KRASG12C inhibition using MRTX1257: a novel radio-sensitizing partner.

BACKGROUND: KRAS activating mutations are considered the most frequent oncogenic drivers and are correlated with radio-resistance in multiple cancers including non-small cell lung cancer (NSCLC) and colorectal cancer. Although KRAS was considered undruggable until recently, several KRAS inhibitors have recently reached clinical development. Among them, MRTX849 (Mirati Therapeutics) showed encouraging clinical outcomes for the treatment of selected patients with KRASG12C mutated NSCLC and colorectal cancers. In this work, we explore the ability of MRTX1257, a KRASG12C inhibitor analogous to MRTX849, to radio-sensitize KRASG12C+/+ mutated cell lines and tumors. METHODS: Both in vitro and in vivo models of radiotherapy (RT) in association with MRTX1257 were used, with different RAS mutational profiles. We assessed in vitro the radio-sensitizing effect of MRTX1257 in CT26 KRASG12C+/+, CT26 WT, LL2 WT and LL2 NRAS KO (LL2 NRAS-/-) cell lines. In vivo, we used syngeneic models of subcutaneous CT26 KRASG12C+/+ tumors in BALB/c mice and T cell deficient athymic nu/nu mice to assess both the radio-sensitizing effect of MRTX1257 and its immunological features. RESULTS: MRTX1257 was able to radio-sensitize CT26 KRASG12C+/+ cells in vitro in a time and dose dependent manner. Moreover, RT in association with MRTX1257 in BALB/c mice bearing CT26 KRASG12C+/+ subcutaneous tumors resulted in an observable cure rate of 20%. However, no durable response was observed with similar treatment in athymic nude mice. The analysis of the immune microenvironment of CT26 KRASG12C+/+ tumors following RT and MRTX1257 showed an increase in the proportion of various cell subtypes including conventional CD4 + T cells, dendritic cells type 2 (cDC2) and inflammatory monocytes. Furthermore, the expression of PD-L1 was dramatically down-regulated within both tumor and myeloid cells, thus illustrating the polarization of the tumor microenvironment towards a pro-inflammatory and anti-tumor phenotype following the combined treatment. CONCLUSION: This work is the first to demonstrate in vitro as in vivo the radio-sensitizing effect of MRTX1257, a potent KRASG12C inhibitor compatible with oral administration, in CT26 KRASG12C mutated cell lines and tumors. This is a first step towards the use of new combinatorial strategies using KRAS inhibitors and RT in KRASG12C mutated tumors, which are the most represented in NSCLC with 14% of patients harboring this mutational profile.

Differential therapeutic effects of PARP and ATR inhibition combined with radiotherapy in the treatment of subcutaneous versus orthotopic lung tumour models.

BACKGROUND: Subcutaneous mouse tumour models are widely used for the screening of novel antitumour treatments, although these models are poor surrogate models of human cancers. METHODS: We compared the antitumour efficacy of the combination of ionising radiation (IR) with two DNA damage response inhibitors, the PARP inhibitor olaparib and the ATR inhibitor AZD6738 (ceralasertib), in subcutaneous versus orthotopic cancer models. RESULTS: Olaparib delayed the growth of irradiated Lewis lung carcinoma (LL2) subcutaneous tumours, in agreement with previous reports in human cell lines. However, the olaparib plus IR combination showed a very narrow therapeutic window against LL2 lung orthotopic tumours, with nearly no additional antitumour effect compared with that of IR alone, and tolerability issues emerged at high doses. The addition of AZD6738 greatly enhanced the efficacy of the olaparib plus IR combination treatment against subcutaneous but not orthotopic LL2 tumours. Moreover, olaparib plus AZD6738 administration concomitant with IR even worsened the response to radiation of head and neck orthotopic tumours and induced mucositis. CONCLUSIONS: These major differences in the responses to treatments between subcutaneous and orthotopic models highlight the importance of using more pathologically relevant models, such as syngeneic orthotopic models, to determine the most appropriate therapeutic approaches for translation to the clinic.

Optimal dosing regimen of CD73 blockade improves tumor response to radiotherapy through iCOS downregulation.

