Reshaping the Tumor Microenvironment of KRASG12D Pancreatic Ductal Adenocarcinoma with combined SOS1 and MEK Inhibition for Improved Immunotherapy Response.

KRAS inhibitors have demonstrated exciting pre-clinical and clinical responses, although resistance occurs rapidly. Here, we investigate the effects of KRAS-targeting therapies on the tumor microenvironment using a library of KRASG12D, p53 mutant, murine PDAC-derived cell lines (KPCY) to leverage immune-oncology combination strategies for long-term tumor efficacy. Our findings show that SOS1 and MEK inhibitors (SOS1i+MEKi) suppressed tumor growth in syngeneic models and increased intra-tumoral CD8+ T cells without durable responses. scRNA-sequencing revealed an increase in inflammatory cancer associated fibroblasts (iCAFs), M2 macrophages, and a decreased dendritic cell quality that ultimately resulted in a highly immunosuppressive microenvironment driven by IL6+ iCAFs. Agonist CD40 treatment was effective to revert macrophage polarization and overcome the lack of mature antigen presenting DCs after SOS1i+MEKi therapy. Treatment increased the overall survival of KPCY tumor-bearing mice. The addition of checkpoint blockade to SOS1i+MEKi combination resulted in tumor free mice with established immune memory. Our data suggests that KRAS inhibition affects myeloid cell maturation and highlights the need for combining KRAS cancer-targeted therapy with myeloid activation to enhance and prolong anti-tumor effects.

SOS1 inhibition enhances the efficacy of and delays resistance to G12C inhibitors in lung adenocarcinoma.

Clinical effectiveness of KRAS G12C inhibitors (G12Cis) is limited both by intrinsic and acquired resistance, necessitating the development of combination approaches. We found that targeting proximal receptor tyrosine kinase (RTK) signaling using the SOS1 inhibitor (SOS1i) BI-3406 both enhanced the potency of and delayed resistance to G12Ci treatment, but the extent of SOS1i effectiveness was modulated by both SOS2 expression and the specific mutational landscape. SOS1i enhanced the efficacy of G12Ci and limited rebound RTK/ERK signaling to overcome intrinsic/adaptive resistance, but this effect was modulated by SOS2 protein levels. Survival of drug-tolerant persister (DTP) cells within the heterogeneous tumor population and/or acquired mutations that reactivate RTK/RAS signaling can lead to outgrowth of tumor initiating cells (TICs) that drive therapeutic resistance. G12Ci drug tolerant persister cells showed a 2-3-fold enrichment of TICs, suggesting that these could be a sanctuary population of G12Ci resistant cells. SOS1i re-sensitized DTPs to G12Ci and inhibited G12C-induced TIC enrichment. Co-mutation of the tumor suppressor KEAP1 limits the clinical effectiveness of G12Cis, and KEAP1 and STK11 deletion increased TIC frequency and accelerated the development of acquired resistance to G12Ci in situ. SOS1i both delayed acquired G12Ci resistance and limited the total number of resistant colonies regardless of KEAP1 and STK11 mutational status. These data suggest that SOS1i could be an effective strategy to both enhance G12Ci efficacy and prevent G12Ci resistance regardless of co-mutations.

Discovery of BI 7446: A Potent Cyclic Dinucleotide STING Agonist with Broad-Spectrum Variant Activity for the Treatment of Cancer.

Activating the stimulator of interferon genes (STING) pathway with STING agonists is an attractive immune oncology concept to treat patients with tumors that are refractory to single-agent anti-PD-1 therapy. For best clinical translatability and broad application to cancer patients, STING agonists with potent cellular activation of all STING variants are desired. Novel cyclic dinucleotide (CDN)-based selective STING agonists were designed and synthesized comprising noncanonical nucleobase, ribose, and phosphorothioate moieties. This strategy led to the discovery of 2',3'-CDN 13 (BI 7446), which features unprecedented potency and activates all five STING variants in cellular assays. ADME profiling revealed that CDN 13 has attractive drug-like properties for development as an intratumoral agent. Injection of low doses of CDN 13 into tumors in mice induced long-lasting, tumor-specific immune-mediated tumor rejection. Based on its compelling preclinical profile, BI 7446 has been advanced to clinical trials (monotherapy and in combination with anti-PD-1 antibody).

Combining SOS1 and MEK Inhibitors in a Murine Model of Plexiform Neurofibroma Results in Tumor Shrinkage.

