Somatostatin receptor 2 (SSTR2) is frequently overexpressed on several types of solid tumors, including neuroendocrine tumors and small-cell lung cancer. Peptide agonists of SSTR2 are rapidly internalized upon binding to the receptor and linking a toxic payload to an SSTR2 agonist is a potential method to kill SSTR2-expressing tumor cells. Herein, we describe our efforts towards an efficacious SSTR2-targeting cytotoxic conjugate; examination of different SSTR2-targeting ligands, conjugation sites, and payloads led to the discovery of 22 (PEN-221), a conjugate consisting of microtubule-targeting agent DM1 linked to the C-terminal side chain of Tyr3-octreotate. PEN-221 demonstrates in vitro activity which is both potent (IC50 = 10 nM) and receptor-dependent (IC50 shifts 90-fold upon receptor blockade). PEN-221 targets high levels of DM1 to SSTR2-expressing xenograft tumors, which has led to tumor regressions in several SSTR2-expressing xenograft mouse models. The safety and efficacy of PEN-221 is currently under evaluation in human clinical trials.

Abstract Here we describe the discovery and the structure of PEN-221, a somatostatin receptor 2 (SSTR2) targeting peptide conjugated to DM1. PEN-221 is the first clinical compound from Tarveda’s Pentarin platform, which utilizes miniaturized drug conjugates that diffuse rapidly and deeply into solid tumors. Antibody drug conjugates (ADCs) have garnered a significant amount of attention in their ability to direct cytotoxic drugs to cancer cells; however, the efficacy of ADCs in solid tumors is limited by the slow diffusion of such large molecules through solid tumor tissue. Pentarins are designed to improve the efficacy of targeted therapies through effective tumor cell targeting and enhanced tumor penetration. SSTR2, a GPCR overexpressed in multiple types of neuroendocrine tumors, including small cell lung cancers, internalizes rapidly upon agonist stimulation, making it an ideal vector for delivering cytotoxic payloads. Examination of a variety of SSTR2 targeting ligands, as well as several potential conjugation sites, led to the identification of the C-terminal side chain of [Tyr3]-octreotate amide as the best conjugation site for a lipophilic payload. The use of DM1 as a payload afforded superior receptor affinity and receptor internalization when compared to other similarly potent microtubule-targeting agents. In vitro studies show that PEN-221 has receptor-dependent cytotoxic effects, and preclinical studies demonstrate PEN-221 induces tumor regression in several SSTR2 expressing xenograft models. Citation Format: Brian H. White, Patrick Bazinet, Kerry Whalen, Michelle DuPont, James M. Quinn, Rossitza Alargova, Tsun Au Yeung, Adam Brockman, James Gifford, Haley Oller, Kristina Kriksciukaite, Charles-Andre Lemelin, Patrick Lim Soo, Benoit Moreau, Samantha Perino, Gitanjali Sharma, Rajesh Shinde, Beata Sweryda-Krawiec, Mary Simcox, Richard Wooster, Mark T. Bilodeau. Discovery of PEN-221, an SSTR2-targeting maytansinoid conjugate with potent activity in vitro and in vivo [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 39. doi:10.1158/1538-7445.AM2017-39

Abstract Purpose: To explore the role of TWEAK in tumor growth and antitumor immune response and the activity and mechanism of RG7212, an antagonistic anti-TWEAK antibody, in tumor models. Experimental Design: TWEAK-induced signaling and gene expression were explored in tumor cell lines and inhibition of these effects and antitumor efficacy with RG7212 treatment was assessed in human tumor xenograft-, patient-derived xenograft, and syngeneic tumor models and phase I patients. Genetic features correlated with antitumor activity were characterized. Results: In tumor cell lines, TWEAK induces proliferation, survival, and NF-κB signaling and gene expression that promote tumor growth and suppress antitumor immune responses. TWEAK-inducible CD274, CCL2, CXCL-10 and -11 modulate T-cell and monocyte recruitment, T-cell activation, and macrophage differentiation. These factors and TWEAK-induced signaling were decreased, and tumor, blood, and spleen immune cell composition was altered with RG7212 treatment in mice. RG7212 inhibits tumor growth in vivo in models with TWEAK receptor, Fn14, expression, and markers of pathway activation. In phase I testing, signs of tumor shrinkage and stable disease were observed without dose-limiting toxicity. In a patient with advanced, Fn14-positive, malignant melanoma with evidence of tumor regression, proliferation markers were dramatically reduced, tumor T-cell infiltration increased, and tumor macrophage content decreased. Antitumor activity, a lack of toxicity in humans and animals and no evidence of antagonism with standard of care or targeted agents in mice, suggests that RG7212 is a promising agent for use in combination therapies in patients with Fn14-positive tumors. Clin Cancer Res; 19(20); 5686–98. ©2013 AACR.

