Genetic analysis of the diabetes-prone C57BLKS/J mouse strain reveals genetic contribution from multiple strains

AbstractThe C57BLKS/J (BKS) inbred mouse strain is a widely used animal model of type 2 diabetes. In the presence of the diabetes (db) mutation, obese BKS-db mice develop severe diabetes. Genetic studies of diabetes-susceptibility in this strain are facilitated by the fact that BKS is a genetic composite between the diabetes-resistant C57BL/6J (B6) and susceptible DBA/2J (DBA) strains. On this basis, it has been hypothesized that diabetes-susceptibility in BKS is conferred by DBA-derived alleles. However, recent studies revealed non-B6/non-DBA genetic material in BKS. To identify the origin of this genetic component, we generated a genomic map of BKS using 537 microsatellite markers. Our results demonstrate that, in addition to B6 and DBA, BKS contains alleles from at least three other strains, including 129, C57BL/10 and an unidentified mouse strain. We also analyzed two congenic strains, B6-db and BKS-db, which are widely used for the genetic mapping of diabetes-susceptibility loci. We identified several donor-derived genomic regions introduced during the generation of these congenic strains. In summary, our study reveals novel aspects of the genetic fine-structure of BKS and related strains and facilitates the identification of diabetes-susceptibility loci in this mouse model

Mena deficiency delays tumor progression and decreases metastasis in polyoma middle-T transgenic mouse mammary tumors

Introduction The actin binding protein Mammalian enabled (Mena), has been implicated in the metastatic progression of solid tumors in humans. Mena expression level in primary tumors is correlated with metastasis in breast, cervical, colorectal and pancreatic cancers. Cells expressing high Mena levels are part of the tumor microenvironment for metastasis (TMEM), an anatomical structure that is predictive for risk of breast cancer metastasis. Previously we have shown that forced expression of Mena adenocarcinoma cells enhances invasion and metastasis in xenograft mice. Whether Mena is required for tumor progression is still unknown. Here we report the effects of Mena deficiency on tumor progression, metastasis and on normal mammary gland development. Methods To investigate the role of Mena in tumor progression and metastasis, Mena deficient mice were intercrossed with mice carrying a transgene expressing the polyoma middle T oncoprotein, driven by the mouse mammary tumor virus. The progeny were investigated for the effects of Mena deficiency on tumor progression via staging of primary mammary tumors and by evaluation of morbidity. Stages of metastatic progression were investigated using an in vivo invasion assay, intravital multiphoton microscopy, circulating tumor cell burden, and lung metastases. Mammary gland development was studied in whole mount mammary glands of wild type and Mena deficient mice. Results Mena deficiency decreased morbidity and metastatic dissemination. Loss of Mena increased mammary tumor latency but had no affect on mammary tumor burden or histologic progression to carcinoma. Elimination of Mena also significantly decreased epidermal growth factor (EGF) induced in vivo invasion, in vivo motility, intravasation and metastasis. Non-tumor bearing mice deficient for Mena also showed defects in mammary gland terminal end bud formation and branching. Conclusions Deficiency of Mena decreases metastasis by slowing tumor progression and reducing tumor cell invasion and intravasation. Mena deficiency during development causes defects in invasive processes involved in mammary gland development. These findings suggest that functional intervention targeting Mena in breast cancer patients may provide a valuable treatment option to delay tumor progression and decrease invasion and metastatic spread leading to an improved prognostic outcome.National Cancer Institute (U.S.). Integrative Cancer Biology Program (grant U54 CA112967)Virginia and D.K. Ludwig Fund for Cancer Researc

Lymph Node Immune Profiles as Predictive Biomarkers for Immune Checkpoint Inhibitor Response

The need for predictive biomarkers that can accurately predict patients who will respond to immune checkpoint inhibitor (ICI) immunotherapies remains a clinically unmet need. The majority of research efforts have focused on expression of immune-related markers on the tumour and its associated tumour microenvironment (TME). However, immune response to tumour neoantigens starts at the regional lymph nodes, where antigen presentation takes place and is regulated by multiple cell types and mechanisms. Knowledge of the immunological responses in bystander lymphoid organs following ICI therapies and their association with changes in the TME, could prove to be a valuable component in understanding the treatment response to these agents. Here, we review the emerging data on assessment of immunological responses within regional lymph nodes as predictive biomarkers for immunotherapies

Supplementary Text and Tables from Myeloid-Derived Suppressor–Cell Dynamics Control Outcomes in the Metastatic Niche

Supplementary Text and Tables</p

Supplementary Figures from Myeloid-Derived Suppressor–Cell Dynamics Control Outcomes in the Metastatic Niche

Supplementary Figures</p

Regulation of the tumor immune microenvironment and vascular normalization in TNBC murine models by a novel peptide

