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International audience; Pancreatic cancer is currently the third leading cause of cancer death in the United States. The clinical hallmarks of this disease include abdominal pain that radiates to the back, the presence of a hypoenhancing intrapancreatic lesion on imaging, and widespread liver metastases. Technologies such as tissue clearing and three-dimensional (3D) reconstruction of digitized serially sectioned hematoxylin and eosin stained slides, can be used to visualize large (up to two to three-centimeter cube) tissues at cellular resolution. When applied to human pancreatic cancers, these 3D visualization techniques have provided novel insights into the basis of a number of the clinical hallmarks of this disease. Here we describe the clinical features of pancreatic cancer, review techniques for clearing and the 3D reconstruction of digitized microscope slides, and provide examples that illustrate how 3D visualization of human pancreatic cancer at the microscopic level has closed the gap between bench and bedside. Compared to animal models and 2D microscopy, studies of human tissues in 3D can reveal the difference between what can happen and what does happen in human cancers.

The treatment of metastatic pancreatic ductal adenocarcinoma (PDAC) is frequently characterized by significant toxicity and rapid development of resistance to current approved therapies. More reliable biomarkers of response are needed to guide clinical decision making. We evaluated cell-free DNA (cfDNA) using a tumor-agnostic platform and traditional biomarkers (CEA and CA19-9) levels in 12 patients treated at Johns Hopkins University on NCT02324543 “Study of Gemcitabine/Nab-Paclitaxel/Xeloda (GAX) in Combination With Cisplatin and Irinotecan in Subjects With Metastatic Pancreatic Cancer.” The pretreatment values, levels after 2 months of treatment, and change in biomarker levels with treatment were compared with clinical outcomes to determine their predictive value. The variant allele frequency (VAF) of KRAS and TP53 mutations in cfDNA after 2 months of treatment was predictive of progression-free survival (PFS) and overall survival (OS). In particular, patients with a lower-than-average KRAS VAF after 2 months of treatment had a substantially longer PFS than patients with higher posttreatment KRAS VAF (20.96 vs. 4.39 months). Changes in CEA and CA19-9 after 2 months of treatment were also good predictors of PFS. Comparison via concordance index demonstrated KRAS or TP53 VAF after 2 months of treatment to be better predictors of PFS and OS than CA19-9 or CEA. This pilot study requires validation but suggests cfDNA measurement is a useful adjunct to traditional protein biomarkers and imaging evaluation and could distinguish between patients who are likely to achieve prolonged responses versus those that will have early progression and may benefit from a change in treatment approach. Significance: We report on the association of cfDNA with response durability for patients undergoing treatment with a novel metronomic chemotherapy regimen (gemcitabine, nab-paclitaxel, capecitabine, cisplatin, irinotecan; GAX-CI) for metastatic PDAC. This investigation offers encouraging evidence that cfDNA may prove to be a valuable diagnostic tool to guide clinical management.

Purpose: To assess the preclinical efficacy, clinical safety and efficacy, and MTD of palbociclib plus nab-paclitaxel in patients with advanced pancreatic ductal adenocarcinoma (PDAC). Experimental Design: Preclinical activity was tested in patient-derived xenograft (PDX) models of PDAC. In the open-label, phase I clinical study, the dose-escalation cohort received oral palbociclib initially at 75 mg/day (range, 50‒125 mg/day; modified 3+3 design; 3/1 schedule); intravenous nab-paclitaxel was administered weekly for 3 weeks/28-day cycle at 100‒125 mg/m2. The modified dose–regimen cohorts received palbociclib 75 mg/day (3/1 schedule or continuously) plus nab-paclitaxel (biweekly 125 or 100 mg/m2, respectively). The prespecified efficacy threshold was 12-month survival probability of ≥65% at the MTD. Results: Palbociclib plus nab-paclitaxel was more effective than gemcitabine plus nab-paclitaxel in three of four PDX models tested; the combination was not inferior to paclitaxel plus gemcitabine. In the clinical trial, 76 patients (80% received prior treatment for advanced disease) were enrolled. Four dose-limiting toxicities were observed [mucositis (n = 1), neutropenia (n = 2), febrile neutropenia (n = 1)]. The MTD was palbociclib 100 mg for 21 of every 28 days and nab-paclitaxel 125 mg/m2 weekly for 3 weeks in a 28-day cycle. Among all patients, the most common all-causality any-grade adverse events were neutropenia (76.3%), asthenia/fatigue (52.6%), nausea (42.1%), and anemia (40.8%). At the MTD (n = 27), the 12-month survival probability was 50% (95% confidence interval, 29.9–67.2). Conclusions: This study showed the tolerability and antitumor activity of palbociclib plus nab-paclitaxel treatment in patients with PDAC; however, the prespecified efficacy threshold was not met. Trial Registration: Pfizer Inc (NCT02501902) Significance: In this article, the combination of palbociclib, a CDK4/6 inhibitor, and nab-paclitaxel in advanced pancreatic cancer evaluates an important drug combination using translational science. In addition, the work presented combines preclinical and clinical data along with pharmacokinetic and pharmacodynamic assessments to find alternative treatments for this patient population.

