Human Pancreatic Tumor Xenografts Research Articles

Real-Time Metabolic Magnetic Resonance Spectroscopy of Pancreatic and Colon Cancer Tumor-Xenografts with Parahydrogen Hyperpolarized 1-13C Pyruvate-d3.

Parahydrogen-induced polarization (PHIP) is an emerging technique to enhance the signal of stable isotope metabolic contrast agents for Magnetic Resonance (MR). The objective of this study is to continue establishing 1-13C-pyruvate-d3, signal-enhanced via PHIP, as a hyperpolarized contrast agent, obtained in seconds, to monitor metabolism in human cancer. Our focus was on human pancreatic and colon tumor xenografts. 1-13C-vinylpyruvate-d6 was hydrogenated using parahydrogen. Thereafter, the polarization of the protons was transferred to 13C. Following a workup procedure, the free hyperpolarized 1-13C-pyruvate-d3 was obtained in clean aqueous solution. After injection into animals bearing either pancreatic or colon cancer xenografts, slice-selective MR spectra were acquired and analyzed to determine rate constants of metabolic conversion into lactate and alanine. 1-13C-pyruvate-d3 proved to follow the increased metabolic rate to lactate and alanine in the tumor xenografts.

Abstract 663: Irreversible covalent azetidine-based small molecule inhibitors of Stat3 activity block growth of human pancreatic tumors

Abstract The incidence and death rates for pancreatic adenocarcinoma in the US have been increasing by about 1% per year and the death rate has increased by approximately 0.2% yearly. The five-year survival rate has remained below 10% for many years. With surgery, radiation, and chemotherapy being the main treatment options to extend survival or for symptom relief, there is an urgent unmet need for effective new treatment options. Signal Transducer and Activator of Transcription (Stat) 3, one of the Stat family members, is aberrantly activated in human pancreatic and many other cancers. Aberrantly-active Stat3 promotes abnormal tumor-cell intrinsic and extrinsic mechanisms, including dysregulation of gene expression and mitochondria energy metabolism in the tumor cells, and suppression of immune cell functions in the tumor microenvironment. Stat3 is therefore an important target for therapeutic development. Herein we present two small molecules, H182 and H333, which are covalent inhibitors of Stat3 and that potently block Stat3 DNA-binding activity in vitro, with IC50 0.66-0.98 µM. H182 and its structural analog, H333 bind Stat3 via irreversible covalent interactions with key cysteine residues in the Stat3 DNA-binding domain to induce a time-dependent inhibition of Stat3 activity. Both compounds inhibit intracellular constitutive Stat3 tyrosine phosphorylation and induce loss of viable cells and apoptosis in vitro of human pancreatic Panc-1 and Mia-Paca2 cells. H333 is 13-fold more specific and H182 is 4-fold more specific against tumor cells over normal cells. Furthermore, MiaPaca-2 cells treated with H182 and analyzed by Seahorse showed dramatically decreased mitochondria respiration. Notably, xenograft models of MiaPaca2 treated with H182 or H333 via i.p. every other day for 27 days showed dramatic growth inhibition. Collectively, our results identify H182 and H333 as therapeutically-viable small molecules with unique irreversible mechanism of Stat3 inhibition which accounts for their strong antitumor effects against human pancreatic tumor xenografts. Citation Format: Yue Chen, Ning Zhai, Peibin Yue, Christine Brotherton-Pleiss, Wenzhen Fu, Kayo Nakamura, Weiliang Chen, Marcus Tius, Francisco Lopez-Tapia, James Turkson. Irreversible covalent azetidine-based small molecule inhibitors of Stat3 activity block growth of human pancreatic tumors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 663.

A polyethylene glycol-conjugate of deoxycytidine analog, DFP-14927, produces potential antitumor effects on pancreatic tumor-xenograft murine models via inducing G2/M arrest

A deoxycytidine analog is a potential agent for the treatment of several cancers, which includes poorly prognostic pancreatic cancer. We previously developed deoxycytidine analog DFP-10917, and long-term/low-dose infusions of this analog has produced antitumor effects in leukemia cancer- and ovarian cancer-xenograft models. DFP-10917 is now undergoing clinical Phase III study in the United States for the treatment of patients with relapsed or refractory acute myeloid leukemia. PEG-drug conjugation has become a promising technique to improve the pharmacokinetic and pharmacodynamic properties of anti-cancer drugs. In the present study, we synthesized a novel PEG-drug conjugate of DFP-10917, referred to hereafter as DFP-14927, using a 4-armed CTPEG system to endow the DFP-10917 drug with favorable long-circulating properties that maximize its utility and antitumor efficacy. Intravenous injection of the synthesized DFP-14927 returned encouraging antitumor effects in a Panc-1 human pancreatic tumor- and a BxPC-3 human pancreatic tumor-xenograft models. These effects were comparable to that of free DFP-10917 as well as to that of gemcitabine, which is considered a standard in the treatment of pancreatic cancer. In vitro studies revealed that DFP-14927 inhibits cell division on human pancreatic cancer cell lines via arrest of the G2/M phase in the cell cycle, which is consistent with the effects of free DFP-10917. Intravenous administration of the newly synthesized DFP-14927 has induced G2/M arrest in human pancreatic tumor-xenograft murine models, which represents an improvement in the pharmacokinetics of DFP-10917. DFP-14927 could be an alternative for patients who cannot accept prolonged or continuous infusions of DFP-10917.