BACKGROUND: Irradiation (IR) and immune checkpoint inhibitor (ICI) combination is a promising treatment modality. However, local and distance treatment failure and resistance can occur. To counteract this resistance, several studies propose CD73, an ectoenzyme, as a potential target to improve the antitumor efficiency of IR and ICI. Although CD73 targeting in combination with IR and ICI has shown attractive antitumor effects in preclinical models, the rationale for CD73 targeting based on CD73 tumor expression level deserves further investigations. METHODS: Here we evaluated for the first time the efficacy of two administration regimens of CD73 neutralizing antibody (one dose vs four doses) in combination with IR according to the expression level of CD73 in two subcutaneous tumor models expressing different levels of CD73. RESULTS: We showed that CD73 is weakly expressed by MC38 tumors even after IR, when compared with the TS/A model that highly expressed CD73. Treatment with four doses of anti-CD73 improved the TS/A tumor response to IR, while it was ineffective against the CD73 low-expressing MC38 tumors. Surprisingly, a single dose of anti-CD73 exerted a significant antitumor activity against MC38 tumors. On CD73 overexpression in MC38 cells, four doses of anti-CD73 were required to improve the efficacy of IR. Mechanistically, a correlation between a downregulation of iCOS expression in CD4+ T cells and an improved response to IR after anti-CD73 treatment was observed and iCOS targeting could restore an impaired benefit from anti-CD73 treatment. CONCLUSIONS: These data emphasize the importance of the dosing regimen for anti-CD73 treatment to improve tumor response to IR and identify iCOS as part of the underlying molecular mechanisms. Our data suggest that the selection of appropriate dosing regimen is required to optimize the therapeutic efficacy of immunotherapy-radiotherapy combinations.

TGFß receptor inhibition unleashes interferon-ß production by tumor-associated macrophages and enhances radiotherapy efficacy.

BACKGROUND: Transforming growth factor-beta (TGFß) can limit the efficacy of cancer treatments, including radiotherapy (RT), by inducing an immunosuppressive tumor environment. The association of TGFß with impaired T cell infiltration and antitumor immunity is known, but the mechanisms by which TGFß participates in immune cell exclusion and limits the efficacy of antitumor therapies warrant further investigations. METHODS: We used the clinically relevant TGFß receptor 2 (TGFßR2)-neutralizing antibody MT1 and the small molecule TGFßR1 inhibitor LY3200882 and evaluated their efficacy in combination with RT against murine orthotopic models of head and neck and lung cancer. RESULTS: We demonstrated that TGFß pathway inhibition strongly increased the efficacy of RT. TGFßR2 antibody upregulated interferon beta expression in tumor-associated macrophages within the irradiated tumors and favored T cell infiltration at the periphery and within the core of the tumor lesions. We highlighted that both the antitumor efficacy and the increased lymphocyte infiltration observed with the combination of MT1 and RT were dependent on type I interferon signaling. CONCLUSIONS: These data shed new light on the role of TGFß in limiting the efficacy of RT, identifying a novel mechanism involving the inhibition of macrophage-derived type I interferon production, and fostering the use of TGFßR inhibition in combination with RT in therapeutic strategies for the management of head and neck and lung cancer.

Dual oxidase 1 limits the IFNγ-associated antitumor effect of macrophages.

BACKGROUND: Macrophages play pivotal roles in tumor progression and the response to anticancer therapies, including radiotherapy (RT). Dual oxidase (DUOX) 1 is a transmembrane enzyme that plays a critical role in oxidant generation. METHODS: Since we found DUOX1 expression in macrophages from human lung samples exposed to ionizing radiation, we aimed to assess the involvement of DUOX1 in macrophage activation and the role of these macrophages in tumor development. RESULTS: Using Duox1-/- mice, we demonstrated that the lack of DUOX1 in proinflammatory macrophages improved the antitumor effect of these cells. Furthermore, intratumoral injection of Duox1-/- proinflammatory macrophages significantly enhanced the antitumor effect of RT. Mechanistically, DUOX1 deficiency increased the production of proinflammatory cytokines (IFNγ, CXCL9, CCL3 and TNFα) by activated macrophages in vitro and the expression of major histocompatibility complex class II in the membranes of macrophages. We also demonstrated that DUOX1 was involved in the phagocytotic function of macrophages in vitro and in vivo. The antitumor effect of Duox1-/- macrophages was associated with a significant increase in IFNγ production by both lymphoid and myeloid immune cells. CONCLUSIONS: Our data indicate that DUOX1 is a new target for macrophage reprogramming and suggest that DUOX1 inhibition in macrophages combined with RT is a new therapeutic strategy for the management of cancers.