Supplemental Figure 1. Pairwise scatterplots comparing protein and cfDNA biomarkers for Post-treatment time point and % change pre-treatment to post-treatment. To determine the relationship between the different protein and cfDNA biomarkers we created pairwise scatterplots for the post-treatment time point (panel A) and % change (panel B). These data demonstrated good agreement between different cfDNA variables while the protein based showed variable correlations with cfDNA variables as outlined in Supplemental Table 3.

Purpose:To assess the preclinical efficacy, clinical safety and efficacy, and MTD of palbociclib plus nab-paclitaxel in patients with advanced pancreatic ductal adenocarcinoma (PDAC).Experimental Design:Preclinical activity was tested in patient-derived xenograft (PDX) models of PDAC. In the open-label, phase I clinical study, the dose-escalation cohort received oral palbociclib initially at 75 mg/day (range, 50‒125 mg/day; modified 3+3 design; 3/1 schedule); intravenous nab-paclitaxel was administered weekly for 3 weeks/28-day cycle at 100‒125 mg/m2. The modified dose–regimen cohorts received palbociclib 75 mg/day (3/1 schedule or continuously) plus nab-paclitaxel (biweekly 125 or 100 mg/m2, respectively). The prespecified efficacy threshold was 12-month survival probability of ≥65% at the MTD.Results:Palbociclib plus nab-paclitaxel was more effective than gemcitabine plus nab-paclitaxel in three of four PDX models tested; the combination was not inferior to paclitaxel plus gemcitabine. In the clinical trial, 76 patients (80% received prior treatment for advanced disease) were enrolled. Four dose-limiting toxicities were observed [mucositis (n = 1), neutropenia (n = 2), febrile neutropenia (n = 1)]. The MTD was palbociclib 100 mg for 21 of every 28 days and nab-paclitaxel 125 mg/m2 weekly for 3 weeks in a 28-day cycle. Among all patients, the most common all-causality any-grade adverse events were neutropenia (76.3%), asthenia/fatigue (52.6%), nausea (42.1%), and anemia (40.8%). At the MTD (n = 27), the 12-month survival probability was 50% (95% confidence interval, 29.9–67.2).Conclusions:This study showed the tolerability and antitumor activity of palbociclib plus nab-paclitaxel treatment in patients with PDAC; however, the prespecified efficacy threshold was not met.Trial Registration:Pfizer Inc (NCT02501902)Significance:In this article, the combination of palbociclib, a CDK4/6 inhibitor, and nab-paclitaxel in advanced pancreatic cancer evaluates an important drug combination using translational science. In addition, the work presented combines preclinical and clinical data along with pharmacokinetic and pharmacodynamic assessments to find alternative treatments for this patient population.

The treatment of metastatic pancreatic ductal adenocarcinoma (PDAC) is frequently characterized by significant toxicity and rapid development of resistance to current approved therapies. More reliable biomarkers of response are needed to guide clinical decision making.We evaluated cell-free DNA (cfDNA) using a tumor-agnostic platform and traditional biomarkers (CEA and CA19-9) levels in 12 patients treated at Johns Hopkins University on NCT02324543 “Study of Gemcitabine/Nab-Paclitaxel/Xeloda (GAX) in Combination With Cisplatin and Irinotecan in Subjects With Metastatic Pancreatic Cancer.” The pretreatment values, levels after 2 months of treatment, and change in biomarker levels with treatment were compared with clinical outcomes to determine their predictive value.The variant allele frequency (VAF) of KRAS and TP53 mutations in cfDNA after 2 months of treatment was predictive of progression-free survival (PFS) and overall survival (OS). In particular, patients with a lower-than-average KRAS VAF after 2 months of treatment had a substantially longer PFS than patients with higher posttreatment KRAS VAF (20.96 vs. 4.39 months). Changes in CEA and CA19-9 after 2 months of treatment were also good predictors of PFS.Comparison via concordance index demonstrated KRAS or TP53 VAF after 2 months of treatment to be better predictors of PFS and OS than CA19-9 or CEA. This pilot study requires validation but suggests cfDNA measurement is a useful adjunct to traditional protein biomarkers and imaging evaluation and could distinguish between patients who are likely to achieve prolonged responses versus those that will have early progression and may benefit from a change in treatment approach.Significance:We report on the association of cfDNA with response durability for patients undergoing treatment with a novel metronomic chemotherapy regimen (gemcitabine, nab-paclitaxel, capecitabine, cisplatin, irinotecan; GAX-CI) for metastatic PDAC. This investigation offers encouraging evidence that cfDNA may prove to be a valuable diagnostic tool to guide clinical management.

