknown. Here, to assess the depigmentation potential of RD, we developed a new mouse model of leukoderma by topically applying RD.. Hairless hk14-SCF Tg mice with melanocytes distributed in the epidermis were used for this study. RD was applied on the dorsal skin of the mice daily for 28 days. Then, immunohistological, biochemical, and electron microscopic analyses were performed on biopsy samples taken from these mice.. The depigmentation in the RD-treated sites appeared on Day 14. Histological examination indicated a loss of epidermal melanocytes at Day 7. On the other hand, the melanocyte number did not decrease in the albino mice having the same background as the hairless hk14-SCF Tg, but without tyrosinase activity. Biochemical analyses showed that the eumelanin content decreased in the RD-treated sites and metabolites of RD-quinone, i.e., non-protein thiol adducts and protein-SH adducts, were produced. Electron microscopic analyses revealed double-membrane-walled structures containing electron-dense material, which might be typical for melanin-containing autophagosomes and a dilated endoplasmic reticulum (ER), which would indicate ER stress.. These data suggested that RD exerted tyrosinase-dependent melanocyte cytotoxicity and that tyrosinase-dependent accumulation of ER stress from activation of the autophagy pathway contributed to melanocyte cytotoxicity.

Topics: Administration, Topical; Animals; Asian People; Autophagy; Butanols; Cell Count; Disease Models, Animal; Endoplasmic Reticulum Chaperone BiP; Endoplasmic Reticulum Stress; Enzyme Inhibitors; Epidermal Cells; Epidermis; Heat-Shock Proteins; Humans; MART-1 Antigen; Melanins; Melanocytes; Mice; Mice, Hairless; Mice, Inbred C57BL; Mice, Transgenic; Models, Animal; Monophenol Monooxygenase; Skin Lightening Preparations; Skin Pigmentation; Vitiligo

We hypothesized that the co-entrapment of melanoma-associated antigens and the Toll-like receptor (TLR) ligands Poly(I:C) and CpG, known to be Th1-immunopotentiators, in mannose-functionalized aliphatic polyester-based nanoparticles (NPs) could be targeted to mannose receptors on antigen-presenting cells and induce anti-tumor immune responses. High entrapment efficiencies of antigens and immunopotentiators in 150nm NPs were obtained. The co-entrapment of the model antigen ovalbumin and the TLR ligands was crucial to induce high IgG2c/IgG1 ratios and high levels of IFN-γ and IL-2. Mannose-functionalization of NPs potentiated the Th1 immune response. The nanoparticulate vaccines decreased the growth rate of murine B16F10 melanoma tumors in therapeutic and prophylatic settings. The combination of mannose-functionalized NPs containing MHC class I- or class II-restricted melanoma antigens and the TLR ligands induced the highest tumor growth delay. Overall, we demonstrate that the multifunctional properties of NPs in terms of targeting and antigen/adjuvant delivery have high cancer immunotherapeutic potential.

Topics: Animals; Cancer Vaccines; Cell Line, Tumor; Cytokines; Disease Models, Animal; Female; gp100 Melanoma Antigen; Granzymes; Immunoglobulin G; Ligands; Male; Mannose; MART-1 Antigen; Melanoma; Mice, Inbred C57BL; Mice, Transgenic; Nanoparticles; Oligodeoxyribonucleotides; Ovalbumin; Peptides; Poly I-C; Polymers; Toll-Like Receptors; Tumor Burden

Although rare, uveal melanoma (UM) is the most common primary intraocular tumor in adults. About half of UM patients develop metastatic disease typically in the liver and die within a short period, due to ineffective systemic therapies. UM has unique and distinct genetic features predictive of metastasis. Animal models are required to improve our understanding of therapeutic options in disseminated UM. Since spontaneous murine UM models are lacking, our aim was to analyze the suitability of the established transgenic melanoma mouse model Tg(Grm1) as a new UM model system. We demonstrated that adult Grm1 transgenic mice develop choroidal thickening and uveal melanocytic neoplasia with expression of the melanocytic markers S100B and MelanA. Further, we showed that GRM1 is expressed in human UM, similar to skin melanoma. This study presents a new mouse model for spontaneous UM and suggests that the glutamate signaling pathway is a possible target for UM therapy.

Topics: Animals; Biomarkers, Tumor; Choroid; Choroid Neoplasms; Disease Models, Animal; Fluorescent Antibody Technique, Indirect; Ki-67 Antigen; MART-1 Antigen; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Mice, Transgenic; Real-Time Polymerase Chain Reaction; Receptors, Metabotropic Glutamate; RNA, Messenger; S100 Calcium Binding Protein beta Subunit; Skin Neoplasms; Tumor Cells, Cultured

Malignant melanoma is one of the deadliest forms of skin cancer and its incidence is expected to rise over the next two decades. At present, there are no effective therapies for advanced melanoma. We have previously shown that administration of whole recombinant yeast expressing human MART-1 (hMART-IT) induces protective antimelanoma immunity in a B16F10 transplantable mouse model. In this study, we examine the effectiveness of the hMART-IT vaccine in a congenic strain of genetically engineered mouse model of melanoma, which recapitulates both the underlying genetics and the proper tumor microenvironment of naturally occurring melanoma. Subcutaneous administration of hMART-IT induced cytotoxicity against melanoma cells and antigen-specific production of Th1-specific cytokines by splenocytes. Weekly administration of hMART-IT significantly delayed the development of melanoma and prolonged the survival of mice compared with controls. Although histological analysis demonstrated diffuse infiltration of CD4(+) T cells and CD8(+) T cells, no reduction of regulatory T cells was observed, suggesting that hMART-IT cannot prevent immunotolerance in the tumor microenvironment. This study provides a proof of concept that genetically engineered mouse models lend valuable insights into immunotherapeutics being tested in the preclinical setting.

