Leslie Curtin

Robot‐assisted approach to ‘W’‐configuration urinary diversion: a step‐by‐step technique

To describe a detailed step‐by‐step approach of our technique for robot‐assisted intracorporeal ‘W’‐configuration orthotopic ileal neobladder.

Influence of the implantation site on the sensitivity of patient pancreatic tumor xenografts to Apo2L/TRAIL therapy.

Objectives We have previously demonstrated activity of Apo2L/TRAIL against patient pancreatic tumor xenografts. Here, we have examined the influence of the tumor implantation site on therapeutic response of orthotopic tumors and their metastases to Apo2L/TRAIL. Methods Sensitivity of 6 patient pancreatic tumor xenografts to Apo2L/TRAIL was determined in a subcutaneous model. To compare the response of orthotopic tumors, cells from subcutaneous xenografts were injected into the pancreas. Tumor growth was confirmed by histological examination of selected mice, and then treatment was started. When all control mice developed externally palpable tumors, the experiment was terminated, and pancreatic weights compared between control and treated groups. Magnetic resonance imaging was used to quantitate the response of orthotopic and metastatic tumors. Results The sensitivity to Apo2L/TRAIL observed in subcutaneous tumors was maintained in orthotopic tumors. Metastatic spread was observed with orthotopic tumor implantation. In an orthotopic model of a sensitive tumor, primary and metastatic tumor burden was significantly reduced, and median survival significantly extended by Apo2L/TRAIL therapy. Conclusions Our data provide evidence that the site of tumor engraftment does not alter the inherent sensitivity of patient xenografts to Apo2L/TRAIL, and these results highlight the potential of Apo2L/TRAIL therapy against primary and metastatic pancreatic cancer.

Spatiotemporal Optoacoustic Mapping of Tumor Hemodynamics in a Clinically Relevant Orthotopic Rabbit Model of Head and Neck Cancer

The purpose of this study was to investigate the usefulness of photoacoustic imaging (PAI) for spatiotemporal mapping of tumor hemodynamics in a rabbit model of head and neck carcinoma. Shope cottontail rabbit papilloma virus associated VX2 carcinomas were established in adult male New Zealand White rabbits (n = 9) by surgical transplantation of tumor tissue in the neck. Noninvasive PAI with co-registered ultrasound (US) was performed to longitudinally monitor tumor growth, oxygen saturation (%sO2), and hemoglobin concentration (HbT). PAI findings were validated with Doppler sonography measures of percent vascularity (PV). Differences in tumor volumes, %sO2, HbT, and PV values over time were analyzed using repeated-measures analysis of variance with multiple comparisons. Two-tailed Spearman correlation analysis was performed to determine the correlation coefficient (r) for comparisons between %sO2, HbT, and tumor volume. US revealed a significant (P < .0001) increase in tumor volume over the 3-week period from 549 ± 260 mm3 on day 7 to 5055 ± 438 mm3 at 21 days postimplantation. Consistent with this aggressive tumor growth, PAI revealed a significant (P < .05) and progressive reduction in %sO2 from day 7 (37.6 ± 7.4%) to day 21 (9.5 ± 2.1%). Corresponding Doppler images also showed a decrease in PV over time. PAI revealed considerable intratumoral spatial heterogeneity with the tumor rim showing two- to three-fold higher %sO2 values compared to the core. Noninvasive PAI based on endogenous contrast provides a label-free method for longitudinal monitoring of temporal changes and spatial heterogeneity in thick head and neck tumors.

Standard Sub-Thermoneutral Caging Temperature Influences Radiosensitivity of Hematopoietic Stem and Progenitor Cells

The production of new blood cells relies on a hierarchical network of hematopoietic stem and progenitor cells (HSPCs). To maintain lifelong hematopoiesis, HSPCs must be protected from ionizing radiation or other cytotoxic agents. For many years, murine models have been a valuable source of information regarding factors that either enhance or reduce the survival of HSPCs after exposure of marrow to ionizing radiation. In a recent series of studies, however, it has become clear that housing-related factors such as the cool room temperature required for laboratory mice can exert a surprising influence on the outcome of experiments. Here we report that the mild, but chronic cold-stress endured by mice housed under these conditions exerts a protective effect on HSPCs after both non-lethal and lethal doses of total body irradiation (TBI). Alleviation of this cold-stress by housing mice at a thermoneutral temperature (30°C) resulted in significantly greater baseline radiosensitivity to a lethal dose of TBI with more HSPCs from mice housed at thermoneutral temperature undergoing apoptosis following non-lethal TBI. Cold-stressed mice have elevated levels of norepinephrine, a key molecule of the sympathetic nervous system that binds to β-adrenergic receptors. We show that blocking this signaling pathway in vivo through use of the β-blocker propanolol completely mitigates the protective effect of cold-stress on HSPC apoptosis. Collectively this study demonstrates that chronic stress endured by the standard housing conditions of laboratory mice increases the resistance of HSPCs to TBI-induced apoptosis through a mechanism that depends upon β-adrenergic signaling. Since β-blockers are commonly prescribed to a wide variety of patients, this information could be important when predicting the clinical impact of HSPC sensitivity to TBI.

