Characterization of immune responses to anti-PD-1 mono and combination immunotherapy in hematopoietic humanized mice implanted with tumor xenografts

Abstract

BackgroundThe success of agents that reverse T-cell inhibitory signals, such as anti-PD-1/PD-L1 therapies, has reinvigorated cancer immunotherapy research. However, since only a minority of patients respond to single-agent therapies, methods to test the potential anti-tumor activity of rational combination therapies are still needed. Conventional murine xenograft models have been hampered by their immune-compromised status; thus, we developed a hematopoietic humanized mouse model, hu-CB-BRGS, and used it to study anti-tumor human immune responses to triple-negative breast cancer (TNBC) cell line and patient-derived colorectal cancer (CRC) xenografts (PDX).MethodsBALB/c-Rag2nullIl2rγnullSIRPαNOD (BRGS) pups were humanized through transplantation of cord blood (CB)-derived CD34+ cells. Mice were evaluated for human chimerism in the blood and assigned into experimental untreated or nivolumab groups based on chimerism. TNBC cell lines or tumor tissue from established CRC PDX models were implanted into both flanks of humanized mice and treatments ensued once tumors reached a volume of ~150mm3. Tumors were measured twice weekly. At end of study, immune organs and tumors were collected for immunological assessment.ResultsHumanized PDX models were successfully established with a high frequency of tumor engraftment. Humanized mice treated with anti-PD-1 exhibited increased anti-tumor human T-cell responses coupled with decreased Treg and myeloid populations that correlated with tumor growth inhibition. Combination therapies with anti-PD-1 treatment in TNBC-bearing mice reduced tumor growth in multi-drug cohorts. Finally, as observed in human colorectal patients, anti-PD-1 therapy had a strong response to a microsatellite-high CRC PDX that correlated with a higher number of human CD8+ IFNγ+ T cells in the tumor.ConclusionHu-CB-BRGS mice represent an in vivo model to study immune checkpoint blockade to human tumors. The human immune system in the mice is inherently suppressed, similar to a tumor microenvironment, and thus allows growth of human tumors. However, the suppression can be released by anti-PD-1 therapies and inhibit tumor growth of some tumors. The model offers ample access to lymph and tumor cells for in-depth immunological analysis. The tumor growth inhibition correlates with increased CD8 IFNγ+ tumor infiltrating T cells. These hu-CB-BRGS mice provide a relevant preclinical animal model to facilitate prioritization of hypothesis-driven combination immunotherapies.Electronic supplementary materialThe online version of this article (10.1186/s40425-019-0518-z) contains supplementary material, which is available to authorized users.

Tumor-treating fields (TTFields) induce immunogenic cell death resulting in enhanced antitumor efficacy when combined with anti-PD-1 therapy

“…In contrast, the combined treatment of TTFields and anti-PD-1 did not result in changes in PD-L1 density in leukocytes from CT-26 tumors, and PD-L1 density was reduced in macrophages from tumors treated with TTFields monotherapy or the combination of the two modalities. This finding can be attributed to the long-term treatment duration in this model and is in line with a recent study by Capasso et al, showing that PD-L1 expression is increased in early stages of treatment with anti-PD-1 and is decreased in later stages [37]. It should also be noted that CT-26 cancer cells express high levels of PD-L1 on their surface while LLC-1 cancer cells do not express PD-L1; therefore, changes in PD-L1 expression in the tumor microenvironment can also potentially be affected by the PD-L1 expression by the cancer cells as well as by drug treatment schedule.…”

RX-5902, a novel β-catenin modulator, potentiates the efficacy of immune checkpoint inhibitors in preclinical models of triple-negative breast Cancer

“…To generate human immune system (HIS) mice, 0.2–0.6 × 10 6 expanded and thawed CD34+ cells were injected into neonate BRGS (BALB/c Rag2 nul l IL2Rγ null Sirpa NOD ) pups that had been irradiated with 300 rad 2–6 h prior to injection [ 18 ]. Mice were injected in the facial vein or liver, as previously described [ 18 ].…”

“…To generate human immune system (HIS) mice, 0.2–0.6 × 10 6 expanded and thawed CD34+ cells were injected into neonate BRGS (BALB/c Rag2 nul l IL2Rγ null Sirpa NOD ) pups that had been irradiated with 300 rad 2–6 h prior to injection [ 18 ]. Mice were injected in the facial vein or liver, as previously described [ 18 ]. Mice were weaned at 3–4 weeks of age and analyzed for human chimerism at 10 and 16 weeks of age by staining of PBMCs, isolated from retro-orbital bleeds, to determine human (hCD45+), T (hCD3, hCD8, hCD5) and B (hCD20) cell chimerism.…”

“…p68 can be phosphorylated at Y593 by growth factors, including platelet-derived growth factor (PDGF) acting through c-Abl signaling, promoting oncogenesis through enhanced β-catenin nuclear localization [ 17 ]. p-p68 functions as a nuclear chaperone for β-catenin, promoting nuclear translocation and activation of cyclin D1, c-JUN and c-MYC [ 17 , 18 ]. In this role, p-p68 regulates cellular proliferation, malignant transformation, epithelial to mesenchymal transition (EMT) and cell migration pointing to a vital role in oncogenesis and cancer progression [ 17 , 19 ].…”

Modifying the tumour microenvironment and reverting tumour cells: New strategies for treating malignant tumours

“…The TME regulates the differentiation and activation of immune cells to reduce their ability to recognize and eliminate tumour cells. Immune cell therapies that more accurately target tumour cells while restoring normal immune cell populations within the TME, enhancing normal immune cell function and disrupting the protective effects of the abnormal immune cell networks have become a new anti‐tumour option 109 . As early as 1998, studies demonstrated that transplanting immune‐activated splenocytes into mice with breast cancer caused tumour shrinkage and conferred long‐term tumour‐free survival, suggesting that adoptive immune cells can be effective anti‐tumour therapeutics 110 .…”