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Background KRAS is mutated in ∼90% of pancreatic ductal adenocarcinomas, ∼35% of colorectal cancers and ∼20% of non-small-cell lung cancers. There has been recent progress in targeting G12CKRAS specifically, but therapeutic options for other mutant forms of KRAS are limited, largely because the complexity of downstream signaling and feedback mechanisms mean that targeting individual pathway components is ineffective. Design The protein kinases RAF and SRC are validated therapeutic targets in KRAS-mutant pancreatic ductal adenocarcinomas, colorectal cancers and non-small-cell lung cancers and we show that both must be inhibited to block growth of these cancers. We describe CCT3833, a new drug that inhibits both RAF and SRC, which may be effective in KRAS-mutant cancers. Results We show that CCT3833 inhibits RAF and SRC in KRAS-mutant tumors in vitro and in vivo, and that it inhibits tumor growth at well-tolerated doses in mice. CCT3833 has been evaluated in a phase I clinical trial (NCT02437227) and we report here that it significantly prolongs progression-free survival of a patient with a G12VKRAS spindle cell sarcoma who did not respond to a multikinase inhibitor and therefore had limited treatment options. Conclusions New drug CCT3833 elicits significant preclinical therapeutic efficacy in KRAS-mutant colorectal, lung and pancreatic tumor xenografts, demonstrating a treatment option for several areas of unmet clinical need. Based on these preclinical data and the phase I clinical unconfirmed response in a patient with KRAS-mutant spindle cell sarcoma, CCT3833 requires further evaluation in patients with other KRAS-mutant cancers., Highlights • We synthesized new drug, CCT3833, that inhibits both RAF and SRC, and so may be effective in KRAS-mutant cancers. • CCT3833 inhibits both signaling pathways in vitro and in vivo, inhibits tumor growth and leads to regressions in mice. • CCT3833 significantly prolonged progression-free survival of a patient with a G12VKRAS spindle cell sarcoma. • CCT3833 is a potential treatment option for several areas of unmet clinical need.

Summary BRAF and MEK inhibitors are effective in BRAF mutant melanoma, but most patients eventually relapse with acquired resistance, and others present intrinsic resistance to these drugs. Resistance is often mediated by pathway reactivation through receptor tyrosine kinase (RTK)/SRC-family kinase (SFK) signaling or mutant NRAS, which drive paradoxical reactivation of the pathway. We describe pan-RAF inhibitors (CCT196969, CCT241161) that also inhibit SFKs. These compounds do not drive paradoxical pathway activation and inhibit MEK/ERK in BRAF and NRAS mutant melanoma. They inhibit melanoma cells and patient-derived xenografts that are resistant to BRAF and BRAF/MEK inhibitors. Thus, paradox-breaking pan-RAF inhibitors that also inhibit SFKs could provide first-line treatment for BRAF and NRAS mutant melanomas and second-line treatment for patients who develop resistance., Graphical Abstract, Highlights • pan-RAF inhibitors also inhibit SRC family kinases • The compounds do not induce paradoxical activation of ERK in RAS mutant cells • The compounds are active in BRAF and NRAS mutant melanomas • The compounds are active in PDXs resistant to BRAF or BRAF plus MEK inhibitors, Girotti et al. describe two pan-RAF inhibitors that also inhibit SRC-family kinases. These compounds do not drive paradoxical MEK/ERK activation and can inhibit MEK in NRAS mutant cells. Moreover, the agents can overcome resistance to clinical BRAF or combination BRAF/MEK inhibitors in patient-derived xenografts.

The tankyrase proteins (TNKS, TNKS2), members of the PARP superfamily of enzymes, are attractive anti-cancer drug targets, particularly as inhibition of their catalytic activity has been shown to antagonise oncogenic WNT signalling. To identify chemical inhibitors of tankyrase we carried out an in silico small molecule screen using a set of ‘PARP-binding’ pharmacophores together with a generated (liganded) tankyrase homology model. This approach identified a structurally diverse set of ~1000 compounds for further study. Subsequent in vitro screening of recombinant tankyrase protein identified a subset of 59 confirmed inhibitors. Early optimisation followed by cell-based studies in WNT-dependent tumour cells, as well as co-crystallisation studies, identified a novel class of 3-aryl-5-substituted isoquinolin-1-ones, such as 21, that exhibit potent inhibition of tankyrase activity as well as growth inhibition of colorectal cancer cells.

