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Activation of apoptosis in malignant cells is an established strategy for controlling cancer and is potentially curative. To assess the impact of concurrently inducing the extrinsic and intrinsic apoptosis-signaling pathways in acute myeloid leukemia (AML), we evaluated activity of the TRAIL receptor agonistic fusion protein eftozanermin alfa (eftoza; ABBV-621) in combination with the B-cell lymphoma protein-2 selective inhibitor venetoclax in preclinical models and human patients. Simultaneously stimulating intrinsic and extrinsic apoptosis-signaling pathways with venetoclax and eftoza, respectively, enhanced their activities in AML cell lines and patient-derived ex vivo/in vivo models. Eftoza activity alone or plus venetoclax required death receptor 4/5 (DR4/DR5) expression on the plasma membrane but was independent of TP53 or FLT3-ITD status. The safety/tolerability of eftoza as monotherapy and in combination with venetoclax was demonstrated in patients with relapsed/refractory AML in a phase 1 clinical trial. Treatment-related adverse events were reported in 2 of 4 (50%) patients treated with eftoza monotherapy and 18 of 23 (78%) treated with eftoza plus venetoclax. An overall response rate of 30% (7/23; 4 complete responses [CRs], 2 CRs with incomplete hematologic recovery, and 1 morphologic leukemia-free state) was reported in patients who received treatment with eftoza plus venetoclax and 67% (4/6) in patients with myoblasts positive for DR4/DR5 expression; no tumor responses were observed with eftoza monotherapy. These data indicate that combination therapy with eftoza plus venetoclax to simultaneously activate the extrinsic and intrinsic apoptosis-signaling pathways may improve clinical benefit compared with venetoclax monotherapy in relapsed/refractory AML with an acceptable toxicity profile. This trial was registered at www.clinicaltrials.gov as #NCT03082209.

Targeting tumor-overexpressed EGFR with an antibody–drug conjugate (ADC) is an attractive therapeutic strategy; however, normal tissue expression represents a significant toxicity risk. The anti-EGFR antibody ABT-806 targets a unique tumor-specific epitope and exhibits minimal reactivity to EGFR in normal tissue, suggesting its suitability for the development of an ADC. We describe the binding properties and preclinical activity of ABT-414, an ABT-806 monomethyl auristatin F conjugate. In vitro, ABT-414 selectively kills tumor cells overexpressing wild-type or mutant forms of EGFR. ABT-414 inhibits the growth of xenograft tumors with high EGFR expression and causes complete regressions and cures in the most sensitive models. Tumor growth inhibition is also observed in tumor models with EGFR mutations, including activating mutations and those with the exon 2–7 deletion [EGFR variant III (EGFRvIII)], commonly found in glioblastoma multiforme. ABT-414 exhibits potent cytotoxicity against glioblastoma multiforme patient-derived xenograft models expressing either wild-type EGFR or EGFRvIII, with sustained regressions and cures observed at clinically relevant doses. ABT-414 also combines with standard-of-care treatment of radiation and temozolomide, providing significant therapeutic benefit in a glioblastoma multiforme xenograft model. On the basis of these results, ABT-414 has advanced to phase I/II clinical trials, and objective responses have been observed in patients with both amplified wild-type and EGFRvIII-expressing tumors. Mol Cancer Ther; 15(4); 661–9. ©2016 AACR.

Amino Acid Sequence of AM-1 VH Region

Table S1: Mouse Strains Used for Xenograft Studies; Table S2: ABT-414 Growth Inhibition of Xenograft Tumors; Figure S1: Efficacy of ABT-414, ABT-806-vcMMAE, and unconjugated MMAE co-dosed with ABT-806 in U87MGde2-7 Tumor-Bearing Mice.

