Treatment planning for MR-guided SBRT of pancreatic tumors on a 1.5 T MR-Linac: A global consensus protocol.

Background and purpose: Treatment planning for MR-guided stereotactic body radiotherapy (SBRT) for pancreatic tumors can be challenging, leading to a wide variation of protocols and practices. This study aimed to harmonize treatment planning by developing a consensus planning protocol for MR-guided pancreas SBRT on a 1.5 T MR-Linac. Materials and methods: A consortium was founded of thirteen centers that treat pancreatic tumors on a 1.5 T MR-Linac. A phased planning exercise was conducted in which centers iteratively created treatment plans for two cases of pancreatic cancer. Each phase was followed by a meeting where the instructions for the next phase were determined. After three phases, a consensus protocol was reached. Results: In the benchmarking phase (phase I), substantial variation between the SBRT protocols became apparent (for example, the gross tumor volume (GTV) D99% ranged between 36.8 - 53.7 Gy for case 1, 22.6 - 35.5 Gy for case 2). The next phase involved planning according to the same basic dosimetric objectives, constraints, and planning margins (phase II), which led to a large degree of harmonization (GTV D99% range: 47.9-53.6 Gy for case 1, 33.9-36.6 Gy for case 2). In phase III, the final consensus protocol was formulated in a treatment planning system template and again used for treatment planning. This not only resulted in further dosimetric harmonization (GTV D99% range: 48.2-50.9 Gy for case 1, 33.5-36.0 Gy for case 2) but also in less variation of estimated treatment delivery times. Conclusion: A global consensus protocol has been developed for treatment planning for MR-guided pancreatic SBRT on a 1.5 T MR-Linac. Aside from harmonizing the large variation in the current clinical practice, this protocol can provide a starting point for centers that are planning to treat pancreatic tumors on MR-Linac systems.

Local Control Following Stereotactic Body Radiation Therapy for Liver Oligometastases: Lessons from a Quarter Century.

The utilization of stereotactic body radiation therapy for the treatment of liver metastasis has been widely studied and has demonstrated favorable local control outcomes. However, several predictive factors play a crucial role in the efficacy of stereotactic body radiation therapy, such as the number and size (volume) of metastatic liver lesions, the primary tumor site (histology), molecular biomarkers (e.g., KRAS and TP53 mutation), the use of systemic therapy prior to SBRT, the radiation dose, and the use of advanced technology and organ motion management during SBRT. These prognostic factors need to be considered when clinical trials are designed to evaluate the efficacy of SBRT for liver metastases.

Inflammatory Cytokines and Radiotherapy in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) remains a therapeutic challenge in clinical oncology. Surgery is the only potentially curative treatment. However, the majority of PDAC patients present with locally advanced/unresectable or metastatic disease, where palliative multiagent chemotherapy is the first-line treatment with the therapeutic intent to delay progression and prolong survival. For locally advanced/unresectable pancreatic cancer patients who are treated with chemotherapy, consolidative radiotherapy in the form concurrent chemoradiation or stereotactic ablative radiotherapy improves locoregional control and pain/symptom control. To improve clinical outcomes of PDAC patients, there is a dire need for discoveries that will shed more light on the pathophysiology of the disease and lead to the development of more efficacious treatment strategies. Inflammatory cytokines are known to play a role in mediating tumor progression, chemoresistance, and radioresistance in PDAC. A PubMed search on published articles related to radiotherapy, inflammatory cytokines, and pancreatic cancer patients in the English language was performed. This article primarily focuses on reviewing the clinical literature that examines the association of inflammatory cytokines with clinical outcomes and the effects of radiotherapy on inflammatory cytokines in PDAC patients.

Benchmarking Outcomes after Ablative Radiotherapy for Molecularly Characterized Intrahepatic Cholangiocarcinoma.