The protein kinase BRAF is mutated ∼40% of human melanomas. BRAF is a component of the RAS/RAF/MEK/ERK pathway and BRAF or MEK inhibitors increase progression-free and overall survival in melanoma patients with BRAF mutations. However, most patients relapse with acquired resistance and ∼20% of patients present intrinsic resistance and do not respond to these drugs. We describe here two novel compounds that target mutant BRAF and wild-type CRAF. Our compounds inhibited the growth of melanoma cells that were resistant to BRAF-selective inhibitors. ERK pathway reactivation is responsible for resistance to BRAF targeted therapies in ∼60% of the patients and in ∼25% of patients resistance is driven by acquisition of mutations in NRAS. We show that our compounds inhibited the growth of melanoma cells that were resistant to BRAF-selective inhibitors due to pathway reactivation mediated by different mechanisms. We show that the drugs were active against patient derived xenografts (PDXs) from patients with acquired or intrinsic resistance to BRAF-selective inhibitors and in whose tumors resistance was associated with ERK pathway reactivation. Further, our compounds are active in a PDX from a patient whose tumor developed acquired resistance to a combination of a BRAF-selective plus a MEK inhibitor and associated with acquisition of an NRAS mutation. Thus, our panRAF inhibitors can inhibit melanomas with different mechanisms of acquired or intrinsic resistance to BRAF-selective and BRAF-selective/MEK inhibitor combinations, potentially providing first-line treatment for naïve patients and second-line treatments for a range of relapsed patients. Citation Format: Maria R. Girotti, Filipa Lopes, Natasha Preece, Dan Niculescu-Duvaz, Alfonso Zambon, Lawrence Davies, Steven Whittaker, Grazia Saturno, Amaya Viros, Malin Pedersen, Bart MJM Suijkerbuijk, Delphine Menard, Robert Mcleary, Louise Johnson, Laura Fish, Sarah Ejiama, Berta Sanchez-Laorden, Neil Carragher, Kenneth Macleod, Garry Ashton, Anna Marusiak, Alberto Fusi, John Brognard, Margaret Frame, Paul Lorigan, Caroline J. Springer, Richard Marais. Novel panRAF inhibitors active in melanomas that are resistant to BRAF-selective, or BRAF-selective/MEK inhibitor combinations. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 3704. doi:10.1158/1538-7445.AM2014-3704

KRAS is mutated in ∼80% of pancreatic ductal adenocarcinoma (PDAC), ∼35% of colorectal cancer (CRC) and ∼20% of non-small-cell lung cancer (NSCLC). KRAS remains an intractable drug target and targeting the downstream pathway component is ineffective because feedback mechanisms or parallel pathways provide alternative routes to cell proliferation and/or survival. Here we show that a new panRAF/SRC inhibitor, CCT3833, is active in KRAS-mutant PDAC, CRC and NSCLC. We demonstrate that CCT3833 inhibits tumor growth in several KRAS-driven cancers via inhibition of RAF and SRC, eliciting therapeutic efficacy at well-tolerated doses in mouse models of human cancer. CCT3833 has already entered clinical trials (NCT02437227) for BRAF mutant and BRAF inhibitor-resistant melanomas and these new data show that it is also effective in KRAS mutant cancers, potentially providing a new therapeutic option for these patients. Citation Format: Grazia Saturno, Filipa Lopes, Maria Romina Girotti, Ion Niculescu-Duvaz, Dan Niculescu-Duvaz, Alfonso Zambon, Lawrence Davies, Louise Johnson, Natasha Preece, Amaya Viros, Malin Pedersen, Robert McLeary, Ruth Knight, Rebecca Lee, Denys Holovanchuk, Paul Lorigan, Nathalie Dhomen, Richard Marais, Caroline Springer. Therapeutic efficacy of the paradox-breaking panRAF and SRC drug CCT3833/BAL3833 in KRAS-driven cancer models. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-212.