Antitumor Activity In Xenografts Research Articles

Abstract A016: NXP800, a novel, small molecule GCN2 kinase activator, demonstrates potent single-agent activity in ARID1A and ARID1B-deficient endometrial cancer xenograft models

Abstract Background: NXP800 is a clinical stage, antineoplastic, oral, small molecule GCN2 kinase activator that was discovered in a phenotypic screen for inhibitors of the heat shock factor 1-regulated stress response. In a panel of human carcinoma cell lines NXP800 induced the expression of genes associated with activation of the integrated stress response and demonstrated robust antiproliferative activity, particularly in ARID1a-mutated ovarian carcinoma cells lines. In ARID1a-mutated ovarian carcinoma xenografts, treatment with NXP800 resulted in substantial tumor growth inhibition and tumor regression. NXP800 is currently being investigated in a Phase 1b clinical trial in patients with ARID1a-mutated ovarian carcinoma. Here we describe an in vivo study of NXP800 in ARID1a/ARID1b-deficient endometrial cancer xenografts, supporting the clinical development of NXP800 in this indication. Methods: Animal strain: CD1 Nude mice (nu/nu, Charles River). Human endometrial cancer cell lines: RL95-2 (ARID1a mutated, ATCC) and KLE (ARID1a wildtype, ARID1b mutated, ATCC), SNG-M (ARID1a mutated, Creative Bioarray). Xenograft tumors were generated by subcutaneous implantation on the right lower flank of the thigh at a cell density of 2 × 106 cells/mouse, at 0.1 ml Matrigel dilution volume/injection. Experiment groups: Vehicle, NXP800 (35 mg/kg); treatment: QD on days 0-4, 7-11, 14-18, 21-25, 28-30. Loss of ARID1A protein expression was confirmed by western blots. Results: In the SNG-M model, baseline tumor volume for the vehicle and NXP800 groups were 130.2, and 139.7 mm3, respectively, and on Day 32, average tumor volumes were 420.4, and 85.4 mm3, respectively, representing TGI of 81%. In the RL-95 model, baseline tumor volume for the vehicle and NXP800 groups were 124.5, and 129.9 mm3, respectively, and on Day 32, average tumor volumes were 773.8, and 190.3 mm3, respectively, representing TGI of 76%. In the KLE model, baseline tumor volume for both the vehicle and NXP800 groups were 88.1 mm3, respectively, and on Day 32, average tumor volumes were 216.4, and 18.9 mm3, for the vehicle and NXP800 groups, respectively, representing TGI of 91.3%. Conclusions: ARID1A and ARID1B genes encode the alternate, but obligatory, DNA-targeting subunit of the switch/sucrose non-fermentable (SWI/SNF) complex. ARID1A is the most frequently mutated SWI/SNF subunit across cancer types and is mutated in approximately 35% of endometrial carcinomas. NXP800 demonstrated robust antitumor activity in xenografts of endometrial carcinoma, a serious condition for which new treatment options are needed, including sustained tumor growth inhibition both in ARID1A and ARID1B mutated models, supporting the clinical development of NXP800 in endometrial cancer. Citation Format: Ramez N. Eskander, Bradley J. Monk, Brian M. Slomovitz, Enrique Poradosu, Allison Woods, Shay Shemesh, Paul Clarke, Robert Te Poele, Marissa Powers, Paul Workman, Shannon N. Westin. NXP800, a novel, small molecule GCN2 kinase activator, demonstrates potent single-agent activity in ARID1A and ARID1B-deficient endometrial cancer xenograft models [abstract]. In: Proceedings of the AACR Special Conference on Endometrial Cancer: Transforming Care through Science; 2023 Nov 16-18; Boston, Massachusetts. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(5_Suppl):Abstract nr A016.