We have previously shown that ablative radiotherapy (A-RT) with a biologically effective dose (BED10) ≥ 80.5 Gy for patients with unresectable intrahepatic cholangiocarcinoma (ICC) is associated with longer survival. Despite recent large-scale sequencing efforts in ICC, outcomes following RT based on genetic alterations have not been described. We reviewed records of 156 consecutive patients treated with A-RT for unresectable ICC from 2008 to 2020. For 114 patients (73%), next-generation sequencing provided molecular profiles. The overall survival (OS), local control (LC), and distant metastasis-free survival (DMFS) were estimated using the Kaplan-Meier method. Univariate and multivariable Cox analyses were used to determine the associations with the outcomes. The median tumor size was 7.3 (range: 2.2-18.2) cm. The portal vein thrombus (PVT) was present in 10%. The RT median BED10 was 98 Gy (range: 81-144 Gy). The median (95% confidence interval) follow-up was 58 (42-104) months from diagnosis and 39 (33-74) months from RT. The median OS was 32 (29-35) months after diagnosis and 20 (16-24) months after RT. The one-year OS, LC, and intrahepatic DMFS were 73% (65-80%), 81% (73-87%), and 34% (26-42%). The most common mutations were in IDH1 (25%), TP53 (22%), ARID1A (19%), and FGFR2 (13%). Upon multivariable analysis, the factors associated with death included worse performance status, larger tumor, metastatic disease, higher CA 19-9, PVT, satellitosis, and IDH1 and PIK3CA mutations. TP53 mutation was associated with local failure. Further investigation into the prognostic value of individual mutations and combinations thereof is warranted.

Dosimetric Uncertainties Resulting From Interfractional Anatomic Variations for Patients Receiving Pancreas Stereotactic Body Radiation Therapy and Cone Beam Computed Tomography Image Guidance.

PURPOSE: To estimate the effects of interfractional anatomic changes on dose to organs at risk (OARs) and tumors, as measured with cone beam computed tomography (CBCT) image guidance for pancreatic stereotactic body radiation therapy. METHODS AND MATERIALS: We evaluated 11 patients with pancreatic cancer whom were treated with stereotactic body radiation therapy (33-40 Gy in 5 fractions) using daily CT-on-rails (CTOR) image guidance immediately before treatment with breath-hold motion management. CBCT alignment was simulated in the treatment planning software by aligning the original planning CT to each fractional CTOR image set via fiducial markers. CTOR data sets were used to calculate fractional doses after alignment by applying the rigid shift of the planning CT and CTOR image sets to the planning treatment isocenter and recalculating the fractional dose. Accumulated dose to the gross tumor volume (GTV), tumor vessel interface, duodenum, small bowel, and stomach were calculated by summing the 5 fractional absolute dose-volume histograms into a single dose-volume histogram for comparison with the original planned dose. RESULTS: Four patients had a GTV D100% of at least 1.5 Gy less than the fractional planned value in several fractions; 4 patients had fractional underestimation of duodenum dose by 1.0 Gy per fraction. The D1.0 cm3 <35 Gy constraint was violated for at least 1 OAR in 3 patients, with either the duodenum (n = 2) or small bowel (n = 1) D1.0 cm3 being higher on the accumulated dose distribution (P = .01). D100% was significantly lower according to accumulated dose GTV (P = .01) and tumor vessel interface (P = .02), with 4 and 2 patients having accumulated D100%  ≥4 Gy lower than the planned value for the GTV and tumor vessel interface, respectively. CONCLUSIONS: For some patients, CBCT image guidance based on fiducial alignment may cause large dosimetric uncertainties for OARs and target structures, according to accumulated dose.

Plasma metabolomic profiles in liver cancer patients following stereotactic body radiotherapy.

BACKGROUND: Stereotactic body radiotherapy (SBRT) is an effective treatment for hepatocellular carcinoma (HCC). This study sought to identify differentially expressed plasma metabolites in HCC patients at baseline and early during SBRT, and to explore if changes in these metabolites early during SBRT may serve as biomarkers for radiation-induced liver injury and/or tumour response. METHODS: Forty-seven HCC patients were treated with SBRT on previously published prospective trials. Plasma samples were collected at baseline and after one to two fractions of SBRT, and analysed by GC/MS and LC/MS for untargeted and targeted metabolomics profiling, respectively. FINDINGS: Sixty-nine metabolites at baseline and 62 metabolites after one to two fractions of SBRT were differentially expressed, and strongly separated the Child Pugh (CP) B from the CP A HCC patients. These metabolites are associated with oxidative stress and alterations in hepatic cellular metabolism. Differential upregulation of serine, alanine, taurine, and lipid metabolites early during SBRT from baseline was noted in the HCC patients who demonstrated the greatest increase in CP scores at three months post SBRT, suggesting that high protein and lipid turnover early during SBRT may portend increased clinical liver toxicity. Twenty annotated metabolites including fatty acids, glycerophospholipids, and acylcarnitines were differentially upregulated early during SBRT from baseline and separated patients with complete/partial response from those with stable disease at three months post SBRT. INTERPRETATION: Dysregulation of amino acid and lipid metabolism detected early during SBRT are associated with subsequent clinical liver injury and tumour response in HCC.