Abstract 4946: Developing anti-angiogenic therapies for human hepatocellular cancer (HCC)- studies of suntinib in the woodchuck model of hepatitis B related HCC

Introduction: Better animal models that recapitulate the liver milieu of human HCC are needed. The eastern woodchuck is an established model of human hepatitis B viral infection and spontaneously develops HCC in the context of chronic woodchuck hepatitis viral infection (WHV). The translational relevance of this model for developing anti-angiogenic therapies was evaluated using sunitinib (S), a potent oral, anti-angiogenic agent. Methods: Woodchucks were bred and inoculated at birth with titered infectious WHV pools obtained from chronic WHV carriers. By 12 months of age, the rate of chronic WHV infection was >60%. Carriers were followed by USG, upon developing HCC, 12 animals were randomized 1:1 to S or placebo (P) given once orally daily for 30 days. From a single treatment S PK study at 4 dose levels, n=3/group, simulations showed 12mg/kg daily was expected to be optimal for achieving steady state serum concentrations between 50- 100 ng/ml in woodchucks. Tumor size and blood flow were assessed using dynamic contrast enhanced magnetic resonance imaging (DCE-MRI) before treatment and on day 28 using standardized protocols. At study completion or when animals were humanely euthanized, tumors and any other small nodules were fixed overnight in 10% buffered formalin. After standard processing and embedding in paraffin, 4 μm sections were prepared, deparaffinized, stained with hematoxylin-eosin (HE 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4946. doi:10.1158/1538-7445.AM2014-4946

Irradiance controls photodynamic efficacy and tissue heating in experimental tumours: implication for interstitial PDT of locally advanced cancer

BackgroundCurrently delivered light dose (J/cm2) is the principal parameter guiding interstitial photodynamic therapy (I-PDT) of refractory locally advanced cancer. The aim of this study was to investigate the impact of light dose rate (irradiance, mW/cm2) and associated heating on tumour response and cure.MethodsFinite-element modeling was used to compute intratumoural irradiance and dose to guide Photofrin® I-PDT in locally advanced SCCVII in C3H mice and large VX2 neck tumours in New Zealand White rabbits. Light-induced tissue heating in mice was studied with real-time magnetic resonance thermometry.ResultsIn the mouse model, cure rates of 70–90% were obtained with I-PDT using 8.4–245 mW/cm2 and ≥45 J/cm2 in 100% of the SCCVII tumour. Increasing irradiance was associated with increase in tissue heating. I-PDT with Photofrin® resulted in significantly (p < 0.05) higher cure rate compared to light delivery alone at same irradiance and light dose. Local control and/or cures of VX2 were obtained using I-PDT with 16.5–398 mW/cm2 and ≥45 J/cm2 in 100% of the tumour.ConclusionIn Photofrin®-mediated I-PDT, a selected range of irradiance prompts effective photoreaction with tissue heating in the treatment of locally advanced mouse tumour. These irradiances were translated for effective local control of large VX2 tumours.

Abstract 5144: Low dose sorafenib delays hepatocellular cancer (HCC) development in the woodchuck model of hepatitis B related HCC