Abstract 4981: The potent quadruplex-binding compound QN-302 down-regulates the S100P gene in in vitro and in vivo models of pancreatic cancer: a potential therapeutic target and biomarker for PDAC

Abstract The compound QN-302, a tetra-substituted naphthalene diimide (ND) derivative has been designed to have high affinity for G-quadruplex (G4) nucleic acids. It has single-digit nM anti-proliferative activity in a panel of human pancreatic cancer cell lines (Ahmed et al., ACS Med Chem Lett, 2020, 11, 1634-1644) and anti-tumor activity in xenograft, orthotopic and genetic (KPC) models for pancreatic cancer (PDAC). Its mode of action involves the targeting of G4-forming sites in promoter regions of cancer-associated genes, where their prevalence is over-represented compared to other genes. Transcriptome analyses of QN-302 in MIA-PACA2 cells have confirmed genes involved in cancer-associated pathways and carry G4 motifs in their promoters as targets. We have found that expression of the S100P gene, a well-authenticated indicator of PDAC disease and progression, is highly down-regulated in QN-302 treated cells. The S100P gene contains at least five putative G4 motifs, including a classic G4 promoter site just upstream of the transcription start site. We have observed that transcriptome analyses on tumor material from human PDAC patients, and those from poorly differentiated as well as more highly differentiated tumors, have shown that the S100P gene is highly over-expressed (especially in poorly differentiated tumors). We now report on gene expression analysis of S100P using control and treated tumor tissues from a therapeutic study in PDAC MIA-PACA2 xenografts in mice, by means of RT-qPCR and Western blot analyses. The data shows statistically significant dose-dependent decreases in expression of S100P at both the mRNA (to ca 25% of control values) and protein levels (to ca 22% of control values), giving added confidence to the concept that the S100P gene is a therapeutic target in PDAC and may also be useful as a marker of QN-302 response in future clinical use. QN-302 is bio-available at therapeutic doses and is well tolerated at these levels in the animal models. It is being developed for clinical evaluation by Qualigen Therapeutics Inc and is currently at the pre-IND stage. Citation Format: Nicole Williams, Jenny Worthington, Ahmed Ahmed, Tariq Arshad, Stephen Neidle. The potent quadruplex-binding compound QN-302 down-regulates the S100P gene in in vitro and in vivo models of pancreatic cancer: a potential therapeutic target and biomarker for PDAC. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4981.

Combination of Talazoparib and Palbociclib as a Potent Treatment Strategy in Bladder Cancer

The use of cyclin-dependent kinase 4/6 (CDK4/6) inhibitors represents a potent strategy for cancer therapy. Due to the complex molecular network that regulates cell cycle progression, cancer cells often acquire resistance mechanisms against these inhibitors. Previously, our group identified molecular factors conferring resistance to CDK4/6 inhibition in bladder cancer (BLCA) that also included components within the DNA repair pathway. In this study, we validated whether a combinatory treatment approach of the CDK4/6 inhibitor Palbociclib with Poly-(ADP-Ribose) Polymerase (PARP) inhibitors improves therapy response in BLCA. First, a comparison of PARP inhibitors Talazoparib and Olaparib showed superior efficacy of Talazoparib in vitro and displayed high antitumor activity in xenografts in the chicken chorioallantoic membrane (CAM) model. Moreover, the combination of Talazoparib and the CDK4/6 inhibitor Palbociclib synergistically reduced tumor growth in Retinoblastoma protein (RB)-positive BLCA in vitro and in a CAM model, an effect that relies on Palbociclib-induced cell cycle arrest in G0/G1-phase complemented by a G2 arrest induced by Talazoparib. Interestingly, Talazoparib-induced apoptosis was reduced by Palbociclib. The combination of Palbociclib and Talazoparib effectively enhances BLCA therapy, and RB is a molecular biomarker of response to this treatment regimen.

Multi-Omic Based Antigen Discovery for the Immunotherapy of Pediatric Acute T Cell Lymphoblastic Leukemia