Single-Fraction Stereotactic Body Radiation Therapy: A Paradigm During the Coronavirus Disease 2019 (COVID-19) Pandemic and Beyond?

PURPOSE: Owing to the coronavirus disease 2019 (COVID-19) pandemic, radiation oncology departments have adopted various strategies to deliver radiation therapy safely and efficiently while minimizing the risk of severe acute respiratory syndrome coronavirus-2 transmission among patients and health care providers. One practical strategy is to deliver stereotactic body radiation therapy (SBRT) in a single fraction, which has been well established for treating bone metastases, although it has been infrequently used for other extracranial sites. METHODS AND MATERIALS: A PubMed search of published articles in English related to single-fraction SBRT was performed. A critical review was performed of the articles that described clinical outcomes of single-fraction SBRT for treatment of primary extracranial cancers and oligometastatic extraspinal disease. RESULTS: Single-fraction SBRT for peripheral early-stage non-small cell lung cancer is supported by randomized data and is strongly endorsed during the COVID-19 pandemic by the European Society for Radiotherapy and Oncology-American Society for Radiation Oncology practice guidelines. Prospective and retrospective studies supporting a single-fraction regimen are limited, although outcomes are promising for renal cell carcinoma, liver metastases, and adrenal metastases. Data are immature for primary prostate cancer and demonstrate excess late toxicity in primary pancreatic cancer. CONCLUSIONS: Single-fraction SBRT should be strongly considered for peripheral early-stage non-small cell lung cancer during the COVID-19 pandemic to mitigate the potentially severe consequences of severe acute respiratory syndrome coronavirus-2 transmission. Although single-fraction SBRT is promising for the definitive treatment of other primary or oligometastatic cancers, multi-fraction SBRT should be the preferred regimen owing to the need for additional prospective evaluation to determine long-term efficacy and safety.

Association of pro-inflammatory soluble cytokine receptors early during hepatocellular carcinoma stereotactic radiotherapy with liver toxicity.

Plasma levels of soluble factors early during hepatocellular carcinoma (HCC) stereotactic body radiotherapy (SBRT) were evaluated in relation to radiation liver injury, tumor response, and risk of early death. No significant differences were found in baseline plasma levels of AFP, CXCL1, and HGF amongst HCC patients with different Child Pugh scores. Higher levels of sTNFRII (P < 0.001), and lower levels of sCD40L (P < 0.001) and CXCL1 (P = 0.01) following one to two fractions of SBRT were noted in patients who developed liver toxicity vs. those who did not. High circulating levels of AFP (HR 2.16, P = 0.04), sTNFRII (HR 2.27, P = 0.01), and sIL-6R (HR 1.99, P = 0.03) early during SBRT were associated with increased risk of death 3 months post treatment. Plasma levels of the studied factors early during SBRT were not associated with tumor response. A pro-inflammatory systemic environment is associated with development of liver toxicity and increased risk of early death following SBRT.

The differential effects of metronomic gemcitabine and antiangiogenic treatment in patient-derived xenografts of pancreatic cancer: treatment effects on metabolism, vascular function, cell proliferation, and tumor growth.