Introduction: The eastern woodchuck is an established model of human hepatitis B viral infection and spontaneously develops HCC in the context of chronic woodchuck hepatitis viral infection (WHV) at a median age of 24 months. In this translational model of angiogenesis driven HCC, we evaluated the impact of sorafenib on delaying HCC development. Methods: Woodchucks were bred and inoculated at birth with titered infectious WHV pools obtained from chronic WHV carriers. By 12 months of age, the rate of chronic WHV infection was >60%. Animals were trained to receive drug orally mixed in a liquid diet. After an initial single dose woodchuck PK study it was predicted that sorafenib given orally M-F at 2.5 mg/kg once daily or 5mg/kg daily would achieve exposure in woodchucks ∼ 2x IC50 and 5X IC50 for all targets of sorafenib. This is within the exposure achieved in humans where the starting dose achieves exposures that are ∼ 10-12X IC50. We hypothesized that for both long term tolerability and delaying of HCC development, lower doses maybe ideal. Viral titers of WHV DNA were serially assessed, PK exposure of sorafenib and serial ultrasound (USG) was done q2 weeks to assess HCC development. Six WHV+ animals of similar age per arm were randomized to Placebo (P), low or high dose sorafenib. Time to tumor development (TTD) was measured from start of therapy to first occurrence of at least one 2 cm HCC was confirmed on 2 serial USGs. Tumor volume of all tumors seen was assessed as tumor burden. Results: Median TTD was 393 days, 490 and 498 days in the P, low and high dose sorafenib groups respectively. No toxicity was seen and PK results showed exposures as predicted in the two sorafenib groups. The tumor burden adjusted for the duration of therapy was significantly lower in the low dose group than the high dose and P groups. We evaluated three possible mechansims for this delay in HCC. WHV titers decreased as tumor burden increased as viral replication halts when malignant transformation occurs. The delay in TTD was not secondary to an antiviral effect that is known in this model and in humans to delay HCC occurrence. Anti-angiogenic effect was measured on H/E following necropsy as tumor necrosis and involved 7% of the total tumor volume in low dose and 13% of total tumor volume in high dose animals and 0% in P animals. High dose sorafenib inhibited the ex-vivo mixed lymphocyte reaction of woodchuck PBMC whereas low dose sorafenib did not which mechanistically may explain the difference seen in tumor burden with low compared to high dose sorafenib. Conclusions: The delay in HCC by 100 days seen in woodchucks would translate into a 3-9 years delay in HCC occurrence in humans. The tolerability at low dose for over 2 years, lower tumor burden and differential immune effects seen at low dose vs higher dose have great translational significance in the light of the recently completed STORM trial. Support: Sorafenib: Bayer. Funding: ACS- MSRG 08-096-01-CCE Citation Format: Renuka V. Iyer, Sandra Sexton, Leslie Curtin, Gerald Fetterly, Orla Maguire, Hans Minderman, Ilia Toshkov, Bud Tennant, Alan Hutson, Donald L. Trump, Candace Johnson. Low dose sorafenib delays hepatocellular cancer (HCC) development in the woodchuck model of hepatitis B related HCC. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5144. doi:10.1158/1538-7445.AM2015-5144

Abstract 4647: Investigation of the pharmacokinetic profile of the novel PIM2 inhibitor, JP_11646

PIM2 is a serine/threonine protein kinase that has roles in cell growth, proliferation, and apoptosis via the regulation of multiple signal transduction cascades. JP_11646 is a novel PIM2 inhibitor that has shown antitumor activity against breast, colon, liver, lung, and pancreatic solid tumors, as well as multiple myeloma and leukemia. The pharmacokinetics (PK) of JP_11646 has not been described. Therefore, a PK study was conducted to characterize JP_11646 disposition in plasma and various tissues. Normal ICR (CD-1) mice received either single day or five consecutive days of 15 - 25 mg/kg JP_11646 via intraperitoneal (IP) or intravenous (IV) administration. Plasma samples were collected at serial time points (0.5, 1, 2, 4, 6, and 8 hrs) following a single dose of 20 mg/kg IV or 25 mg/kg IP JP_11646 administration. Plasma and various tissues (brain, heart, kidney, lung, liver, muscle, colon, pancreas, and spleen) were collected at serial time points following multiple daily doses of 15 mg/kg IP JP_11646 on day five. Three mice were euthanized per time point on each collection day. JP_11646 concentrations were determined by a validated LC-MS/MS method, with a LLOQ of 1 ng/ml. Noncompartmental PK analysis was performed using Phoenix 64 (Pharsight, WinNonlin 6.3). Following single dose administration of JP_11646 of 20 (IV) and 25 mg/kg (IP), mean plasma Cmax was 6,951 and 6,630 ng/ml, respectively. Mean plasma drug exposures (AUCinf) were 11,065 and 11,024 ng-hr/ml, respectively, resulting in similar drug exposure between the IV and IP route. To compare plasma and tissue exposure following multiple dose administration, JP_11646 was given at 15 mg/kg IP. The mean plasma Cmax was 4,601 ng/ml., while the mean Cmax in colon, pancreas, and spleen were 24,523, 77,811, and 18,835 ng/ml, respectively. The half-life of JP_11646 in plasma and tissues ranged from 0.8 - 3 hrs. Tissue exposure in colon, pancreas, and spleen was 48,792, 157,273, and 40,511 ng-hr/ml, respectively. The pancreas to plasma partition coefficient (Kp) of 20.6 demonstrates that the pancreas receives the highest exposure to JP_11646. The drug also distributes to the colon and spleen, with Kp values of 6.4 and 5.3, respectively. JP_11646 PK appears linear and the half-life ranges from 0.8 - 3.0 hrs. JP_11646 distributes widely to tissues, with the highest drug exposure in the pancreas, spleen, and colon. These results give insight into tumor types with potential for optimal antitumor therapy with JP_11646. Citation Format: Laura B. Pitzonka, Allison Gaudy, Sarah Schihl, Leslie Curtin, Sandra Sexton, Carmen M. Baldino, Justin Caserta, Yvonne Flanders, Stephane Dumas, Gerald Fetterly. Investigation of the pharmacokinetic profile of the novel PIM2 inhibitor, JP_11646. [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 4647. doi:10.1158/1538-7445.AM2014-4647

