In vivo evaluation of pharmacokinetic drug-drug interactions between fluorinated pyrimidine anticancer drugs, 5-fluorouracil and capecitabin, and an anticoagulant, warfarin.

Warfarin is a common anticoagulant and has demonstrated interactions with several drugs. Among them, as a serious adverse event, a case of death due to the enhanced warfarin action owing to its combined use with a fluoropyrimidine anticancer drug has been reported, but the detailed mechanism has not been elucidated.Some reports have advocated that fluorinated pyrimidine anticancer drugs reduce cytochrome P450 2C9 expression, leading to the enhanced pharmacological effects of warfarin.The purpose of this study was to clarify the mechanisms of drug-drug interactions between warfarin and 5-fluorouracil (5-FU) and capecitabine in vivo using rats. Rats were administered warfarin in combination with 5-FU (15 mg/kg/d) or capecitabine (15 mg/kg/d) for 7 d. Prothrombin time (PT) and activated partial thromboplastin time were significantly prolonged in the warfarin plus 5-FU or capecitabine groups compared with those in the warfarin alone group. No significant difference was observed in the area under the plasma concentration-time curve of the warfarin alone group compared with the warfarin with 5-FU or capecitabine groups.These data suggest that the enhancement of warfarin efficacy caused by the combination of 5-FU or capecitabine was due to a pharmacological interaction rather than a pharmacokinetic interaction.

Neoadjuvant concurrent chemoradiotherapy using infusional gemcitabine in locally advanced rectal cancer: A phase II trial.

INTRODUCTION: Gemcitabine is a well-known radiosensitizer. Herein, we tested the efficacy and toxicity of preoperative concurrent infusional gemcitabine and radiotherapy in locally advanced rectal cancer. PATIENTS AND METHODS: This was a phase II, single-arm trial. Eligible patients had a diagnosis of rectal adenocarcinoma with clinical stage T3-T4 and/or nodal involvement, age ≥18 years, and no prior chemotherapy or radiotherapy. Patients received preoperative radiation at a dose of 50.4-54 Gy over 28 days with concurrent infusional gemcitabine administered at a dose of 100 mg/m2 over the course of 24 h weekly for 6 weeks. The primary endpoint was pathological complete response (pCR). RESULTS: Forty patients were recruited. Only one patient did not complete therapy due to death. Eight patients did not undergo surgery, one died, two progressed to nonresectable disease, and five withdrew consent. Five patients progressed prior to surgery, with two having unresectable metastases and three having resectable liver metastases. One was found to have peritoneal metastasis during surgery. Out of the 32 patients who underwent surgery, seven achieved pCR at a rate of 20%. With a median follow-up of 30 months, four additional patients had a distant relapse (one had a subsequent local relapse). The 3-year event-free and overall survival rates were 70% and 85%, respectively. The commonest preoperative grade 3-4 toxicity included lymphopenia (50%), neutropenia (41%), anemia (15%), diarrhea (12%), abdominal pain (12%), and proctitis (8%). CONCLUSION: Concurrent preoperative chemoradiotherapy using infusional gemcitabine for locally advanced rectal cancer achieved an encouraging degree of local control with manageable toxicity.

The efficacy and safety of gemcitabine-based induction chemotherapy for locally advanced nasopharyngeal carcinoma treated with concurrent chemoradiation: A meta-analysis.

