Diagnostic and predictive factors in pancreatic cancer : clinical and translational studies

Aims: The aim of Study I was to assess the effectiveness of chemotherapy for patients with advanced pancreatic cancer in a real-world setting. Study II aimed to evaluate the utility of a clinical decision support system to identify precision oncology opportunities for pancreatic cancer. The aim of Study III was to identify the blood component most enriched with pancreatic cancer-derived circulating DNA; that of Study IV was to characterize long and short tumor-derived circulating DNA fragments. Methods: Study I is a single-institution retrospective cohort study of prospectively generated clinical data. We included a total of 595 patients and used univariate and multivariate models, including flexible parametric models to analyze overall survival and time to treatment failure according to different first-line chemotherapy regimens. Exploratory analyses included the adherence to different protocols, adverse events, and second-line chemotherapy. Study II is a prospective observational study of 39 patients with pancreatic cancer who were enrolled in the PePaCaKa-01 study. Archival tumor tissue was sequenced, and data was processed with the proprietary clinical decision support system MH Guide and results were evaluated by a study-specific molecular tumor board. Endpoints of the study were the frequency of successful generation of support system reports, the frequency of actionable molecular targets, and their evaluation by the tumor board. We performed a post-hoc analysis to determine the proportion of patients who received molecular informed therapies. Studies III– IV analyzed blood samples from a prospective cohort of patients with advanced pancreatic cancer. We systematically separated whole blood into red and white blood cells, platelets, apoptotic bodies, large and small extracellular vesicles, and soluble protein using differential centrifugation. We confirmed efficient separation with protein assays (Western blotting and multiplex bead-based extracellular vesicle flow cytometry), nanoparticle tracking analysis and transmission electron microscopy, and extracted DNA from all components. We used digital PCR to quantify the abundance of KRASmut DNA, a hallmark of pancreatic cancerderived DNA. In Study IV we additionally used automated electrophoresis to quantify the lengths of circulating DNA fragments and ligation-based sequencing library preparation and tagmentation to selectively target short and long DNA fragments, respectively. Results: In Study I, we observed similar overall survival for gemcitabine/capecitabine, gemcitabine/nab-paclitaxel, and FOLFIRINOX (including modified regimens) compared to gemcitabine. Combinations of 5-fluorouracil/oxaliplatin and best supportive care were associated with poorer outcomes. Models adjusting for other demographic and clinical variables showed a survival benefit for gemcitabine-combinations and FOLFIRINOX. Exploratory analyses revealed differences in protocol adherence across different treatments, a relatively low frequency of AEs, and a difference between different sequences of first- and second-line therapy. In Study II, a CDSS report was generated for 31/39 patients, 28/31 reports were evaluated at the study-specific molecular tumor board The clinical decision support system made 80 individual recommendations to use molecularly informed therapies based on 61 genomic variants. In 21/28 cases, the tumor board classified at least one molecularly informed therapy as a potential clinical option. The post-hoc analysis revealed that six patients received molecularly informed treatment in routine care. In Study III, KRASmut DNA had the highest concentrations in the soluble protein and small vesicles blood fractions at late stages of PDAC. At early stages, it was highest in large and small extracellular vesicles. Small extracellular vesicles also contained the highest ratio of the concentrations of mutant : wild type KRAS DNA at this stage. In Study IV, blood from PDAC patients had significantly higher concentrations of short cell-free DNA in the soluble protein fraction than that of healthy individuals. The mutant allele frequency of KRASmut was highest in this blood component. Long genomic DNA fragments were most reliably measured in association with apoptotic bodies but KRASmut genomic DNA occurred in all assessed blood fractions. Conclusions: Chemotherapy in clinical routine use can result in outcomes that reflect relevant randomized controlled trials and gemcitabine-based regimens are highly effective this setting. Differences between different treatments might be related to how they are applied (Study I). The clinical decision support system MH Guide could identify clinically relevant opportunities for molecularly informed treatments of advanced pancreatic cancer (Study II). At early stages of pancreatic cancer, tumor-derived DNA is mostly associated with small extracellular vesicles; in more advanced disease it is mainly a feature of the soluble protein fraction (Study III). Short DNA fragments in this fraction are a more robust source of tumorderived DNA than longer genomic DNA fragments (Study IV)

TLR9 signalling inhibits Plasmodium liver infection by macrophage activation.

