Friday, September 28, 2018 10:30 AM–12:00 PM abstracts: innovation, surface technology and biomechanics: 182. In vivo synergistic effect of checkpoint blockade and radiation therapy against chordomas in a humanized mouse model.

BACKGROUND CONTEXT With the advent of immunotherapy (IT) against various cancers, its applications to other cancers have been extensively investigated. However, it has been a challenge to apply IT to chordomas, due to lack of clinically-translatable in vivo models. Unlike othercancers that have been extensively studied from the immunotherapeutic standpoint such as melanomas, currently, there are no well-established murine chordoma cell lines that can be injected to syngeneic mice or transgenic mouse models that develop chordomas spontaneously, which prevents us from investigating the interaction between murine chordomas and murine immune cells. Meanwhile, in the field of immunology, the development of humanized mouse models, where a human immune system is incorporated into immunodeficient mice by engrafting them with human hematopoietic stem cells and fetal thymus, has emerged as a state-of-the-art technique, which allows researchers to investigate human immune system in vivo in the context of several immunological disorders including cancer. To scrutinize IT against chordoma, the development of this model is imperative. PURPOSE We aimed to develop a humanized mouse model engrafted with chordoma cells to study the interaction between humanimmune system and human chordomas. We also sought to utilize it to investigate synergistic effect between IT and radiation therapy (RT) against chordoma. STUDY DESIGN/SETTING Animal experiments. OUTCOME MEASURES Tumor size, cell subpopulations of tumor infiltratinglymphocytes, and cytokine levels in blood and tumors in each treatment group. METHODS Fifteen 10-12-week-old NSG mice, which lacks mouse T cells, B cells, and NK cells as well as functional mouse macrophages, were sub-lethally (1.5Gy) irradiated and then implanted with human fetal thymic tissue and CD34+ stem cells that had been harvested from a fetus, whose HLA-type is partially-matched with that of the U-CH1 chordoma cell line. Reconstitution of immune cells in NSG mice was confirmed 8 weeks posttransplantation and then each animal (15 humanized NSG mice and 12 naive NSG mice) was injected with U-CH1 cell suspension bilaterally and subcutaneously. Next, they were treated for 4 weeks as follows: (A) control, isotype antibodies (Abs) injection (n=3), (B) anti-human-PD-1 Abs (n=4), (C) RT + isotype Abs (n=3, unilaterally to the left-sided tumor, 8Gy × 4), (D) anti-human-PD-1 Abs and RT (n=5), (E) naive NSG mice (n=6, without the engraftment of human immune cells) + isotype, and (F) naive NSG mice (n=6) + anti-human-PD-1 Abs. During and after the treatment, anti-tumor activities were monitored via tumor size, flow cytometry, qRT-PCR, and immunohistochemistry. The data were statistically analyzed and p < 0.05 was set as statistically significant. RESULTS On average, human peripheral blood mononuclear cells (PBMCs) of 43.8% among all PBMCs (human + mouse), human T cells of 23.4% among human PBMCs, human CD8+ T cells of 24.3% among human T cells, and other lymphocytes such as B cells, macrophages, and NK cells were observed in peripheral blood of humanized mice via flow cytometry. One week after the treatment, on the irradiated side, (D) demonstrated lowest tumor volume ((A) 2220.0mm3, (B) 615.8mm3, (C) 1259.0mm3, (D)144.0mm3, p<.001), highest number of human PBMCs, highest % of CD8+ human T cells ((A) 31.0%, (B) 60.3%, (C) 43.0%, (D) 71.2%, p<.001), highest % of CD45RO+CD4+ (memory) human T cells ((A) 47.0%, (B) 56.5%, (C) 48.0%, (D) 65.4%, p<.001), and lowest % of PD−1+CD8+ (exhausted) human T cells ((A) 61.0%, (B) 42.5%, (C) 60.0%, (D) 36.6%, p<.001) in the tumors via flow cytometry, and highest IFN-gamma in the tumors via qRT-PCR, compared to the other five groups with statistical significance. On the nonirradiated side, similarly (D) had the smallest tumor compared to the others (P=.09). There were no statistically significant differences in tumor volume amongst (A), (E), and (F), suggesting that HLA-partially-mismatched immune cells from the fetus did not influence U-CH1 growth (immune tolerance) and that anti-human-PD-1 Abs we administered were efficacious only in the humanized mice. Currently, immunohistochemistry is being performed to further confirm the findings via the other modalities as described above. CONCLUSIONS We demonstrated that this humanized mouse model could be a revolutionary platform to investigate IT against rare cancers such as chordomas, where murine equivalent cell lines are not available to date, which hinders us from utilizing syngeneic or transgenic mouse models to study IT against cancers. The direct synergistic effect between IT and RT against chordoma as well as the potential abscopal effect was observed. [ABSTRACT FROM AUTHOR]