The YUMM lines: a series of congenic mouse melanoma cell lines with defined genetic alterations

Abstract

The remarkable success of immune therapies emphasizes the need for immune-competent cancer models. Elegant genetically engineered mouse models of a variety of cancers have been established, but their effective use is limited by cost and difficulties in rapidly generating experimental data. Some mouse cancer cell lines are transplantable to immunocompetent host mice and have been utilized extensively to study cancer immunology. Here, we describe the Yale University Mouse Melanoma (YUMM) lines, a comprehensive system of mouse melanoma cell lines that are syngeneic to C57BL/6, have well-defined human-relevant driver mutations, and are genomically stable. This will be a useful tool for the study of tumor immunology and genotype-specific cancer biology.

Keywords: YUMM; cancer immunology; cell lines; melanoma; mouse models.

Figure 1. Derivation of YUMM lines

(A) Mice containing the appropriate allele combination were activated with 4-hydroxytamoxifen (4-OHT) for tumor induction at the site of application. (B) Depending on the allele combinations, the mice develop tumors from 1–6 months after the application of 4-OHT. (C) Approximately 1×1×1 mm tumor pieces were removed from the tumor, washed by with 70% ethanol and sequential 1× PBS containing 2% pen-strep washes. (D) Tumors were minced finely and added to flasks for containing DMEM-F12 with 10% FBS.

Figure 2. Growth characteristics of YUMM lines

(A) In vitro growth curves over a three day window were determined by a CyQuant DNA-based assay. A range of proliferatiion rates of the YUMM lines is observed. In vivo tumor growth curves are show in (B) with the lines showing variable tumor growth rates. (C) and (D) are H& E comparisons of the YUMM lines (C) and the GEMM model (D) from which the cell lines were derived. All images are at 60× magnification.

Figure 3. Response to Braf inhibition

YUMM cells with Braf driver mutations are able to respond to vemurafenib in vivo (A). Select YUMM cells were tested for their response and the corresponding IC50 curves were generated over a three day CyQuant DNA-based proliferation assay. (B) A resistant YUMM1.1 line was generated at 5 μM vemurafenib (YUMM1.1 5R) and the IC50 increased to 7.86 μM. (C) Analysis of the downstream signaling pathways by Western Blot analysis shows decreased p-ERK in sensitive YUMM lines and restoration of p-ERK signal in YUMM1.1 5R. YUMM1.7 was injected in C57BL/6 mice and half of the mice were treated with PLX4720 chow diet one week after injection of the YUMM cells and the non-treated mice were fed a control diet. The PLX4720 treated mice eventually develop resistant to PLX4720 around 45 days after the start of treatment.

Figure 4. Immune characteristics of YUMM tumors

(A) YUMM lines were injected into C57BL/6J mice with healthy immune systems and RAG mice without B and T cells. At day 30 after injection, no significant difference was found between the tumor volumes. (B) Select YUMM grafts were harvested at the tumor endpoint and a histological analysis was performed to determine the fraction of nucleated cells that were CD3 T cells, F4/80 macrophages, and CD45 white blood cells. (C) Representative images from the immunhistochemical stains are displayed (400×). Positive cells (brown) were counted based on the characteristic staining of the cells (surrounding the nuclei).