Depatuxizumab mafodotin (depatux-m, ABT-414) is a tumor-selective antibody drug conjugate (ADC) comprised of the anti-EGFR antibody ABT-806 and the monomethyl auristatin F (MMAF) warhead. Depatux-m has demonstrated promising clinical activity in glioblastoma multiforme (GBM) patients and is currently being evaluated in clinical trials in first-line and recurrent GBM disease settings. Depatux-m responses have been restricted to patients with amplified EGFR, highlighting the need for therapies with activity against tumors with nonamplified EGFR overexpression. In addition, depatux-m dosing has been limited by corneal side effects common to MMAF conjugates. We hypothesized that a monomethyl auristatin E (MMAE) ADC utilizing an EGFR-targeting antibody with increased affinity may have broader utility against tumors with more modest EGFR overexpression while mitigating the risk of corneal side effects. We describe here preclinical characterization of ABBV-221, an EGFR-targeting ADC comprised of an affinity-matured ABT-806 conjugated to MMAE. ABBV-221 binds to a similar EGFR epitope as depatux-m and retains tumor selectivity with increased binding to EGFR-positive tumor cells and greater in vitro potency. ABBV-221 displays increased tumor uptake and antitumor activity against wild-type EGFR-positive xenografts with a greatly reduced incidence of corneal side effects relative to depatux-m. ABBV-221 has similar activity as depatux-m against an EGFR-amplified GBM patient derived xenograft (PDX) model and is highly effective alone and in combination with standard-of-care temozolomide in an EGFRvIII-positive GBM xenograft model. Based on these results, ABBV-221 has advanced to a phase I clinical trial in patients with advanced solid tumors associated with elevated levels of EGFR. Mol Cancer Ther; 17(4); 795–805. ©2018 AACR.

TRAIL can activate cell surface death receptors, resulting in potent tumor cell death via induction of the extrinsic apoptosis pathway. Eftozanermin alfa (ABBV-621) is a second generation TRAIL receptor agonist engineered as an IgG1-Fc mutant backbone linked to two sets of trimeric native single-chain TRAIL receptor binding domain monomers. This hexavalent agonistic fusion protein binds to the death-inducing DR4 and DR5 receptors with nanomolar affinity to drive on-target biological activity with enhanced caspase-8 aggregation and death-inducing signaling complex formation independent of FcγR-mediated cross-linking, and without clinical signs or pathologic evidence of toxicity in nonrodent species. ABBV-621 induced cell death in approximately 36% (45/126) of solid cancer cell lines in vitro at subnanomolar concentrations. An in vivo patient-derived xenograft (PDX) screen of ABBV-621 activity across 15 different tumor indications resulted in an overall response (OR) of 29% (47/162). Although DR4 (TNFSFR10A) and/or DR5 (TNFSFR10B) expression levels did not predict the level of response to ABBV-621 activity in vivo, KRAS mutations were associated with elevated TNFSFR10A and TNFSFR10B and were enriched in ABBV-621–responsive colorectal carcinoma PDX models. To build upon the OR of ABBV-621 monotherapy in colorectal cancer (45%; 10/22) and pancreatic cancer (35%; 7/20), we subsequently demonstrated that inherent resistance to ABBV-621 treatment could be overcome in combination with chemotherapeutics or with selective inhibitors of BCL-XL. In summary, these data provide a preclinical rationale for the ongoing phase 1 clinical trial (NCT03082209) evaluating the activity of ABBV-621 in patients with cancer.Significance:This study describes the activity of a hexavalent TRAIL-receptor agonistic fusion protein in preclinical models of solid tumors that mechanistically distinguishes this molecular entity from other TRAIL-based therapeutics.

