Neal Huang

Glioblastoma-infiltrated innate immune cells resemble M0 macrophage phenotype

Glioblastomas are highly infiltrated by diverse immune cells, including microglia, macrophages, and myeloid-derived suppressor cells (MDSCs). Understanding the mechanisms by which glioblastoma-associated myeloid cells (GAMs) undergo metamorphosis into tumor-supportive cells, characterizing the heterogeneity of immune cell phenotypes within glioblastoma subtypes, and discovering new targets can help the design of new efficient immunotherapies. In this study, we performed a comprehensive battery of immune phenotyping, whole-genome microarray analysis, and microRNA expression profiling of GAMs with matched blood monocytes, healthy donor monocytes, normal brain microglia, nonpolarized M0 macrophages, and polarized M1, M2a, M2c macrophages. Glioblastoma patients had an elevated number of monocytes relative to healthy donors. Among CD11b+ cells, microglia and MDSCs constituted a higher percentage of GAMs than did macrophages. GAM profiling using flow cytometry studies revealed a continuum between the M1- and M2-like phenotype. Contrary to current dogma, GAMs exhibited distinct immunological functions, with the former aligned close to nonpolarized M0 macrophages.

Immune modulatory nanoparticle therapeutics for intracerebral glioma

Background Previously we showed therapeutic efficacy of unprotected miR-124 in preclinical murine models of glioblastoma, including in heterogeneous genetically engineered murine models by exploiting the immune system and thereby negating the need for direct tumor delivery. Although these data were promising, to implement clinical trials, we required a scalable formulation that afforded protection against circulatory RNases. Methods We devised lipid nanoparticles that encapsulate and protect the miRs from degradation and provide enhanced delivery into the immune cell compartment and tested in vivo antitumor effects. Results Treatment with nanoparticle-encapsulated miR-124, LUNAR-301, demonstrated a median survival exceeding 70 days, with an associated reversal of tumor-mediated immunosuppression and induction of immune memory. In both canine and murine models, the safety profile of LUNAR-301 was favorable. Conclusions For the first time, we show that nanoparticles can direct a therapeutic response by targeting intracellular immune pathways. Although shown in the context of gliomas, this therapeutic approach would be applicable to other malignancies.

Abstract 4291: An optimized therapeutic nanoparticle delivery platform of miRNA in preclinical murine models of malignancy

INTRODUCTION: We have previously shown robust therapeutic efficacy of miRNAs in preclinical murine models of glioblastoma and were one of the first groups to deliver therapeutic miRNAs intravenously. However a major hurdle to clinical translation is a scalable formulation that affords protection against circulatory RNAses. Nanoparticles can encapsulate and protect the miRNA from degradation and enhance delivery into the immune cell compartment facilitating antitumor effects, in part through the reversal of tumor-mediate immune suppression and increased expression of effector cytokines - thus, overcoming the need for direct tumor delivery of the therapeutic agent. METHODS: FDA acceptable lipid nanoparticles were devised to enhance delivery of miRNA into the peripheral blood mononuclear cells (PBMCs) and verified by in vivo compartmental pharmacokinetic analysis and functional immune monitoring. Nanoparticle test articles contain an active immune modulatory agent - miR-124, which inhibits the signal transducer and activator of transcript 3 (STAT3) pathway. The lead candidate was designated LUNAR-301, and further refinements included unlocking the nucleic acids (LUNAR-302) to enhance efficacy. Nanoparticle formulations were tested in multiple murine models of malignancy including established intracerebral gliomas. RESULTS: In non-tumor bearing mice dosed with intravenous LUNAR-301, miR-124 was delivered to the peripheral blood mononuclear cells (PBMCs) with no clinical signs of toxicity or organ damage on histopathologic exam. In an intracerebral GL261 model, lower pSTAT3 expression was observed in mice treated with LUNAR-301 compared to both empty nanoparticle treated mice or untreated mice, p = 0.0081 and p = 0.0001 respectively. Similarly, lower Foxp3 expression was observed in the LUNAR-301 treated mice, p = 0.0057 and p = 0.0223 respectively. Median survival time for mice treated with LUNAR-301 exceeded 70 days, compared to only 32.5 days for mice treated with the previous gold-standard, miR-124 + lipofectamine. The cure rate difference between LUNAR-301 (9 out of 15 mice) and LUNAR-302 (2 out of 10 mice) was 40% (P = 0.0576); the difference in cure rates between LUNAR-301 and miR-124 + lipofectamine (4 out of 16 mice) was 35% (P = 0.0532). In a subcutaneous murine model of melanoma, tumor growth rate per day without treatment was 44% (i.e., tumor volume was expected to increase 44% cumulatively on a daily basis), while it was reduced to 26.1% in the LUNAR-301-treated group (P = 0.007), and to 16.2% in the LUNAR-302-treated group (P CONCLUSIONS: Nanoparticle delivery of miR-124 has a favorable safety and efficacy profile to justify implementation in client-owned canines or human clinical trials for the treatment of gliomas. Citation Format: Nasser K. Yaghi, Jun Wei, Ling-Yuan Kong, Yuuri Hashimoto, Edjah K. Nduom, Neal Huang, Xiaoyang Ling, Shouhao Zhou, Jonathan M. Levine, Virginia R. Fajt, Kiyoshi Tachikawa, Padmanabh Chivukula, David C. Webb, Joseph E. Payne, Amy B. Heimberger. An optimized therapeutic nanoparticle delivery platform of miRNA in preclinical murine models of malignancy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 4291. doi:10.1158/1538-7445.AM2015-4291