OBJECTIVES: To assess the efficacy and toxicity of gemcitabine-based induction chemotherapy followed by concurrent chemoradiotherapy (CCRT) in locally advanced nasopharyngeal carcinoma (LA-NPC). METHODS: Both observational studies (OBS) and randomized controlled trials (RCT) were included in the meta-analysis. Systematic online searches were conducted in Web of Sciences, PubMed, Embase, meeting proceedings and ClinicalTrials.gov from the inception to May 25, 2020. The primary endpoint of interest was overall survival. RESULTS: five OBSs and 2 RCTs including 1680 patients were incorporated in the analysis. The evidence from the RCTs showed that adding gemcitabine-based induction chemotherapy to CCRT significantly improved progression free survival (hazard ratio (HR): 0.60, 95% confidence interval (CI): 0.40-0.88; P = .010; chi square P = .25; I2 = 24%) and overall survival (HR: 0.47; 95% CI: 0.28-0.80; P = 0.005; chi square P = .49, I2 = 0%) and was related to a higher risk of hematological toxicities. Furthermore, based on the data of OBSs, overall survival (HR: 0.52; 95% CI: 0.31-0.88; P = .02; chi square P = .37, I2 = 6%) was significantly improved in patients treated with gemcitabine-based induction chemotherapy compared to those treated with taxane-based induction chemotherapy. However, the progression free survival (HR: 0.67; 95% CI: 0.45-1.01; P = .06; chi square P = .74; I2 = 0%) showed no significant difference. CONCLUSIONS: For LA-NPC patients, adding gemcitabine-based induction chemotherapy to CCRT significantly improved overall survival and progression free survival with a higher risk of hematological toxicities when compared to CCRT alone. Also, gemcitabine-based regimen could be used as an alternative induction chemotherapy regimen to taxane-based regimen in the treatment of LA-NPC.

Enhanced gemcitabine cytotoxicity with knockdown of multidrug resistance protein genes in human cholangiocarcinoma cell lines.

BACKGROUND AND AIM: Cholangiocarcinoma (CCA) is an often fatal primary cancer of the liver that tends to be resistant to chemotherapy. Multidrug resistance proteins (MRPs) contribute to the chemoresistance of these tumors. The objectives of the study were to document MRP expression profiles in two representative human intrahepatic and extrahepatic CCA cells lines (HuCCT1 and KMBC, respectively) and gemcitabine-induced cytotoxicity prior to and following MRP knockdown. METHODS: Multidrug resistance protein mRNA and protein expression were documented by real-time reverse transcription-polymerase chain reaction and western blots, respectively. MRP knockdown was achieved with lentivirus small hairpin RNA constructs. RESULTS: Prior to gemcitabine exposure, MRP1, MRP2, MRP4, MRP5, and MRP6 mRNA were expressed in HuCCT1 cells and MRP1, MRP3, MRP4, and MRP5 in KMBC cells. Following gemcitabine exposure, MRP5 and MRP6 expressions were significantly upregulated in HuCCT1 cells and MRP5 in KMBC cells. In HuCCT1 cells, although MRP5 knockdown had no effect, MRP6 knockdown significantly increased gemcitabine-induced cytotoxicity. In KMBC cells, MRP5 knockdown significantly increased gemcitabine cytotoxicity. CONCLUSIONS: Inhibition of MRP6 expression in intra-hepatic and MRP5 in extra-hepatic should be explored as potential treatments for CCA in humans.

Pterostilbene Enhances Cytotoxicity and Chemosensitivity in Human Pancreatic Cancer Cells.

Gemcitabine (GEM) drug resistance causes high mortality rates and poor outcomes in pancreatic ductal adenocarcinoma (PDAC) patients. Receptor for advanced glycation end products (RAGE) involvement in the GEM resistance process has been demonstrated. Therefore, finding a safe and effective way to inhibit receptors for RAGE-initiated GEM resistance is urgent. Pterostilbene (PTE), a natural methoxylated analogue derived from resveratrol and found in grapes and blueberries, has diverse bioactivities, such as antioxidative, anti-inflammatory, and anticancer qualities. The overall research objective was to determine the potential of PTE to enhance tumor cytotoxicity and chemosensitivity in PDAC cells. Our results have demonstrated that PTE induced S-phase cell cycle arrest, apoptosis, and autophagic cell death and inhibited multidrug resistance protein 1 (MDR1) expression by downregulating RAGE/PI3K/Akt signaling in both MIA PaCa-2 and MIA PaCa-2 GEMR cells (GEM-resistant cells). Remarkably, convincing evidence was established by RAGE small interfering RNA transfection. Taken together, our study demonstrated that PTE promoted chemosensitivity by inhibiting cell proliferation and MDR1 expression via the RAGE/PI3K/Akt axis in PDAC cells. The observations in these experiments indicate that PTE may play a crucial role in MDR1 modulation for PDAC treatment.