Supplementary file containing all supplementary figures and tables

TRAIL can activate cell surface death receptors, resulting in potent tumor cell death via induction of the extrinsic apoptosis pathway. Eftozanermin alfa (ABBV-621) is a second generation TRAIL receptor agonist engineered as an IgG1-Fc mutant backbone linked to two sets of trimeric native single-chain TRAIL receptor binding domain monomers. This hexavalent agonistic fusion protein binds to the death-inducing DR4 and DR5 receptors with nanomolar affinity to drive on-target biological activity with enhanced caspase-8 aggregation and death-inducing signaling complex formation independent of FcγR-mediated cross-linking, and without clinical signs or pathologic evidence of toxicity in nonrodent species. ABBV-621 induced cell death in approximately 36% (45/126) of solid cancer cell lines in vitro at subnanomolar concentrations. An in vivo patient-derived xenograft (PDX) screen of ABBV-621 activity across 15 different tumor indications resulted in an overall response (OR) of 29% (47/162). Although DR4 (TNFSFR10A) and/or DR5 (TNFSFR10B) expression levels did not predict the level of response to ABBV-621 activity in vivo, KRAS mutations were associated with elevated TNFSFR10A and TNFSFR10B and were enriched in ABBV-621–responsive colorectal carcinoma PDX models. To build upon the OR of ABBV-621 monotherapy in colorectal cancer (45%; 10/22) and pancreatic cancer (35%; 7/20), we subsequently demonstrated that inherent resistance to ABBV-621 treatment could be overcome in combination with chemotherapeutics or with selective inhibitors of BCL-XL. In summary, these data provide a preclinical rationale for the ongoing phase 1 clinical trial (NCT03082209) evaluating the activity of ABBV-621 in patients with cancer. Significance: This study describes the activity of a hexavalent TRAIL-receptor agonistic fusion protein in preclinical models of solid tumors that mechanistically distinguishes this molecular entity from other TRAIL-based therapeutics.

Cancer cells are highly dependent on NAD+/NADH produced via the nicotinamide salvage pathway. The rate-limiting enzyme in this pathway is the nicotinamide phosphoribosyltransferase (NAMPT), which we have targeted with novel NAMPT inhibitors. NAMPT inhibition elicits depletion of total cellular NAD+ levels and ultimately cytotoxicity via depletion of cellular ATP levels. 18F-fluorodeoxyglucose– positron emission tomography (FDG-PET) is a translational imaging tool to assess glucose utilization in tumors and normal tissue. We used FDG-PET to understand the timing of ATP depletion in vivo and better understand the pharmacology of NAMPT inhibitors. Because of the intimate relationship between cellular ATP levels and cell viability, we developed an in-depth understanding of our NAMPT inhibitor pharmacology and the relationship with changes in tumor FDG uptake. Taken together, we show that FDG-PET could be used as a biomarker in clinical studies to understand dose and provide proof of mechanism for NAMPT inhibitors. SIGNIFICANCE STATEMENT Our imaging data suggest that tumor 18F-fluorodeoxyglucose uptake can provide insight into the ATP status inside the tumor after nicotinamide phosphoribosyltransferase (NAMPT) therapy, with a novel NAMPT inhibitor. Such an approach could be used clinically as a pharmacodynamic biomarker to help understand the implications of dose, schedule, rescue strategy, or other clinical biomarkers.

In continuation of our previous research towards the discovery of potent, selective and drug-like Wee1 inhibitors, 2 novel series of biaryl heterocycles were designed, synthesized and evaluated. The new biaryl cores were designed to enable structure−activity exploration of substituents at C-8 or N-8 which were used for tuning compound properties and to improve compound profiles. The lead molecule 33 demonstrated a desirable pharmacokinetic profile and potentiated the anti-proliferative activity of irinotecan in vivo when dosed orally in the human breast MX-1 xenograft model.

The dynamic and reversible acetylation of proteins, catalysed by histone acetyltransferases (HATs) and histone deacetylases (HDACs), is a major epigenetic regulatory mechanism of gene transcription and is associated with multiple diseases. Histone deacetylase inhibitors are currently approved to treat certain cancers, but progress on the development of drug-like histone actyltransferase inhibitors has lagged behind. The histone acetyltransferase paralogues p300 and CREB-binding protein (CBP) are key transcriptional co-activators that are essential for a multitude of cellular processes, and have also been implicated in human pathological conditions (including cancer). Current inhibitors of the p300 and CBP histone acetyltransferase domains, including natural products, bi-substrate analogues and the widely used small molecule C646, lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like catalytic inhibitor of p300 and CBP. We present a high resolution (1.95 Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 competes with acetyl coenzyme A (acetyl-CoA). A-485 selectively inhibited proliferation in lineage-specific tumour types, including several haematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen-sensitive and castration-resistant prostate cancer and inhibited tumour growth in a castration-resistant xenograft model. These results demonstrate the feasibility of using small molecule inhibitors to selectively target the catalytic activity of histone acetyltransferases, which may provide effective treatments for transcriptional activator-driven malignancies and diseases.