Preventing Radiation-Induced Injury by Topical Application of an Amifostine Metabolite-Loaded Thermogel.

PURPOSE: Despite the development of high-precision radiation therapy, ionizing radiation inevitably damages healthy tissues. Radiodermatitis and radioinduced oral mucositis are frequent and significant side effects among patients with breast and head and neck cancer, respectively. These radiation-related injuries negatively affect patient quality of life and can lead to unplanned therapeutic breaks and compromise treatment outcomes. Currently, no preventive or mitigating agent has emerged to address these issues. Although amifostine, a well-known free radical scavenger, has proven efficacy against specific radio- and chemo-induced toxicities, severe adverse side effects (reversible hypotension, nausea, emesis, etc) combined with logistical hurdles are associated with its recommended intravenous route of administration, limiting its use. METHODS AND MATERIALS: We developed a thermogel containing the active thiol metabolite of amifostine (CPh-1014) that polymerizes at body temperature and serves as a matrix for topical application onto the skin or mucosa. RESULTS: Applied before irradiation, CPh-1014 greatly reduced the severity of oral mucositis and dermatitis induced by either a single dose or fractionated irradiation regimens in in vivo mouse models. The cytoprotective effect of CPh-1014 was confirmed by the decrease in DNA double-strand breaks in the irradiated epithelium. Noticeably, CPh-1014 did not affect radiation therapy efficacy against tumors grafted at submucosal and subcutaneous sites. In contrast to the intravenous administration of amifostine, CPh-1014 oral application did not induce hypotension in dogs. CONCLUSIONS: CPh-1014 confers radioprotective effects in healthy tissues with reduced systemic side effects without compromising radiation therapy efficacy. We propose CPh-1014 as an easy-to-implement therapeutic approach to alleviate radiation therapy toxicity in patients with breast and head and neck cancer.

The MET/AXL/FGFR Inhibitor S49076 Impairs Aurora B Activity and Improves the Antitumor Efficacy of Radiotherapy.

Several therapeutic agents targeting HGF/MET signaling are under clinical development as single agents or in combination, notably with anti-EGFR therapies in non-small cell lung cancer (NSCLC). However, despite increasing data supporting a link between MET, irradiation, and cancer progression, no data regarding the combination of MET-targeting agents and radiotherapy are available from the clinic. S49076 is an oral ATP-competitive inhibitor of MET, AXL, and FGFR1-3 receptors that is currently in phase I/II clinical trials in combination with gefitinib in NSCLC patients whose tumors show resistance to EGFR inhibitors. Here, we studied the impact of S49076 on MET signaling, cell proliferation, and clonogenic survival in MET-dependent (GTL16 and U87-MG) and MET-independent (H441, H460, and A549) cells. Our data show that S49076 exerts its cytotoxic activity at low doses on MET-dependent cells through MET inhibition, whereas it inhibits growth of MET-independent cells at higher but clinically relevant doses by targeting Aurora B. Furthermore, we found that S49076 improves the antitumor efficacy of radiotherapy in both MET-dependent and MET-independent cell lines in vitro and in subcutaneous and orthotopic tumor models in vivo In conclusion, our study demonstrates that S49076 has dual antitumor activity and can be used in combination with radiotherapy for the treatment of both MET-dependent and MET-independent tumors. These results support the evaluation of combined treatment of S49076 with radiation in clinical trials without patient selection based on the tumor MET dependency status. Mol Cancer Ther; 16(10); 2107-19. ©2017 AACR.