Pediatric T-cell acute lymphoblastic leukemia (T-ALL) is a high-risk disease due to treatment related complications and poor prognosis of patients with relapsed disease. Immunotherapy with monoclonal antibodies (MAbs) and/or chimeric antigen receptor (CAR) T-cells for T-ALL is limited by identification of tumor specific target antigens. Differential expression is necessary to prevent on-target/off-tumor toxicities and fratricide of activated T-cells. Targeting multiple antigens can bypass immune escape and result in improved T-cell effector function, since antigen density correlates with T-cell activation. Here we designed a pipeline (Figure 1) to identify unique surface antigens expressed in T-ALL using proteomic and transcriptomic analyses followed by flow cytometry validation, and functional studies with CAR T cells targeting the identified antigens. We generated an Illumina total stranded RNAseq library from healthy donor myeloid and lymphoid cells of bone marrow, peripheral blood and cord blood (N= 116). We compared data to 265 St. Jude pediatric T-ALL samples and against 53 normal tissue expression data from the GTEx (Genotype-Tissue Expression) project. To analyze the T-cell surface proteome, we isolated plasma membrane fractions from 11 samples including healthy T-cells and T-ALL cell lines using a differential centrifugation-based method. The purity of the plasma membrane fraction was confirmed by western blot. Na+/K+ ATPase and GAPDH were used as controls for the plasma membrane and cytosolic fractions respectively. Following plasma membrane enrichment, the membrane proteins were applied for proteomic analysis using an advanced TMT-L/LC-MS/MS pipeline, and the acquired proteomic data were further processed via the JUMP software suite. 997 unique proteins were quantified from the membrane fractions. Integrated analysis the transcriptomic and proteomic datasets showed significant correlation and yielded a list of candidate genes, which were validated by flow cytometry on a panel of T-ALL cell lines (CCRF, RPMI8402, and MOLT3) and resting and activated T-cells from healthy donors. We identified GRP78 as one of the differentially expressed cell surface antigens and further confirmed its expression on additional T-ALL cell lines (KE37, PF382, PEER, CEMC7) and 3 PDX samples. Finally, we generated GRP78-CAR T cells and demonstrate that GRP78-CAR T cells recognize and kill GRP78+ T-ALL cells and have potent antitumor activity in xenograft and PDX models. We have established an unbiased pipeline to identify differentially expressed antigens on the cell surface of T-ALL blasts and created a healthy tissue RNAseq library. The results from our analyses are encouraging and interrogation of our pipeline has yielded differentially expressed immunotherapy targets for the treatment of relapsed refractory T-ALL. Our results highlight the importance of integrated surface proteomics and transcriptomics analysis. Figure 1: Outline of strategy for target selection: Figure Disclosures Hebbar: St. Jude: Patents & Royalties. Epperly:St. Jude: Patents & Royalties. Gottschalk:Inmatics and Tidal: Membership on an entity's Board of Directors or advisory committees; TESSA Therapeutics: Other: research collaboration; Patents and patent applications in the fields of T-cell & Gene therapy for cancer: Patents & Royalties; Merck and ViraCyte: Consultancy. Mullighan:AbbVie, Inc.: Research Funding; Illumina: Consultancy, Honoraria, Speakers Bureau; Pfizer: Honoraria, Research Funding, Speakers Bureau. Velasquez:Rally! Foundation: Membership on an entity's Board of Directors or advisory committees; St. Jude: Patents & Royalties.

Abstract 5861: RX-3117 promotes epigenetic effects in cancer cells through enhanced degradation of DNMT1