Supplementary Materials and Methods

Supplementary Table S3 from A Preclinical and Phase Ib Study of Palbociclib plus Nab-Paclitaxel in Patients with Metastatic Adenocarcinoma of the Pancreas

Table S1: Patient demographics. Table S2: Summary of TP53 VAF and KRAS VAF data for patient cohort (N=12). Table S3: Correlation between TP53 VAF, KRAS VAF, or median VAF and CEA and CA19-9 biomarkers using Spearman rank correlation coefficient (R). Table S4: Comparing predictive value of TP53 VAF, KRAS VAF, or median VAF versus CEA and CA19-9 for OS using C-index

Pancreatic ductal adenocarcinoma (PDAC) is considered a “nonimmunogenic” neoplasm. Single-agent immunotherapies have failed to demonstrate significant clinical activity in PDAC and other “nonimmunogenic” tumors, in part due to a complex tumor microenvironment (TME) that provides a formidable barrier to immune infiltration and function. We designed a neoadjuvant and adjuvant clinical trial comparing an irradiated, granulocyte-macrophage colony-stimulating factor (GM-CSF)–secreting, allogeneic PDAC vaccine (GVAX) given as a single agent or in combination with low-dose cyclophosphamide to deplete regulatory T cells (Treg) as a means to study how the TME is altered by immunotherapy. Examination of resected PDACs revealed the formation of vaccine-induced intratumoral tertiary lymphoid aggregates in 33 of 39 patients 2 weeks after vaccine treatment. Immunohistochemical analysis showed these aggregates to be regulatory structures of adaptive immunity. Microarray analysis of microdissected aggregates identified gene-expression signatures in five signaling pathways involved in regulating immune-cell activation and trafficking that were associated with improved postvaccination responses. A suppressed Treg pathway and an enhanced Th17 pathway within these aggregates were associated with improved survival, enhanced postvaccination mesothelin-specific T-cell responses, and increased intratumoral Teff:Treg ratios. This study provides the first example of immune-based therapy converting a “nonimmunogenic” neoplasm into an “immunogenic” neoplasm by inducing infiltration of T cells and development of tertiary lymphoid structures in the TME. Post-GVAX T-cell infiltration and aggregate formation resulted in the upregulation of immunosuppressive regulatory mechanisms, including the PD-1–PD-L1 pathway, suggesting that patients with vaccine-primed PDAC may be better candidates than vaccine-naïve patients for immune checkpoint and other immunomodulatory therapies. Cancer Immunol Res; 2(7); 616–31. ©2014 AACR.

Figure S1 Schema of the Neoadjuvant and Adjuvant Study of Panc GVAX. Figure S2 Pre- and post-vaccination mesothelin peptide-specific T cell levels in responders versus nonresponders. Figure S3 Intratumoral tertiary lymphoid aggregate in PDACs from patients treated with GVAX and lymphocyte cluster in PDACs from patients not treated with GVAX. Figure S4 The development of post-vaccination intratumoral lymphoid aggregates is associated with survival. Figure S5 Relatively more Granzyme B expressing cells observed outside of the lymphoid aggregates. Figure S6 Microdissection of Lymphoid Aggregates for Microarray Analysis. Figure S7 Microarray analysis of lymphoid aggregates revealed gene signatures in four pathways involved in regulating immune cell trafficking and function. Figure S8 Decreased Treg and increased TH17 pathway gene expression signatures are associated with improved post-vaccination responses. Figure S9 Downregulation of Treg pathway gene expression in TIL following treatment with PDAC GVAX is associated with longer survival. Figure S10 IL17A-producing T cells are present in low levels in TIL from PDAC GVAX treated patients. Figure S11 Treatment with low dose Cy increases post-vaccination densities of intratumoral lymphoid aggregates and reduces Treg levels in the PDAC TME. Figure S12 T cell infiltration in PDACs with low versus high CD8/Foxp3 ratio. Table S1 Immunohistochemistry protocols. Table S2 Quantitative real-time RT-PCR primers. Table S3 Scores of TH1, TH2, Treg, and TH17 pathway gene expression in PDAC TIL analyzed by quantitative real-time RT-PCR. Table S4 Numbers of TIL specimens with detectable expression of immune related genes.