Cancer cells are highly dependent on NAD

Targeting tumor-overexpressed EGFR with an antibody–drug conjugate (ADC) is an attractive therapeutic strategy; however, normal tissue expression represents a significant toxicity risk. The anti-EGFR antibody ABT-806 targets a unique tumor-specific epitope and exhibits minimal reactivity to EGFR in normal tissue, suggesting its suitability for the development of an ADC. We describe the binding properties and preclinical activity of ABT-414, an ABT-806 monomethyl auristatin F conjugate. In vitro, ABT-414 selectively kills tumor cells overexpressing wild-type or mutant forms of EGFR. ABT-414 inhibits the growth of xenograft tumors with high EGFR expression and causes complete regressions and cures in the most sensitive models. Tumor growth inhibition is also observed in tumor models with EGFR mutations, including activating mutations and those with the exon 2–7 deletion [EGFR variant III (EGFRvIII)], commonly found in glioblastoma multiforme. ABT-414 exhibits potent cytotoxicity against glioblastoma multiforme patient-derived xenograft models expressing either wild-type EGFR or EGFRvIII, with sustained regressions and cures observed at clinically relevant doses. ABT-414 also combines with standard-of-care treatment of radiation and temozolomide, providing significant therapeutic benefit in a glioblastoma multiforme xenograft model. On the basis of these results, ABT-414 has advanced to phase I/II clinical trials, and objective responses have been observed in patients with both amplified wild-type and EGFRvIII-expressing tumors. Mol Cancer Ther; 15(4); 661–9. ©2016 AACR.

Depatuxizumab mafodotin (depatux-m, ABT-414) is a tumor-selective antibody drug conjugate (ADC) comprised of the anti-EGFR antibody ABT-806 and the monomethyl auristatin F (MMAF) warhead. Depatux-m has demonstrated promising clinical activity in glioblastoma multiforme (GBM) patients and is currently being evaluated in clinical trials in first-line and recurrent GBM disease settings. Depatux-m responses have been restricted to patients with amplified EGFR, highlighting the need for therapies with activity against tumors with nonamplified EGFR overexpression. In addition, depatux-m dosing has been limited by corneal side effects common to MMAF conjugates. We hypothesized that a monomethyl auristatin E (MMAE) ADC utilizing an EGFR-targeting antibody with increased affinity may have broader utility against tumors with more modest EGFR overexpression while mitigating the risk of corneal side effects. We describe here preclinical characterization of ABBV-221, an EGFR-targeting ADC comprised of an affinity-matured ABT-806 conjugated to MMAE. ABBV-221 binds to a similar EGFR epitope as depatux-m and retains tumor selectivity with increased binding to EGFR-positive tumor cells and greater in vitro potency. ABBV-221 displays increased tumor uptake and antitumor activity against wild-type EGFR-positive xenografts with a greatly reduced incidence of corneal side effects relative to depatux-m. ABBV-221 has similar activity as depatux-m against an EGFR-amplified GBM patient derived xenograft (PDX) model and is highly effective alone and in combination with standard-of-care temozolomide in an EGFRvIII-positive GBM xenograft model. Based on these results, ABBV-221 has advanced to a phase I clinical trial in patients with advanced solid tumors associated with elevated levels of EGFR. Mol Cancer Ther; 17(4); 795–805. ©2018 AACR.

Aided by molecular modeling, compounds with a pyrimidine-based tricyclic scaffold were designed and confirmed to inhibit Wee1 kinase. Structure-activity studies identified key pharmacophores at the aminoaryl and halo-benzene regions responsible for binding affinity with sub-nM K i values. The potent inhibitors demonstrated sub-μM activities in both functional and mechanism-based cellular assays and also possessed desirable pharmacokinetic profiles. The lead molecule, 31, showed oral efficacy in potentiating the antiproliferative activity of irinotecan, a cytotoxic agent, in a NCI-H1299 mouse xenograft model.

