Ellen H. Filvaroff

A technology platform to assess multiple cancer agents simultaneously within a patient’s tumor

Simultaneous in vivo assessment of multiple cancer drugs and drug combinations using microinjection technology predicts systemic response in model tumors and has shown feasibility for assessment of drug efficacy in a pilot study in cancer patients. There’s no place like the human Animal models of human tumors and dish cultures of cancer cells are not sufficient to predict an individual patient’s response to therapy. In the emerging era of personalized medicine, why limit ourselves to rodent models and engineered in vitro tumor models when we can study a drug directly in the patient’s tumor? This question was answered by Klinghoffer et al. by creating a microinjection system called CIVO that delivers small doses of up to eight different drugs simultaneously, directly into the tumor. The tumors could then be removed and evaluated for various markers of cancer response; in short, the authors looked for markers of cell death and drug-related mechanisms of action. By using an injection-tracking dye, Klinghoffer and colleagues could see where the drug was deposited and then use an automated analyzer for quantitative image processing along the 6-mm injection tract. In mouse models of human lymphoma, the authors were able to correctly predict systemic responsiveness to doxorubicin or vincristine—or not, in the case of resistant lymphomas. They also uncovered unexpected drug sensitivities, which were not picked up by traditional cell culture, including to novel anticancer agents, and confirmed these in vivo. The authors pilot-tested the device in dog and human patients, demonstrating the ability of CIVO to inject and track local tumor response to chemotherapies. Ultimately, such a personalized approach to drug sensitivity testing will allow for rational selection of therapeutics while sparing patients the pain—and time—associated with ineffective treatments. A fundamental problem in cancer drug development is that antitumor efficacy in preclinical cancer models does not translate faithfully to patient outcomes. Much of early cancer drug discovery is performed under in vitro conditions in cell-based models that poorly represent actual malignancies. To address this inconsistency, we have developed a technology platform called CIVO, which enables simultaneous assessment of up to eight drugs or drug combinations within a single solid tumor in vivo. The platform is currently designed for use in animal models of cancer and patients with superficial tumors but can be modified for investigation of deeper-seated malignancies. In xenograft lymphoma models, CIVO microinjection of well-characterized anticancer agents (vincristine, doxorubicin, mafosfamide, and prednisolone) induced spatially defined cellular changes around sites of drug exposure, specific to the known mechanisms of action of each drug. The observed localized responses predicted responses to systemically delivered drugs in animals. In pair-matched lymphoma models, CIVO correctly demonstrated tumor resistance to doxorubicin and vincristine and an unexpected enhanced sensitivity to mafosfamide in multidrug-resistant lymphomas compared with chemotherapy-naïve lymphomas. A CIVO-enabled in vivo screen of 97 approved oncology agents revealed a novel mTOR (mammalian target of rapamycin) pathway inhibitor that exhibits significantly increased tumor-killing activity in the drug-resistant setting compared with chemotherapy-naïve tumors. Finally, feasibility studies to assess the use of CIVO in human and canine patients demonstrated that microinjection of drugs is toxicity-sparing while inducing robust, easily tracked, drug-specific responses in autochthonous tumors, setting the stage for further application of this technology in clinical trials.

Dual TORK/DNA-PK inhibition blocks critical signaling pathways in chronic lymphocytic leukemia

Inhibition of B-cell receptor (BCR) signaling pathways in chronic lymphocytic leukemia (CLL) provides significant clinical benefit to patients, mainly by blocking adhesion of CLL cells in the lymph node microenvironment. The currently applied inhibitors ibrutinib and idelalisib have limited capacity however to induce cell death as monotherapy and are unlikely to eradicate the disease. Acquired resistance to therapy in CLL is often caused by mutations in the response network being targeted, both for DNA damage or BCR signaling pathways. Thus, drugs with dual targeting capacity could offer improved therapeutic value. Here, the potency of CC-115, a novel inhibitor of mammalian target of rapamycin kinase (TORK) and DNA-dependent protein kinase (DNA-PK), was evaluated in primary CLL cells in vitro and in CLL patients. Combined TORK and DNA-PK inhibition in vitro resulted in caspase-dependent cell killing irrespective of p53, ATM, NOTCH1, or SF3B1 status. Proliferation induced by CD40(+) interleukin-21 stimulation was completely blocked by CC-115, and CD40-mediated resistance to fludarabine and venetoclax could be reverted by CC-115. BCR-mediated signaling was inhibited by CC-115 and also in CLL samples obtained from patients with acquired resistance to idelalisib treatment. Clinical efficacy of CC-115 was demonstrated in 8 patients with relapsed/refractory CLL/small lymphocytic lymphoma harboring ATM deletions/mutations; all but 1 patient had a decrease in lymphadenopathy, resulting in 1 IWCLL partial response (PR) and 3 PRs with lymphocytosis. In conclusion, these preclinical results, along with early promising clinical activity, suggest that CC-115 may be developed further for treatment of CLL. The trial was registered at www.clinicaltrials.gov as #NCT01353625.