Abstract RX-3117 is a novel, investigational, oral, small molecule nucleoside compound that shows promising antitumor activity in xenografts including patient-derived xenografts resistant to gemcitabine. RX-3117 is currently being evaluated in a Phase IIa multi-center, open-label clinical study in patients with relapsed or refractory pancreatic cancer and advanced bladder cancer. RX-3117 can downregulate DNMT1 and activate genes controlled by methylation, such as E-cadherin and p16. We aimed to elucidate the mechanism of RX-3117 mediated downregulation of DNMT1. Expression of DNMT1 was evaluated by western blotting. Binding of DNMT1 to DNA was measured after DNA isolation, DNAse treatment and gel electrophoresis. DNMT1 trapping in the nucleus was also studied with ImageStream FACS analysis. DNMT1 ubiquitination was measured by immunoprecipitation and quantification of the ubiquitin tag K48. Translocation of DNMT1 was studied by confocal microscopy. Modeling studies were performed with Quantum Mechanics (QM) and Molecular Dynamics (MD). DNMT1 acts on the DNA during the so-called base flipping necessary for methyl transfer. To determine whether RX-3117 treatment would lead to trapping of DNMT1, NSCLC A549 and PDAC SUIT-028 cells were treated for 24 hr with 1 µM RX-3117. Immunostaining of DNMT1-DNA and ImageStream FACS analysis demonstrated binding of DNMT1 to DNA. Treatment with RX-3117 also resulted in DNMT1 ubiquitination, as shown by an increase in the ubiquitin tag K48, reaching the highest level after 48 hr. Incubation of A549 and SUIT-028 cells with 80 nM of the specific proteasome inhibitor bortezomib prevented RX-3117 mediated degradation of DNMT1 (at 24 and 24 hr exposure to 1 µM RX-3117). This underlines the role of the proteasome in DNMT1 degradation. With confocal microscopy it was demonstrated that DNMT1 and the proteasome co-localized and translocated to the cytosol of A549 and SUIT-028 cells treated for 48 hours with 1 µM RX-3117. With molecular modelling using QM and MD, changes in nucleophilicity of C4-C5 double bound were observed indicating that DNMT1 would initiate methyl transfer but cannot proceed. In summary we conclude that RX-3117 induces epigenetic changes in cancer cells by trapping DNMT1 in the nucleus followed by translocation of the protein to the proteasome for degradation. Citation Format: Dzjemma Sarkisjan, Ferry Pronk, Rosan Kuin, Btissame el Hassouni, Young B. Lee, Deog J. Kim, Marc Dijk, Daan P. Geerke, G. J. Peters. RX-3117 promotes epigenetic effects in cancer cells through enhanced degradation of DNMT1 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5861.

UniPR1331, a small molecule targeting Eph/ephrin interaction, prolongs survival in glioblastoma and potentiates the effect of antiangiogenic therapy in mice.

Glioblastoma multiforme (GBM) is the most malignant brain tumor, showing high resistance to standard therapeutic approaches that combine surgery, radiotherapy, and chemotherapy. As opposed to healthy tissues, EphA2 has been found highly expressed in specimens of glioblastoma, and increased expression of EphA2 has been shown to correlate with poor survival rates. Accordingly, agents blocking Eph receptor activity could represent a new therapeutic approach. Herein, we demonstrate that UniPR1331, a pan Eph receptor antagonist, possesses significant in vivo anti-angiogenic and anti-vasculogenic properties which lead to a significant anti-tumor activity in xenograft and orthotopic models of GBM. UniPR1331 halved the final volume of tumors when tested in xenografts (p<0.01) and enhanced the disease-free survival of treated animals in the orthotopic models of GBM both by using U87MG cells (40 vs 24 days of control, p<0.05) or TPC8 cells (52 vs 16 days, p<0.01). Further, the association of UniPR1331 with the anti-VEGF antibody Bevacizumab significantly increased the efficacy of both monotherapies in all tested models. Overall, our data promote UniPR1331 as a novel tool for tackling GBM.

Anticancer Therapy by Tumor Vessel Infarction with Polyethylene Glycol Conjugated Retargeted Tissue Factor

tTF-NGR consists of the extracellular domain of tissue factor and the peptide GNGRAHA, a ligand of the surface protein aminopeptidase N and of integrin αvβ3. Both surface proteins are upregulated on endothelial cells of tumor vessels. tTF-NGR shows antitumor activity in xenografts and inhibition of tumor blood flow in cancer patients. We performed random TMS(PEG)12 PEGylation of tTF-NGR to improve the antitumor profile of the molecule. PEGylation resulted in an approximately 2-log step decreased procoagulatory activity of the molecule. Pharmacokinetic studies in mice showed a more than 1-log step higher mean area under the curve. Comparison of the LD10 values for both compounds and their lowest effective antitumor dose against human tumor xenografts showed an improved therapeutic range (active/toxic dose in mg/kg body weight) of 1/5 mg/kg for tTF-NGR and 3/>160 mg/kg for TMS(PEG)12 tTF-NGR. Results demonstrate that PEGylation can significantly improve the therapeutic range of tTF-NGR.