Pancreatic cancer growth promoted by bone marrow mesenchymal stromal cell–derived IL-6 is reversed predominantly by IL-6 blockade

Pancreatic cancer is a highly lethal cancer characterized by local invasiveness, early metastasis, recurrence and high resistance to current therapies. Extensive stroma or desmoplasia is a key histological feature of the disease, and interactions between cancer and stromal cells are critical for pancreatic cancer development and progression. Mesenchymal stromal cells [MSCs] exhibit preferential tropism to primary and metastatic tumor sites and may either suppress or support tumor growth. Although MSCs represent a potential source of pancreatic cancer stroma, their contribution to pancreatic tumor growth remains poorly known. Here, we show that bone marrow MSCs significantly contribute to pancreatic cancer growth in vitro and in vivo. Furthermore, MSCs create a pro-carcinogenic microenvironment through the release of key factors mediating growth and angiogenesis, including interleukin (IL)-6, IL-8, vascular endothelial growth factor and activation of STAT3 signaling in tumor cells. IL-6 released by MSCs was largely responsible for the pro-tumorigenic effects of MSCs. Knockdown of IL-6 expression in MSCs by small interfering RNA (siRNA) abolished the MSC growth-promoting effect in vitro, reducing tumor cell proliferation and clonogenic potential. In addition, in a heterotopic nude mouse model of human pancreatic tumor xenografts, blockade of IL-6 with the anti-IL-6 receptor antibody, tocilizumab, or of its downstream effector STAT3 with the small molecule STAT3 inhibitor S3I-201, abrogated MSC-mediated tumor promotion and delayed tumor formation significantly. Our data demonstrate that MSCs promote pancreatic cancer growth, with IL-6 produced by MSCs playing a pivotal role.

Two Laser Treatments Can Improve Tumor Ablation Efficiency of Chemophototherapy.

Chemophototherapy is an emerging tumor ablation modality that can improve local delivery of chemotherapeutic agents. Long circulating doxorubicin (Dox) in porphyrin-phospholipid (PoP) liposomes (LC-Dox-PoP) has previously been developed as an effective chemophototherapy agent. In the present study, we observed that in mice, LC-Dox-PoP showed enhanced accumulation in human pancreatic tumor xenografts even with suboptimal light doses, as assessed by fluorometric analysis of tissue homogenates and microscopic imaging of Dox and PoP in tumor slices. A second laser treatment, at a time point in which tumors had greater drug accumulation as a result of the first laser treatment, induced potent tumor ablation. Efficacy studies were carried out in two human pancreatic cancer subcutaneous mouse tumor models; MIA PaCa-2 or low-passage patient derived pancreatic cancer xenografts. A single treatment of 3 mg/kg LC-Dox-PoP and an initial 150 J/cm2 laser treatment 1 h after drug administration, followed by second laser treatment of 50 J/cm2 8 h after drug administration, was more effective than a single laser treatment of 200 J/cm2 at either of those time points. Thus, this study presents proof-of-principle and rationale for using two discrete laser treatments to enhance the efficacy of chemophototherapy.

Eradicating mesothelin-positive human gastric and pancreatic tumors in xenograft models with optimized anti-mesothelin antibody\u2013drug conjugates from synthetic antibody libraries

Mesothelin (MSLN) is an attractive candidate of targeted therapy for several cancers, and hence there are increasing needs to develop MSLN-targeting strategies for cancer therapeutics. Antibody–drug conjugates (ADCs) targeting MSLN have been demonstrated to be a viable strategy in treating MSLN-positive cancers. However, developing antibodies as targeting modules in ADCs for toxic payload delivery to the tumor site but not to normal tissues is not a straightforward task with many potential hurdles. In this work, we established a high throughput engineering platform to develop and optimize anti-MSLN ADCs by characterizing more than 300 scFv CDR-variants and more than 50 IgG CDR-variants of a parent anti-MSLN antibody as candidates for ADCs. The results indicate that only a small portion of the complementarity determining region (CDR) residues are indispensable in the MSLN-specific targeting. Also, the enhancement of the hydrophilicity of the rest of the CDR residues could drastically increase the overall solubility of the optimized anti-MSLN antibodies, and thus substantially improve the efficacies of the ADCs in treating human gastric and pancreatic tumor xenograft models in mice. We demonstrated that the in vivo treatments with the optimized ADCs resulted in almost complete eradication of the xenograft tumors at the treatment endpoints, without detectable off-target toxicity because of the ADCs’ high specificity targeting the cell surface tumor-associated MSLN. The technological platform can be applied to optimize the antibody sequences for more effective targeting modules of ADCs, even when the candidate antibodies are not necessarily feasible for the ADC development due to the antibodies’ inferior solubility or affinity/specificity to the target antigen.