BACKGROUND: Metronomic chemotherapy has shown promising activity against solid tumors and is believed to act in an antiangiogenic manner. The current study describes and quantifies the therapeutic efficacy, and mode of activity, of metronomic gemcitabine and a dedicated antiangiogenic agent (DC101) in patient-derived xenografts of pancreatic cancer. METHODS: Two primary human pancreatic cancer xenograft lines were dosed metronomically with gemcitabine or DC101 weekly. Changes in tumor growth, vascular function, and metabolism over time were measured with magnetic resonance imaging, positron emission tomography, and immunofluorescence microscopy to determine the anti-tumor effects of the respective treatments. RESULTS: Tumors treated with metronomic gemcitabine were 10-fold smaller than those in the control and DC101 groups. Metronomic gemcitabine, but not DC101, reduced the tumors' avidity for glucose, proliferation, and apoptosis. Metronomic gemcitabine-treated tumors had higher perfusion rates and uniformly distributed blood flow within the tumor, whereas perfusion rates in DC101-treated tumors were lower and confined to the periphery. DC101 treatment reduced the tumor's vascular density, but did not change their function. In contrast, metronomic gemcitabine increased vessel density, improved tumor perfusion transiently, and decreased hypoxia. CONCLUSION: The aggregate data suggest that metronomic gemcitabine treatment affects both tumor vasculature and tumor cells continuously, and the overall effect is to significantly slow tumor growth. The observed increase in tumor perfusion induced by metronomic gemcitabine may be used as a therapeutic window for the administration of a second drug or radiation therapy. Non-invasive imaging could be used to detect early changes in tumor physiology before reductions in tumor volume were evident.

The involvement of insulin-like growth factor 2 binding protein 3 (IMP3) in pancreatic cancer cell migration, invasion, and adhesion.

BACKGROUND: Over-expression of insulin-like growth factor 2 mRNA binding protein 3 (IMP3) is correlated with poor prognosis in pancreatic ductal adenocarcinoma (PDAC). Previous studies examining other cancer types have implicated IMP3 in the regulation of several cellular functions that are characteristic of tumour cells. However, the role of this oncofetal protein in PDAC progression remained unclear. METHODS: Using siRNA, we examined the effect of IMP3 inhibition on the motility, invasive ability, and matrix adhesion of PDAC cells. In addition, we also evaluated the expression of cytoskeleton-associated genes following IMP depletion. RESULTS: Knockdown of IMP3 significantly decreased the motility, invasion, and extracellular matrix adhesion of select PDAC cells in vitro. In addition, IMP3-depleted cells exhibited lower levels of CD44 protein and KIF11 mRNA. Moreover, we also observed a reduction in downstream RhoA signaling following IMP3 knockdown, indicating that IMP3 modulates the levels of proteins involved in cytoskeletal organization. CONCLUSIONS: These results suggest that IMP3 facilitates PDAC progression by enhancing the pro-metastatic behaviour of tumour cells.

Irinophore C™, a lipid nanoparticulate formulation of irinotecan, improves vascular function, increases the delivery of sequentially administered 5-FU in HT-29 tumors, and controls tumor growth in patient derived xenografts of colon cancer.

PURPOSE: A liposomal formulation of irinotecan, Irinophore C™ (IrC™) is efficacious in a panel of tumor models, normalizes tumor vasculature, and increases the accumulation of a second drug in the same tumor. We now show that Irinophore C™ is also effective against patient derived xenografts (PDX) of colon cancer, and examine the kinetics of vascular normalization in the HT-29 tumor model and assess how these changes might be used with 5-FU sequentially. MATERIALS AND METHODS: Rag2M mice bearing HT-29 tumors were treated with IrC™ (25mg/kg; Q7D×3) for up to three weeks. Groups of tumors were harvested for analysis at 7, 14 and 21days after the start of treatment. Drug and lipid levels in the tumor were evaluated using HPLC and scintillation counts, respectively. Changes in tumor morphology (H&E), vasculature (CD31), perfusion (Hoechst 33342) and apoptosis (TUNEL) were quantified using microscopy. The accumulation of a second drug ([(14)C]-5-FU, 40mg/kg) given 3h before sacrifice was determined using liquid scintillation. The efficacy of IrC™ (Q7D×3) followed by 5-FU treatment (Q7D×3) was assessed in mice bearing established HT-29 tumors. The efficacy of IrC™ was also evaluated in primary human colorectal tumors grown orthotopically in NOD-SCID mice. RESULTS: Following a single dose of IrC™ the active lactone forms of irinotecan and its metabolite SN-38 were measurable in HT-29 tumors after 7days. The treatment reduced tumor cell density and increased apoptosis. Hoechst 33342 perfusion and accumulation of [(14)C]-5-FU in the treated tumors increased significantly on days 7 and 14. This was accompanied by an increase in the number of endothelial cells relative to total nuclei in the tumor sections. Pre-treatment with IrC™ (Q7D×3) followed by 5-FU (Q7D×3) delayed the time taken for tumors to reach 1cm(3) by 9days (p<0.05). IrC™ was just as effective as free irinotecan when used on patient derived xenografts of colorectal cancer. CONCLUSIONS: Treatment with IrC™ reduces tumor cell viability and appears to normalize the vascular function of the tumor after a single treatment cycle. A concomitant increase in the accumulation of a second drug (5-FU) in the tumor was observed in tumors from IrC™ treated animals and this was correlated with changes in vascular structure consistent with normalization. The treatment effects of sequential 5-FU dosing following IrC™ are additive with no additional toxicity in contrast to previous studies where concurrent 5-FU and IrC™ treatment exacerbated 5-FU toxicity. The studies with PDX tumors also indicate that IrC™ is just as effective as free irinotecan on PDX tumors even though the delivered dose is halved.