Gemcitabine induced cytotoxicity, DNA damage and hepatic injury in laboratory mice.

The present study was conducted to demonstrate cytotoxicity, apoptosis and hepatic damage induced by gemcitabine in laboratory mice. Animals were treated with a single dose of gemcitabine (415 mg/kg body wt), equivalent to a human therapeutic dose, and sacrificed after 1, 2 and 3 weeks. A significant decrease in mean body weight and absolute liver weight was registered. The levels of alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) were increased as a result of this induced stress. Various structural changes were observed in the liver tissue of treated mice, as evident in the histological sections. Specifically, gemcitabine exposure was able to induce apoptosis in liver cells, and the incidence of TUNEL positive liver cells was increased compared to the control group. DNA fragmentation appeared on agarose gel and flow cytometry analysis confirmed the induction of apoptosis. These findings in gemcitabine-treated animal tissues suggest that inhibition or disruption of cells' DNA synthesis may be the mechanism by which this drug induces toxicity in the animal body.

In vitro and molecular chemosensitivity in human cholangiocarcinoma tissues.

Adjuvant chemotherapy is required for cholangiocarcinoma (CCA) patients after surgical treatment. Gemcitabine and gemcitabine plus cisplatin are considered the appropriate regimen; however, the response spectrum to chemotherapy differs between patients. Thus, the present study aims to evaluate the response pattern of individual CCA patients by using an in vitro method, histoculture drug response assay (HDRA), to predict the chemosensitivity of individual patients in a prospective study. Moreover, we also investigate the expression of gemcitabine and cisplatin sensitivity factors in CCA tissues in the same cases. Based on the dose response curve, 1000 and 1500 µg/ml of gemcitabine were used as the testing concentrations. For cisplatin, concentrations of 20 and 25 µg/ml were selected for testing and for the combination regimen, 1000 µg/ml of gemcitabine and 20 µg/ml of cisplatin were chosen. The median %IR of each drug was measured as the cut-off to categorize the response pattern into response and non-response groups. In addition, we compared the effectiveness of the chemotherapy regimens between gemcitabine alone and gemcitabine plus cisplatin. The %IR of the combination of gemcitabine and cisplatin was significantly higher than gemcitabine alone. The relationship between the expression level of gemcitabine and cisplatin sensitive factors and the individual response pattern as well as clinicopathological data of CCA patients were analyzed. The results indicated that a low expression of the gemcitabine sensitive factor hENT-1 was significantly associated with the non-response group in vitro (p = 0.002). Moreover, the low expression of hENT-1 was also significantly associated with advanced stages CCA in the patients (p = 0.025). A low expression of MT and ERCC1 was significantly correlated with the response group in the in vitro experiments (p = 0.015 and p = 0.037 for MT and ERCC1, respectively). Therefore, HDRA may serve as an aid to selecting chemotherapy, and the expression of hNET-1, MT and ERCC1 may serve as biomarkers for predicting chemotherapy success.

Synergistic and Pharmacotherapeutic Effects of Gemcitabine and Cisplatin Combined Administration on Biliary Tract Cancer Cell Lines.

Gemcitabine (GEM) and cisplatin (CDDP) combination therapy (GC) is the standard chemotherapy for advanced biliary tract cancer (BTC); however, its pharmacotherapeutic efficacy remains unclear. To investigate the effects of GC, we selected 11 from 17 BTC cell lines, according to their GEM sensitivity, to be assessed using the MTS assay. The presence of synergistic effects of GC was determined using the Bliss additivism model (BM) and the combination index (CI) at a GEM:CDDP molar ratio of 7:1; this ratio was based on the respective human renal clearances of the two drugs. The pharmacotherapeutic effects were evaluated by comparing the IC50 values for administrations of GEM alone and GC in combination. All cell lines showed synergistic effects when analyzed using the BM. Based on the CI values, strong synergism, synergism, and additive effects were seen in four, five, and two cell lines, respectively. For all four GEM-resistant cell lines, on which GC had strong synergistic effects, the pharmacotherapeutic effects of GC were disappointing, with all IC50 values > 1 µM. For the GEM-effective cell lines, on which GC had synergistic or additive effects, the IC50 values were all <1 µM, and the differences were small between the IC50s for administration of GEM alone and GC in combination. Our results suggest that GC has synergistic effects on BTC cell lines but that its pharmacotherapeutic effects are inadequate.