Purpose: To identify a predictive molecular “signature” for sensitivity to retinoic acid in pancreatic cancer.Experimental Design: Fourteen patient-derived, low-passage pancreatic ductal adenocarcinoma (PDAC) lines with varied expression of fatty acid–binding protein 5 (FABP5) and cellular retinoic acid–binding protein 2 (CRABP2) were used to evaluate the response to all-trans retinoic acid (ATRA). Cell proliferation, apoptosis, and migration/invasion assays were used to measure the in vitro response. Tumor growth was monitored in subcutaneous xenografts in athymic nude mice for 4 weeks.Results: Response to ATRA was observed to be dependent upon differential expression of FABP5 versus CRABP2. Thus, elevated FABP5 expression was associated with minimal cytotoxicity and tumor growth inhibition and a paradoxical increase in migration and invasion. Conversely, CRABP2 expression in the absence of FABP5 was associated with significant tumor growth inhibition with ATRA, even in gemcitabine-resistant tumors. The ATRA-resistant phenotype of FABP5highCRABP2null cells could be circumvented by ectopic expression of CRABP2. Alternatively, reexpression of endogenous CRABP2 could be enabled in FABP5highCRABP2null PDAC lines by exposure to decitabine and trichostatin A, thereby relieving epigenetic silencing of the CRABP2 gene promoter. Immunohistochemical staining for FABP5 in archival human tissue microarrays identifies a subset of cases (13 of 63, ∼20%) which are negative for FABP5 expression and might be candidates for ATRA therapy.Conclusions: The widely used agent ATRA deserves a “second look” in PDAC, but needs to be targeted to patient subsets with biopsy-proven FABP5-negative tumors, or be combined with a chromatin-modifying agent to reexpress endogenous CRABP2. Clin Cancer Res; 18(1); 280–9. ©2011 AACR.

Observed genome equivalents, wild-type DNA fragments, mutant DNA fragments, and mutant allele frequency in K2EDTA tubes at 4{degree sign}C and Streck{trade mark, serif} BCT tubes at room temperature.

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Pharmacokinetic parameters of guadecitabine and decitabine after guadecitabine injection

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Purpose: The goal of this study was to evaluate prospectively the engraftment rate, factors influencing engraftment, and predictability of clinical outcome of low-passage xenografts from patients with resectable pancreatic ductal adenocarcinoma (PDA) and to establish a bank of PDA xenografts.Experimental Design: Patients with resectable PDA scheduled for resection at the Johns Hopkins Hospital were eligible. Representative pieces of tumor were implanted in nude mice. The status of the SMAD4 gene and content of tumor-generating cells were determined by immunohistochemistry. Gene expression was carried out by using a U133 Plus 2.0 array. Patients were followed for progression and survival.Results: A total of 94 patients with PDA were resected, 69 tumors implanted in nude mice, and 42 (61%) engrafted. Engrafted carcinomas were more often SMAD4 mutant, and had a metastatic gene expression signature and worse prognosis. Tumors from patients resistant to gemcitabine were enriched in stroma-related gene pathways. Tumors sensitive to gemcitabine were enriched in cell cycle and pyrimidine gene pathways. The time to progression for patients who received treatment with gemcitabine for metastatic disease (n = 7) was double in patients with xenografts sensitive to gemcitabine.Conclusion: A successful xenograft was generated in 61% of patients attempted, generating a pool of 42 PDA xenografts with significant biological information and annotated clinical data. Patients with PDA and SMAD4 inactivation have a better engraftment rate. Engraftment is a poor prognosis factor, and engrafted tumors have a metastatic gene expression signature. Tumors from gemcitabine-resistant patients were enriched in stromal pathways. Clin Cancer Res; 17(17); 5793–800. ©2011 AACR.