Discovery of novel KRAS‒PDEδ inhibitors with potent activity in patient-derived human pancreatic tumor xenograft models

KRAS‒PDEδ interaction is revealed as a promising target for suppressing the function of mutant KRAS. The bottleneck in clinical development of PDEδ inhibitors is the poor antitumor activity of known chemotypes. Here, we identified novel spiro-cyclic PDEδ inhibitors with potent antitumor activity both in vitro and in vivo. In particular, compound 36l (KD = 127 ± 16 nmol/L) effectively bound to PDEδ and interfered with KRAS–PDEδ interaction. It influenced the distribution of KRAS in Mia PaCa-2 cells, downregulated the phosphorylation of t-ERK and t-AKT and promoted apoptosis of the cells. The novel inhibitor 36l exhibited significant in vivo antitumor potency in pancreatic cancer patient-derived xenograft (PDX) models. It represents a promising lead compound for investigating the druggability of KRAS‒PDEδ interaction.

Reduction-Responsive and Multidrug Deliverable Albumin Nanoparticles: An Antitumor Drug to Abraxane against Human Pancreatic Tumor-Bearing Mice.

Many macromolecular antitumor drugs were developed based on the enhanced permeability and retention (EPR) effect, for example, albumin-bound paclitaxel nanoparticles (nab-PTX and Abraxane) and pegylated liposomal doxorubicin (Doxil). However, these EPR effect-based therapeutic systems are less effective in malignant tumors with low vascular permeability, such as pancreatic tumors. Because the EPR effect depends on nanoparticles' size, we first determined nanoparticles' size associated with a high tumor-targeting rate in a human pancreatic tumor xenograft model with low vascular permeability. Abraxane appears to behave as an albumin monomer (7 nm) in the blood circulation following intravenous injection. The in vitro and in vivo tumor-targeted delivery and antitumor activity of PTX-loaded albumin nanoparticles were significantly improved by optimizing the mean nanoparticle diameter to 30 nm. Furthermore, nitric oxide was added to 30 nm PTX-loaded albumin nanoparticles to examine the feasibility of albumin nanoparticles as a platform for multiple drug delivery. Their antitumor effect was evaluated in an orthotopic transplantation mouse model of a human pancreatic tumor. The nitric oxide PTX-loaded 30 nm albumin nanoparticle treatment on model mice achieved a significantly higher survival rate than Abraxane treatment. These findings suggest that 30 nm albumin nanoparticles have a high therapeutic effect as a useful platform for multiple drugs against human pancreatic tumors.

Molecular Imaging of the Tumor Microenvironment Reveals the Relationship between Tumor Oxygenation, Glucose Uptake, and Glycolysis in Pancreatic Ductal Adenocarcinoma.

Molecular imaging approaches for metabolic and physiologic imaging of tumors have become important for treatment planning and response monitoring. However, the relationship between the physiologic and metabolic aspects of tumors is not fully understood. Here, we developed new hyperpolarized MRI and electron paramagnetic resonance imaging procedures that allow more direct assessment of tumor glycolysis and oxygenation status quantitatively. We investigated the spatial relationship between hypoxia, glucose uptake, and glycolysis in three human pancreatic ductal adenocarcinoma tumor xenografts with differing physiologic and metabolic characteristics. At the bulk tumor level, there was a strong positive correlation between 18F-FDG-PET and lactate production, while pO2 was inversely related to lactate production and 18F-2-fluoro-2-deoxy-D-glucose (18F-FDG) uptake. However, metabolism was not uniform throughout the tumors, and the whole tumor results masked different localizations that became apparent while imaging. 18F-FDG uptake negatively correlated with pO2 in the center of the tumor and positively correlated with pO2 on the periphery. In contrast to pO2 and 18F-FDG uptake, lactate dehydrogenase activity was distributed relatively evenly throughout the tumor. The heterogeneity revealed by each measure suggests a multimodal molecular imaging approach can improve tumor characterization, potentially leading to better prognostics in cancer treatment. SIGNIFICANCE: Novel multimodal molecular imaging techniques reveal the potential of three interrelated imaging biomarkers to profile the tumor microenvironment and interrelationships of hypoxia, glucose uptake, and glycolysis.

Monitoring Dual VEGF Inhibition in Human Pancreatic Tumor Xenografts With Dynamic Contrast-Enhanced Ultrasound.