The DNA repair helicase RECQ1 has a checkpoint-dependent role in mediating DNA damage responses induced by gemcitabine.

The response of cancer cells to therapeutic drugs that cause DNA damage depends on genes playing a role in DNA repair. RecQ-like helicase 1 (RECQ1), a DNA repair helicase, is critical for genome stability, and loss-of-function mutations in the RECQ1 gene are associated with increased susceptibility to breast cancer. In this study, using a CRISPR/Cas9-edited cell-based model, we show that the genetic or functional loss of RECQ1 sensitizes MDA-MB-231 breast cancer cells to gemcitabine, a nucleoside analog used in chemotherapy for triple-negative breast cancer. RECQ1 loss led to defective ATR Ser/Thr kinase (ATR)/checkpoint kinase 1 (ChK1) activation and greater DNA damage accumulation in response to gemcitabine treatment. Dual deficiency of MUS81 structure-specific endonuclease subunit (MUS81) and RECQ1 increased gemcitabine-induced, replication-associated DNA double-stranded breaks. Consistent with defective checkpoint activation, a ChK1 inhibitor further sensitized RECQ1-deficient cells to gemcitabine and increased cell death. Our results reveal an important role for RECQ1 in controlling cell cycle checkpoint activation in response to gemcitabine-induced replication stress.

A case report of a severe fluoropyrimidine-related toxicity due to an uncommon DPYD variant.

INTRODUCTION: Fluoropyrimidines such as 5-fluorouracil (5-FU) and its orally active prodrug, capecitabine, are widely used in the treatment of gastrointestinal cancer, including colorectal cancer. Dihydropyrimidine dehydrogenase (DPD) plays an important role in the 5-FU metabolism. Dihydropyrimidine dehydrogenase gene (DPYD) is a highly polymorphic gene with several hundreds of reported genetic variants and DPD activity levels vary considerably among individuals, with different 5-FU-related efficacy and toxicity. About 5% of the population is deficient in DPD enzyme activity. The most well studied DPYD variant is the IVS14+1G>A, also known as DPYD *2A. In this report, we present a case of a patient with a double heterozygote DPYD variant (DPYD activity score: 0,5 according to Clinical Pharmacogenetics Implementation Consortium) who experienced a severe fluoropyrimidine-related toxicity resolved without any consequence. PATIENT CONCERNS: A 46-years-old Caucasian man with diagnosis of left colon adenocarcinoma underwent left hemicolectomy on July 2017: pT3 G3 N1c M0. According to the disease stage, he started an adjuvant therapy with XELOX using capecitabine at 50% of total dose, because of his DPYD IVS14+1G>A variant, detected before the treatment. DIAGNOSIS: After few days, despite of this dose reduction, he experienced life-threatening adverse events such as mucositis G3, diarrhea G3, neutropenia G4, thrombocytopenia G4, and hyperbilirubinemia G3 according to Common Terminology Criteria for Adverse Events v 5.0. INTERVENTIONS: As first, we set up an intensive rehydration therapy, antibiotic and antifungal prophylaxis, Granulocyte-Colony Stimulating Factors, and supportive blood transfusions. Additional genetic tests revealed a double heterozygote variant of DPYD gene (DPYD IVS14+1G>A and 2846A>T) which is a very rare situation and only 3 cases are described in literature, all of them concluded with patient's death. OUTCOMES: After 3 weeks of intensive therapy, the patient was fully recovered. Furthermore, all the whole-body CT scans performed since discharge from the hospital until now, have confirmed no evidence of disease. CONCLUSIONS: Recent studies demonstrated that screening strategy for the most common DPYD variants allowed for avoiding toxicities and saving money. This report underlines the importance of genotyping DPYD before treatment and emphasizes the role of genotype-guided dose individualization.