B7-H4 expression in normal and diseased human islet ß cells.

OBJECTIVES: B7-H4 is a negative coregulatory molecule known to be involved in immune response. We study here B7-H4 expression and its possible role in diabetes and cancer development. METHODS: Formalin-fixed, paraffin-processed pancreas samples from patients with type 1 diabetes (T1D), insulinoma, pancreatic ductal adenocarcinoma (PDAC), and normal organ donors were studied by bright-field and multifluorescence immunohistochemistry to examine B7-H4 expression and its colocalization with islet endocrine hormones. Quantitative RT-PCR and Western blot assay were used to examine B7-H4 mRNA and protein expression in the islet and exocrine tissues from normal donors and pancreatic cancer cell lines. RESULTS: B7-H4 protein expression in islet ß cells is decreased in T1D and PDAC, but increased in insulinoma patients when compared to normal controls; the changes in B7-H4 expression are concomitant with insulin expression on the islet ß cells. The insulin/B7-H4 colocalization on the ß cells, expressed in colocalization coefficient Pearson r, is also changed in these islets. CONCLUSIONS: Our observation of altered B7-H4 expression, concomitant with insulin expression, in the pancreatic islets of T1D, PDAC, and insulinoma patients when compared to normal controls suggests that B7-H4 pathway might play an important role in maintenance of ß-cell function, but its exact role remains to be explored.

The autophagy inhibitor verteporfin moderately enhances the antitumor activity of gemcitabine in a pancreatic ductal adenocarcinoma model.

Pancreatic ductal adenocarcinoma (PDAC) is highly resistant to chemotherapy. It has been described as requiring elevated autophagy for growth and inhibiting autophagy has been proposed as a treatment strategy. To date, all preclinical reports and clinical trials investigating pharmacological inhibition of autophagy have used chloroquine or hydroxychloroquine, which interfere with lysosomal function and block autophagy at a late stage. Verteporfin is a newly discovered autophagy inhibitor that blocks autophagy at an early stage by inhibiting autophagosome formation. Here we report that PDAC cell lines show variable sensitivity to verteporfin in vitro, suggesting cell-line specific autophagy dependence. Using image-based and molecular analyses, we show that verteporfin inhibits autophagy stimulated by gemcitabine, the current standard treatment for PDAC. Pharmacokinetic and efficacy studies in a BxPC-3 xenograft mouse model demonstrated that verteporfin accumulated in tumors at autophagy-inhibiting levels and inhibited autophagy in vivo, but did not reduce tumor volume or increase survival as a single agent. In combination with gemcitabine verteporfin moderately reduced tumor growth and enhanced survival compared to gemcitabine alone. While our results do not uphold the premise that autophagy inhibition might be widely effective against PDAC as a single-modality treatment, they do support autophagy inhibition as an approach to sensitize PDAC to gemcitabine.

Evaluation of (111)In labeled antibodies for SPECT imaging of mesothelin expressing tumors.