Background:Association of drugs acting against different antiangiogenic mechanisms may increase therapeutic effect and reduce resistance. Noninvasive monitoring of changes in the antiangiogenic response of individual tumors could guide selection and administration of drug combinations. Noninvasive detection of early therapeutic response during dual, vertical targeting of the vascular endothelial growth factor pathway was investigated in an ectopic subcutaneous xenograft model for human pancreatic tumor.Methods:Dynamic contrast-enhanced ultrasound 12 MHz was used to monitor tumor-bearing Naval Medical Research Institute mice beginning 15 days after tumor implantation. Mice received therapy from 15 to 29 days with sorafenib (N = 9), ziv-aflibercept (N = 11), combined antiangiogenic agents (N = 11), and placebo control (N = 14). Sorafenib (BAY 43-9006; Nexavar), a multikinase inhibitor acting on Raf kinase and receptor tyrosine kinases—including vascular endothelial growth factor receptors 2 and 3—was administered daily (60 mg/kg, per os). Ziv-aflibercept (ZALTRAP), a high-affinity ligand trap blocking the activity of vascular endothelial growth factor A, vascular endothelial growth factor B, and placental growth factor was administered twice per week (40 mg/kg, intraperitoneally).Results:Functional evaluation with dynamic contrast-enhanced ultrasound indicated stable tumor vascularization for the control group while revealing significant and sustained reduction after 1 day of therapy in the combined group (P = .007). There was no survival benefit or penalty due to drug combination. The functional progression-free survival assessed with dynamic contrast-enhanced ultrasound was significantly higher for the 3 treated groups; whereas, the progression-free survival based on tumor size did not discriminate therapeutic effect.Conclusions:Dynamic contrast-enhanced ultrasound, therefore, presents strong potential to monitor microvascular modifications during antiangiogenic therapy, a key role to monitoring antiangiogenic combining therapy to adapt dose range drug.

Abstract 3090: In vivo characterization of AMG 510 - a potent and selective KRASG12Ccovalent small molecule inhibitor in preclinical KRASG12Ccancer models

Abstract Somatic activating mutations of RAS family members are tumor driver mutations found in an estimated 21% of all cancers. Oncogenic KRAS mutations at residues G12, G13, and Q61 represent the most common RAS mutations found in solid malignancies. The prevalence of KRAS p.G12C tumors is ~13% of lung adenocarcinoma (including NSCLC), 3% of colorectal carcinoma (CRC), and 1% to 2% of numerous other solid tumors, representing an unmet medical need. We have developed AMG 510, an orally bioavailable, covalent inhibitor of KRASG12C with potent biochemical and cellular activity, and robust in vivo efficacy. AMG 510 inhibited SOS-catalyzed nucleotide exchange of recombinant mutant KRASG12C/C118A but had minimal effect on KRASC118A, which is wildtype at position 12. In cellular assays, AMG 510 covalently modified KRASG12C and inhibited KRASG12C signaling as measured by phosphorylation of ERK1/2 (p-ERK) in all KRAS p.G12C-mutant cell lines tested but did not inhibit p-ERK in cell lines with various other KRAS mutations. AMG 510 also selectively impaired viability of KRAS p.G12C mutant cell lines but did not affect cell lines with other KRAS mutations. In vivo pharmacodynamic assays demonstrated dose- and time-dependent inhibition of KRASG12Csignaling in human pancreatic and NSCLC tumor xenografts. Covalent modification of KRASG12C by AMG 510 was measured by mass spectrometry and correlated with p-ERK inhibition in tumors. AMG 510 significantly inhibited the growth of KRAS p.G12C xenografts and resulted in tumor regression. Combination treatment of AMG 510 with standard-of-care and targeted agents demonstrated enhanced tumor growth inhibition compared to either single agent. In a syngeneic model of KRAS p.G12C mutant cancer, AMG 510 treatment significantly inhibited tumor growth and caused regression. AMG 510 is currently in Phase 1, first-in-human clinical trials for patients with advanced solid tumors harboring a KRAS p.G12C mutation. Citation Format: Karen Rex, Anne Y. Saiki, Ji-Rong Sun, Tyler Holt, Neelima Koppada, Brian A. Lanman, Laurie P. Volak, Robert S. Foti, Victor J. Cee, J. Russell Lipford, Jude Canon. In vivo characterization of AMG 510 - a potent and selective KRASG12Ccovalent small molecule inhibitor in preclinical KRASG12Ccancer models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3090.

Correlation of two distinct metastasis-associated proteins, MTA1 and S100A4, in angiogenesis for promoting tumor growth.