Polygemcitabine nanogels with accelerated drug activation for cancer therapy.

To overcome the slow activation of gemcitabine, we synthesized a DNA-like polygemcitabine (Ge10) strand through solid-phase synthesis, which not only undergoes rapid intracellular degradation to generate active gemcitabine derivatives, but can also self-assemble into nanogels through molecular recognition, rendering them as promising self-delivered nanodrugs for cancer therapy.

Genome-scale CRISPR/Cas9 screen determines factors modulating sensitivity to ProTide NUC-1031.

Gemcitabine is a fluoropyrimidine analogue that is used as a mainstay of chemotherapy treatment for pancreatic and ovarian cancers, amongst others. Despite its widespread use, gemcitabine achieves responses in less than 10% of patients with metastatic pancreatic cancer and has a very limited impact on overall survival due to intrinsic and acquired resistance. NUC-1031 (Acelarin), a phosphoramidate transformation of gemcitabine, was the first anti-cancer ProTide to enter the clinic. We find it displays important in vitro cytotoxicity differences to gemcitabine, and a genome-wide CRISPR/Cas9 genetic screening approach identified only the pyrimidine metabolism pathway as modifying cancer cell sensitivity to NUC-1031. Low deoxycytidine kinase expression in tumour biopsies from patients treated with gemcitabine, assessed by immunostaining and image analysis, correlates with a poor prognosis, but there is no such correlation in tumour biopsies from a Phase I cohort treated with NUC-1031.

Randomized, phase I/II study of gemcitabine plus IGF-1R antagonist (MK-0646) versus gemcitabine plus erlotinib with and without MK-0646 for advanced pancreatic adenocarcinoma.

BACKGROUND: Binding of insulin-like growth factor-I (IGF-1) to its receptor (IGF-1R) initiates downstream signals that activate PI3K/Akt/mTOR and MEK/Erk pathways, which stimulate cancer cell proliferation and induce drug resistance. Cross talk between IGF-1R and epidermal growth factor receptor (EGFR) mediates resistance to anti-EGFR agents. We studied safety, tolerability, and outcomes of MK-0646, IGF-1 monoclonal antibody, in combination with gemcitabine (G) ± erlotinib (E) in metastatic pancreatic cancer. METHODS: Our study included a phase I dose escalation and phase II randomization and expansion cohorts. A 3 + 3 dose escalation protocol was used to determine MK-0646 maximum tolerable dose (MTD) in combination with G ± E standard doses. For phase II, patients were randomized to arm A (G + MK), arm B (G + MK + E), or arm C (G + E). Primary endpoint was progression-free survival (PFS). Secondary endpoints were overall survival (OS), disease control rate, toxicity, and correlation between OS and IGF-1 in patients treated with MK-0646. RESULTS: MK-0646 MTD was 10 mg/kg in combination with G and 5 mg/kg in combination with G + E. In randomization cohort, 15 patients were treated in each arm. Disease control rates were 50, 60, and 40% respectively. PFS was not different between the three arms. OS was significantly different between arm A (10.4 months) and C (5.7 months) (P = 0.02). However, addition of erlotinib in arm B yielded no OS benefit compared to arm A (P = 0.6). Plasma and tissue IGF-1 levels did not correlate with OS (P = 0.64, 0.87). Grade 3-4 toxicity during phase II cohorts were neutropenia (10/arm A, 14/arm B, 5/arm C), leukopenia (5/A, 5/B, 7/C), thrombocytopenia (8/A, 9/B, 2/C), hyponatremia (1/A, 3/B), and hyperglycemia (8/A, 1/B). CONCLUSIONS: MK-0646 was tolerable in combination with G and associated with improvement in OS but not PFS as compared with G + E. Tissue and serum IGF-1 did not correlate with clinical outcome. TRIAL REGISTRATION: This trial is registered in ClinicalTrial.gov under the Identifier NCT00769483 and registration date was October 9, 2008.

Licoricidin enhances gemcitabine-induced cytotoxicity in osteosarcoma cells by suppressing the Akt and NF-κB signal pathways.