Individuals with neurofibromatosis type 1 develop rat sarcoma virus (RAS)-mitogen-activated protein kinase-mitogen-activated and extracellular signal-regulated kinase (RAS-MAPK-MEK)-driven nerve tumors called neurofibromas. Although MEK inhibitors transiently reduce volumes of most plexiform neurofibromas in mouse models and in neurofibromatosis type 1 (NF1) patients, therapies that increase the efficacy of MEK inhibitors are needed. BI-3406 is a small molecule that prevents Son of Sevenless (SOS)1 interaction with Kirsten rat sarcoma viral oncoprotein (KRAS)-GDP, interfering with the RAS-MAPK cascade upstream of MEK. Single agent SOS1 inhibition had no significant effect in the DhhCre;Nf1 fl/fl mouse model of plexiform neurofibroma, but pharmacokinetics (PK)-driven combination of selumetinib with BI-3406 significantly improved tumor parameters. Tumor volumes and neurofibroma cell proliferation, reduced by MEK inhibition, were further reduced by the combination. Neurofibromas are rich in ionized calcium binding adaptor molecule 1 (Iba1)+ macrophages; combination treatment resulted in small and round macrophages, with altered cytokine expression indicative of altered activation. The significant effects of MEK inhibitor plus SOS1 inhibition in this preclinical study suggest potential clinical benefit of dual targeting of the RAS-MAPK pathway in neurofibromas. SIGNIFICANCE STATEMENT: Interfering with the RAS-mitogen-activated protein kinase (RAS-MAPK) cascade upstream of mitogen activated protein kinase kinase (MEK), together with MEK inhibition, augment effects of MEK inhibition on neurofibroma volume and tumor macrophages in a preclinical model system. This study emphasizes the critical role of the RAS-MAPK pathway in controlling tumor cell proliferation and the tumor microenvironment in benign neurofibromas.

Combined KRASG12C and SOS1 inhibition enhances and extends the anti-tumor response in KRASG12C-driven cancers by addressing intrinsic and acquired resistance.

Efforts to improve the anti-tumor response to KRASG12C targeted therapy have benefited from leveraging combination approaches. Here, we compare the anti-tumor response induced by the SOS1-KRAS interaction inhibitor, BI-3406, combined with a KRASG12C inhibitor (KRASG12Ci) to those induced by KRASG12Ci alone or combined with SHP2 or EGFR inhibitors. In lung cancer and colorectal cancer (CRC) models, BI-3406 plus KRASG12Ci induces an anti-tumor response stronger than that observed with KRASG12Ci alone and comparable to those by the other combinations. This enhanced anti-tumor response is associated with a stronger and extended suppression of RAS-MAPK signaling. Importantly, BI-3406 plus KRASG12Ci treatment delays the emergence of acquired adagrasib resistance in both CRC and lung cancer models and is associated with re-establishment of anti-proliferative activity in KRASG12Ci-resistant CRC models. Our findings position KRASG12C plus SOS1 inhibition therapy as a promising strategy for treating both KRASG12C-mutated tumors as well as for addressing acquired resistance to KRASG12Ci.

Pharmacodynamic and Antitumor Activity of BI 836880, a Dual Vascular Endothelial Growth Factor and Angiopoietin 2 Inhibitor, Alone and Combined with Programmed Cell Death Protein-1 Inhibition.