The anti-Fn14 antibody BIIB036 inhibits tumor growth in xenografts and patient derived primary tumor models and enhances efficacy of chemotherapeutic agents in multiple xenograft models

Agonistic antibodies targeting Fn14, the receptor for TWEAK, have demonstrated anti-tumor activity in xenograft models. Herein, we further explore the therapeutic potential of the humanized anti-Fn14 agonistic antibody, BIIB036, as a single agent and in combination with standard of care cancer therapeutics. Pharmacokinetic studies of BIIB036 in tumor-bearing mice revealed a half-life of approximately three days suggesting twice a week dosing would be necessary to maintain efficacy. However, in multiple xenograft models, BIIB036 treatment resulted in extended tumor growth inhibition up to 40–50 d following cessation of dosing, suggesting that frequent administration of BIIB036 may not be necessary to maintain prolonged anti-tumor activity. Subsequent xenograft studies revealed that maximal efficacy was achieved with BIIB036 dosing once every two weeks, by either intraperitoneal or subcutaneous administration. Xenograft tumors that were initially treated with BIBI036 and then re-grew up to 1000 mm3 following cessation of the first cycle of treatment remained sensitive to a second cycle of treatment. BIIB036 was also evaluated in patient derived primary colon tumor models, where efficacy compared favorably with a standard of care agent. Lastly, BIIB036 enhanced the efficacy of several standard of care chemotherapeutics, including paclitaxel in MDA-MBA-231 breast tumor xenografts, paclitaxel or carboplatin in HOP62 non-small cell lung xenografts, and 5-FU in NCI-N87 gastric xenografts, with no overlapping toxicities. These studies thus establish BIIB036 as a promising therapeutic agent with durable anti-tumor activity in human xenografts as well as patient derived primary tumor models, and enhanced activity and tolerability in combination with standard of care chemotherapeutics. Taken together, the data presented herein suggest that BIIB036 warrants evaluation in the clinic.

Abstract 2617: MPC-3100, a synthetic Hsp90 inhibitor, induces biomarker changes in vitro and in vivo

Abstract Introduction: MPC-3100 is a fully synthetic, orally bioavailable, Hsp90 inhibitor in clinical development. It is broadly active in xenograft models with anti-tumor activity ranging from tumor regression to tumor growth inhibition in many cancer types including colon, gastric, ovarian, prostate, breast, lung and myeloid leukemia. Here we evaluate the effect of MPC-3100 on stability of client proteins in cells and xenograft tumors. We also determine the effect on Hsp70 protein levels, a biomarker of Hsp90 inhibition, in peripheral blood mononuclear cells (PBMCs) from cancer patients receiving MPC-3100. Methods: Her2, Akt, Cdk4, c-Raf and Hsp70 client protein levels were monitored in cell culture with protein immunoblots. Formalin-fixed, paraffin-embedded sections of xenograft tumors from mice dosed orally with MPC-3100 were analyzed by immunohistochemistry (IHC) to monitor changes in Her2, Akt and Hsp70 protein levels. To monitor changes in Hsp70 in cancer patients treated with MPC-3100, PBMCs were collected prior to drug administration, 8 and 24 hours post-dose on Day 1 and 24 hours post-dose on Days 7 and 21 of the first treatment cycle. Hsp70 protein levels were determined by ELISA. Results: Exposure of HCT-116, NCI-N87 and DU-145 cells to MPC-3100 in vitro resulted in a time-dependent reduction in client protein levels with maximal reduction by 24 hours. The IC50 values for client protein reduction ranged from 0.1 μM to 0.5 μM, comparable to the cellular cytotoxicity values of MPC-3100 at 72 hours for the various cell lines. IHC revealed reduction in Her2 and Akt protein in N87 xenografts in mice given a single oral dose of 200 mg/kg MPC-3100 relative to tumors from animals dosed with vehicle. Healthy volunteer PBMCs exposed to 1 μM MPC-3100 for 24 hours ex vivo revealed a reduction in Akt, c-Raf and Cdk4 protein levels ranging from 50% to 90%. PBMCs from cancer patients receiving MPC-3100 showed an increase of 28 to 589 ng of Hsp70 protein per mg total protein over baseline by Day 8. The increase in Hsp70 expression was seen as early as 8 hours after the first dose and sustained through at least Day 22. Conclusions: The changes in client proteins and biomarkers observed in cells and tumor xenografts exposed to MPC-3100 confirm that the cellular cytotoxic activity and anti-tumor activity in xenografts are a result of Hsp90 inhibition. The consistent increase in Hsp70 expression in PBMCs from cancer patients receiving MPC-3100 indicates that Hsp90 function is inhibited in patients at doses that have been well tolerated in the clinic. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2617. doi:10.1158/1538-7445.AM2011-2617