INTRODUCTION: Mesothelin is expressed in many cancers, especially in mesothelioma and lung, pancreatic and ovarian cancers. In the present study, we evaluate (111)In labeled antimesothelin antibodies as an imaging bioprobe for the SPECT imaging of mesothelin-expressing tumors. METHODS: We radiolabeled the antimesothelin antibodies mAbMB and mAbK1 with (111)In using the p-SCN-bn-DTPA chelator. The immunoreactivity, affinity (K(d)) and internalization properties of the resulting two (111)In labeled antibodies were evaluated in vitro using mesothelin-expressing A431K5 cells. The biodistribution and microSPECT/CT imaging studies with (111)In labeled antibodies were performed in mice bearing both mesothelin positive (A431K5) and mesothelin negative (A431) tumors. RESULTS: In vitro studies demonstrated that (111)In-mAbMB bound with a higher affinity (K(d)=3.6±1.7 nM) to the mesothelin-expressing A431K5 cells than did the (111)In-mAbK1 (K(d)=29.3±2.3 nM). (111)In-mAbMB was also internalized at a greater rate and extent into the A431K5 cells than was the (111)In-mAbK1. Biodistribution studies showed that (111)In-mAbMB was preferentially localized in A431K5 tumors when compared to A431 tumors. At the low dose, the peak A431K5 tumor uptake of 9.65±2.65% ID/g (injected dose per gram) occurred at 48 h, while at high dose tumor uptake peaked with 14.29±6.18% ID/g at 72 h. Non-specific localization of (111)In-mAbMB was mainly observed in spleen.(111)In-mAbK1 also showed superior localization in A431K5 tumors than in A431 tumors, but the peak uptake was only 3.04±0.68% ID/g at 24 h. MicroSPECT/CT studies confirmed better visualization of A431K5 tumors with (111)In-mAbMB, than with (111)In-mAbK1. CONCLUSION: SPECT imaging of mesothelin expressing tumors was demonstrated successfully. Our findings indicate that the antimesothelin antibody mAbMB has the potential to be developed into a diagnostic agent for imaging mesothelin-expressing cancers.

Comparison of bifunctional chelates for (64)Cu antibody imaging.

PURPOSE: Improved bifunctional chelates (BFCs) are needed to facilitate efficient (64)Cu radiolabeling of monoclonal antibodies (mAbs) under mild conditions and to yield stable, target-specific agents. The utility of two novel BFCs, 1-Oxa-4,7,10-triazacyclododecane-5-S-(4-isothiocyanatobenzyl)-4,7,10-triacetic acid (p-SCN-Bn-Oxo-DO3A) and 3,6,9,15-tetraazabicyclo[9.3.1]pentadeca-1(15),11,13-triene-4-S-(4-isothiocyanatobenzyl)-3,6,9-triacetic acid (p-SCN-Bn-PCTA), for mAb imaging with (64)Cu were compared to the commonly used S-2-(4-isothiocyanatobenzyl)-1,4,7,10-tetraazacyclododecane-tetraacetic acid (p-SCN-Bn-DOTA). METHODS: The BFCs were conjugated to trastuzumab, which targets the HER2/neu receptor. (64)Cu radiolabeling of the conjugates was optimized. Receptor binding was analyzed using flow cytometry and radioassays. Finally, PET imaging and biodistribution studies were done in mice bearing either HER2/neu-positive or HER2/neu-negative tumors. RESULTS: (64)Cu-Oxo-DO3A- and PCTA-trastuzumab were prepared at room temperature in >95% radiochemical yield (RCY) in <30 min, compared to only 88% RCY after 2 h for the preparation of (64)Cu-DOTA-trastuzumab under the same conditions. Cell studies confirmed that the immunoreactivity of the mAb was retained for each of the bioconjugates. In vivo studies showed that (64)Cu-Oxo-DO3A- and PCTA-trastuzumab had higher uptake than the (64)Cu-DOTA-trastuzumab at 24 h in HER2/neu-positive tumors, resulting in higher tumor to background ratios and better tumor images. By 40 h all three of the (64)Cu-BFC-trastuzumab conjugates allowed for clear visualization of the HER2/neu-positive tumors but not the negative control tumor. CONCLUSION: The antibody conjugates of PCTA and Oxo-DO3A were shown to have superior (64)Cu radiolabeling efficiency and stability compared to the analogous DOTA conjugate. In addition, (64)Cu-PCTA and Oxo-DO3A antibody conjugates may facilitate earlier imaging with greater target to background ratios than the analogous (64)Cu-DOTA antibody conjugates.

Insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3) overexpression in pancreatic ductal adenocarcinoma correlates with poor survival.