Vascular endothelial growth factor (VEGF) and angiopoietin (ANG)-2 have complementary roles in angiogenesis and promote an immunosuppressive tumor microenvironment. It is anticipated that the combination of VEGF and ANG2 blockade could provide superior activity to the blockade of either pathway alone and that the addition of VEGF/ANG2 inhibition to an anti-programmed cell death protein-1 (PD-1) antibody could change the tumor microenvironment to support T-cell-mediated tumor cytotoxicity. Here, we describe the pharmacologic and antitumor activity of BI 836880, a humanized bispecific nanobody comprising two single-variable domains blocking VEGF and ANG2, and an additional module for half-life extension in vivo. BI 836880 demonstrated high affinity and selectivity for human VEGF-A and ANG2, resulting in inhibition of the downstream signaling of VEGF/ANG2 and a decrease in endothelial cell proliferation and survival. In vivo, BI 836880 exhibited significant antitumor activity in all patient-derived xenograft models tested, showing significantly greater tumor growth inhibition (TGI) than bevacizumab (VEGF inhibition) and AMG386 (ANG1/2 inhibition) in a range of models. In a Lewis lung carcinoma syngeneic tumor model, the combination of PD-1 inhibition with VEGF inhibition showed superior efficacy versus the blockade of either pathway alone. TGI was further increased with the addition of ANG2 inhibition to VEGF/PD-1 blockade. VEGF/ANG2 inhibition had a strong antiangiogenic effect. Our data suggest that the blockade of VEGF and ANG2 with BI 836880 may offer improved antitumor activity versus the blockade of either pathway alone and that combining VEGF/ANG2 inhibition with PD-1 blockade can further enhance antitumor effects. SIGNIFICANCE STATEMENT: Vascular endothelial growth factor (VEGF) and angiopoietin (ANG)-2 play key roles in angiogenesis and have an immunosuppressive effect in the tumor microenvironment. This study shows that BI 836880, a bispecific nanobody targeting VEGF and ANG2, demonstrates substantial antitumor activity in preclinical models. Combining VEGF/ANG2 inhibition with the blockade of the PD-1 pathway can further improve antitumor activity.

Expanding the Reach of Precision Oncology by Drugging All KRAS Mutants.

KRAS is the most frequently mutated oncogene, harboring mutations in approximately one in seven cancers. Allele-specific KRASG12C inhibitors are currently changing the treatment paradigm for patients with KRASG12C-mutated non-small cell lung cancer and colorectal cancer. The success of addressing a previously elusive KRAS allele has fueled drug discovery efforts for all KRAS mutants. Pan-KRAS drugs have the potential to address broad patient populations, including KRASG12D-, KRASG12V-, KRASG13D-, KRASG12R-, and KRASG12A-mutant or KRAS wild-type-amplified cancers, as well as cancers with acquired resistance to KRASG12C inhibitors. Here, we review actively pursued allele-specific and pan-KRAS inhibition strategies and their potential utility. SIGNIFICANCE: Mutant-selective KRASG12C inhibitors target a fraction (approximately 13.6%) of all KRAS-driven cancers. A broad arsenal of KRAS drugs is needed to comprehensively conquer KRAS-driven cancers. Conceptually, we foresee two future classes of KRAS medicines: mutant-selective KRAS drugs targeting individual variant alleles and pan-KRAS therapeutics targeting a broad range of KRAS alterations.

KRAS Secondary Mutations That Confer Acquired Resistance to KRAS G12C Inhibitors, Sotorasib and Adagrasib, and Overcoming Strategies: Insights From In Vitro Experiments.

INTRODUCTION: KRAS mutations have been recognized as undruggable for many years. Recently, novel KRAS G12C inhibitors, such as sotorasib and adagrasib, are being developed in clinical trials and have revealed promising results in metastatic NSCLC. Nevertheless, it is strongly anticipated that acquired resistance will limit their clinical use. In this study, we developed in vitro models of the KRAS G12C cancer, derived from resistant clones against sotorasib and adagrasib, and searched for secondary KRAS mutations as on-target resistance mechanisms to develop possible strategies to overcome such resistance. METHODS: We chronically exposed Ba/F3 cells transduced with KRASG12C to sotorasib or adagrasib in the presence of N-ethyl-N-nitrosourea and searched for secondary KRAS mutations. Strategies to overcome resistance were also investigated. RESULTS: We generated 142 Ba/F3 clones resistant to either sotorasib or adagrasib, of which 124 (87%) harbored secondary KRAS mutations. There were 12 different secondary KRAS mutations. Y96D and Y96S were resistant to both inhibitors. A combination of novel SOS1 inhibitor, BI-3406, and trametinib had potent activity against this resistance. Although G13D, R68M, A59S and A59T, which were highly resistant to sotorasib, remained sensitive to adagrasib, Q99L was resistant to adagrasib but sensitive to sotorasib. CONCLUSIONS: We identified many secondary KRAS mutations causing resistance to sotorasib, adagrasib, or both, in vitro. The differential activities of these two inhibitors depending on the secondary mutations suggest sequential use in some cases. In addition, switching to BI-3406 plus trametinib might be a useful strategy to overcome acquired resistance owing to the secondary Y96D and Y96S mutations.

One Atom Makes All the Difference: Getting a Foot in the Door between SOS1 and KRAS.