Growth and Metastases of Human Lung Cancer Are Inhibited in Mouse Xenografts by a Transition State Analogue of 5′-Methylthioadenosine Phosphorylase

The S-adenosylmethionine (AdoMet) salvage enzyme 5'-methylthioadenosine phosphorylase (MTAP) has been implicated as both a cancer target and a tumor suppressor. We tested these hypotheses in mouse xenografts of human lung cancers. AdoMet recycling from 5'-methylthioadenosine (MTA) was blocked by inhibition of MTAP with methylthio-DADMe-Immucillin-A (MTDIA), an orally available, nontoxic, picomolar transition state analogue. Blood, urine, and tumor levels of MTA increased in response to MTDIA treatment. MTDIA treatment inhibited A549 (human non-small cell lung carcinoma) and H358 (human bronchioloalveolar non-small cell lung carcinoma cells) xenograft tumor growth in immunodeficient Rag2(-/-)γC(-/-) and NCr-nu mice. Systemic MTA accumulation is implicated as the tumor-suppressive metabolite because MTDIA is effective for in vivo treatment of A549 MTAP(-/-) and H358 MTAP(+/+) tumors. Tumors from treated mice showed increased MTA and decreased polyamines but little alteration in AdoMet, methionine, or adenine levels. Gene expression profiles of A549 tumors from treated and untreated mice revealed only modest alterations with 62 up-regulated and 63 down-regulated mRNAs (≥ 3-fold). MTDIA antitumor activity in xenografts supports MTAP as a target for lung cancer therapy.

Abstract 4497: NVP-BKM120, a novel inhibitor of phosphoinosotide 3-kinase in Phase I/II clinical trials, shows significant antitumor activity in xenograft and primary tumor models

Abstract A substantial number of epidemiological and experimental studies support an important role for PI3K in the biology of human cancer. The activation of PI3K, and its downstream effectors, has been clearly validated as an essential step for the initiation and maintenance of the tumorigenic phenotype. Parallel to the clinical development of our dual pan-PI3K/mTOR modulators (e.g., NVP-BEZ235), we continued our drug discovery activities to identify PI3K inhibitors with distinct biological and pharmacological profiles. Following a structure-based design strategy, we have identified a new clinical candidate, NVP-BKM120. This 2,6-dimorpholino pyrimidine derivative is a potent pan-PI3K (e.g., IC50= 35 nM, p110α) that does not significantly inhibit other protein or lipid kinases (e.g., IC50= 4.6 μM, mTOR). The compound exhibits potent antiproliferative activity against a broad panel of tumour cell lines by specifically blocking the biological function of PI3K signaling components (e.g. IC50 = 93 nM S473P-Akt in Rat1-p110α cells). The activity of this pan-PI3K inhibitor in cellular settings translates well in in vivo models of human cancer. NVP-BKM120 shows good oral bioavailability in preclinical species and demonstrated significant antitumor activity (growth inhibition or regression) at tolerated doses in xenografts of diverse cancer lineage in mice and rat models. Consistent with its mechanism of action, transient increases in plasma insulin and glucose levels are observed in these studies. Analyses of tumor tissues after acute dosing or at the end of efficacy studies showed a good correlation between compound exposure, PI3K pathway blockade (reduction in P-Akt levels) and antitumor activity. Dose dependent tumor growth delay or regression is also observed in primary tumor models, and, as demonstrated previously with our dual pan-PI3K/mTOR inhibitor NVP-BEZ235, in vivo synergistic activity is observed when NVP-BKM120 is combined with a MEK inhibitor in K-Ras human cancer models. NVP-BKM120 is currently undergoing Phase I/II human clinical trials for the treatment of solid tumors and hematological malignancies. 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 4497.