Extensive studies on metastasis-associated proteins, S100A4 and MTA1, have been carried out for over two decades, but correlation of both proteins remains obscure. Here we show evidence for the correlation in angiogenesis. First, silencing of each protein by siRNA-mediated knockdown in mouse endothelial MSS31 cells resulted in the inhibition of tube formation. Unexpectedly, the knockdown of MTA1 affected not only its own expression but also the expression of S100A4, whereas silencing of S100A4 did not affect the MTA1 expression. Additionally, non-muscle myosin IIA (NMIIA) phosphorylation, which was partly controlled by S100A4, was found to be upregulated by knockdown of both proteins in MSS31 cells. Moreover, cycloheximide treatment of MSS31 cells revealed that the rate of S100A4 degradation was accelerated by MTA1 knockdown. This finding, together with our observation that cytoplasmic MTA1, but not nuclear MTA1, was colocalized with S100A4, suggested the involvement of MTA1 in S100A4 stability. The direct in vivo angiogenesis assay showed that both protein siRNAs provoked a significant inhibition of new blood vessel formation induced by angiogenic factors, indicating their anti-angiogenic activities. Treatment of human pancreatic tumor (PANC-1) xenograft in mice with mMTA1 siRNA resulted in tumor regression via suppression of angiogenesis in vivo, as also observed in the case of human prostate cancer xenograft treated with mS100A4 siRNA. Taken together, these data led us to conclude that the MTA1-S100A4-NMIIA axis exists in endothelial cells as a novel pathway in promoting tumor vascular formation and could be a target for suppressing tumor growth and metastasis.

Resibufogenin suppresses transforming growth factor-β-activated kinase 1-mediated nuclear factor-κB activity through protein kinase C-dependent inhibition of glycogen synthase kinase 3.

Resibufogenin (RB), one of the major active compounds of the traditional Chinese medicine Chansu, has received considerable attention for its potency in cancer therapy. However, the anticancer effects and the underlying mechanisms of RB on pancreatic cancer remain elusive. Here, we found that RB inhibited the viability and induces caspase‐dependent apoptosis in human pancreatic cancer cells Panc‐1 and Aspc. Resibufogenin‐induced apoptosis was through inhibition of constitutive nuclear factor‐κB (NF‐κB) activity and its target genes’ expression, which was caused by downregulation of transforming growth factor‐β‐activated kinase 1 (TAK1) levels and suppression of IκB kinase activity in Panc‐1 and Aspc cells. This induction of TAK1‐mediated NF‐κB inactivation by RB was associated with increased glycogen synthase kinase‐3 (GSK‐3) phosphorylation and subsequent suppression of its activity. Moreover, RB‐induced GSK‐3 phosphorylation/inactivation acted through activation of protein kinase C but not Akt. Finally, RB suppressed human pancreatic tumor xenograft growth in athymic nude mice. Thus, our findings reveal a novel mechanism by which RB suppresses TAK1‐mediated NF‐κB activity through protein kinase C‐dependent inhibition of GSK‐3. Our findings provide a rationale for the potential application of RB in pancreatic cancer therapy.

Abstract B68: Targeted inhibition of Tgfβr2 reduces IL-6 production from cancer-associated fibroblasts, suppresses Stat3 activation in pancreatic cancer cells and reverses immunosuppression

Abstract Pancreatic cancer is characterized by dense stroma consisting of myofibroblasts, abundant extracellular matrix (ECM) and inflammatory/immune cells. Elevated levels of transforming growth factor β (TGFβ), a prominent driver of stromal accumulation, is a negative prognostic indicator for pancreatic cancer patients. However, inhibition of TGFβ has been challenging due to the context-dependent tumor suppressor function of TGFβ in pancreatic cancer. We reported previously that neutralizing the activity of murine TGFβ receptor 2 (Tgfβr2) using a monoclonal antibody (2G8), has potent antitumor activity in orthotopic human pancreatic tumor xenografts. The findings in xenografts highlight that TGFβ induction of stromal cell function is critical for its impact on tumor cell behavior and PDA progression and supports that inhibition of TGFβ-dependent effector functions on stromal cells has potential efficacy in PDA. Herein we demonstrate that inhibition of murine Tgfβr2 reduces IL-6 production from cancer-associated fibroblasts (CAFs), resulting in a reduction of Stat3 activation in tumor cells and reversion of the immunosuppressive landscape of murine models of PDA. Moreover, the reduction of stromal IL-6 results in activation of NK cells. In summary, we found TGFβ induced IL-6 secretion from CAFs in PDA contributes to tumorigenesis and immunosuppression and can be abrogated by inhibition of stromal Tgfβr2. Citation Format: Huocong Huang, Moriah Hagopian, Wenting Du, Yuqing Zhang, Kyla E. Driscoll, Rolf A. Brekken. Targeted inhibition of Tgfβr2 reduces IL-6 production from cancer-associated fibroblasts, suppresses Stat3 activation in pancreatic cancer cells and reverses immunosuppression [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2017 Oct 1-4; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2018;6(9 Suppl):Abstract nr B68.