KRAS, the most common oncogenic driver in human cancers, is controlled and signals primarily through protein-protein interactions (PPIs). The interaction between KRAS and SOS1, crucial for the activation of KRAS, is a typical, challenging PPI with a large contact surface area and high affinity. Here, we report that the addition of only one atom placed between Y884SOS1 and A73KRAS is sufficient to convert SOS1 activators into SOS1 inhibitors. We also disclose the discovery of BI-3406. Combination with the upstream EGFR inhibitor afatinib shows in vivo efficacy against KRASG13D mutant colorectal tumor cells, demonstrating the utility of BI-3406 to probe SOS1 biology. These findings challenge the dogma that large molecules are required to disrupt challenging PPIs. Instead, a "foot in the door" approach, whereby single atoms or small functional groups placed between key PPI interactions, can lead to potent inhibitors even for challenging PPIs such as SOS1-KRAS.

BI-3406, a Potent and Selective SOS1-KRAS Interaction Inhibitor, Is Effective in KRAS-Driven Cancers through Combined MEK Inhibition.

KRAS is the most frequently mutated driver of pancreatic, colorectal, and non-small cell lung cancers. Direct KRAS blockade has proved challenging, and inhibition of a key downstream effector pathway, the RAF-MEK-ERK cascade, has shown limited success because of activation of feedback networks that keep the pathway in check. We hypothesized that inhibiting SOS1, a KRAS activator and important feedback node, represents an effective approach to treat KRAS-driven cancers. We report the discovery of a highly potent, selective, and orally bioavailable small-molecule SOS1 inhibitor, BI-3406, that binds to the catalytic domain of SOS1, thereby preventing the interaction with KRAS. BI-3406 reduces formation of GTP-loaded RAS and limits cellular proliferation of a broad range of KRAS-driven cancers. Importantly, BI-3406 attenuates feedback reactivation induced by MEK inhibitors and thereby enhances sensitivity of KRAS-dependent cancers to MEK inhibition. Combined SOS1 and MEK inhibition represents a novel and effective therapeutic concept to address KRAS-driven tumors. SIGNIFICANCE: To date, there are no effective targeted pan-KRAS therapies. In-depth characterization of BI-3406 activity and identification of MEK inhibitors as effective combination partners provide an attractive therapeutic concept for the majority of KRAS-mutant cancers, including those fueled by the most prevalent mutant KRAS oncoproteins, G12D, G12V, G12C, and G13D.See related commentary by Zhao et al., p. 17.This article is highlighted in the In This Issue feature, p. 1.

Antitumor Activity of the IGF-1/IGF-2-Neutralizing Antibody Xentuzumab (BI 836845) in Combination with Enzalutamide in Prostate Cancer Models.

Androgen deprivation therapy and second-generation androgen receptor signaling inhibitors such as enzalutamide are standard treatments for advanced/metastatic prostate cancer. Unfortunately, most men develop resistance and relapse; signaling via insulin-like growth factor (IGF) has been implicated in castration-resistant prostate cancer. We evaluated the antitumor activity of xentuzumab (IGF ligand-neutralizing antibody), alone and in combination with enzalutamide, in prostate cancer cell lines (VCaP, DuCaP, MDA PCa 2b, LNCaP, and PC-3) using established in vitro assays, and in vivo, using LuCaP 96CR, a prostate cancer patient-derived xenograft (PDX) model. Xentuzumab + enzalutamide reduced the viability of phosphatase and tensin homolog (PTEN)-expressing VCaP, DuCaP, and MDA PCa 2b cells more than either single agent, and increased antiproliferative activity and apoptosis induction in VCaP. Xentuzumab or xentuzumab + enzalutamide inhibited IGF type 1 receptor and AKT serine/threonine kinase (AKT) phosphorylation in VCaP, DuCaP, and MDA PCa 2b cells; xentuzumab had no effect on AKT phosphorylation and proliferation in PTEN-null LNCaP or PC-3 cells. Knockdown of PTEN led to loss of antiproliferative activity of xentuzumab and reduced activity of xentuzumab + enzalutamide in VCaP cells. Xentuzumab + enzalutamide inhibited the growth of castration-resistant LuCaP 96CR PDX with acquired resistance to enzalutamide, and improved survival in vivo The data suggest that xentuzumab + enzalutamide combination therapy may overcome castration resistance and could be effective in patients who are resistant to enzalutamide alone. PTEN status as a biomarker of responsiveness to combination therapy needs further investigation.

Selective and Potent CDK8/19 Inhibitors Enhance NK-Cell Activity and Promote Tumor Surveillance.