Recognition of pathogen-associated molecular patterns (PAMPs) through Toll-like receptors (TLRs) plays a pivotal role in first-line pathogen defense. TLRs are also likely triggered during a Plasmodium infection in vivo by parasite-derived components. However, the contribution of innate responses to liver infection and to the subsequent clinical outcome of a blood infection is not well understood. To assess the potential effects of enhanced TLR-signalling on Plasmodium infection, we systematically examined the effect of agonist-primed immune responses to sporozoite inoculation in the P. berghei/C57Bl/6 murine malaria model. We could identify distinct stage-specific effects on the course of infection after stimulation with two out of four TLR-ligands tested. Priming with a TLR9 agonist induced killing of pre-erythrocytic stages in the liver that depended on macrophages and the expression of inducible nitric oxide synthase (iNOS). These factors have previously not been recognized as antigen-independent effector mechanisms against Plasmodium liver stages. Priming with TLR4 and -9 agonists also translated into blood stage-specific protection against experimental cerebral malaria (ECM). These insights are relevant to the activation of TLR signalling pathways by adjuvant systems of antimalaria vaccine strategies. The protective role of TLR4-activation against ECM might also explain some unexpected clinical effects observed with pre-erythrocytic vaccine approaches

Fosmidomycin Uptake into Plasmodium and Babesia-Infected Erythrocytes Is Facilitated by Parasite-Induced New Permeability Pathways

., a mouse malaria parasite. and related parasites. Our data provide further evidence that parasite-induced new permeability pathways may be exploited as routes for drug delivery

Survival Benefits of Chemotherapy for Patients with Advanced Pancreatic Cancer in A Clinical Real-World Cohort

Clinical outcomes of chemotherapy for patients with advanced pancreatic adenocarcinoma in a real-world setting might differ from outcomes in randomized clinical trials (RCTs). Here we show in a single-institution cohort of 595 patients that median overall survival (OS) of patients who received gemcitabine alone (n = 185; 6.6 months (95% CI; 5.5–7.7)) was the same as in pivotal RCTs. Gemcitabine/capecitabine (n = 60; 10.6 months (95% CI; 7.8–13.3)) and gemcitabine/nab-paclitaxel (n = 66; 9.8 months (95% CI; 7.9–11.8)) resulted in a longer median OS and fluorouracil/oxaliplatin/irinotecan (n = 31, 9.9 months (95% CI; 8.1–11.7)) resulted in a shorter median OS than previously reported. Fluorouracil/oxaliplatin (n = 35, 5.8 months (95% CI; 4.5–7)) and best supportive care (n = 206, 1.8 months (95% CI; 1.5–2.1)) could not be benchmarked against any RCTs. The degree of protocol adherence explained differences between real-world outcomes and the respective RCTs, while exposure to second-line treatments did not

Recent Discoveries of Diagnostic, Prognostic and Predictive Biomarkers for Pancreatic Cancer

Simple Summary Biomarkers for cancer diagnosis, prognosis and prediction are important tools and an urgent need in precision medicine for pancreatic cancer. In recent years, many experimental and clinical studies aimed at identifying new biomarkers for pancreatic ductal adenocarcinoma. In the review, we summarized current investigations on using novel protein markers, cell-free DNA, metabolome compounds, immune and stroma signatures and microbiome compositions as biomarkers for pancreatic cancer. Our comprehensive overview shows that although there are new promising biomarkers, CA 19-9 remains currently the only regularly used and validated biomarker for pancreatic cancer in clinical routine. Pancreatic ductal adenocarcinoma (PDAC) is an aggressive disease with a dismal prognosis that is frequently diagnosed at an advanced stage. Although less common than other malignant diseases, it currently ranks as the fourth most common cause of cancer-related death in the European Union with a five-year survival rate of below 9%. Surgical resection, followed by adjuvant chemotherapy, remains the only potentially curative treatment but only a minority of patients is diagnosed with locally resectable, non-metastatic disease. Patients with advanced disease are treated with chemotherapy but high rates of treatment resistance and unfavorable side-effect profiles of some of the used regimens remain major challenges. Biomarkers reflect pathophysiological or physiological processes linked to a disease and can be used as diagnostic, prognostic and predictive tools. Thus, accurate biomarkers can allow for better patient stratification and guide therapy choices. Currently, the only broadly used biomarker for PDAC, CA 19-9, has multiple limitations and the need for novel biomarkers is urgent. In this review, we highlight the current situation, recent discoveries and developments in the field of biomarkers of PDAC and their potential clinical applications