The identification of new biomarkers is essential in the implementation of personalized health care strategies that offer new therapeutic approaches with optimized and individualized treatment. In support of hypothesis generation and testing in the course of our biomarker research an online portal and respective function-tested reverse transcription quantitative real-time PCR assays (RT-qPCR) facilitated the selection of relevant biomarker genes. We have established workflows applicable for convenient high throughput gene expression analysis in biomarker research with cell lines (in vitro studies) and xenograft mouse models (in vivo studies) as well as formalin-fixed paraffin-embedded tissue (FFPET) sections from various human research and clinical tumor samples. Out of 92 putative biomarker candidate genes selected in silico, 35 were shown to exhibit differential expression in various tumor cell lines. These were further analysed by in vivo xenograft mouse models, which identified 13 candidate genes including potential response prediction biomarkers and a potential pharmacodynamic biomarker. Six of these candidate genes were selected for further evaluation in FFPET samples, where optimized RNA isolation, reverse transcription and qPCR assays provided reliable determination of relative expression levels as precondition for differential gene expression analysis of FFPET samples derived from projected clinical studies. Thus, we successfully applied function tested RT-qPCR assays in our biomarker research for hypothesis generation with in vitro and in vivo models as well as for hypothesis testing with human FFPET samples. Hence, appropriate function-tested RT-qPCR assays are available in biomarker research accompanying the different stages of drug development, starting from target identification up to early clinical development. The workflow presented here supports the identification and validation of new biomarkers and may lead to advances in efforts to achieve the goal of personalized health care. (C) 2012 Elsevier Inc. All rights reserved.

Abstract Solid tumor growth requires new blood vessel formation or angiogenesis. Inhibition of angiogenesis as a therapeutic strategy in oncology has been validated by treatments that block vascular endothelial growth factor (VEGF) or its receptors, such as bevacizumab (Avastin®), which is an anti-VEGF antibody that prolongs survival in colorectal, lung and other cancer patients in combination with chemotherapy. However, since not all tumors are sensitive to VEGF blockade and resistance mechanisms to VEGF therapies can develop, inhibition of additional targets may be necessary in order to control tumor growth and achieve better clinical effects. PlGF is a member of the VEGF family that is found only in very low levels under normal physiological conditions, but is up-regulated in almost all major malignant diseases. PlGF expression has shown to correlate with tumor stages and patient survival in breast cancer, CRC and gastric cancer [1-3]. Pre-clinical data support a role for PlGF in tumor angiogenesis, and demonstrate that blocking PlGF can inhibit tumor growth [4]. RO5323441, a humanized IgG1 monoclonal antibody directed against PlGF, has demonstrated anti-tumor activity in human tumor xenograft models in mice, has a benign preclinical toxicology profile and is being developed for the treatment of multiple advanced cancer indications. RO5323441 binds to both PlGF-1 and PlGF-2 in a dose dependent manner, and is not cross-reactive with murine PlGF or human VEGF. RO5323441 inhibits the binding of human PlGF-1 or PlGF-2 to VEGFR −1 with IC50 values of 0.1 and 0.2 nM, respectively. RO5323441 blocks PlGF-induced VEGFR-1 phosphorylation in Flt-1-transfected HEK293 cells. Antitumor activity has been demonstrated in mutliple tumor models including ACHN and Caki-1 renal cell carcinoma and Huh-7, hepatocellular carcinoma xenografts. Inhibition of established tumors ranged from 43-97% with twice weekly dosing. A refractory model of non-small cell lung cancer has also been identified. Studies to understand the mechanism of action and to identify pharmacodynamic markers and have been carried out in ACHN-tumor bearing mice. References 1. Wei SC, Tsao PN, Yu SC, et al. Placenta growth factor expression is correlated with survival of patients with colorectal cancer. Gut. 2005 May; 54(5):666-72. 2. Parr C, Watkins G, Boulton M et al. Placenta growth factor is over-expressed and has prognostic value in human breast cancer. Eur J Cancer. 2005 Dec;41(18):2819-27. 3. Chen CN, Hsieh FJ, Cheng YM, et al. The significance of placenta growth factor in angiogenesis and clinical outcome of human gastric cancer. Cancer Lett. 2004 Sep 15;213(1):73-82. 4. Fischer C, Jonckx B, Mazzone M, et al. Anti-PlGF inhibits growth of VEGF(R)-inhibitor-resistant tumors without affecting healthy vessels. Cell. 2007. 131 : 463-75. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 1370.