Triple-negative breast cancer (TNBC) is a highly metastatic and aggressive disease with limited treatment options. Recently, the combination of the immune checkpoint inhibitor (ICI) atezolizumab (anti-PD-L1) with nab-paclitaxel was approved following a clinical trial that showed response rates in at least 43% of patients. While this approval marks a major advance in the treatment of TNBC it may be possible to improve the efficacy of ICI therapies through further modulation of the suppressive tumor immune microenvironment (TIME). Several factors may limit immune response in TNBC including aberrant growth factor signaling, such as VEGFR2 and cMet signaling, inefficient vascularization, poor delivery of drugs and immune cells, and the skewing of immune cell populations toward immunosuppressive phenotypes. Here we investigate the immune-modulating properties of AXT201, a novel 20 amino-acid integrin-binding peptide in two syngeneic mouse TNBC models: 4T1-BALB/c and NT4-FVB. AXT201 treatment improved survival in the NT4 model by 20% and inhibited the growth of 4T1 tumors by 47% over 22 days post-inoculation. Subsequent immunohistochemical analyses of 4T1 tumors also showed a 53% reduction in vascular density and a 184% increase in pericyte coverage following peptide treatment. Flow cytometry analyses demonstrated evidence of a more favorable anti-tumor immune microenvironment following treatment with AXT201, including significant decreases in the populations of T regulatory cells, monocytic myeloid-derived suppressor cells, and PD-L1 expressing cells and increased expression of T cell functional markers. Together, these findings demonstrate immune-activating properties of AXT201 that could be developed in combination with other immunomodulatory agents in the treatment of TNBC

Mena invasive (Mena[superscript INV]) and Mena11a isoforms play distinct roles in breast cancer cell cohesion and association with TMEM

Mena, an actin regulatory protein, functions at the convergence of motility pathways that drive breast cancer cell invasion and migration in vivo. The tumor microenvironment spontaneously induces both increased expression of the Mena invasive (Mena[superscript INV]) and decreased expression of Mena11a isoforms in invasive and migratory tumor cells. Tumor cells with this Mena expression pattern participate with macrophages in migration and intravasation in mouse mammary tumors in vivo. Consistent with these findings, anatomical sites containing tumor cells with high levels of Mena expression associated with perivascular macrophages were identified in human invasive ductal breast carcinomas and called TMEM. The number of TMEM sites positively correlated with the development of distant metastasis in humans. Here we demonstrate that mouse mammary tumors generated from EGFP-Mena[superscript INV] expressing tumor cells are significantly less cohesive and have discontinuous cell–cell contacts compared to Mena11a xenografts. Using the mouse PyMT model we show that metastatic mammary tumors express 8.7 fold more total Mena and 7.5 fold more Mena[superscript INV] mRNA than early non-metastatic ones. Furthermore, Mena[superscript INV] expression in fine needle aspiration biopsy (FNA) samples of human invasive ductal carcinomas correlate with TMEM score while Mena11a does not. These results suggest that Mena[superscript INV] is the isoform associated with breast cancer cell discohesion, invasion and intravasation in mice and in humans. They also imply that Mena[superscript INV] expression and TMEM score measure related aspects of a common tumor cell dissemination mechanism and provide new insight into metastatic risk.Virginia and Daniel K. Ludwig Graduate FellowshipMassachusetts Institute of Technology. Ludwig Center for Molecular OncologyNational Cancer Institute (U.S.). Integrative Cancer Biology Program (Grant U54 CA112967

A Mena Invasion Isoform Potentiates EGF-Induced Carcinoma Cell Invasion and Metastasis

The spread of cancer during metastatic disease requires that tumor cells subvert normal regulatory networks governing cell motility to invade surrounding tissues and migrate toward blood and lymphatic vessels. Enabled (Ena)/vasodilator-stimulated phosphoprotein (VASP) proteins regulate cell motility by controlling the geometry of assembling actin networks. Mena, an Ena/VASP protein, is upregulated in the invasive subpopulation of breast cancer cells. In addition, Mena is alternately spliced to produce an invasion isoform, Mena[superscript INV]. Here we show that Mena and Mena[superscript INV] promote carcinoma cell motility and invasiveness in vivo and in vitro, and increase lung metastasis. Mena and Mena[superscript INV] potentiate epidermal growth factor (EGF)-induced membrane protrusion and increase the matrix degradation activity of tumor cells. Interestingly, Mena[superscript INV] is significantly more effective than Mena in driving metastases and sensitizing cells to EGF-dependent invasion and protrusion. Upregulation of Mena[superscript INV] could therefore enable tumor cells to invade in response to otherwise benign EGF stimulus levels.Anna Fuller Molecular Oncology FundMassachusetts Institute of Technology. Ludwig Center for Molecular OncologyNational Cancer Institute (U.S.). Integrative Cancer Biology Program (Grant 1-U54-CA112967)National Institutes of Health (U.S.) (Grant GM58801