Polycomb repressive complex 2 (PRC2) is a regulator of epigenetic states required for development and homeostasis. PRC2 trimethylates histone H3 at lysine 27 (H3K27me3), which leads to gene silencing, and is dysregulated in many cancers. The embryonic ectoderm development (EED) protein is an essential subunit of PRC2 that has both a scaffolding function and an H3K27me3-binding function. Here we report the identification of A-395, a potent antagonist of the H3K27me3 binding functions of EED. Structural studies demonstrate that A-395 binds to EED in the H3K27me3-binding pocket, thereby preventing allosteric activation of the catalytic activity of PRC2. Phenotypic effects observed in vitro and in vivo are similar to those of known PRC2 enzymatic inhibitors; however, A-395 retains potent activity against cell lines resistant to the catalytic inhibitors. A-395 represents a first-in-class antagonist of PRC2 protein-protein interactions (PPI) for use as a chemical probe to investigate the roles of EED-containing protein complexes.

The dynamic and reversible acetylation of proteins catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs) is a major epigenetic regulatory mechanism of gene transcription 1 associated with multiple diseases. While HDAC inhibitors are approved to treat certain cancers, progress on the development of drug-like HAT inhibitors has lagged 2. The HAT paralogs p300 and CBP (p300/CBP) are key transcriptional co-activators essential for a multitude of cellular processes and also implicated in human pathological conditions, including cancer 3. Current p300/CBP HAT domain inhibitors including natural products, 4 bi-substrate analogs (Lys-CoA) 5 and the widely utilized C646 6, 7 lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like p300/CBP catalytic inhibitor. We show the first high resolution (1.95Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 is acetyl-CoA competitive. A-485 selectively inhibited proliferation across lineage-specific tumor types, including several hematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen sensitive and castrate resistant prostate cancer and inhibited tumor growth in a castration resistant xenograft model. These results demonstrate the feasibility of selectively targeting the catalytic activity of histone acetyltransferases.

The dynamic and reversible acetylation of proteins catalyzed by histone acetyltransferases (HATs) and histone deacetylases (HDACs) is a major epigenetic regulatory mechanism of gene transcription associated with multiple diseases. While HDAC inhibitors are approved to treat certain cancers, progress on the development of drug-like HAT inhibitors has lagged. The HAT paralogs p300 and CBP (p300/CBP) are key transcriptional co-activators essential for a multitude of cellular processes and also implicated in human pathological conditions, including cancer. Current p300/CBP HAT domain inhibitors including natural products, bi-substrate analogs (Lys-CoA) and the widely utilized C646 lack potency or selectivity. Here, we describe A-485, a potent, selective and drug-like p300/CBP catalytic inhibitor. We show the first high resolution (1.95Å) co-crystal structure of a small molecule bound to the catalytic active site of p300 and demonstrate that A-485 is acetyl-CoA competitive. A-485 selectively inhibited proliferation across lineage-specific tumor types, including several hematological malignancies and androgen receptor-positive prostate cancer. A-485 inhibited the androgen receptor transcriptional program in both androgen sensitive and castrate resistant prostate cancer and inhibited tumor growth in a castration resistant xenograft model. These results demonstrate the feasibility of selectively targeting the catalytic activity of histone acetyltransferases. Citation Format: Kenneth D. Bromberg, Loren M. Lasko, Clarissa G. Jakob, Wei Qiu, Debra Montgomery, Enrico L. Digiammarino, Todd M. Hansen, Roberto M. Risi, Robin R. Frey, Vlasios Manaves, Bailin Shaw, Mikkel Algire, Paul Hessler, Lloyd T. Lam, Tamar Uziel, Emily Favire, Debra Ferguson, Fritz G. Buchanan, Ruth L. Martin, Maricel Torrent, Saul H. Rosenberg, Michael R. Michaelides, Albert Lai. Discovery of a potent catalytic p300/CBP inhibitor that targets lineage-specific tumors [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr LB-A23.

