FF-10101

Acute myeloid leukaemia (AML) is a hard to treat blood cancer. Mutations in FLT3 are common among the genetic aberrations that characterise the cancer. Patients initially react to FLT3 inhibitors but drug resistance is a hinder to successful therapy. To better understand the mechanisms leading to drug resistance, we generated four AML cell lines resistant to the inhibitors gilteritinib or FF-10101, and explored their resistance mechanisms. We further tested whether the novel inhibitor Chen-9u could be used to limit cell growth. The results showed that each of the four resistant cell lines became resistant through a different mechanism. Resistant cells showed decreased FLT3 and increased NRAS pathway activity and reduced DNA synthesis due to decrease in CDK4 activity. Resistance mechanisms included resistance mutations in FLT3 (C695F and N701K), and a novel mutation in NRAS (G12C). In a fourth line, resistance might have developed through a MYCN mutation. Cell growth was inhibited by Chen-9u and resistant clones could not be obtained with this inhibitor. The results highlight opportunities and limitations. On the one hand, resistant cells were produced due to different mechanisms, showing the versatility of tumour cells. Furthermore, resistance developed to the most advanced inhibitors, one of which is covalent and the other non-covalent but highly specific. On the other hand, it is shown that DNA synthesis is reduced, which means that resistance has evolutionary consequences. Finally, the novel drug-resistant cell lines may serve as useful models for better understanding of the cellular events associated with inherent and acquired drug resistance.

Tumor-associated macrophages (TAMs) are one of the key immunosuppressive components in the tumor microenvironment (TME) and contribute to tumor development, progression, and resistance to cancer immunotherapy. Several reagents targeting TAMs have been tested in preclinical and clinical studies, but they have had limited success. Here, we show that a unique reagent, FF-10101, exhibited a sustained inhibitory effect against colony-stimulating factor 1 receptor by forming a covalent bond and reduced immunosuppressive TAMs in the TME, which led to strong antitumor immunity. In preclinical animal models, FF-10101 treatment significantly reduced immunosuppressive TAMs and increased antitumor TAMs in the TME. In addition, tumor antigen-specific CD8+ T cells were increased; consequently, tumor growth was significantly inhibited. Moreover, combination treatment with an anti-programmed cell death 1 (anti-PD-1) antibody and FF-10101 exhibited a far stronger antitumor effect than either treatment alone. In human cancer specimens, FF-10101 treatment reduced programmed cell death 1 ligand 1 (PD-L1) expression on TAMs, as observed in animal models. Thus, FF-10101 acts as an immunomodulatory agent that can reduce immunosuppressive TAMs and augment tumor antigen-specific T cell responses, thereby generating an immunostimulatory TME. We propose that FF-10101 is a potential candidate for successful combination cancer immunotherapy with immune checkpoint inhibitors, such as PD-1/PD-L1 blockade.