BACKGROUND: Pancreatic ductal adenocarcinoma is a lethal disease with a 5-year survival rate of 4% and typically presents in an advanced stage. In this setting, prognostic markers identifying the more aggressive tumors could aid in management decisions. Insulin-like growth factor 2 mRNA binding protein 3 (IGF2BP3, also known as IMP3 or KOC) is an oncofetal RNA-binding protein that regulates targets such as insulin-like growth factor-2 (IGF-2) and ACTB (beta-actin). METHODS: We evaluated the expression of IGF2BP3 by immunohistochemistry using a tissue microarray of 127 pancreatic ductal adenocarcinomas with tumor grade 1, 2 and 3 according to WHO criteria, and the prognostic value of IGF2BP3 expression. RESULTS: IGF2BP3 was found to be selectively overexpressed in pancreatic ductal adenocarcinoma tissues but not in benign pancreatic tissues. Nine (38%) patient samples of tumor grade 1 (n = 24) and 27 (44%) of tumor grade 2 (n = 61) showed expression of IGF2BP3. The highest rate of expression was seen in poorly differentiated specimen (grade 3, n = 42) with 26 (62%) positive samples. Overall survival was found to be significantly shorter in patients with IGF2BP3 expressing tumors (P = 0.024; RR 2.3, 95% CI 1.2-4.8). CONCLUSIONS: Our data suggest that IGF2BP3 overexpression identifies a subset of pancreatic ductal adenocarcinomas with an extremely poor outcome and supports the rationale for developing therapies to target the IGF pathway in this cancer.

Irinophore C, a novel nanoformulation of irinotecan, alters tumor vascular function and enhances the distribution of 5-fluorouracil and doxorubicin.

PURPOSE: To examine the antitumor effects of Irinophore C, a nanopharmaceutical formulation of irinotecan, on the tissue morphology and function of tumor vasculature in HT-29 human colorectal tumors. EXPERIMENTAL DESIGN: Fluorescence microscopy was used to map and quantify changes in tissue density, tumor vasculature, hypoxia, and the distribution of Hoechst 33342, a perfusion marker, and the anticancer drug, doxorubicin. Noninvasive magnetic resonance imaging was used to quantify Ktrans, the volume transfer constant of a solute between the blood vessels and extracellular tissue compartment of the tumor, as a measure of vascular function. Following treatment with Irinophore C, 19F magnetic resonance spectroscopy was used to monitor the delivery of 5-fluorouracil (5-FU) to the tumor tissue, whereas scintigraphy was used to quantify the presence of bound [14C]5-FU. RESULTS: Irinophore C decreased cell density (P = 8.42 x 10(-5)), the overall number of endothelial cells in the entire section (P = 0.014), tumor hypoxia (P = 5.32 x 10(-9)), and K(trans) (P = 0.050). However, treatment increased the ratio of endothelial cells to cell density (P = 0.00024) and the accumulation of Hoechst 33342 (P = 0.022), doxorubicin (P = 0.243 x 10(-5)), and 5-FU (P = 0.0002) in the tumor. Vascular endothelial growth factor and interleukin-8, two proangiogenic factors, were down-regulated, whereas the antiangiogenic factor TIMP-1 was up-regulated in Irinophore C-treated tumors. CONCLUSIONS: Irinophore C treatment improves the vascular function of the tumor, thereby reducing tumor hypoxia and increasing the delivery and accumulation of a second drug. Reducing hypoxia would enhance radiotherapy, whereas improving delivery of a second drug to the tumor should result in higher cell kill.

Assessment of vessel size by MRI in an orthotopic model of human pancreatic cancer.

Pancreatic cancer is a devastating disease with no cure. Therapies that target the tumor vasculature are promising new treatment strategies. Magnetic resonance imaging (MRI) can non-invasively determine a vessel size index and a blood volume fraction to characterize the vascular compartment in a tumor. The changes in the T2 and T2* relaxation rate constants after the administration of superparamagnetic iron oxide (SPIO) particles are dependent on the size and morphology of tissue blood vessels. In this study, MRI was used to investigate changes in the tumor vasculature in an orthotopic primary human pancreatic cancer xenograft model during tumor progression. The SPIO contrast agent Feridex I.V. was first validated as an intravascular contrast agent over the course of the imaging session, and shown to remain in the blood for at least 1.5 h. The average vessel size index was not correlated to the tumor area within an image slice, but the average blood volume fraction was significantly and negatively correlated to the tumor area (p<0.05). Blood volume fraction may serve as a non-invasive biomarker for changes in the tumor vasculature due to tumor growth Further investigation is needed to evaluate this promising technique as a tool to monitor tumor vascular changes in response to antiangiogenic therapies in pancreatic cancer.

