Microporous scaffolds loaded with immunomodulatory lentivirus to study the contribution of immune cell populations to tumor cell recruitment in vivo

Metastases are preceded by stochastic formation of a hospitable microenvironment known as the premetastatic niche, which has been difficult to study. Herein, we employ implantable polycaprolactone scaffolds as an engineered premetastatic niche to independently investigate the role of interleukin‐10 (IL10), CXCL12, and CCL2 in recruiting immune and tumor cells and impacting breast cancer cell phenotype via lentiviral overexpression. Lentivirus delivered from scaffolds in vivo achieved sustained transgene expression for 56 days. IL10 lentiviral expression, but not CXCL12 or CCL2, significantly decreased tumor cell recruitment to scaffolds in vivo. Delivery of CXCL12 enhanced CD45+ immune cell recruitment to scaffolds while delivery of IL10 reduced immune cell recruitment. CCL2 did not alter immune cell recruitment. Tumor cell phenotype was investigated using conditioned media from immunomodulated scaffolds, with CXCL12 microenvironments reducing proliferation, and IL10 microenvironments enhancing proliferation. Migration was enhanced with CCL2 and reduced with IL10‐driven microenvironments. Multiple linear regression identified populations of immune cells associated with tumor cell abundance. CD45+ immune and CD8+ T cells were associated with reduced tumor cell abundance, while CD11b+Gr1+ neutrophils and CD4+ T cells were associated with enhanced tumor cell abundance. Collectively, biomaterial scaffolds provide a tool to probe the formation and function of the premetastatic niche.Metastases are preceded by stochastic formation of a hospitable microenvironment known as the premetastatic niche, which has been difficult to study. Herein, we employ implantable polycaprolactone scaffolds as an engineered premetastatic niche to independently investigate the role of interleukin‐10 (IL10), CXCL12, and CCL2 in recruiting immune and tumor cells and impacting breast cancer cell phenotype via lentiviral overexpression.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/153114/1/bit27179.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/153114/2/bit27179_am.pd

Biomaterial Scaffolds as Pre‐metastatic Niche Mimics Systemically Alter the Primary Tumor and Tumor Microenvironment

Primary tumor (PT) immune cells and pre‐metastatic niche (PMN) sites are critical to metastasis. Recently, synthetic biomaterial scaffolds used as PMN mimics are shown to capture both immune and metastatic tumor cells. Herein, studies are performed to investigate whether the scaffold‐mediated redirection of immune and tumor cells would alter the primary tumor microenvironment (TME). Transcriptomic analysis of PT cells from scaffold‐implanted and mock‐surgery mice identifies differentially regulated pathways relevant to invasion and metastasis progression. Transcriptomic differences are hypothesized to result from scaffold‐mediated modulations of immune cell trafficking and phenotype in the TME. Culturing tumor cells with conditioned media generated from PT immune cells of scaffold‐implanted mice decrease invasion in vitro more than two‐fold relative to mock surgery controls and reduce activity of invasion‐promoting transcription factors. Secretomic characterization of the conditioned media delineates interactions between immune cells in the TME and tumor cells, showing an increase in the pan‐metastasis inhibitor decorin and a concomitant decrease in invasion‐promoting chemokine (C‐C motif) ligand 2 (CCL2) in scaffold‐implanted mice. Flow cytometric and transcriptomic profiling of PT immune cells identify phenotypically distinct tumor‐associated macrophages (TAMs) in scaffold‐implanted mice, which may contribute to an invasion‐suppressive TME. Taken together, this study demonstrates biomaterial scaffolds systemically influence metastatic progression through manipulation of the TME.Biomaterial implants that mimic the pre‐metastatic niche are shown to redirect immune and tumor cell populations in vivo. However, the systemic effects of pre‐metastatic niche mimics on metastasis progression have yet to be characterized. In this work, synthetic biomaterial implants were shown to systemically alter the primary tumor and the tumor microenvironment to promote an invasion‐suppressive phenotype.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/144244/1/adhm201700903-sup-0001-S1.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/144244/2/adhm201700903_am.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/144244/3/adhm201700903.pd

Microporous scaffolds loaded with immunomodulatory lentivirus to study the contribution of immune cell populations to tumor cell recruitment in vivo

Metastases are preceded by stochastic formation of a hospitable microenvironment known as the premetastatic niche, which has been difficult to study. Herein, we employ implantable polycaprolactone scaffolds as an engineered premetastatic niche to independently investigate the role of interleukin‐10 (IL10), CXCL12, and CCL2 in recruiting immune and tumor cells and impacting breast cancer cell phenotype via lentiviral overexpression. Lentivirus delivered from scaffolds in vivo achieved sustained transgene expression for 56 days. IL10 lentiviral expression, but not CXCL12 or CCL2, significantly decreased tumor cell recruitment to scaffolds in vivo. Delivery of CXCL12 enhanced CD45+ immune cell recruitment to scaffolds while delivery of IL10 reduced immune cell recruitment. CCL2 did not alter immune cell recruitment. Tumor cell phenotype was investigated using conditioned media from immunomodulated scaffolds, with CXCL12 microenvironments reducing proliferation, and IL10 microenvironments enhancing proliferation. Migration was enhanced with CCL2 and reduced with IL10‐driven microenvironments. Multiple linear regression identified populations of immune cells associated with tumor cell abundance. CD45+ immune and CD8+ T cells were associated with reduced tumor cell abundance, while CD11b+Gr1+ neutrophils and CD4+ T cells were associated with enhanced tumor cell abundance. Collectively, biomaterial scaffolds provide a tool to probe the formation and function of the premetastatic niche.Metastases are preceded by stochastic formation of a hospitable microenvironment known as the premetastatic niche, which has been difficult to study. Herein, we employ implantable polycaprolactone scaffolds as an engineered premetastatic niche to independently investigate the role of interleukin‐10 (IL10), CXCL12, and CCL2 in recruiting immune and tumor cells and impacting breast cancer cell phenotype via lentiviral overexpression.Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/153114/1/bit27179.pdfhttps://deepblue.lib.umich.edu/bitstream/2027.42/153114/2/bit27179_am.pd