Average DNA Yield and Range of Plasma Isolated in Healthy Donors vs. Diseased Donors

Schematic of sample collection and analyses for the comparative studies between K2EDTA (EDTA) and Streck{trade mark, serif} BCT (Streck{trade mark, serif}) tubes.

LINE1 changes in tumor CpG 1-3 on cycle 1 day 8 from baseline. LINE1 changes in tumor DNA in individual patient biopsy samples.

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PDF file - 70K, We examined a gene signature of response to gemcitabine that included genes involved in the uptake (nucleoside transporters) and activation of gemcitabine (deoxycytin kinase) in tumor samples from patient JH034. We included samples from primary tumor, subcutaneous and orthotopic xenografts. Our analysis shows that primary and orthotopic xenografts are clustered together and differ from subcutaneous xenografts.

LINE1 changes in circulating DNA CpG 1-3 as compared to baseline.

Purpose: Analysis of genomic alterations in cell-free DNA (cfDNA) is evolving as an approach to detect, monitor, and genotype malignancies. Methods to separate the liquid from the cellular fraction of whole blood for circulating tumor DNA (ctDNA) analyses have been largely unstudied, although these may be a critical consideration for assay performance.Experimental Design: To evaluate the influence of blood processing on cfDNA and ctDNA quality and yield, we compared the cfDNA levels in serum with those in plasma. Given the limitations of serum for ctDNA analyses, we evaluated the effects of two plasma processing approaches, K2EDTA and Cell-Free DNA BCT (BCT) tubes, on cfDNA and ctDNA recovery. A total of 45 samples from nine patients with cancer were collected in both tube types. Once collected, blood was processed into plasma immediately or kept at room temperature and processed into plasma at 1, 3, 5, or 7 days.Results: As early as 24 hours after collection, plasma isolated from blood collected in K2EDTA tubes contained an elevated level of cfDNA that increased over time compared with BCT tubes where no significant increase in cfDNA levels was observed. When samples from an additional six patients with cancer, collected in the same manner, were stored at 4°C in K2EDTA tubes over the course of 3 days, total cfDNA and ctDNA levels were comparable between samples collected in BCT tubes. At day 3, there was a trend toward a decrease in ctDNA levels in both tubes that was more pronounced when measuring the mutant allele fraction for cases stored at 4°C in K2EDTA tubes.Conclusions: In summary, methods of blood processing have a strong influence on cfDNA and ctDNA levels and should be a consideration when evaluating ctDNA in peripheral circulation. Clin Cancer Res; 23(10); 2471–7. ©2016 AACR.

Purpose:Chemotherapeutic resistance eventually develops in all patients with metastatic colorectal cancer (mCRC). Gene silencing through promoter demethylation is one potential reversible mechanism of resistance with administration of hypomethylating agents. We evaluated the safety and tolerability of guadecitabine and irinotecan in patients with mCRC previously treated with irinotecan.Patients and Methods:In this 3+3 dose-escalation study, patients with mCRC previously exposed to irinotecan received guadecitabine days 1 to 5 of a 28-day cycle and irinotecan 125 mg/m2 days 8 and 15 [dose level (DL) 1, guadecitabine 45 mg/m2; DL −1: guadecitabine 30 mg/m2; DL −1G: guadecitabine 30 mg/m2 with growth factor support (GFS); DL 1G: guadecitabine 45 mg/m2 with GFS].Results:Twenty-two patients were treated across four DLs. Dose-limiting toxicities were neutropenic fever (DL 1 and −1G), biliary drain infection (DL −1), colonic obstruction (DL −1), and severe dehydration (DL 1G). Most common toxicities were neutropenia (82% any grade, 77% Grade 3/4), neutropenic fever (23%), leukopenia (73% any grade, 50% Grade 3/4), and injection site reactions (64% total, 0% Grade 3/4). Patients received a median of 4.5 cycles of treatment; 12/17 evaluable patients had stable disease as best response, with one having initial disease progression but subsequently durable partial response. Circulating tumor DNA showed decrease in global demethylation by LINE-1 after treatment.Conclusions:We report the first study of chemo-priming with epigenetic therapy in gastrointestinal cancers. Guadecitabine 45 mg/m2 and irinotecan 125 mg/m2 with GFS was safe and tolerable in patients with mCRC, with early indication of benefit. These data have provided the basis for an ongoing phase II randomized, multicenter trial.

Summary of clinical characteristics, plasma volume and DNA yield in 15 cancer patients.