Abstract 2443: Tetraspanin CD151 as a target for antibody-based cancer therapy

Abstract The tetraspanin CD151 is known to be functionally linked to cancer metastasis. In human, increased expression of CD151 is indicative of a poor prognosis in different cancer types. Whereas its precise mechanism of action remains obscure, CD151 was shown to regulate cell motility and adhesion through association with laminin-binding integrins such as α3β1 or α6β4, and more recently the communication between endothelial and tumor cells. Overexpression of CD151 in tumor cells enhances cell migration, cell invasion and experimental metastasis. Silencing of CD151 with specific siRNA was shown to delay tumour formation in vivo in a mouse model of breast cancer. Several anti-CD151 monoclonal antibodies (Mabs) were shown to display anti-metastatic activity in vivo. Inhibition of metastasis was not attributed to any effect of these Mabs on tumor cell growth but was essentially linked to inhibition of cell motility. However, a more direct effect could be expected. For these reasons, we generated different anti-CD151 Mabs via immunisation with transfectant cells and evaluated their anti-tumor activity in xenograft and orthotopic mouse models to further investigate the role of CD151 in cancer progression in vivo. Interestingly, anti-CD151 Mabs inhibited potently primary tumor growth in different xenograft models including prostate, lung and breast cancer models. These results demonstrate that CD151 could be of importance for controlling tumor cell proliferation. In addition, as anticipated, we showed that anti-CD151 Mabs were also able to inhibit metastasis in vivo in an orthotopic A549 lung cancer model with well characterized lung metastases. The mechanism of action of these Mabs on tumor cells is under investigation and preliminary in vitro data will be presented. Altogether, our results strongly suggest that CD151 could function at multiple cancer stages, including not only metastasis cascade steps but also earlier steps of primary tumor growth, reinforcing thus the interest of this innovative target in oncology. Mabs targeting CD151 may be of significant interest for cancer biotherapy. 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 2443.

A phase 2 study of ARQ 501 in combination with gemcitabine in adult patients with treatment naïve, unresectable pancreatic adenocarcinoma

15017 Background: ARQ 501 is a DNA damage checkpoint pathway activator whose effect is to induce selective cell death in cancer cells, independent of the tumor cell’s p53 status. Current evidence implicates a rapid and sustained increase of the pro-apoptotic protein E2F-1 by ARQ 501 as the mechanism of action. Cancer cells are selectively affected due to their pre-existing DNA damage. Induction of E2F-1 in combination with gemcitabine demonstrated potent anti-tumor activity in xenografts of human pancreatic cancer (PACA-2). A recommended phase 2 dose of 400 mg/m2 of ARQ 501 and 800 mg/m2 gemcitabine was identified. Methods: A Phase 2 study in adult patients with treatment naïve, non-resectable adenocarcinoma of the pancreas was initiated to assess the effect of ARQ 501 and gemcitabine on ORR in approximately 66 patients. Cycles consisting of weekly ARQ 501 (400mg/m2) and gemcitabine (800 mg/m2 were repeated every 4 weeks until progression, unacceptable toxicity, or another discontinuation criterion was met. Results: 73 patients were enrolled, with 58 treated to date with at least one infusion. Data is available for 27 patients (16M/11F, median age, 63 years). Of these 27, 5 did not reach a post baseline assessment (3 deaths, 2 PD prior to week 8), 20 are evaluable for response at eight weeks (1 PR, 12 SD of 8–24+ weeks), 7 PD) and 2 have yet to be assessed but are active. Tumor regressions have been observed in 6/20 patients assessed to date. This includes 1 PR, 4 MR (14.3% - 24%) and 1 patient with significant reduction (29.6%) in pancreatic lesions although new superficial liver lesions were observed. Adverse events (N=235) the most common being: anemia (54, 22%), hemolysis (17, 7%), fatigue (15, 6%), edema (7, 3%), and nausea (5, 2%). Conclusions: ARQ 501 has been administered to 58 patients with treatment-naïve, non-resectable adenocarcinoma of the pancreas. Enrollment is complete and early efficacy data is encouraging. A high proportion of patients (13/20) have achieved a minimum SD at the first tumor evaluation. In addition, objective tumor regressions has been observed in this study, providing preliminary confirmation of the utility of induction of E2F-1 in combination with gemcitabine. [Table: see text]