Abstract 3129: A platform to assess multiple therapy options simultaneously in a patient's own tumor

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Proper selection of anti-cancer agents at the earliest stage of patient treatment following diagnosis of disease relapse is expected to substantially impact clinical response to treatment. Currently, genomic approaches to personalized cancer treatments have been yielded mixed results, while empirical tests to assess tumor responsiveness have been limited to ex vivo systems that disrupt the native tumor microenvironment and show limited predictive value. To address the need for multiplexed in vivo chemosensitivity testing, we have developed a technology that allows simultaneous assessment of multiple cancer therapeutics directly in a patients tumor. This technology could provide a valuable decision-making tool to prioritize effective treatments in the oncology clinic. Data herein highlight how this technology enables controlled and reliable microinjection of multiple drugs simultaneously in preclinical tumor models, canine lymphoma, and human lymphoma patients. Consistent with the controlled drug delivery of this system, spatially localized, readily detectable, and mechanism-specific cellular changes were observed around sites of microinjection in response to classic chemotherapy drugs (vincristine and doxorubicin) as well as to a small molecule inhibitor of TOR kinase. Importantly, localized response (or lack thereof) to individual components of CHOP combination therapy correlated with response to long-term systemic drug administration across multiple cell line and patient-derived xenograft models of lymphoma. Underscoring the importance of assessing drug efficacy in the context of an intact in vivo system, tumor responses to vincristine were impacted by the local tumor microenvironment. Our results also emphasize the importance of selecting effective therapies early in the course of treatment, as drug resistance mechanisms induced cross-resistance to otherwise efficacious drugs. These studies set the stage for use of this platform in oncology drug development, where the ability to more rapidly assess drug efficacy using clinically relevant in vivo tumors may decrease the current reliance on in vitro cell-based models of cancer and possibly increase the likelihood of clinical success. This platform may thus be useful a clinical decision-making tool for selection of patient-specific anti-cancer therapies. Citation Format: Richard Klinghoffer, Alicia Moreno-Gonzalez, Michael Carleton, Jason Frazier, Marc Grenley, Ilona Tretyak, Nathan Hedin, Joyoti Dey, Joseph Casalini, Beryl Hatton, Sally Ditzler, James Olson, Daniel Pierce, Ellen Filvaroff, Nathan Caffo. A platform to assess multiple therapy options simultaneously in a patients own tumor. [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 3129. doi:10.1158/1538-7445.AM2014-3129

Abstract LB-94: Highlights of innovative preclinical studies which guided the rapid bench to bedside development of nab-paclitaxel plus gemcitabine combination for the treatment of pancreatic cancer

Proceedings: AACR Annual Meeting 2014; April 5-9, 2014; San Diego, CA Background and Aim: Advanced pancreatic cancer is both deadly and difficult to treat with success. Here, we disseminate the results of comprehensive preclinical studies which laid a strong foundation for the rapid bench to bedside development of nab -paclitaxel, an albumin-bound formulation of paclitaxel, in combination with gemcitabine, a regimen recently approved by U.S. Food and Drug Administration as a first-line treatment for patients with metastatic pancreatic cancer. Materials and Methods : We enrolled a total of 650 mice with established pancreatic tumors originated from a collection of patient-derived pancreatic cancer xenografts. The present study investigated the anti-tumor activity, survival advantage and mechanism of action of nab -paclitaxel or Cremophor EL™ (CreEL)-based paclitaxel monotherapy and in combination with gemcitabine. Results : When tested in mice with subcutaneous tumors originating from 11 separate individual patient xenografts, nab -paclitaxel plus gemcitabine treatment demonstrated superior tumor regression response, robustly depleted the tumor desmoplatic stroma, leading to enhanced gemcitabine uptake (2.8-fold) in the tumor compared to tumors in the gemcitabine alone treated mice. In orthotopic models, nab -paclitaxel treatment leads to an average of 3.64-fold decrease in primary tumor volumes compared to CreEL-based paclitaxel. Intra-tumor stromal collapse combined with decreased tumor cell proliferation was clearly evident in the primary tumors of nab -paclitaxel treated mice compared to CreEL-based paclitaxel, when the mice were sacrificed immediately after five consecutive days treatment or three weeks after the final dose of the agents. In a highly aggressive orthotopic model, nab -paclitaxel plus gemcitabine treatment prevented primary tumor progression, and metastatic spread to liver, lymph nodes and diaphragm. While CreEL-based paclitaxel plus gemcitabine treatment failed to enhance mouse survival compared to gemcitabine monotherapy, the nab -paclitaxel plus gemcitabine combination proved statistically significant ( p =0.0133) in enhancing survival. nab -paclitaxel monotherapy demonstrated statistically significant survival advantage compared to CreEL-based paclitaxel monotherapy ( p =0.0072). Remarkably, nab -paclitaxel monotherapy was equivalent to nab -paclitaxel plus gemcitabine in providing survival advantage in a highly aggressive metastatic model of pancreatic cancer. Conclusion: Our results demonstrated that co-treatment with nab-paclitaxel and gemcitabine resulted in superior tumor regression response, stromal depletion and enhanced intra-tumoral gemcitabine uptake compared with either single agent alone. nab -paclitaxel demonstrated superior anti-tumor activity and provided a statistically significant survival advantage compared to CreEL-based paclitaxel. Our results provide further rationale for future preclinical and clinical trials in pancreatic cancer using nab -paclitaxel as a backbone therapy in combination with novel experimental and targeted agents. Acknowledgements : The study was supported by funding from Celgene Corporation and AACR-Stand Up To Cancer Dream Team Translational Cancer Research Grant (SU2C-AACR-DT0509). Citation Format: N.V Rajeshkumar, Shinichi Yabuuchi, Shweta G. Pai, Scott Bateman, Ellen Filvaroff, Daniel W. Pierce, Carla Heise, Daniel D. Von Hoff, Anirban Maitra, Manuel Hidalgo. Highlights of innovative preclinical studies which guided the rapid bench to bedside development of nab-paclitaxel plus gemcitabine combination for the treatment of pancreatic cancer. [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 LB-94. doi:10.1158/1538-7445.AM2014-LB-94