Gemcitabine treatment promotes immunosuppressive microenvironment in pancreatic tumors by supporting the infiltration, growth, and polarization of macrophages

Chemotherapy-induced immunosuppression poses an additional challenge to its limited efficacy in pancreatic cancer (PC). Here we investigated the effect of gemcitabine on macrophages, which are the first line of immune-defense mechanisms. We observed an increased presence of macrophages in orthotopic human pancreatic tumor xenografts from mice treated with gemcitabine as compared to those from vehicle only-treated mice. Conditioned media from gemcitabine-treated PC cells (Gem-CM) promoted growth, migration and invasion of RAW264.7 macrophage. In addition, Gem-CM also induced upregulation of M2-polarized macrophage markers, arginase-1 and TGF-β1. Cytokine profiling of gemcitabine-treated PC cells identified IL-8 as the most differentially-expressed cytokine. Incubation of Gem-CM with IL-8 neutralizing antibody diminished its ability to induce growth, migration and invasion of RAW264.7 macrophages, but did not abrogate their M2 polarization. Together, our findings identify IL-8 as an important mediator in the gemcitabine-induced infiltration of macrophages within the pancreatic tumor microenvironment and suggest the requirement of additional mechanism(s) for macrophage polarization.

Abstract 3036: PEGylated hyaluronidase increases tumor uptake of 89Zr-DFO-HuMab-5B1 (MVT-2163) in a CA19-9 positive hyaluronan-accumulating pancreatic cancer model

Abstract Hyaluronan (HA) accumulates in the extracellular matrix of many solid tumors, and is associated with poor prognosis in pancreatic ductal adenocarcinoma. In non-clinical models, enzymatic degradation of HA with PEGylated recombinant human hyaluronidase PH20 (PEGPH20) has been shown to remodel the tumor stroma, reduce tumor interstitial fluid pressure, and expand tumor blood vessels, resulting in enhanced delivery of therapeutic and imaging agents, such as monoclonal antibodies. 89Zr-DFO-HuMab-5B1 (MVT-2163) is a targeted ImmuoPET imaging agent for CA19-9 positive malignancies currently in clinical evaluation. CA19-9 plays a role in tumor adhesion and metastasis, and is an independent prognostic indicator of cancer survival. CA19-9 is expressed in pancreatic and other cancers, including small cell lung and colon. In this study, we aimed to show the effects of PEGPH20 on the biodistribution of MVT-2163 in a CA19-9 positive HA-rich human pancreatic tumor xenograft model. Nude mice were peritibially implanted on the right hind limb with 5x106 human pancreatic tumor cells engineered to overexpress hyaluronan synthase 3 (BxPC3/HAS3). Tumor volumes were measured by MR imaging, and mice were staged when average tumor size reached 320 mm3. The control group received intravenous (IV) vehicle, and the test group received PEGPH20 (IV, 1 mg/kg). Both control and test groups also received MVT-2163 (IV, 3 mg/kg) 24 hours later. PET images were captured from 2 to 120 hours post MVT-2163, and region of interest (ROI) of tumor and liver were analyzed (N=6/group). After the final PET image, tumors and livers were harvested for ex vivo gamma counting. Ex vivo gamma counting demonstrated improved MVT-2163 accumulation in animals treated with PEGPH20, with a 50.9% increase in counts in the excised tumors. ROI analysis of the tumors showed increased tumor SUV of 8.0, 9.4, and 24.1% at 72, 96, and 120 hours post-injection, respectively, with PEGPH20 compared to vehicle. ROI analysis of liver regions showed an 11.4 to 26.4% reduction in SUV in the PEGPH20 group compared to control. Ex vivo counts of liver tissue confirmed the PET signal reduction with PEGPH20. Analysis of tumor-to-liver ratios showed average increases of 34.1, 35.7, and 58.5% at 72, 96, and 120 hours, respectively, post MVT-2163 injection in PEGPH20 treated mice. In summary, as measured by SUV, PEGPH20 increased both the tumor uptake and the tumor-to-liver ratios of MVT-2163 in a CA19-9 positive xenograft mouse model of HA accumulating pancreatic cancer. Ex vivo analysis confirmed in vivo results. Taken together, the increased tumor uptake and the decreased liver uptake support further investigation into the potential clinical utility for the combination of PEGPH20 and MVT-2163. Citation Format: Jonah Rainey, Paul Maffuid, Wolfgang W. Scholz, Jack Ostrowski, H Toni Jun, Paul Resnick, Xiaoming Li, Jesse D. Bahn, Susan Zimmerman, Kelly Chen, Barbara Blouw, Curtis B. Thompson, Daniel C. Maneval, David W. Kang. PEGylated hyaluronidase increases tumor uptake of 89Zr-DFO-HuMab-5B1 (MVT-2163) in a CA19-9 positive hyaluronan-accumulating pancreatic cancer model [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 3036.

Antifibrotic effects of pentoxifylline improve the efficacy of gemcitabine in human pancreatic tumor xenografts.