ProbodyTM therapeutics are antibody prodrugs designed to remain largely inactive until proteolytically activated in the tumor microenvironment (TME), potentially enabling the safer targeting of antigens that are highly expressed in both tumor and normal tissue. CD71 (transferrin receptor) is an example of an ideal Probody Drug Conjugate (PDC) target, not only because it efficiently internalizes and can deliver a cytotoxic payload intracellularly, but also because it is expressed at high levels both in many different tumor types as well as in dividing normal cells. We have previously demonstrated that while an anti-CD71 antibody drug conjugate (ADC) is highly toxic, a CD71-targeting PDC is both efficacious in mouse tumor models and well tolerated in nonhuman primates. Two key components of a Probody therapeutic prodomain that reduce its binding to normal tissue and allow for its tumor-specific activation are 1) a mask that reduces the ability of the antibody binding site to interact with target antigen and 2) a protease-activatable substrate that is cleaved in the TME, resulting in removal of the mask. Here, we demonstrate how modulating mask strength and substrate cleavability can optimize efficacy and safety of a CD71-targeting PDC in preclinical models. Through this process, we have selected a lead molecule, CX-2029, for further development. CX-2029 is a CD71-targeting PDC conjugated to vcMMAE with a Drug to Probody Ratio (DPR) of 2, achieved by purification. At dose levels consistent with those expected in humans, a more strongly masked PDC, CX-2018, was less efficacious in a mouse xenograft tumor model compared to PDC CX-2016, which has a weaker mask, demonstrating that mask strength affects antitumor activity. Further, PDC CX-2019, which has the same mask but a less cleavable substrate than CX-2016, was similarly efficacious in a mouse xenograft tumor model, demonstrating that both substrates are sufficiently cleaved in the TME to activate the PDCs. However, CX-2019 was better tolerated in NHP at 6 mg/kg than CX-2016, suggesting that the less cleavable substrate in CX-2019 leads to a better therapeutic index. Using a LC/MS/MS method, we showed lower levels of circulating activated CX-2019 compared with circulating activated CX-2016, which is consistent with CX-2019’s improved tolerability. Lead CX-2029 contains the same mask and substrate as CX-2019 but differs in having a DPR of 2 versus ~3 for CX-2019. Up to 6 mg/kg of CX-2029 as a single dose produced complete regressions and durable responses in mouse xenograft tumor models encompassing multiple indications, and was tolerated in monkeys at doses of up to 12 mg/kg. These data demonstrate that, in preclinical models, tuning of mask strength and substrate cleavability can optimize the efficacy and tolerability of Probody Therapeutics and have the potential to enable the safe and effective targeting of highly expressed tumor antigens like CD-71. CX-2029 is currently under development, with an IND filing expected in 2018. PROBODY is a trademark of CytomX Therapeutics, Inc. Citation Format: Shweta Singh, Laura Serwer, Niharika Chauhan, Amy DuPage, Michael Krimm, Ken Wong, Yuanhui Huang, Andrew Jang, Eric Ureno, Adam Miller, Sarah Patrick, Shanti Duvur, Fritz Buchanan, Matthew M. Ravn, Rob Leanna, Ilaria Badagnani, Tracy Henriques, Shouchun Liu, Claus Krebber, Sridhar Viswanathan, Jennifer Richardson, Susan Morgan-Lappe, Michael Kavanaugh. Optimizing a CD71-targeting Probody drug conjugate (PDC) for activity in multiple solid tumor and lymphoma models and for tolerability in nonhuman primates [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr B116.

Despite clinical efficacy, current approved agents targeting EGFR are associated with on-target toxicities as a consequence of disrupting normal EGFR function. MAb 806 is a novel EGFR antibody that selectively targets a tumor-selective epitope suggesting that a mAb 806-based therapeutic would retain antitumor activity without the on-target toxicities associated with EGFR inhibition. To enable clinical development, a humanized variant of mAb 806 designated ABT-806 was generated and is currently in phase 1 trials. We describe the characterization of binding and functional properties of ABT-806 compared with the clinically validated anti-EGFR antibody cetuximab. ABT-806 binds the mutant EGFRvIII with high affinity and, relative to cetuximab, exhibits increased potency against glioblastoma multiforme cell line and patient-derived xenografts expressing this form of the receptor. ABT-806 also inhibits the growth of squamous cell carcinoma xenograft models expressing high levels of wild-type EGFR, associated with inhibition of EGFR signaling, although higher doses of ABT-806 than cetuximab are required for similar activity. ABT-806 enhances in vivo potency of standard-of-care therapies used to treat glioblastoma multiforme and head and neck squamous cell carcinoma. An indium-labeled version of ABT-806, [111In]-ABT-806, used to investigate the relationship between dose and receptor occupancy, revealed greater receptor occupancy at lowers doses in an EGFRvIII-expressing model and significant uptake in an orthotopic model. Collectively, these results suggest that ABT-806 may have antitumor activity superior to cetuximab in EGFRvIII-expressing tumors, and similar activity to cetuximab in tumors highly overexpressing wild-type EGFR with reduced toxicity. Mol Cancer Ther; 14(5); 1141–51. ©2015 AACR.