e14629 Background: R1530 is a multikinase inhibitor currently in clinical phase I testing. Its inhibitory profile includes several kinases that play critical roles in cancer cell growth and division as well as tumor angiogenesis. These properties translate into a potent cytotoxicity in a wide range of cancer cell lines in vitro and tumor growth inhibition in human tumor xenografts. Preclinical studies were conducted to evaluate the effects of R alone and in combination with B and P in the Caki-1 RCC xenograft model. Methods: We initially evaluated the antitumor activity of optimal dose (OD) and 1/2 OD R alone and with OD B. This was followed up with testing of OD & 1/2 OD P ± OD B, along with triplets of 1/2 OD P + OD B + 1/2 OD R and OD P + OD B + 1/2 OD R. A final study compared 1/2 OD R to OD R triplets to attempt to increase tumor growth inhibition (TGI) and increase life span (ILS). Results: No doublets or triplets tested showed antagonism or enhanced toxicity. Antitumor activity and survival results are listed below (Table). Conclusions: 1/2 OD R + OD B or OD R + OD B doublets gave better TGI and ILS than monotherapy. Comparing these two doublets, TGI is better in the high dose combination but ILS is equivalent. All TGI and ILS are better in doublet P + B combinations over respective single agent arms except for TGI (but not ILS) for 1/2 OD P vs its correlative doublet with B. The OD P + B doublet gave better TGI and ILS than 1/2 OD P + B doublet. TGI and ILS do not differ between triplets containing OD R + 1/2 OD or OD P or for triplets containing the OD P + 1/2 OD or OD R. Therefore, either agent can be alternatively dose reduced without a loss of tumor response or detriment to survival in this preclinical model of RCC. [Table: see text] [Table: see text]

1440 Tubulin is a clinically validated target for anti-cancer therapy. Taxanes (microtubule stabilizers) are amongst the most powerful chemotherapeutic agents in NSCLC, breast cancer, ovarian cancer and prostate cancer. However, the clinical benefit obtained with these drugs has been limited by drug resistance acquired by tumors and the safety profile. Mechanisms of resistance to taxanes include mutations in the tubulin genes and overexpression of transmembrane drug efflux pumps (e.g. MDR1 and BCRP). Adverse events associated with taxanes include hematological toxicity, neurotoxicity and hypersensitivity reactions. For patients who acquire resistance to, or who failed prior treatment with taxanes, the success of salvage chemotherapy has been reported to be modest. Therefore, there is a need to develop new drugs targeting tubulin structure with better safety profile and less resistance. We previously presented the potent antitumor activity of our synthetic dolastatin analogue (Compound 1) in a broad range of human xenograft models of NSCLC, breast cancer, colorectal cancer, prostate cancer and hepatocellular cancers including a taxane resistant tumor cell line. In our studies, we examined the antitumor activity of the combination of Compound 1 plus existing antitumor agents such as capecitabine, irinotecan, cisplatin, trastuzumab and pertuzumab in several human cancer xenograft models compared to single agent activity. Compound 1 exhibited synergistic/additive antitumor effects in combination with irinotecan (CPT-11) or with cisplatin (CDDP) in NSCLC models, with capecitabine in CRC models and with trastuzumab or pertuzumab in a Her2 positive breast cancer model. Tumor growth inhibition (TGI) in these combinations was statistically significant compared to those in single agent administration. Notably, the combination of Compound 1 with trastuzumab resulted in a high antitumor efficacy and also showed complete regression in some animals. Thus, this new tubulin polymerization inhibitor not only shows significantly improved TGI as single agent compared to other tubulin interacting drugs on the market, but also in combination with existing antitumor drugs including molecular targeting agents. Based on this favorable pre-clinical profile, clinical studies are ongoing.

RO4396686 is a small molecule KDR, FGFR, and PDGFR inhibitor with good pharmacokinetic properties in rodents. In a mouse corneal neovascularization assay, this compound inhibited VEGF-induced angiogenesis. Tested in a H460a xenograft tumor model this agent effected significant tumor growth inhibition at doses as low as 50 mg/kg.

(+/-)-1-(anti-3-Hydroxy-cyclopentyl)-3-(4-methoxy-phenyl)-7-phenylamino-3,4-dihydro-1H-pyrimido[4,5-d]pyrimidin-2-one (RO4383596) is a potent and selective inhibitor of the pro-angiogenic receptor tyrosine kinases KDR, FGFR, and PDGFR. This agent has an excellent pharmacokinetic profile and is highly efficacious in rodent models of angiogenesis upon oral administration.