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Next-Generation Sequencing Performance of Plasma Isolated in Different Blood Collection Tubes

Background A family history of pancreatic cancer is associated with increased pancreatic cancer risk. However, risk estimates for individuals in kindreds with an aggregation of pancreatic cancer (>1 relative) are imprecise because of small samples sizes or potentially impacted by biases inherent in retrospective data. Objective The objective of this study is to determine the age-specific pancreatic cancer risk as a function of family history using prospective data. Methods We compared pancreatic cancer incidence (n = 167) in 21 141 individuals from 4433 families enrolled in the National Familial Pancreatic Cancer Registry with that expected based on Surveillance Epidemiology and End Results data and estimated the cumulative probability of pancreatic cancer using competing risk regression. Results Familial pancreatic kindred members (kindreds with pancreatic cancer in 2 first-degree relatives [FDRs] or a pathogenic variant) had a standardized incidence ratio of 4.86 (95% confidence interval [CI] = 4.01 to 5.90), and sporadic kindred members (kindreds not meeting familial criteria) had a standardized incidence ratio of 2.55 (95% CI = 1.95 to 3.34). Risk in familial pancreatic cancer kindreds increased with an increasing number of FDRs with pancreatic cancer, with a standardized incidence ratio of 3.46 (95% CI = 2.52 to 4.76), 5.44 (95% CI = 4.07 to 7.26), and 10.78 (95% CI = 6.87 to 16.89) for 1, 2, and 3 or more FDRs with pancreatic cancer, respectively. Risk was also higher among individuals with a family history of young-onset (aged younger than 50 years) pancreatic cancer. Conclusion Pancreatic cancer risk is strongly dependent on family history, including both the degree of relationship(s) and age of onset of pancreatic cancer in relatives. These risk estimates will help inform the design of early detection studies and the risk and benefit analysis of screening trials.

Purpose: Mistletoe extract is widely used in cancer patients to support therapy and to improve quality of life (QoL). However, its use is controversial due to suboptimal trials and a lack of data supporting its intravenous administration. Patients and Methods: This phase I trial of intravenous Mistletoe (Helixor M) aimed to determine the recommended phase 2 dosing and to evaluate safety. Solid tumor patients progressing on at least one line of chemotherapy received escalating doses of Helixor M three times a week. Assessments were also made of tumor marker kinetics and QoL. Results: Twenty-one patients were recruited. The median follow-up duration was 15.3 weeks. The maximum tolerated dose was 600mg. Treatment-related adverse events (AEs) occurred in 13 patients (61.9%), with the most common being fatigue (28.6%), nausea (9.5%), and chills (9.5%). Grade 3+ treatment-related AEs were noted in three patients (14.8%). Stable disease was observed in five patients who had 1-6 prior therapies. Reductions in baseline target lesions were observed in three patients who had 2-6 prior therapies. Objective responses were not observed. The disease control rate (percentage of complete/partial response and stable disease) was 23.8%. The median stable disease was 15 weeks. Serum cancer antigen-125 or carcinoembryonic antigen showed a slower rate of increase at higher dose levels. The median QoL by FACT-G increased from 79.7 at week 1 to 93 at week 4. Conclusions: Intravenous mistletoe demonstrated manageable toxicities with disease control and improved QoL in a heavily pretreated solid tumor population. Future phase II trials are warranted. Citation Format: Channing J. Paller, Lin Wang, Wei Fu, Rajedra Kumar, Jennifer Durham, Nilofer Azad, Daniel Laheru, Ilene Browner, Sushant Kachhap, Kavya Boyapati, Thomas Odeny, Deborah Armstrong, Christian Meyer, Stephanie Gaillard, Julie Brahmer, Ivelisse Page, Hao Wang, Luis A. Diaz. Phase I trial of intravenous mistletoe extract in advanced cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 1600.