We investigated the combinatorial effects of pentoxifylline (PTX) on the efficacy of gemcitabine (GEM) in a human pancreatic tumor xenograft model. PTX significantly improved the efficacy of GEM, as shown by a 50% reduction in tumor growth rate at 4 weeks of treatment compared with that in animals given GEM alone. The fluorescent drug doxorubicin (DOX) was used to test whether drug delivery was improved by PTX, contributing to the improved efficacy of GEM. PTX given for 2 weeks prior to giving DOX improved drug distribution by 1.8‐ to 2.2‐fold with no changes in vessel density, suggesting that improvement in drug delivery was not related to the vascular mechanism. Instead, collagen I content in tumor stroma was significantly reduced, as was the expression of alpha‐smooth muscle actin of cancer‐associated fibroblasts and connective tissue growth factor (CTGF) by PTX pretreatment. Overall, our data demonstrated that increased efficacy of GEM by PTX was associated with improved drug delivery to tumor tissue, which may be attributed to decreased expression of CTGF and subsequent reduction in the stromal collagen matrix in the pancreatic ductal adenocarcinoma tumor. These results support the usefulness of PTX in combination with chemotherapy for targeting drug delivery barriers associated with the stromal matrix, which should be further evaluated for clinical development.

Abstract 5204: Preclinical development of MVT-1075 as radioimmunotherapy for pancreatic cancer and other CA19-9 positive malignancies

Abstract Rationale: The CA19-9 antigen is frequently overexpressed in pancreatic and other GI tumors. MVT-5873 (HuMab-5B1), a fully human monoclonal antibody currently in phase I study, targets the sialyl Lewis A (sLea) epitope on CA19-9, and is a promising platform for development of a targeted radioimmunotherapy (RIT). MVT-5873 was conjugated with the chelator CHX-A″-DTPA and radiolabeled with the beta-emitting isotopes Lutetium -177 (177Lu) or Yttrium-90 (90Y) to form the RIT agents MVT-1075 (177Lu- CHX-A″-DTPA-HuMAb-5B1) and MVT-1916 (90Y- CHX-A″-DTPA-HuMAb-5B1), respectively. The antitumor efficacy of each of the constructs was studied in nude mice bearing BxPC3 human pancreatic tumor xenografts, known to express CA19-9. Methods: The initial dose-finding studies utilized doses of MVT-1075 of 75-450 μCi and MVT-1916 of 25-250 μCi, administered to groups of mice (n = 8) bearing subcutaneous (subQ) BxPC3 tumors (~ 150 mm3). Further studies focused on MVT-1075 and assessed antitumor effect in an orthotopic xenograft model, the effect of dose fractionation, and biodistribution in nontumor bearing (normal) and BxPC3 tumor-bearing mice. Results: A single dose of MVT-1075 at 75, 150, 300, or 450 μCi significantly inhibited subQ BxPC3 tumor growth at all dose levels, with sustained suppression with higher doses. MVT-1916 produced similar results. MVT-1075 was selected based on the favorable half-life of 177Lu (6.7 d) and its utility for clinical biodistribution assessments. In an orthotopic BxPC3 tumor model, treatment with a single dose of MVT-1075 at 300 μCi significantly inhibited tumor growth, with Day 20 tumor volume approximately 50% that of the initial starting volume. A third BxPC3 xenograft study evaluated fractionated dosing schedules, (150 μCi x 1, 75 μCi x 2, 50 μCi x3), with both single-dose and fractionated schedules effectively inhibiting subQ BxPC3 tumor growth. Biodistribution studies in normal mice showed an expected gradually decreasing activity in blood, heart, and lungs, with low uptake in normal pancreas. In subQ BxPC3 tumor-bearing mice, tumor uptake was rapid, reaching 69% ID/g by 24 h and 86% ID/g by 120 h. Otherwise, the biodistribution pattern paralleled that of normal mice, with relative %ID/g values within about ± 25% of normal mice across all time points comparing blood, heart, lungs, kidneys, and pancreas, with slightly higher uptake in liver and slightly lower uptake in spleen. Conclusions: MVT-1075 demonstrates promising antitumor activity in a human pancreatic cancer xenograft model, with efficacy shown in both single dose and fractionated schedules. Biodistribution shows rapid and substantial tumor uptake, with much lower uptake in normal organs. These findings support the phase I clinical trial of MVT-1075 in patients with CA19-9 positive pancreatic cancers planned to begin in early 2017. Citation Format: Jacob L. Houghton, Ryan Lanning, Dayla Abdel-atti, Toni Jun, Christine M. Kearns, Michael Schlosser, Wolfgang Scholz, Jason S. Lewis, Paul W. Maffuid. Preclinical development of MVT-1075 as radioimmunotherapy for pancreatic cancer and other CA19-9 positive malignancies [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 5204. doi:10.1158/1538-7445.AM2017-5204

Abstract A73: Antitumor activity of MVT-5873, a monoclonal antibody targeting sialyl Lewisa, alone and in combination with gemcitabine/nab-paclitaxel in a BxPC3 human pancreatic cancer xenograft model