Osteosarcoma (OS) is the most common bone malignancy in children and adolescents. Combined treatments of anti-cancer drugs can remarkably improve chemotherapeutic outcomes. Gemcitabine and licoricidin both have potential anti-tumor activity in several cancers. However, the combined therapeutic efficiency of gemcitabine and licoricidin for OS has not been explored. Here, we found that licoricidin or gemcitabine inhibited OS cell viability in a dose-dependent manner. Cotreatment with licoricidin and gemcitabine enhanced gemcitabine-induced cytotoxicity in OS cells. Licoricidin suppressed activation of the Akt and nuclear factor-kappa B (NF-κB) pathways. Gemcitabine had no effect on Akt signal, but facilitated the activation of NF-κB signal in OS cells. Moreover, combined treatment of licoricidin and gemcitabine markedly curbed the activation of Akt and NF-κB pathways in OS cells. Inhibition of the Akt and NF-κB pathways enhanced gemcitabine-induced cytotoxicity in OS cells. In vivo assay further manifested that licoricidin enhanced gemcitabine-induced cytotoxicity in tumor xenograft models of OS via inactivation of the Akt and NF-κB pathways. In conclusion, licoricidin enhanced gemcitabine-induced cytotoxicity in OS cells by inactivation of the Akt and NF-κB pathways in vitro and in vivo.

A mixed treatment comparison of toxicity of gemcitabine combined with different targeted drugs in the treatment of advanced or metastatic pancreatic cancer.

The mixed treatment comparison study was performed in order to compare the toxicities of Gemcitabine and different targeted drug combinations in the treatment of advanced/metastatic pancreatic cancer (PC). Searches were performed from the inception of PubMed and Cochrane Library databases to February 2017. This study included randomized controlled trials (RCTs) of Gemcitabine and different targeted drug combinations in the treatment of advanced/metastatic PC. Odds ratio (OR) values were calculated by direct and indirect comparisons, and the surface under the cumulative ranking curves (SUCRA) were drawn. A total of six RCTs were finally incorporated into the study. These studies included six therapy regimens: Gemcitabine + Axitinib, Gemcitabine + Trametinib, Gemcitabine + Sorafenib, Gemcitabine + Bevacizumab, Gemcitabine + Erlotinib and Gemcitabine + Tipifarnib. The results showed that Gemcitabine + Axitinib combinations showed lower incidence rates of rashes (all grades) in comparison to Gemcitabine + Trametinib and Gemcitabine + Erlotinib combinations. Compared with Gemcitabine+ Trametinib combinations, Gemcitabine + Axitinib combinations showed lower incidence rates of diarrhea (grade ≥ 3). Moreover, the cluster analyses results revealed that Gemcitabine + Axitinib combinations and Gemcitabine + Sorafenib combinations showed lower incidence rates of hematotoxicity, while Gemcitabine + Axitinib combinations showed lower incidence rates of non-hematotoxicity. Collectively, the data provided strong evidence of Gemcitabine + Axitinib combinations showing lower incidence rates of non-hematotoxicity, and Gemcitabine + Axitinib and Gemcitabine + Sorafenib combinations may have lower incidence rates of hematotoxicity in the treatment of advanced/metastatic PC.

Linc-DYNC2H1-4 promotes EMT and CSC phenotypes by acting as a sponge of miR-145 in pancreatic cancer cells.