Gene variants that encode pancreatic enzymes with impaired secretion can induce pancreatic acinar endoplasmic reticulum (ER) stress, cellular injury and pancreatitis. The role of such variants in pancreatic cancer risk has received little attention. We compared the prevalence of ER stress-inducing variants in CPA1 and CPB1 in patients with pancreatic ductal adenocarcinoma (PDAC cases), enrolled in the National Familial Pancreas Tumor Registry, to their prevalence in non-cancer controls in the Genome Aggregation Database (gnomAD). Variants of unknown significance were expressed and variants with reduced secretion assessed for ER stress induction. In vitro assessments were compared with software predictions of variant function. Protein variant software was used to assess variants found in only one gnomAD control (“n-of-one” variants). A meta-analysis of prior PDAC case/control studies was also performed. Of the 1385 patients with PDAC, 0.65% were found to harbor an ER stress-inducing variant in CPA1 or CPB1, compared to 0.17% of the 64,026 controls (Odds Ratio (OR): 3.80 [1.92–7.51] P=0.0001). ER stress-inducing variants in the CPA1 gene were identified in 4 of 1385 PDAC cases versus 77 of 64,026 gnomAD controls (OR: 2.4 [0.88–6.58], p=0.087), and variants in CPB1 were detected in 5 of 1385 cases versus 33 of 64,026 controls (OR: 7.02 [2.74–18.01], p=0.0001). Meta-analysis demonstrated strong associations for pancreatic cancer and ER-stress inducing variants for both CPA1 (OR: 3.65 [1.58, 8.39], p

Background: Genome-wide association studies (GWAS) identify associations of individual single-nucleotide polymorphisms (SNPs) with cancer risk but usually only explain a fraction of the inherited variability. Pathway analysis of genetic variants is a powerful tool to identify networks of susceptibility genes. Methods: We conducted a large agnostic pathway-based meta-analysis of GWAS data using the summary-based adaptive rank truncated product method to identify gene sets and pathways associated with pancreatic ductal adenocarcinoma (PDAC) in 9040 cases and 12 496 controls. We performed expression quantitative trait loci (eQTL) analysis and functional annotation of the top SNPs in genes contributing to the top associated pathways and gene sets. All statistical tests were two-sided. Results: We identified 14 pathways and gene sets associated with PDAC at a false discovery rate of less than 0.05. After Bonferroni correction (P

Long-term outcomes for patients with locally advanced hepatocellular carcinoma (HCC) are poor. Even if clear margins are obtained at the time of surgical resection, most patients with large tumors (>5 cm) relapse due to micrometastatic disease. No therapy has demonstrated clinical benefit in the perioperative setting, underscoring the need for novel therapeutic strategies for resectable HCC. We hypothesize that neoadjuvant therapy resulting in downstaging and elimination of micrometastasis may increase the probability of successful surgical resection. This ongoing open label, single arm Phase Ib trial (NCT03299946) aims to assess the feasibility of neoadjuvant cabozantinib plus nivolumab (CaboNivo) in patients with locally advanced HCC. Targets of cabozantinib include MET and VEGFR. Nivolumab is an immune checkpoint inhibitor targeting PD-1. This study will also allow us to test the hypothesis that cabozantinib primes the tumor microenvironment for anti-PD1 therapy and to study mechanisms of response and resistance to combination therapy. To our knowledge, this is the first neoadjuvant clinical trial of an immune checkpoint inhibitor and the first study of neoadjuvant combination therapy in HCC. 15 patients aged ≥ 18 with potentially resectable HCC that is at high risk of recurrence, ECOG ≤ 1, normal organ and marrow function, and adequate future liver remnant will be enrolled in the study. After 2 weeks of 40 mg daily cabozantinib monotherapy lead-in, patients will receive concurrent 240 mg nivolumab infusions once every 2 weeks for 4 doses. After 8 weeks of neoadjuvant therapy and restaging, patients eligible for resection will proceed to definitive surgical resection ≥ 28 days after the last dose of cabozantinib. Primary outcomes are feasibility and safety of neoadjuvant cabozantinib plus nivolumab. Secondary outcomes are percentage of patients obtaining an R0 resection, pathologic complete response and major pathologic response rates, objective response rate, median overall survival, and median disease free survival. Tumor biopsies from patients at baseline and after 2 weeks of cabozantinib lead-in as well as tumor tissue collected from surgical resection samples will be used to determine the effect of cabozantinib monotherapy and combination therapy with nivolumab on tumor-infiltrating lymphocytes, immune activation and suppression pathways, and cytokine/chemokine signaling. Citation Format: Aleksandra Popovic, Elizabeth Sugar, Anna Ferguson, Bradley Wilt, Jennifer N. Durham, Ihab R. Kamel, Amy Kim, Benjamin Philosophe, Robert A. Anders, Elizabeth M. Jaffee, Daniel Laheru, Matthew J. Weiss, Mark Yarchoan. Feasibility of neoadjuvant cabozantinib plus nivolumab followed by definitive resection for patients with locally advanced hepatocellular carcinoma: A Phase Ib trial (NCT03299946) [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 CT207.