We have developed a series of phenylpyrrolidine- and phenylpiperidine-substituted benzimidazole carboxamide poly(ADP-ribose) polymerase (PARP) inhibitors with excellent PARP enzyme potency as well as single-digit nanomolar cellular potency. These efforts led to the identification of (S)-2-(2-fluoro-4-(pyrrolidin-2-yl)phenyl)-1H-benzimidazole-4-carboxamide (22b, A-966492). Compound 22b displayed excellent potency against the PARP-1 enzyme with a K(i) of 1 nM and an EC(50) of 1 nM in a whole cell assay. In addition, 22b is orally bioavailable across multiple species, crosses the blood-brain barrier, and appears to distribute into tumor tissue. It also demonstrated good in vivo efficacy in a B16F10 subcutaneous murine melanoma model in combination with temozolomide and in an MX-1 breast cancer xenograft model both as a single agent and in combination with carboplatin.

BackgroundThe insulin-like growth factor (IGF) axis is an important signaling pathway in the growth and survival of many cell and tissue types. This pathway has also been implicated in many aspects of cancer progression from tumorigenesis to metastasis. The multiple roles of IGF signaling in cancer suggest that inhibition of the pathway might yield clinically effective therapeutics.MethodsWe describe A-928605, a novel pyrazolo [3,4-d]pyrimidine small molecule inhibitor of the receptor tyrosine kinases (IGF1R and IR) responsible for IGF signal transduction. This compound was first tested for its activity and selectivity via conventionalin vitrokinome profiling and cellular IGF1R autophosphorylation. Additionally, cellular selectivity and efficacy of A-928605 were analyzed in an IGF1R oncogene-addicted cell line by proliferation, signaling and microarray studies. Finally,in vivoefficacy of A-928605 was assessed in the oncogene-addicted cell line and in a neuroblastoma model as a single agent as well as in combination with clinically approved therapeutics targeting EGFR in models of pancreatic and non-small cell lung cancers.ResultsA-928605 is a selective IGF1R inhibitor that is able to abrogate activation of the pathway bothin vitroandin vivo. This novel compound dosed as a single agent is able to produce significant growth inhibition of neuroblastoma xenograftsin vivo. A-928605 is also able to provide additive effects when used in combination with clinically approved agents directed against EGFR in non-small cell lung and human pancreatic tumor models.ConclusionThese results suggest that a selective IGF1R inhibitor such as A-928605 may provide a useful clinical therapeutic for IGF pathway affected tumors and warrants further investigation.

