Modulation of T Cell Compartment in a Preclinical CLL Murine Model By a Selective PI3K Delta Inhibitor, TGR-1202

INTRODUCTION: In the microenvironment of CLL, T cells are characteristically dysfunctional compared to a normal, healthy setting. Idelalisib, an approved PI3Kδ inhibitor has shown successful clinical response in relapsed/refractory CLL, but is associated with a high rate of discontinuation because of likely immune-related severe adverse events (SAE). More recently, increased toxicity has been reported in treatment-naïve patient trials. The described hepatotoxicity, colitis, and pneumonitis appear to be associated with changes in the T cell compartment and specifically regulatory T cells (Tregs). In the phase I trials of the next generation PI3Kδ inhibitor, TGR-1202, a comparable rate of clinical responses have been reported with apparently less toxicity than prior PI3Kδ inhibitors, even with long term follow-up (Burris et al, ASCO 2016). We previously demonstrated in vitrothat TGR-1202 relatively preserved the number and function of normal human T cell subsets, including Tregs, when compared to the other clinically available PI3Kδ inhibitors (Maharaj et al AACR 2016) Herein, we aimed to further investigate how TGR-1202 regulates T cell subsets in a preclinical murine model of CLL.

METHODS: For the preclinical model of CLL, we adoptively transferred 25x10^6 leukemic euTCL1 splenocytes into wildtype mice. Following establishment of disease, we orally administered TGR-1202 or duvelisib (PI3Kδγ inhibitor) and quantified immune changes over time using flow cytometry to detect T cell subsets and associated functional markers. Duvelisb was chosen as a comparator since idelalisib is not suitable for murine models, and duvelisib has been shown to have similar efficacy and toxicity as idelalisib in preclinical and clinical studies.

RESULTS: First, we confirmed general immune changes typical of CLL progression. Peripheral white blood cell (WBC) count was significantly higher in leukemic mice compared to wildtype (p=0.02). Following oral TGR-1202 or duvelisib treatment, WBC count decreased significantly over time (p<0.001 respectively), suggesting comparable efficacy in disease eradication. Both inhibitors also significantly reduced overall circulating CD3+ T cell number, however TGR-1202 differentially regulated T cell subset ratios previously reported to regulate autoimmunity. These effects were more pronounced in the CLL setting than wildtype. While Tcon:Teff (CD4:CD8) ratio remained intact, Th1:Th2 ratio was increased. Total Treg number was significantly decreased following treatment, leading to increased Tcon:Treg and Teff:Treg ratios. On the Treg population, TGR-1202 maintained expression of functional markers PD-1 and CTLA-4 closer to control when compared to duvelisib (TGR-1202 vs. control PD-1: p=ns, CTLA-4: p=ns ; duvelisib vs. control PD-1: p=0.02* CTLA-4: p=0.03*). This indicated greater retention of Treg suppressive capacity in vivo inTGR-1202 group. Interestingly, overall Th17 number was reduced in both treatments over time but there was greater disruption of Treg:Th17 ratio in duvelisib group (duvelisib vs. control Treg/Th17 p=0.04* ; TGR-1202 vs. control Treg/Th17 p=ns ). In perspective, maintenance of this balance has been extensively reported to mediate autoimmunity. In order to determine susceptibility to apoptosis, we quantified CD95 (FASL) expression. Lower CD95 mean fluorescence intensity (MFI) was observed on CD4, CD8 and Treg populations in both periphery and spleen of TGR-1202 group compared to those treated with duvelisib. Pathology studies are ongoing to assess the toxicity after TGR-1202 or duvelisib treatment in these mice.

CONCLUSIONS: In conclusion, we have demonstrated the efficacy of two PI3Kδinhibitors in a murine CLL model and their differential impact on Treg and Th17 compartments, which may be of relevance to study the toxicity in this class of drugs. Extensive studies of the immune modulating effects of TGR-1202 are particularly of interest given its current clinical development and considering the lower rate of SAE observed in trials.