Immune Cell Response Profiling

Various immunological modulators have the potential to provide an anti-tumor immune-driven response, which is evaluated through in vivo efficacy across valuable and relevant in vivo models, and thus offer promising approaches for cancer therapy. Efficiency of many of these modulators is usually underlied by a strong impact on the immune system which needs to be depicted through relevant models and immunological readouts, in order to see whether the modulation of immune subset responses correlates with the impact of therapeutics on tumor growth in untreated and challenged mice. Notably, depending on their responsiveness degree to different immune checkpoint inhibitors and therapies, tumor models differ in extent and type of immune cell modulation, while the in vivo tumor responses can be similar. Profiling immune cell function permits for building a complete package with in vivo data for a deeper understanding of how a cancer therapy performs.

Explicyte has developed and validated a comprehensive panel of tools aiming at evaluating the modulation of the immune system response by novel therapies in syngeneic tumor-bearing mice, in the presence of a functionally immunocompetent system. Profiling of immune function of key cell subsets can be performed both at the tumor site level and in peripheral compartments e.g. blood, spleen, tumor-draining lymph nodes (TDLNs)…, through assaying cell surface markers, cell activation markers, antigen-specific T lymphocyte markers, MDSC (myeloid-derived suppressor cells) markers, cytokines release, and immunological marker-encoding genes, using appropriate methodologies and platforms (e.g. flow cytometry, RT-qPCR, immunohistology-based imaging…).

Illustrative data

Immune response profiling of MCA205 tumor-bearing model to anti-PDL1 therapy
Expression analysis of key immune markers encoding genes in MCA205 tumor-bearing model of anti-PDL1 therapy.

Intratumoral expression of selected immune marker encoding genes is analyzed on intratumoral biopsies collected from vehicle and anti-PDL1 treated mice. PDL1 antibody treatment modulates proinflammatory marker encoding genes, including Tnfa and Il6. 

 

Expression analysis of key immune markers encoding genes in MCA205 tumor-bearing model of ...

Tumor infiltrating lymphocytes profiling of MCA205 tumor-bearing mice upon anti-PDL1 blockade
Flow cytometry analysis of MCA205 tumors demonstrated a higher immune cell infiltration within the tumor upon PDL1 blockade.

Anti-PDL1 antibody treatment of MCA205-timor bearing mice leads to a higher number of Tumor infiltrating lymphocytes (as shown by CD45+). This feature is associated with an increase in T cells (CD3+) and particularly with an accumulation of effector T cells (CD8+). In contrast, CD4+ T cells infiltration of the tumor is slightly decreased upon treatment.

Flow cytometry analysis of MCA205 tumors demonstrated a higher immune cell infiltration within ...

Characterization of tumor immune cell infiltration in orthotopic 4T1 mammary fat pad model
Orthotopic mammary fat pad model is characterized by tumor-infiltrating immune cells with immune suppressive properties.

Hematoxylin-associated immunohistochemistry staining of tumor sections of 4T1 mammary fat pad (left image) shows the presence of F4/80+ macrophages and of CD8+ cytotoxic lymphocytes infiltrating the tumor. Presence of tumor-infiltrating PD1-expressing immune cells is highlighted.

Orthotopic mammary fat pad model is characterized by tumor-infiltrating immune cells with ...

Immune response profiling for MOA deconvolution

  • Immune function can be evaluated to delineate a mechanism of action and see how a therapy performs in the presence of a functional immune system where tumor-host immunity interactions are preserved.
  • Standardly, 4 mice are used per experimental group to characterize the immune response underlying anti-tumor effects.
    • Standard package includes 4 experimental groups comprised of 4 mice each exposed to i) vehicle (controls), ii) a reference treatment, iii) a test compound, and iv) test compound combined with the reference treatment.
    • Immune profiling can be performed in 2 compartments; either within the tumor and/or at the periphery (TDLNs, spleen, blood…).
  • Quantitative multiplex immunological analysis is performed by assaying relevant immune markers to address the immune function modulation.
    • Immunological markers including specific antigens, activation markers, and soluble mediators can be evaluated using adapted methodologies with appropriate platforms, e.g. flow cytometry, RT-qPCR, immunohistology-based imaging, ELISA…
    • Validated traditional but also custom marker panels can be generated to address specific program-related needs.