Natural killer (NK) cells play a pivotal role in controlling cancer. Multiple extracellular receptors and internal signaling nodes tightly regulate NK activation. Cyclin-dependent kinases of the mediator complex (CDK8 and CDK19) were described as a signaling intermediates in NK cells. Here, we report for the first time the development and use of CDK8/19 inhibitors to suppress phosphorylation of STAT1S727 in NK cells and to augment the production of the cytolytic molecules perforin and granzyme B (GZMB). Functionally, this resulted in enhanced NK-cell-mediated lysis of primary leukemia cells. Treatment with the CDK8/19 inhibitor BI-1347 increased the response rate and survival of mice bearing melanoma and breast cancer xenografts. In addition, CDK8/19 inhibition augmented the antitumoral activity of anti-PD-1 antibody and SMAC mimetic therapy, both agents that promote T-cell-mediated antitumor immunity. Treatment with the SMAC mimetic compound BI-8382 resulted in an increased number of NK cells infiltrating EMT6 tumors. Combination of the CDK8/19 inhibitor BI-1347, which augments the amount of degranulation enzymes, with the SMAC mimetic BI-8382 resulted in increased survival of mice carrying the EMT6 breast cancer model. The observed survival benefit was dependent on an intermittent treatment schedule of BI-1347, suggesting the importance of circumventing a hyporesponsive state of NK cells. These results suggest that CDK8/19 inhibitors can be combined with modulators of the adaptive immune system to inhibit the growth of solid tumors, independent of their activity on cancer cells, but rather through promoting NK-cell function.

The novel BET bromodomain inhibitor BI 894999 represses super-enhancer-associated transcription and synergizes with CDK9 inhibition in AML.

Bromodomain and extra-terminal (BET) protein inhibitors have been reported as treatment options for acute myeloid leukemia (AML) in preclinical models and are currently being evaluated in clinical trials. This work presents a novel potent and selective BET inhibitor (BI 894999), which has recently entered clinical trials (NCT02516553). In preclinical studies, this compound is highly active in AML cell lines, primary patient samples, and xenografts. HEXIM1 is described as an excellent pharmacodynamic biomarker for target engagement in tumors as well as in blood. Mechanistic studies show that BI 894999 targets super-enhancer-regulated oncogenes and other lineage-specific factors, which are involved in the maintenance of the disease state. BI 894999 is active as monotherapy in AML xenografts, and in addition leads to strongly enhanced antitumor effects in combination with CDK9 inhibitors. This treatment combination results in a marked decrease of global p-Ser2 RNA polymerase II levels and leads to rapid induction of apoptosis in vitro and in vivo. Together, these data provide a strong rationale for the clinical evaluation of BI 894999 in AML.

The IGF1R/INSR Inhibitor BI 885578 Selectively Inhibits Growth of IGF2-Overexpressing Colorectal Cancer Tumors and Potentiates the Efficacy of Anti-VEGF Therapy.

Clinical studies of pharmacologic agents targeting the insulin-like growth factor (IGF) pathway in unselected cancer patients have so far demonstrated modest efficacy outcomes, with objective responses being rare. As such, the identification of selection biomarkers for enrichment of potential responders represents a high priority for future trials of these agents. Several reports have described high IGF2 expression in a subset of colorectal cancers, with focal IGF2 amplification being responsible for some of these cases. We defined a novel cut-off value for IGF2 overexpression based on differential expression between colorectal tumors and normal tissue samples. Analysis of two independent colorectal cancer datasets revealed IGF2 to be overexpressed at a frequency of 13% to 22%. An in vitro screen of 34 colorectal cancer cell lines revealed IGF2 expression to significantly correlate with sensitivity to the IGF1R/INSR inhibitor BI 885578. Furthermore, autocrine IGF2 constitutively activated IGF1R and Akt phosphorylation, which was inhibited by BI 885578 treatment. BI 885578 significantly delayed the growth of IGF2-high colorectal cancer xenograft tumors in mice, while combination with a VEGF-A antibody increased efficacy and induced tumor regression. Besides colorectal cancer, IGF2 overexpression was detected in more than 10% of bladder carcinoma, hepatocellular carcinoma and non-small cell lung cancer patient samples. Meanwhile, IGF2-high non-colorectal cancer cells lines displayed constitutive IGF1R phosphorylation and were sensitive to BI 885578. Our findings suggest that IGF2 may represent an attractive patient selection biomarker for IGF pathway inhibitors and that combination with VEGF-targeting agents may further improve clinical outcomes. Mol Cancer Ther; 16(10); 2223-33. ©2017 AACR.