Multiplexing Engineered Receptors for Multiparametric Evaluation of Environmental Ligands

Engineered cell-based therapies comprise a promising, emerging biomedical technology. Broad utilization of this strategy will require new approaches for implementing sophisticated functional programs, such as sensing and responding to the environment in a defined fashion. Toward this goal, we investigated whether our self-contained receptor and signal transduction system (MESA) could be multiplexed to evaluate extracellular cues, with a focus on elucidating principles governing the integration of such engineered components. We first developed a set of hybrid promoters that exhibited AND gate activation by two transcription factors. We then evaluated these promoters when paired with two MESA receptors and various ligand combinations. Unexpectedly, although the multiplexed system exhibited distinct responses to ligands applied individually and in combination, the same synergy was not observed as when promoters were characterized with soluble transcription factors. Therefore, we developed a mechanistic computational model leveraging these observations, to both improve our understanding of how the receptors and promoters interface and to guide the design and implementation of future systems. Notably, the model explicitly accounts for the impact of intercellular variation on system characterization and performance. Model analysis identified key factors that affect the current receptors and promoters, and enabled an <i>in silico</i> exploration of potential modifications that inform the design of improved logic gates and their robustness to intercellular variation. Ultimately, this quantitative design-driven approach may guide the use and multiplexing of synthetic receptors for diverse custom biological functions beyond the case study considered here

Multiplexing Engineered Receptors for Multiparametric Evaluation of Environmental Ligands

Engineered cell-based therapies comprise a promising, emerging biomedical technology. Broad utilization of this strategy will require new approaches for implementing sophisticated functional programs, such as sensing and responding to the environment in a defined fashion. Toward this goal, we investigated whether our self-contained receptor and signal transduction system (MESA) could be multiplexed to evaluate extracellular cues, with a focus on elucidating principles governing the integration of such engineered components. We first developed a set of hybrid promoters that exhibited AND gate activation by two transcription factors. We then evaluated these promoters when paired with two MESA receptors and various ligand combinations. Unexpectedly, although the multiplexed system exhibited distinct responses to ligands applied individually and in combination, the same synergy was not observed as when promoters were characterized with soluble transcription factors. Therefore, we developed a mechanistic computational model leveraging these observations, to both improve our understanding of how the receptors and promoters interface and to guide the design and implementation of future systems. Notably, the model explicitly accounts for the impact of intercellular variation on system characterization and performance. Model analysis identified key factors that affect the current receptors and promoters, and enabled an <i>in silico</i> exploration of potential modifications that inform the design of improved logic gates and their robustness to intercellular variation. Ultimately, this quantitative design-driven approach may guide the use and multiplexing of synthetic receptors for diverse custom biological functions beyond the case study considered here

Pre‐Metastatic Niche: Biomaterial Scaffolds as Pre‐metastatic Niche Mimics Systemically Alter the Primary Tumor and Tumor Microenvironment (Adv. Healthcare Mater. 10/2018)

Peer Reviewedhttps://deepblue.lib.umich.edu/bitstream/2027.42/144278/1/adhm201870040.pd

Metastatic Conditioning of Myeloid Cells at a Subcutaneous Synthetic Niche Reflects Disease Progression and Predicts Therapeutic Outcomes

Monitoring metastatic events in distal tissues is challenged by their sporadic occurrence in obscure and inaccessible locations within these vital organs. A synthetic biomaterial scaffold can function as a synthetic metastatic niche to reveal the nature of these distal sites. These implanted scaffolds promote tissue ingrowth, which upon cancer initiation is transformed into a metastatic niche that captures aggressive circulating tumor cells. We hypothesized that immune cell phenotypes at synthetic niches reflect the immunosuppressive conditioning within a host that contributes to metastatic cell recruitment and can identify disease progression and response to therapy. We analyzed the expression of 632 immune-centric genes in tissue biopsied from implants at weekly intervals following inoculation. Specific immune populations within implants were then analyzed by single-cell RNA-seq. Dynamic gene expression profiles in innate cells, such as myeloid-derived suppressor cells, macrophages, and dendritic cells, suggest the development of an immunosuppressive microenvironment. These dynamics in immune phenotypes at implants was analogous to that in the diseased lung and had distinct dynamics compared with blood leukocytes. Following a therapeutic excision of the primary tumor, longitudinal tracking of immune phenotypes at the implant in individual mice showed an initial response to therapy, which over time differentiated recurrence versus survival. Collectively, the microenvironment at the synthetic niche acts as a sentinel by reflecting both progression and regression of disease. SIGNIFICANCE: Immune dynamics at biomaterial implants, functioning as a synthetic metastatic niche, provides unique information that correlates with disease progression. GRAPHICAL ABSTRACT: http://cancerres.aacrjournals.org/content/canres/80/3/602/F1.large.jpg