Abstract Introduction: MVT-5873 (HuMab-5B1) is a fully human monoclonal antibody that specifically targets the sialyl Lewis A (sLea) epitope on CA19-9, a carbohydrate antigen frequently overexpressed in pancreatic and other gastrointestinal malignancies. The antitumor activity of MVT-5873 as a single agent and in combination with gemcitabine (Gem) + nab-paclitaxel (nab-P, Abraxane®), a standard of care combination for pancreatic cancer, was evaluated in SCID mice bearing subcutaneous BxPC3 human pancreatic cancer tumor xenografts, known to express CA19-9. Methods: Treatment was initiated on Day 9 after BxPC3 cell inoculation; mean tumor volume having reached 150 mm3. Groups of mice (n= 5) were given MVT-5873 at dose levels of 5, 15, and 30 mg/kg alone or with Gem 80 mg/kg + nab-P 20 mg/kg intraperitoneally 2x/week for 5 weeks. Control groups received human IgG 30 mg/kg, Gem/nab-P, or saline on the same schedule. Tumor volumes were measured 2x/week through Day 41, trough (Cmin) serum samples for MVT-5873 pharmacokinetic (PK) analysis were obtained Days 13, 16, 23, 30, 37, & 44. Tumor tissue samples for IHC analysis were obtained upon study termination, Day 44. Results: Day 41 tumor volumes were substantially reduced with single-agent MVT-5873 at doses of 5, 15, and 30 mg/kg, with relative mean volumes of 68%, 76%, and 64%, respectively, that of IgG control. Combination of Gem/nab-P with MVT-5873 at 5, 15, or 30 mg/kg further enhanced activity, resulting relative tumor volumes of 39%, 32%, and 31%, respectively, that of IgG control (p <.01 for all). Using Gem/nab-P as comparative baseline, relative Day 41 tumor volumes were 82%, 67%, and 65%, respectively, with MVT-5873 at 5, 15, or 30 mg/kg + Gem/nab-P (p <.05 for the 15 and 30 mg/kg groups). Tumor Growth Inhibition (TGI) Index values for single-agent MVT-5873 at 5, 15, and 30 mg/kg were 37%, 28%, and 42%, respectively, while combinations of MVT-5873 at 5, 15, and 30 mg/kg with Gem/nab-P produced TGI values of 71%, 80%, and 81%, respectively. Time to 50% tumor growth increased approximately 2-fold across all MVT-5873 + Gem/nab-P groups. Serial MVT-5873 Cmin PK values remained relatively constant in the absence of tumor but declined over time in tumor-bearing animals, with an inverse relationship between serum concentrations and tumor volume suggested. IHC analysis of tumor-associated MVT-5873 antibody showed intensified uptake relative to MVT-5873 dose and an indication of higher tumor uptake with the addition of chemotherapy. Conclusions: MVT-5873 demonstrates antitumor activity as a single agent and enhances the activity of Gem/nab-P chemotherapy in a BxPC3 human pancreatic xenograft model. These findings support the current Phase I clinical investigation of MVT-5873 as a single agent and in combination with Gem/nab-P in patients with advanced pancreatic cancer and other CA19-9 positive malignancies. (NCT02672917) Citation Format: Govind Ragupathi, Xiaohong Wu, Philip Livingston, Wolfgang Scholz, Christine Kearns, Paul Maffuid.{Authors}. Antitumor activity of MVT-5873, a monoclonal antibody targeting sialyl Lewisa, alone and in combination with gemcitabine/nab-paclitaxel in a BxPC3 human pancreatic cancer xenograft model. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr A73.

Bazedoxifene as a Novel GP130 Inhibitor for Pancreatic Cancer Therapy.

The IL6/GP130/STAT3 pathway is crucial for tumorigenesis in multiple cancer types, including pancreatic cancer, and presents as a viable target for cancer therapy. We reported Bazedoxifene, which is approved as a selective estrogen modulator by FDA, as a novel inhibitor of IL6/GP130 protein-protein interactions using multiple ligand simultaneous docking and drug repositioning approaches. STAT3 is one of the major downstream effectors of IL6/GP130. Here, we observed Bazedoxifene inhibited STAT3 phosphorylation and STAT3 DNA binding, induced apoptosis, and suppressed tumor growth in pancreatic cancer cells with persistent IL6/GP130/STAT3 signaling in vitro and in vivo In addition, IL6, but not INFγ, rescued Bazedoxifene-mediated reduction of cell viability. Bazedoxifene also inhibited STAT3 phosphorylation induced by IL6 and IL11, but not by OSM or STAT1 phosphorylation induced by INFγ in pancreatic cancer cells, suggesting that Bazedoxifene inhibits the GP130/STAT3 pathway mediated by IL6 and IL11. Furthermore, Bazedoxifene combined with paclitaxel or gemcitabine synergistically inhibited cell viability and cell migration in pancreatic cancer cells. These results indicate that Bazedoxifene is a potential agent and can generate synergism when combined with conventional chemotherapy in human pancreatic cancer cells and tumor xenograft in mice. Therefore, our results support that Bazedoxifene as a novel inhibitor of GP130 signaling and may be a potential and safe therapeutic agent for human pancreatic cancer therapy. Mol Cancer Ther; 15(11); 2609-19. ©2016 AACR.