An allometric pharmacokinetic/pharmacodynamics model for BI 893923, a novel IGF-1 receptor inhibitor.

PURPOSE: BI 893923 is a novel IGF1R/INSR inhibitor with promising anti-tumor efficacy. Dose-limiting hyperglycemia has been observed for other IGF1R/INSR inhibitors in clinical trials. To counterbalance anti-tumor efficacy with the risk of hyperglycemia and to determine the therapeutic window, we aimed to develop a translational pharmacokinetic/pharmacodynamics model for BI 893923. This aimed to translate pharmacokinetics and pharmacodynamics from animals to humans by an allometrically scaled semi-mechanistic model. METHODS: Model development was based on a previously published PK/PD model for BI 893923 in mice (Titze et al., Cancer Chemother Pharmacol 77:1303-1314, 13). PK and blood glucose parameters were scaled by allometric principles using body weight as a scaling factor along with an estimation of the parameter exponents. Biomarker and tumor growth parameters were extrapolated from mouse to human using the body weight ratio as scaling factor. RESULTS: The allometric PK/PD model successfully described BI 893923 pharmacokinetics and blood glucose across mouse, rat, dog, minipig, and monkey. BI 893923 human exposure as well as blood glucose and tumor growth were predicted and compared for different dosing scenarios. A comprehensive risk-benefit analysis was conducted by determining the net clinical benefit for each schedule. An oral dose of 2750 mg BI 893923 divided in three evenly distributed doses was identified as the optimal human dosing regimen, predicting a tumor growth inhibition of 90.4% without associated hyperglycemia. CONCLUSION: Our model supported human therapeutic dose estimation by rationalizing the optimal efficacious dosing regimen with minimal undesired effects. This modeling approach may be useful for PK/PD scaling of other IGF1R/INSR inhibitors.

A comprehensive pharmacokinetic/pharmacodynamics analysis of the novel IGF1R/INSR inhibitor BI 893923 applying in vitro, in vivo and in silico modeling techniques.

PURPOSE: BI 893923 is a novel IGF1R/INSR tyrosine kinase inhibitor demonstrating anti-tumor efficacy and good tolerability. We aimed to characterize the relationship between BI 893923 plasma concentration, tumor biomarker modulation, tumor growth and hyperglycemia in mice using in silico modeling analyses. METHODS: In vitro molecular and cellular assays were used to demonstrate the potency and selectivity of BI 893923. Diverse in vitro DMPK assays were used to characterize the compound's drug-like properties. Mice xenografted with human GEO tumors were treated with different doses of BI 893923 to demonstrate the compound's efficacy, biomarker modulation and tolerability. PK/PD analyses were performed using nonlinear mixed-effects modeling. RESULTS: BI 893923 demonstrated potent and selective molecular inhibition of the IGF1R and INSR and demonstrated attractive drug-like properties (permeability, bioavailability). BI 893923 dose-dependently reduced GEO tumor growth and demonstrated good tolerability, characterized by transient hyperglycemia and normal body weight gain. A population PK/PD model was developed, which established relationships between BI 893923 pharmacokinetics, hyperglycemia, pIGF1R reduction and tumor growth. CONCLUSION: BI 893923 demonstrates molecular properties consistent with a highly attractive inhibitor of the IGF1R/INSR. A generic PK/PD model was developed to support preclinical drug development and dose finding in mice.

Structure-Based Design of an in Vivo Active Selective BRD9 Inhibitor.

Components of the chromatin remodelling switch/sucrose nonfermentable (SWI/SNF) complex are recurrently mutated in tumors, suggesting that altering the activity of the complex plays a role in oncogenesis. However, the role that the individual subunits play in this process is not clear. We set out to develop an inhibitor compound targeting the bromodomain of BRD9 in order to evaluate its function within the SWI/SNF complex. Here, we present the discovery and development of a potent and selective BRD9 bromodomain inhibitor series based on a new pyridinone-like scaffold. Crystallographic information on the inhibitors bound to BRD9 guided their development with respect to potency for BRD9 and selectivity against BRD4. These compounds modulate BRD9 bromodomain cellular function and display antitumor activity in an AML xenograft model. Two chemical probes, BI-7273 (1) and BI-9564 (2), were identified that should prove to be useful in further exploring BRD9 bromodomain biology in both in vitro and in vivo settings.