Understanding the roles of epigenetic alterations in cancer development and maintenance holds great promise for cancer prevention, detection, and therapy. Cancer can be considered as a pathogenic state where cellular differentiation is suppressed (i.e. stem cell-like) and aberrant epigenetic patterning is commonly observed in tumors. Histone methyltransferases play a key role in epigenetics by modifying key lysine and arginine residues on histones and thereby influencing biological processes. Previous studies have suggested that the histone lysine methyltransferase G9a (EHMT2) is required to perpetuate malignant phenotypes through over-expression in a variety of cancer types. These reports have shown that pharmacologic inhibition or genetic ablation of G9a leads to retardation of tumor cell growth and cellular invasion in vitro as well as inhibition of metastasis in vivo. To further elucidate the enzymatic role of G9a in cancer, we describe herein the discovery of a novel histone methyltransferase inhibitor, A-366, that selectively inhibits G9a and the closely related GLP (EHMT1). A-366 is a peptide competitive inhibitor of G9a/GLP with in vitro enzymatic IC50 of ∼ 3 nM and cellular activity of ∼ 100 nM and > 100-fold selectivity over other methyltransferases and other non-epigenetic targets. A-366 has significantly less cytotoxic effects on the growth of solid tumor cell lines compared to other known G9a/GLP small molecule inhibitors despite roughly equivalent cellular activity on methylation of H3K9me2. However, the excellent selectivity profile of A-366 has aided in the discovery of an important role for G9a/GLP in lineage maintenance of a subset of leukemias. Treatment of various leukemia cell lines in vitro resulted in marked differentiation and morphological changes of these tumors in the absence of cytotoxicity resulting in cytostasis. Furthermore, treament of MV4;11 flank xenografts with A-366 resulted in growth inhibition in vivo consistent with the profile of H3K9me2 reduction observed. In summary, A-366 is a novel and highly selective peptide-competitive inhibitor of G9a/GLP that has enabled the discovery of a role for G9a/GLP enzymatic activity in the epigenetic maintenance of a subset of leukemia cells. Citation Format: Jun Guo, Marina Pliushchev, Yupeng He, Debra Ferguson, Sujatha Jagadeeswaran, Andrew Petros, Chaohong Sun, Niru B. Soni, Bailin Shaw, Alla Korepanova, David Maag, Ramzi Sweis, Fritz G. Buchanan, Michael Michaelides, Alex Shoemaker, Chris Tse, Gary G. Chiang, William N. Pappano. Discovery of A-366, a novel small molecule inhibitor that uncovers an unappreciated role for G9a/GLP in the epigenetics of leukemia. [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 5532. doi:10.1158/1538-7445.AM2014-5532

PARP's role in DNA damage recognition/repair makes PARP inhibition an attractive cancer therapeutic target. Veliparib (ABT-888) is a potent PARP inhibitor with excellent oral bioavailability that readily crosses the blood-brain barrier and is currently in Phase 2 clinical trials. Veliparib has shown significant ability to potentiate multiple DNA damaging agents (cisplatin, carboplatin, cyclophosphamide, irinotecan, radiation and temozolomide) in a spectrum of tumors from multiple histological types. In studies using cytotoxic combinations with veliparib (cisplatin, cyclophosphamide, TMZ) there was a marked increase in efficacy over cytotoxic monotherapies, especially in the MX-1 breast model. Since DNA repair deficiencies (e.g. HR-defective) are known to render cells very sensitive to PARP inhibition, we genotyped the MX-1 breast line and found it to be HR-deficient. Subsequent studies with both the MX-1 (BRCA1-deleted and BRCA2-mutated breast line) and the Capan-1 (BRCA2-deficient pancreatic cancer cell line), showed that veliparib (25 mg/kg/day, p.o., b.i.d.×5) demonstrated enhancement of TMZ (50 mg/kg/day, p.o., q.d.×5) activity, with regressions compared to TMZ alone (81–97% TGI, tumor growth inhibition). Continuous dosing at 4×-8× higher doses (100 and 200 mg/kg/day, p.o., b.i.d.×21) demonstrated a significant single agent, dose-responsive activity (26–87% TGI). Similar results were observed with single agent veliparib treatment of BRCA isogenic cell lines in vitro. Further, the combination treatment of veliparib (100 and 200 mg/kg/day, p.o., b.i.d.×21) with carboplatin, carbotaxol, radiation, gemcar, cyclophosphamide, topotecan and TMZ also demonstrated a measurable advantage over either veliparib or cytotoxic agent alone. Both high dose single agent veliparib and the cytotoxic combination therapy were well tolerated and there were no observable animal health concerns; this indicated the feasibility of using the single agent and cytotoxic combination regimen in the clinic. Analysis of tumors for the reduction of PAR after veliparib treatment demonstrated a correlative and significant reduction by western blot indicating the ability of veliparib to inhibit PARP activity in vivo. Altogether, our studies show the strong activity of veliparib as a single agent in BRCA-deficient xenografts as well as the enhanced activity and tolerability of veliparib in combination with various cytotoxic agents. These results warrant investigation of single agent therapy and cytotoxic combination regimens in BRCA-deficient patients to further the clinical development of veliparib. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B62.