Phase I and pharmacological studies of the cryptophycin analogue LY355703 administered on a single intermittent or weekly schedule

LY355703 is a synthetic derivative of the marine cryptophycins, cytotoxic agents which induce mitotic arrest by binding at the microtubule vinca binding domain. Promising preclinical features of LY355703 were the 40–400 greater potency than paclitaxel or vinca alkaloids, the broad spectrum of antitumor activity in xenografts and the antitumour activity in multidrug resistant (MDR)-expressing murine tumours. Aims of this study were to define the maximum tolerated dose (MTD) and the dose recommended for Phase II, the pattern of toxicity, the pharmacokinetic profile and to document hints of antitumour activity of LY355703 given as 2-h infusion on day 1 every 3 weeks (Study 1) or, later on, on days 1, 8 and 15 every 4 weeks (Study 2). The latter weekly regimen was selected because of the acute dose-related toxicity reported in Study 1. The dose was escalated using a modified Continual Reassessment Method. Pharmacokinetic studies were performed on day 1 of cycle 1 in both studies; LY355703 plasma concentrations were assessed by liquid chromatography with tandem mass spectrometry. A total of 35 adult patients with solid tumours entered Study 1; the dose was escalated from 0.1 to 1.92 mg/m2; at this dose 2 of 5 patients presented grade 3 neuropathy and myalgias; 1.48 mg/m2 was then recommended for Phase II study. A total of 8 patients were treated in Study 2 at 1 mg/m2; cumulative long-lasting neuroconstipation and neurosensory toxicity precluded the completion of the cycle in 9 out of 15 cycles; the clinical development of the weekly regimen was then discontinued. Other toxicities included cardiac dysrhythmia and mild alopecia. Pharmacokinetics of LY355703 appeared to be linear over the dose range studied. The administration of LY355703 on a 3-week schedule is associated with an acute dose-dependent peripheral neuropathy and myalgia of high interpatient variability for which possible risk factors and pharmacokinetic correlates could not be identified.

Relationship between tumor extracellular fluid exposure to topotecan and tumor response in human neuroblastoma xenograft and cell lines.

We have reported a 6-fold difference in the topotecan (TPT) lactone systemic exposure achieving a complete response in the human neuroblastoma xenografts NB-1691 and NB-1643. However, the relationship between tumor extracellular fluid (ECF) exposure to TPT and the antitumor activity in xenograft and in vitro models has not been established. TPT was given i.v. to mice bearing NB-1691 and NB-1643 tumors. Prior to dosing, microdialysis probes were placed in tumors of mice bearing NB-1691 and NB-1643 tumors. Plasma and tumor ECF concentrations of TPT lactone were assayed by high performance liquid chromatography. The inhibitory concentration (IC50) was determined for NB-1691 and NB-1643 cell lines in vitro. The TPT AUC(ECF) values determined for NB-1691 (n = 10) and NB-1643 (n = 11) were 7.3 +/- 0.84 and 25.6 +/- 0.76 ng h ml(-1), respectively (P < 0.05). TPT tumor ECF penetration in NB-1691 and NB-1643 was 0.04 +/- 0.04 and 0.15 +/- 0.11 (P < 0.05), respectively. The IC50 values recorded after 6 h of TPT exposure daily for 5 consecutive days for NB-1691 and NB-1643 were 2.7 +/- 1.1 and 0.53 +/- 0.19 ng/ml, respectively (P < 0.05). NB-1643 was more sensitive in vitro than NB-1691, and at similar plasma TPT exposures, NB-1643 had a greater degree of TPT tumor ECF exposure and penetration as compared with NB-1691. Potential factors affecting tumor TPT ECF disposition include tumor vascularity, capillary permeability, and interstitial pressure. The clinical importance of this study is underscored by the need to select anticancer agents with a high capacity for tumor penetration and to optimize drug administration to increase tumor penetration.