Efficacy of Chemotherapy, Phd-Inhibitor Molidustat or BRD4 Inhibitor JQ1 in Combination with Targeted Inhibition of Mutated IDH1 in Human AML In Vivo
Authors: Chaturvedi Anuhar , Gupta Charu , Kaulfuss Stefan , Othman Basem , Görlich Kerstin , Schottmann Renate , Panknin Olaf , Wagner Markus , Ganser Arnold , Jeffers Michael , Haegebarth Andrea , Heuser Michael 
Published: November 2019

Background: Approximately 40% of AML patients with IDH1 mutation respond to monotherapy with the IDH1 inhibitor ivosidenib with a median duration of response of 8.2 months, suggesting that IDH1 inhibitors should be rationally combined with other agents to improve efficacy. We have previously shown the synergistic activity of the mutant IDH1 (mIDH1) inhibitor BAY1436032 with azacitidine. We have also shown that the leukemogenic activity of the mIDH1 protein depends on PHD3 independent of R-2HG and its inhibition as a novel therapeutic strategy (Chaturvedi et al. 2018). Inhibition of Brd4 has been shown to induce rapid differentiation and death of IDH2 mutated AML mouse models (Chen et al., 2013). In the present study, we assessed the combination of either conventional chemotherapy, prolyl hydroxylase (PHD) inhibitor molidustat or bromodomain inhibitor JQ1 with BAY1436032 (BAY) in a preclinical patient-derived xenograft (PDX) model of mIDH1 AML.
Methods and Results: Leukemic cells from an AML patient with mutated IDH1, NPM1, FLT3-TKD and NRAS were xenografted in immunocompromised mice. We investigated the effects of BAY in sequential (seq) or simultaneous (sim) combination with cytarabine plus doxorubicin in our mIDH1 PDX model. The control groups were treated with either vehicle, BAY (150 mg/kg once daily p.o. continuously), or chemotherapy, which consisted of cytarabine (50 mg/kg once daily days 1-5 i.v.) and doxorubicin (1 mg/kg once daily days 1-3 i.v.). The treatment was repeated once after 29 days. The test groups were treated with BAY and chemotherapy in the doses mentioned above either starting both drugs on day 1 (sim group) or starting chemotherapy on day 1 and BAY on day 6 (seq group). Treatment with BAY was stopped after 12 weeks. Leukemic cells in peripheral blood constantly increased in vehicle and chemotherapy-treated mice with median time to 50% engraftment (MT 50%) of 84 and 112 days respectively. After stop of treatment, the percentage of leukemic cells increased in the group receiving BAY1436032 (MT 50%: 252 days) and sequential combination (MT 50%: 280 days), however, the MT50% was not reached with the simultaneous treatment (P<0.001). Median survival for vehicle-treated mice was 173 days, 206 days for chemotherapy-treated mice, 325 days for BAY treated mice, and 340 days for the sequential combination treatment. Strikingly, 5/8 mice treated simultaneously with BAY and chemotherapy survived until the end of the study at 400 days and the median survival was not reached.
We hypothesized that combination therapy with BAY and molidustat may demonstrate superior anti-leukemic activity in comparison to single agents in the treatment of IDH1-mutant AML. mIDH1 PDX mice were treated with either vehicle, BAY at a dose of 150 mg/kg or molidustat at a dose of 10 mg/kg p.o. as monotherapy or in combination. The MT50% for Vehicle, Molidustat and BAY was 70, 70 and 182 days respectively,. However, the MT50% was significantly delayed (322 days) for the combination treatment (P<0.001). Interestingly, all mice treated with the combination had normal blood counts until week 40 and survived significantly longer than in the BAY group (median latency: 249 vs 179 days, P<0.001).
Mutant IDH1 PDX mice were treated with either vehicle, BAY, BRD4 inhibitor JQ1 (50mg/kg i.p. once daily) or the combination of BAY and JQ1 for 12 weeks. JQ1 monotherapy showed a significant delay in leukemia engraftment compared to vehicle-treated mice in the first 12 weeks of treatment. During the treatment, the leukemic cells remained low in both BAY and combination treated mice but relapsed at the same time 4 weeks after the treatment had been stopped. Median survival for vehicle-treated mice was 139 days, 164 days for JQ1-treated mice, 220 days for BAY treated mice, and 242 days for the combination treatment (BAY vs combination; P=0.03).
Conclusion: Similar to the combination treatment of BAY with azacitidine the concurrent administration of BAY with chemotherapy significantly improves efficacy of this combination compared to sequential administration. In addition, the combination of an IDH1 inhibitor with molidustat is a promising therapeutic approach, while the combination with the BRD4 inhibitor JQ1 did not improve the outcome compared to treatment with an IDH1 inhibitor alone. Our findings support ongoing and future clinical investigations and suggest that IDH1 inhibitors should be applied concurrently with chemotherapy.

Disclosures
Chaturvedi: Bayer Pharma AG, Berlin, Germany: Research Funding. Kaulfuss:Bayer Pharma AG, Berlin, Germany: Employment. Panknin:Bayer Pharma AG, Berlin, Germany: Employment. Wagner:Bayer Pharma AG, Berlin, Germany: Employment, Equity Ownership. Jeffers:Bayer Pharma AG, Whippany, NJ, USA: Employment. Haegebarth:Bayer Pharma AG, Berlin, Germany: Employment, Equity Ownership. Heuser:Synimmune: Research Funding; Bayer Pharma AG, Berlin: Research Funding.