BI 885578, a Novel IGF1R/INSR Tyrosine Kinase Inhibitor with Pharmacokinetic Properties That Dissociate Antitumor Efficacy and Perturbation of Glucose Homeostasis.

Inhibition of the IGF1R, INSRA, and INSRB receptor tyrosine kinases represents an attractive approach of pharmacologic intervention in cancer, owing to the roles of the IGF1R and INSRA in promoting cell proliferation and survival. However, the central role of the INSRB isoform in glucose homeostasis suggests that prolonged inhibition of this kinase could result in metabolic toxicity. We describe here the profile of the novel compound BI 885578, a potent and selective ATP-competitive IGF1R/INSR tyrosine kinase inhibitor distinguished by rapid intestinal absorption and a short in vivo half-life as a result of rapid metabolic clearance. BI 885578, administered daily per os, displayed an acceptable tolerability profile in mice at doses that significantly reduced the growth of xenografted human GEO and CL-14 colon carcinoma tumors. We found that treatment with BI 885578 is accompanied by increases in circulating glucose and insulin levels, which in turn leads to compensatory hyperphosphorylation of muscle INSRs and subsequent normalization of blood glucose within a few hours. In contrast, the normalization of IGF1R and INSR phosphorylation in GEO tumors occurs at a much slower rate. In accordance with this, BI 885578 led to a prolonged inhibition of cell proliferation and induction of apoptosis in GEO tumors. We propose that the remarkable therapeutic window observed for BI 885578 is achieved by virtue of the distinctive pharmacokinetic properties of the compound, capitalizing on the physiologic mechanisms of glucose homeostasis and differential levels of IGF1R and INSR expression in tumors and normal tissues.

Efficacy and mechanism of action of volasertib, a potent and selective inhibitor of Polo-like kinases, in preclinical models of acute myeloid leukemia.

Polo-like kinase 1 (Plk1), a member of the Polo-like kinase family of serine/threonine kinases, is a key regulator of multiple steps in mitosis. Here we report on the pharmacological profile of volasertib, a potent and selective Plk inhibitor, in multiple preclinical models of acute myeloid leukemia (AML) including established cell lines, bone marrow samples from AML patients in short-term culture, and subcutaneous as well as disseminated in vivo models in immune-deficient mice. Our results indicate that volasertib is highly efficacious as a single agent and in combination with established and emerging AML drugs, including the antimetabolite cytarabine, hypomethylating agents (decitabine, azacitidine), and quizartinib, a signal transduction inhibitor targeting FLT3. Collectively, these preclinical data support the use of volasertib as a new therapeutic approach for the treatment of AML patients, and provide a foundation for combination approaches that may further improve and prolong clinical responses.

Pharmacodynamic and antineoplastic activity of BI 836845, a fully human IGF ligand-neutralizing antibody, and mechanistic rationale for combination with rapamycin.

Insulin-like growth factor (IGF) signaling is thought to play a role in the development and progression of multiple cancer types. To date, therapeutic strategies aimed at disrupting IGF signaling have largely focused on antibodies that target the IGF-I receptor (IGF-IR). Here, we describe the pharmacologic profile of BI 836845, a fully human monoclonal antibody that utilizes an alternative approach to IGF signaling inhibition by selectively neutralizing the bioactivity of IGF ligands. Biochemical analyses of BI 836845 demonstrated high affinity to human IGF-I and IGF-II, resulting in effective inhibition of IGF-induced activation of both IGF-IR and IR-A in vitro. Cross-reactivity to rodent IGFs has enabled rigorous assessment of the pharmacologic activity of BI 836845 in preclinical models. Pharmacodynamic studies in rats showed potent reduction of serum IGF bioactivity in the absence of metabolic adverse effects, leading to growth inhibition as evidenced by reduced body weight gain and tail length. Moreover, BI 836845 reduced the proliferation of human cell lines derived from different cancer types and enhanced the antitumor efficacy of rapamycin by blocking a rapamycin-induced increase in upstream signaling in vitro as well as in human tumor xenograft models in nude mice. Our data suggest that BI 836845 represents a potentially more effective and tolerable approach to the inhibition of IGF signaling compared with agents that target the IGF-I receptor directly, with potential for rational combinations with other targeted agents in clinical studies.