Topics: Animals; Cancer Vaccines; Cytotoxicity, Immunologic; Disease Models, Animal; Genetic Engineering; MART-1 Antigen; Melanoma, Experimental; Mice; Mice, Inbred C57BL; Saccharomyces cerevisiae; Vaccines, Synthetic

The muted (mu) mouse is a model for Hermansky-Pudlak Syndrome (HPS), an inherited disorder of humans causing hypopigmentation, hemorrhaging and early death due to lung abnormalities. The mu gene regulates the synthesis of specialized mammalian organelles such as melanosomes, platelet dense granules and lysosomes. Further, balance defects indicate that it controls the synthesis of otoliths of the inner ear. The mu gene has been identified by a positional/candidate approach involving large mouse interspecific backcrosses. It encodes a novel ubiquitously expressed transcript, specifying a predicted 185 amino acid protein, whose expression is abrogated in the mu allele which contains an insertion of an early transposon (ETn) retrotransposon. Expression is likewise expected to be lost in the mu( J) allele which contains a deletion of a single base pair within the coding region. The presence of structurally aberrant melanosomes within the eyes of mutant mice together with localization of the muted protein within vesicles in both the cell body and dendrites of transfected melan-a melanocytes emphasizes the role of the mu gene in vesicle trafficking. The mu gene is present only in mice and humans among analyzed genomes. As is true for several other recently identified mouse HPS genes, the mu gene is absent in lower eukaryotes. Therefore, the mu gene is a member of the novel gene set that has evolved in higher eukaryotes to regulate the synthesis/function of highly specialized subcellular organelles such as melanosomes and platelet dense granules.

Topics: Amino Acid Sequence; Animals; Antigens, Neoplasm; Choroid; Chromosomes, Artificial, Bacterial; Cytoplasmic Granules; Cytoplasmic Vesicles; Disease Models, Animal; DNA Primers; Drosophila; Drosophila Proteins; Eye Proteins; Hermanski-Pudlak Syndrome; Homeodomain Proteins; Humans; MART-1 Antigen; Melanosomes; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Molecular Sequence Data; Mutation; Neoplasm Proteins; Pigment Epithelium of Eye; Polymerase Chain Reaction; RNA, Messenger; Sequence Homology, Amino Acid

Dendritic cells (DCs) are potent antigen-presenting cells that have been shown to play a critical role in the initiation of host immune responses against tumor antigens. In this study, a recombinant adenovirus vector encoding the melanoma-associated antigen, MART-1, was used to transduce murine DCs, which were then tested for their ability to activate cytotoxic T lymphocytes (CTLs) and induce protective immunity against B16 melanoma tumor cells implanted intracranially.. Genetic modification of murine bone marrow-derived DCs to express MART-1 was achieved through the use of an E1-deficient, recombinant adenovirus vector (AdVMART1). Sixty-two C57BL/6 mice were immunized by subcutaneous injection of AdVMART-1-transduced DCs (23 mice), untransduced DCs (17 mice), or sterile saline (22 mice). Using the B16 murine melanoma, which naturally expresses the MART-1 antigen, all the mice were then challenged intracranially with viable, unmodified syngeneic B16 tumor cells 7 days later. Splenocytes obtained from representative animals in each group were harvested for standard cytotoxicity and enzyme-linked immunospot assays. The remaining mice were followed for survival. Immunization of C57BL/6 mice with DCs transduced with AdVMART1-DC elicited the development of antigenspecific CTL responses. As evidenced by a prolonged survival curve when compared with control-immunized mice harboring intracranial B16 tumors, AdMART1-DC vaccination was able to elicit partial protection against central nervous system (CNS) tumor challenge in vivo. However, this CNS antitumor immunity was weaker than that previously demonstrated against subcutaneous B16 tumors in which the same vaccination strategy was used.. These data suggest that immune responses generated against CNS tumors by DC-based vaccines may be different from those obtained against subcutaneous tumors.

Topics: Adenoviridae; Animals; Antigens, Neoplasm; Brain Neoplasms; Cells, Cultured; Cytokines; Dendritic Cells; Disease Models, Animal; Female; Humans; Immunization; MART-1 Antigen; Melanoma; Mice; Mice, Inbred C57BL; Neoplasm Proteins; Neoplasm Transplantation; Spleen; T-Lymphocytes, Cytotoxic; Transduction, Genetic; Xenograft Model Antitumor Assays