Project description:Melanoma is one of the most commonly diagnosed malignancies and serves as a model for studying immunotherapy. The B16 melanoma model is characterized by low T cell infiltration in the tumor, and blocking the PD-1 pathway shows no significant anti-tumor activity, mirroring melanoma patients with a cold tumor immunophenotype. Therefore, understanding the molecular basis that prevents T cell-mediated anti-tumor activity in B16 melanoma is of great significance. In this study, we generated Tyr knockout B16 melanoma cells using CRISPR/Cas9 and discovered that tyrosinase in melanoma significantly inhibits the anti-tumor activity of T cells. Tyrosinase deficiency significantly increases T-cell infiltration and activation within the tumor. Single-cell RNA sequencing reveals an altered cold tumor immunophenotype in tyrosinase-deficient B16 melanoma. In wild-type mice, T cells in tyrosinase-deficient tumors express elevated levels of PD-1 and Foxp3. However, strikingly, in PD-1 deficient mice, the loss of tyrosinase in B16 melanoma unleashes the anti-tumor activity of PD-1 deficient T cells. This enhanced anti-tumor activity is explained by significantly increased tumor T cell infiltration accompanied by reduced frequencies of Tregs in PD-1 knockout mice. Targeting tyrosinase may enhance the anti-tumor efficacy of PD-1 blockade in cold tumors, offering a novel strategy to render cold tumors responsive to immunotherapy. Clinically, higher levels of tyrosinase expression in human melanoma are associated with a poorer prognosis, indicating that our findings could potentially improve the efficacy of immunotherapy in melanoma patients.
Project description:Analysis of gene expression patterns of B-1 cells from C57BL6 wild type mice versus B-1 cells fromIL-10 Knockout mice. This study will help in the identification of factors in B-1 cells involved in changing of metastatic behavior of B16 melanoma after contacting with B-1 lymphocytes.
Project description:We have shown that C57BL/6J CCR5 knockout mice develop 30.4% ± 8.6% fewer B16 F10 lung nodules compared to wild type mice after the intravenous injection of 100,000 B16 F10 cells. We sought to understand this phenomenon by comparing gene expression in the lungs of these mice at 6, 24, and 48 hours after tumor injection. There were 6 groups or conditions (CCR5 - 6 hours, WT - 6 hours, CCR5 - 24 hours, WT - 24 hours, CCR5 - 48 hours, WT - 48 hour); each group had 4 to 5 mice. All of the mice were injected with 100,000 B16 F10 melanoma cells by tail vein. At the designated time, the mice were anesthetized with Avertin and their lungs were perfused with PBS. After this, the lungs were harvested and snap frozen in liquid nitrogen. They were kept at minus 80 degrees C until processing. mRNA was extracted as described below and equivalent amounts of mRNA (by weight) was pooled by group.
Project description:We have shown that C57BL/6J CCR5 knockout mice develop 30.4% ± 8.6% fewer B16 F10 lung nodules compared to wild type mice after the intravenous injection of 100,000 B16 F10 cells. We sought to understand this phenomenon by comparing gene expression in the lungs of these mice at 6, 24, and 48 hours after tumor injection.
Project description:The FOXC2 transcription factor regulates a variety of developmental and biological processes in both embryonic and adult tissues. Importantly, overexpression or dysregulation of FOXC2 is also associated with oncogenic activity in numerous cancer types, though the function of FOXC2 in the context of melanoma has not been previously investigated. Therefore, the goal of this study was to assess FOXC2's regulation of gene expression in melanoma cells. To this end, we employed CRISPR-Cas9 gene editing technology to disrupt the Foxc2 gene in B16-F1 melanoma, and we performed RNA-seq analysis to assess differential gene expression between the wild-type B16-F1 melanoma cell line and our novel FOXC2-deficient B16-F1ΔFOXC2 gene-edited variant cell line.
Project description:Plasmodium berghei transcriptomes of wild-type and ap2-z knockout parasites [ap2-z(-)] were analyzed at 6 hours post starting ookinete culture (hpoc).
Project description:The goal of this study was to compare the gene expression profiles of a highly versus poorly tumorigenic murine melanoma. B16-F1 is a well-characterized murine melanoma cell line that grows progressively in hosts, both as primary tumors in subcutaneous tissue and as metastatic lesions in internal organs. D5.1G4 is a chemically mutated variant of B16 melanoma that grows with significantly slower kinetics than its wild-type counterpart. Whole genome expression microarray analysis of RNA isolated from these murine melanoma lines was performed to provide insights into factors that regulate the growth and metastasis of these tumors, which are useful models for investigating in a murine system the outgrowth, progression, and control of a cancer type that is prevalent in humans worldwide.
Project description:To investigate the impact of Card11 on TIL differentiation, we isolated TILs from B16-F10 tumors of WT mice, E134G mice, and K215M mice and performed scRNA seq. We also sorted tumor-infiltrating lymphocytes (TILs) from melanoma(B16-F10) of WT mice and conducted scRNA and scTCR sequencing. To further investigate the impact of Card11 on TCR clonal diversity, we sorted CD8 T cells from melanoma WT mice or K215M mice and performed scRNA and scTCR sequencing.
Project description:Whole Genome Sequencing of the murine breast cancer cell line 4T1 and of the murine melanoma cell line B16-ova was carried out with the aim of identifying somatic mutations. We also ran deep Mass Spectrometry proteomics analysis on the same cell lines, aiming to determine which somatic mutations carry over to the protein expression level. Further, we tested these cancer specific protein epitopes (putative neoantigens) for immunogenicity using mouse models. Finally, the putative neoantigens that showed good immunogenic potential were used in tumor growth control experiments with mice engrafted with the two tumor cell lines. In these experiments we tested whether cancer vaccines based on individual neoantigen peptides (MHC-I) restricted the growth of the tumor compared to adequate controls. The overall aim of the project is to validate the ability of our multi-omics/bioinformatics pipeline to identify and deliver neoantigens that can be used to suppress tumor growth. File names Sample names P10859_101_S1_L001_R1_001_BHKWV3CCXY 4T1_S1_L001_R1_001_BHKWV3CCXY P10859_101_S1_L001_R2_001_BHKWV3CCXY 4T1_S1_L001_R2_001_BHKWV3CCXY P10859_101_S1_L002_R1_001_BHKWV3CCXY 4T1_S1_L002_R1_001_BHKWV3CCXY P10859_101_S1_L002_R2_001_BHKWV3CCXY 4T1_S1_L002_R2_001_BHKWV3CCXY P10859_102_S2_L003_R1_001_BHKWV3CCXY B16-OVA_S2_L003_R1_001_BHKWV3CCXY P10859_102_S2_L003_R2_001_BHKWV3CCXY B16-OVA_S2_L003_R2_001_BHKWV3CCXY P10859_102_S2_L004_R1_001_BHKWV3CCXY B16-OVA_S2_L004_R1_001_BHKWV3CCXY P10859_102_S2_L004_R2_001_BHKWV3CCXY B16-OVA_S2_L004_R2_001_BHKWV3CCXY