The acquisition of epithelial-mesenchymal transition (EMT) and/or existence of a sub-population of cancer stem-like cells (CSC) are associated with malignant behavior and chemoresistance. To identify which factor could promote EMT and CSC formation and uncover the mechanistic role of such factor is important for novel and targeted therapies. In the present study, we found that the long intergenic non-coding RNA linc-DYNC2H1-4 was upregulated in pancreatic cancer cell line BxPC-3-Gem with acquired gemcitabine resistance. Knockdown of linc-DYNC2H1-4 decreased the invasive behavior of BxPC-3-Gem cells while ectopic expression of linc-DYNC2H1-4 promoted the acquisition of EMT and stemness of the parental sensitive cells. Linc-DYNC2H1-4 upregulated ZEB1, the EMT key player, which led to upregulation and downregulation of its targets vimentin and E-cadherin respectively, as well as enhanced the expressions of CSC makers Lin28, Nanog, Sox2 and Oct4. Linc-DYNC2H1-4 is mainly located in the cytosol. Mechanically, it could sponge miR-145 that targets ZEB1, Lin28, Nanog, Sox2, Oct4 to restore these EMT and CSC-associated genes expressions. We proved that MMP3, the nearby gene of linc-DYNC2H1-4 in the sense strand, was also a target of miR-145. Downregulation of MMP3 by miR-145 was reverted by linc-DYNC2H1-4, indicating that competing with miR-145 is one of the mechanisms for linc-DYNC2H1-4 to regulate MMP3. In summary, our results explore the important role of linc-DYNC2H1-4 in the acquisition of EMT and CSC, and the impact it has on gemcitabine resistance in pancreatic cancer cells.

The novel phospholipid mimetic KPC34 is highly active against preclinical models of Philadelphia chromosome positive acute lymphoblastic leukemia.

Philadelphia chromosome positive B cell acute lymphoblastic leukemia (Ph+ ALL) is an aggressive cancer of the bone marrow. The addition of tyrosine kinase inhibitors (TKIs) has improved outcomes but many patients still suffer relapse and novel therapeutic agents are needed. KPC34 is an orally available, novel phospholipid conjugate of gemcitabine, rationally designed to overcome multiple mechanisms of resistance, inhibit the classical and novel isoforms of protein kinase C, is able to cross the blood brain barrier and is orally bioavailable. KPC34 had an IC50 in the nanomolar range against multiple ALL cell lines tested but was lowest for Ph+ lines. In mice bearing either naïve or resistant Ph+ ALL, KPC34 treatment resulted in significantly improved survival compared to cytarabine and gemcitabine. Treatment with KPC34 and doxorubicin was more effective than doxorubicin and cytarabine. Mice with recurrence of their ALL after initial treatment with cytarabine and doxorubicin saw dramatic improvements in hind limb paralysis after treatment with KPC34 demonstrating activity against established CNS disease. Consistent with this KPC34 was better than gemcitabine at reducing CNS leukemic burden. These promising pre-clinical results justify the continued development of KPC34 for the treatment of Ph+ALL.

Unique cellular interactions between pancreatic cancer cells and the omentum.

Pancreatic cancer is a common cause of cancer-related mortality. Omental spread is frequent and usually represents an ominous event, leading to patient death. Omental metastasis has been studied in ovarian cancer, but data on its role in pancreatic cancer are relatively scarce and the molecular biology of this process has yet to be explored. We prepared tissue explants from human omental fat, and used conditioned medium from the explants for various in vitro and in vivo experiments designed to evaluate pancreatic cancer development, growth, and survival. Mass spectrometry identified the fat secretome, and mRNA array identified specific fat-induced molecular alternations in tumor cells. Omental fat increased pancreatic cancer cellular growth, migration, invasion, and chemoresistance. We identified diverse potential molecules secreted by the omentum, which are associated with various pro-tumorigenic biological processes. Our mRNA array identified specific omental-induced molecular alternations that are associated with cancer progression and metastasis. Omental fat increased the expression of transcription factors, mRNA of extracellular matrix proteins, and adhesion molecules. In support with our in vitro data, in vivo experiments demonstrated an increased pancreatic cancer tumor growth rate of PANC-1 cells co-cultured for 24 hours with human omental fat conditioned medium. Our results provide novel data on the role of omental tissue in omental metastases of pancreatic cancer. They imply that omental fat secreted factors induce cellular reprogramming of pancreatic cancer cells, resulting in increased tumor aggressiveness. Understanding the mechanisms of omental metastases may enable us to discover new potential targets for therapy.

Genotoxicity kinetics in murine normoblasts as an approach for the in vivo action of difluorodeoxycytidine.