Risk of Developing Pancreatic Cancer in Patients with Chronic Pancreatitis

Background: Patients with chronic pancreatitis (CP) have an increased risk of developing pancreatic ductal adenocarcinoma (PDAC). We present data on PDAC in one of the most extensive European single-centre cohort studies of patients with CP. Methods: Retrospective analysis of prospectively collected data of patients with CP was performed. Aetiology of CP was determined according to the M-ANNHEIM classification system and only patients with definite CP > 18 years at data analysis were included. The final dataset included 581 patients with definite CP diagnosed between 2003 and 2018. Results: At CP diagnosis, there were 371 (63.9%) males and 210 (36.1%) females (median age 57 years, range 2–86). During 3423 person-years of observation, six pancreatic cancers were diagnosed (0.2% year). The mean time between diagnosis of CP and the occurrence of PDAC was 5.0 years (range 2.7–8.6). None of the cancer patients had a family history of PDAC. Diabetes mellitus (DM) was present in five of six (83.3%) patients with PDAC: in three patients before and in two after CP diagnosis. Clinical/laboratory signs of pancreatic exocrine insufficiency (PEI) were present in five of six (83.3%) patients with PDAC: in two at diagnosis of CP and in three after diagnosis. The mean survival time was 4 months after the diagnosis of PDAC (range 0.5–13). PDAC occurred significantly more often (p < 0.001) in two groups of patients without previous acute pancreatitis (AP): 2 of 20 patients (10%) with low body mass index (BMI) and PEI and in 3 of 10 (30%) patients with high BMI and DM at diagnosis of CP. Conclusions: Patients with CP have a high risk of developing PDAC, although risk is low in absolute terms. Our data suggest the possibility of defining subgroups of patients with a particularly elevated risk of PDAC. Such a possibility would open a path to personalised decision making on initiation of PDAC surveillance of patients with no previous episode of AP, (i) with low BMI and PEI, or (ii) elevated BMI and DM

Plasma protein biomarkers for early detection of pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) has a poor prognosis, mainly due to late diagnosis at advanced tumor stages. In this study, we aimed to identify plasma protein biomarkers for early detection of PDAC. Totally, 135 PDAC patients (early PDAC, Stage I/II, n = 71; advanced PDAC, Stage III/IV, n = 64), 13 benign lesions/chronic pancreatitis patients and 72 healthy individuals, with corresponding plasma samples from a case-control study in Sweden were included. A proximity extension assay was used to detect 92 cancer-related proteins, and an enzyme-linked immunosorbent assay/electrochemiluminescence immunoassay was used to detect CA19-9. Predictive features were selected from these 93 candidate proteins and three covariates in the Swedish participants, and then validated in Spanish participants, including 37 early PDAC patients, 38 advanced PDAC patients, 19 chronic pancreatitis patients and 36 healthy controls. A panel of eight proteins discriminating early PDAC from healthy individuals was identified, and the cross-validated area under the curves (AUCs) were 0.85 (95% confidence interval, 95% CI, 0.78-0.91) and 0.81 (95% CI, 0.70-0.92) in the Swedish and Spanish participants, respectively. Another eight-protein panel was predictive for classifying advanced PDAC from healthy controls in two populations, with cross-validated AUCs of 0.89 (95% CI, 0.83-0.95) and 0.90 (95% CI, 0.83-0.98), respectively. In conclusion, eight protein biomarkers were identified and externally validated, potentially allowing early detection of PDAC patients if validated in additional prospective studies