Abstract 3372: Pharmacokinetic analysis of CFAK-C4 in dogs and estimation of first dose in man.

Purpose: Inhibition of the FAK-VEGFR-3 interaction by the small molecule inhibitor chloropyramine hydrochloride (CFAK-C4) has been shown to reduce tumor growth both alone and synergistically with doxorubicin and gemcitabine in an animal tumor model. A multiple dose study of CFAK-C4 in dogs was investigated to gain knowledge in estimating a first in man dose. Methods: CFAK-C4 was given to two female and two male dogs as an IV infusion at 1.25 or 2.5 mg/kg for five minutes, on three consecutive days, days 1, 2 and 3. Single- and multiple-dose PK samples were collected on days 1 and 3 of dosing. Plasma samples were collected at 0.5, 1, 2, 4, 8, and 24 hours post-dose. CFAK-C4 concentrations were determined using LC-MS/MS, with a LLOQ of 2.5 pg/ml. Noncompartmental PK analysis was performed using WinNonlin (Pharsight, version 5.3). Results: The mean CFAK-C4 plasma concentration-time profiles revealed a biexponential decline of drug following IV infusion, independent of day of dosing. The median (range) half-life was 3.42 hours (2.75-4.06 hours) and 3.79 (3.5-5.0) on days 1 and 3 following administration of 1.25 mg/kg CFAK-C4. At the 2.5 mg/kg dose level, the median half-life was similar. The median (range) Cmax of 1.25 mg/kg CFAK-C4 was determined to be 75 ng/ml (41-87 ng/ml) on day 1 and 99 ng/ml (84-102 ng/ml) on day 3. After the 2.5 mg/kg dose the Cmax was 185 ng/ml (165-244 ng/ml) and 171 ng/ml (153-181 ng/ml) on days 1 and 3 respectively. The AUC0-24 at the 1.25 mg/kg dose level was similar on both days 1 and 3 with a median (range) of 167 ng*hr/ml (117-231 ng*hr/ml) and 178 ng*hr/ml (170-194 ng*hr/ml), respectively. The AUC0-24 was also similar after the 2.5 mg/kg dose on days 1 and 3 with a median (range) of 447 ng*hr/ml (434-495 ng*hr/ml) and 406 ng*hr/ml (400-460 ng*hr/ml), respectively. The median (range) clearance for dose 1.25 mg/kg was 7.46 L/hr/kg (5.36-10.59 L/hr/kg) on day 1 and 6.47 L/hr/kg (6.07-6.91L/hr/kg) on day 3. For the 2.5 mg/kg dose level, the median (range) clearance was slightly lower compared with the 1.25 mg/kg dose, with values of 5.48 L/hr/kg (5.02-5.66 L/hr/kg) and 5.85 L/hr/kg (5.04-5.93 L/hr/kg) on days 1 and 3, respectively. Utilizing allometric scaling techniques of PK parameters from mice and dogs for CFAK-C4, the estimated human PK parameters for volume of distribution, clearance and half-life were 1695 L, 126 L/hr, and 9.34 hours, respectively. Subsequently, using a no observable adverse event level (NOAEL) of 44.5 mg/kg IP for mice and a NOAEL of 2.5 mg/kg IV for dog, the human starting dose (assuming a 70 kg human) would be a daily dose of 25 mg and 10 mg based on mouse or dog, respectively. Given that dogs are the most sensitive species, 10 mg daily is the suggested first dose in man for CFAK-C4. Conclusions: These results demonstrate that CFAK-C4 has linear kinetics and that no accumulation of drug occurs after multiple dosing. Allometric scaling of mouse and dog data allowed for the estimation of the first in man dose of CFAK-C4. Citation Format: Allison Gaudy, John Wilton, Leslie Curtin, Elena Kurenova, Sandra Buitrago, William Cance, Gerald Fetterly. Pharmacokinetic analysis of CFAK-C4 in dogs and estimation of first dose in man. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3372. doi:10.1158/1538-7445.AM2013-3372