Gemcitabine is considered the standard first-line therapy for patients with advanced pancreatic cancer. More recent strategies have focused on improving the efficacy of gemcitabine by either improving the method of delivery or by combining gemcitabine with other non-cross-resistant chemotherapy agents or with small-molecule drugs. However, the clinical benefits, response rates, and duration of responses have been modest. Deoxycytidine kinase (dCK) is the rate-limiting enzyme involved in the metabolism of gemcitabine. The expression of dCK has been postulated to be correlative of gemcitabine resistance. We determined the relationship of dCK immunohistochemical protein expression and/or genetic status of dCK in a panel of human pancreatic cancer tissues and pancreatic cancer cell lines and determined the relationship of these variables to the clinical outcome of patients treated with gemcitabine. We report that dCK protein expression is expressed in the majority of pancreatic cancers analyzed (40 of 44 cases, 91%) and showed a range of labeling intensities ranging from 1+ (labeling weaker in intensity than normal lymphocytes present in same section) to 3+ (labeling greater in intensity than normal lymphocytes present in same section). When labeling intensity was compared with survival, low dCK expression (1+ labeling) was correlated with both overall survival (P < 0.009) and progression-free survival following gemcitabine treatment (P < 0.04). Low dCK labeling intensity was also significantly correlated with patient age (70.3 ± 8.1 versus 59.8 ± 7.4 years; P < 0.0006), suggesting that age-related methylation of the dCK gene may account in part for the observed differences. Sequencing of the entire dCK coding sequence in 17 cell lines and 9 patients' cancer tissues with disease progression while on gemcitabine did not identify any mutations, suggesting that genetic alterations of dCK are not a common mechanism of resistance to gemcitabine for this tumor type. Moreover, dCK labeling showed similar patterns and intensities of labeling among matched pretreatment and post-treatment tissues. In summary, pretreatment levels of dCK protein are most correlated with overall survival following gemcitabine treatment and are stable even after resistance to gemcitabine is clinically documented.

The purpose of this study was to evaluate preoperative treatment with full-dose gemcitabine, oxaliplatin, and radiation therapy (RT) in patients with localized pancreatic cancer.Eligibility included confirmation of adenocarcinoma, resectable or borderline resectable disease, a performance status ≤2, and adequate organ function. Treatment consisted of two 28-day cycles of gemcitabine (1 g/m(2) over 30 minutes on days 1, 8, and 15) and oxaliplatin (85 mg/m(2) on days 1 and 15) with RT during cycle 1 (30 Gray [Gy] in 2-Gy fractions). Patients were evaluated for surgery after cycle 2. Patients who underwent resection received 2 cycles of adjuvant chemotherapy.Sixty-eight evaluable patients received treatment at 4 centers. By central radiology review, 23 patients had resectable disease, 39 patients had borderline resectable disease, and 6 patients had unresectable disease. Sixty-six patients (97%) completed cycle 1 with RT, and 61 patients (90%) completed cycle 2. Grade ≥3 adverse events during preoperative therapy included neutropenia (32%), thrombocytopenia (25%), and biliary obstruction/cholangitis (14%). Forty-three patients underwent resection (63%), and complete (R0) resection was achieved in 36 of those 43 patients (84%). The median overall survival was 18.2 months (95% confidence interval, 13-26.9 months) for all patients, 27.1 months (95% confidence interval, 21.2-47.1 months) for those who underwent resection, and 10.9 months (95% confidence interval, 6.1-12.6 months) for those who did not undergo resection. A decrease in CA 19-9 level after neoadjuvant therapy was associated with R0 resection (P = .02), which resulted in a median survival of 34.6 months (95% confidence interval, 20.3-47.1 months). Fourteen patients (21%) are alive and disease free at a median follow-up of 31.4 months (range, 24-47.6 months).Preoperative therapy with full-dose gemcitabine, oxaliplatin, and RT was feasible and resulted in a high percentage of R0 resections. The current results are particularly encouraging, because the majority of patients had borderline resectable disease.

Pancreatic cancer, like most other cancers, is fundamentally caused by mutations in specific genes. Many of the genes targeted in pancreatic cancer have been identified in the past decade, and an understanding of these genes and their function has helped identify familial forms of pancreatic cancer, define the precursor lesions from which invasive pancreatic cancers arise, and will soon lead to gene-specific therapies for this disease. This article reviews the tumor-suppressor genes, oncogenes, and DNA repair genes targeted in pancreatic cancer, and presents clinical applications based on our understanding of the genetic basis of pancreatic cancer.