PURPOSE: This study analyzed the kinetics of in vivo micronucleus induction in normoblasts by determining the kinetics of difluorodeoxycytidine (dFdC)-induced micronucleated polychromatic erythrocytes (MN-PCEs) in the peripheral blood of mice. The kinetic indexes of MN-PCE induction of dFdC were correlated with the previously reported mechanisms DNA damage induction by this compound. In general, this study aimed to establish an in vivo approach for discerning the processes underlying micronucleus induction by antineoplastic agents or mutagens in general. METHODS: The frequencies of PCEs and MN-PCEs in the peripheral blood of mice were determined prior to treatment and after treatment using dFdC at doses of 95, 190, or 380 µmol/kg at 8 h intervals throughout a 72 h post-treatment. RESULTS: The area beneath the curve (ABC) for MN-PCE induction as a function of time, which is an index of the total effect, indicated that the dose response was directly proportional and that the effect of dFdC on micronucleus induction was reduced compared with that of aneuploidogens and monofunctional and bifunctional alkylating agents but increased compared with that of promutagens, which is consistent with our previous results. The ABC showed a single peak with a small broadness index, which indicates that dFdC has a single mechanism or concomitant mechanisms for inducing DNA breaks. The time of the relative maximal induction (T rmi) indicated that dFdC requires more time to achieve MN-PCE induction compared with aneugens and monofunctional and bifunctional alkylating agents, although it requires a similar time to achieve MN-PCE induction as azacytidine, which is consistent with evidence showing that both agents must be incorporated into DNA for their action to be realized. The timing of maximal cytotoxicity observed with the lowest dFdC dose was correlated with the timing of the main genotoxic effect. However, early and late cytotoxic effects were detected, and these effects were independent of the genotoxic response. CONCLUSIONS: A correlation analysis indicated that dFdC appears to induce MN-PCEs through only one mechanism or mechanisms that occur concomitantly, which could be explained by the previously reported concurrent inhibitory effects of dFdC on DNA polymerase alpha, polymerase epsilon, and/or topoisomerase. The timing of maximal cytotoxicity was correlated with the timing of maximal genotoxicity; however, an early cytotoxic effect that appeared to occur prior to the incorporation of dFdC into DNA was likely related to a previously reported inhibitory effect of dFdC on thymidylate synthase and/or ribonucleotide reductase.

Preclinical Evaluation of the Short-Term Toxicity of 4-(N)-Docosahexaenoyl 2´, 2´- Difluorodeoxycytidine (DHA-dFdC).

PURPOSE: This study was designed to test the short-term toxicity of DHA-dFdC in a mouse model and its efficacy in a mouse model of leukemia at or below its repeat-dose maximum tolerated dose (RD-MTD). METHOD: A repeat-dose dose-ranging toxicity study was designed to determine the tolerability of DHA-dFdC when administered to DBA/2 mice by intravenous (i.v.) injection on a repeat-dose schedule (i.e. injections on days 0, 3, 7, 10, and 13). In order to determine the effect of a lethal dose of DHA-dFdC, mice were injected i.v. with three doses of DHA-dFdC at 100 mg/kg on days 0, 3, and 5 (i.e. a lethal-RD). The body weight of mice was recorded two or three times a week. At the end of the study, major organs (i.e. heart, liver, spleen, kidneys, lung, and pancreas) of mice that received the lethal-RD or RD-MTD were weighed, and blood samples were collected for analyses. Finally, DHA-dFdC was i.v. injected into DBA/2 mice with syngeneic L1210 mouse leukemia cells to evaluate its efficacy at or below RD-MTD. RESULTS: The RD-MTD of DHA-dFdC is 50 mg/kg. At 100 mg/kg, a lethal-RD, DHA-dFdC decreases the weights of mouse spleen and liver and significantly affected certain blood parameters (i.e. white blood cells, lymphocytes, eosinophils, and neutrophil segmented). At or below its RD-MTD, DHA-dFdC significantly prolonged the survival of L1210 leukemia-bearing mice. CONCLUSION: DHA-dFdC has dose-dependent toxicity, affecting mainly spleen at a lethal-RD. At or below its RD-MTD, DHA-dFdC is effective against leukemia in a mouse model.