Influence of formulation vehicle on metronomic taxane chemotherapy: albumin-bound versus cremophor EL-based paclitaxel.

PURPOSE: Low-dose metronomic chemotherapy treatments, especially when combined with 'dedicated' antiangiogenic agents, can induce significant antitumor activity without serious toxicity in various preclinical models. It remains unclear, however, whether some cytotoxic drugs are better suited for metronomic regimens than others. Paclitaxel appears to be a strong candidate for metronomic chemotherapy given its ability to inhibit endothelial cell functions relevant to angiogenesis in vitro at extraordinarily low concentrations and broad-spectrum antitumor activity. Clinically relevant concentrations of the formulation vehicle cremophor EL in Taxol, however, were previously reported to nullify the antiangiogenic effect of paclitaxel, the result of which would hamper its usefulness in metronomic regimens. We hypothesized that ABI-007, a cremophor EL-free, albumin-bound, 130-nm form of paclitaxel, could potentially alleviate this problem. EXPERIMENTAL DESIGN: The antiangiogenic activity of ABI-007 was assessed by multiple in vitro assays. The in vivo optimal dose of ABI-007 for metronomic chemotherapy was determined by measuring circulating endothelial progenitors in peripheral blood. The antitumor effects of metronomic and maximum tolerated dose ABI-007 and Taxol were then evaluated and compared in severe combined immunodeficient mice bearing human MDA-MD-231 breast cancer and PC3 prostate cancer xenografts. RESULTS: ABI-007 significantly inhibited rat aortic microvessel outgrowth, human endothelial cell proliferation, and tube formation. The optimal metronomic dose of ABI-007 was determined to be between 3 and 10 mg/kg. Metronomic ABI-007 but not Taxol, significantly suppressed tumor growth in both xenograft models. Furthermore, the antitumor effect of minimally toxic metronomic ABI-007 approximated that of the maximum tolerated dose of Taxol. CONCLUSIONS: Our results underscore the influence of formulation vehicles on the selection of cytotoxic drugs for metronomic chemotherapy.

Antitumor effects of thalidomide analogs in human prostate cancer xenografts implanted in immunodeficient mice.

PURPOSE: Thalidomide has demonstrated clinical activity in various malignancies including androgen-independent prostate cancer. The development of novel thalidomide analogs with better activity/toxicity profiles is an ongoing research effort. Our laboratory previously reported the in vitro antiangiogenic activity of the N-substituted thalidomide analog CPS11 and the tetrafluorinated analogs CPS45 and CPS49. The current study evaluated the therapeutic potential of these analogs in the treatment of prostate cancer in vivo. EXPERIMENTAL DESIGN: Severely combined immunodeficient mice bearing s.c. human prostate cancer (PC3 or 22Rv1) xenografts were treated with the analogs at their maximum tolerated doses. Tumors were then excised and processed for ELISA and CD31 immunostaining to determine the levels of various angiogenic factors and microvessel density (MVD), respectively. RESULTS: CPS11, CPS45, and CPS49 induced prominent and modest growth inhibition in PC3 and 22Rv1 tumors, respectively. Thalidomide had no effect on tumor growth in either xenograft. Vascular endothelial growth factor and basic fibroblast growth factor levels were not significantly altered by any of the thalidomide analogs or thalidomide in both PC3 and 22Rv1 tumors. CPS45, CPS49, and thalidomide significantly reduced PC3 tumor platelet-derived growth factor (PDGF)-AA levels by 58-82% (P < 0.05). Interestingly, treatment with the analogs and thalidomide resulted in differential down-regulation (>/=1.5-fold) of genes encoding PDGF and PDGF receptor isoforms as determined by DNA microarray analysis. Intratumoral MVD of 22Rv1 xenografts was significantly decreased by CPS45 and CPS49. CPS49 also reduced MVD in PC3 xenografts. CONCLUSIONS: Thalidomide analogs CPS11 and 49 are promising anti-cancer agents. PDGF signaling pathway may be a potential target for these thalidomide analogs. Detailed microarray and functional analyses are under way with the aim of elucidating the molecular mechanism(s) of action of these thalidomide analogs.