Nonlinear association between body mass index and overall survival in advanced NSCLC patients treated with immune checkpoint blockade.

BACKGROUND: We investigated the association of body mass index (BMI) modeled as a continuous variable with survival outcomes in advanced non-small cell lung cancer (NSCLC) patients treated with immune checkpoint inhibitors (ICI). METHODS: We performed a single-institution retrospective analysis of consecutively diagnosed locally advanced or metastatic NSCLC patients treated with single-agent ICI in the first line or recurrent setting. The primary outcome was overall survival (OS). Secondary outcomes were progression-free survival (PFS) and objective response rate (ORR). BMI was modeled using a four-knot restricted cubic spline. Multiple Cox regression was used for survival analysis. RESULTS: Two hundred patients were included (female 54%; never smoker 12%). Adenocarcinoma was the most common histology (61%). Median age was 67 years, median BMI was 25.9 kg/m2, and 65% of patients had Eastern Cooperative Oncology Group performance status (ECOG PS) of 0-1. On multivariable analysis, only BMI and ECOG PS were independently associated with OS (p < 0.01). Mortality risk decreased as the BMI increased from 20 to 30 kg/m2 (HR 0.49, 95% CI 0.28-0.84); however, it was reversed as the BMI surpassed ~ 30 kg/m2. Compared to ECOG PS ≥ 2, patients with ECOG PS of 0-1 had a longer OS (HR 0.42, 95% CI 0.28-0.63). Similar trends were observed with PFS and ORR, but the strength of the association was weaker. CONCLUSION: We observed a nonlinear association between BMI and OS following treatment with ICI in advanced NSCLC. Risk of death increases at both extremes of BMI with a nadir that exists around 30 kg/m2.

SFPQ and Its Isoform as Potential Biomarker for Non-Small-Cell Lung Cancer.

Cancer markers are measurable molecules in the blood or tissue that are produced by tumor cells or immune cells in response to cancer progression. They play an important role in clinical diagnosis, prognosis, and anti-drug monitoring. Although DNA, RNA, and even physical images have been used, proteins continue to be the most common marker. There are currently no specific markers for lung cancer. Metastatic lung cancer, particularly non-small-cell lung cancer (NSCLC), is one of the most common causes of death. SFPQ, YY1, RTN4, RICTOR, LARP6, and HELLS are expressed at higher levels in cells from NSCLC than in control or cells from inflammatory diseases. SFPQ shows the most difference between the three cell types. Furthermore, the cytoplasmic isoform of SFPQ is only found in advanced cancers. We have developed ELISAs to detect SFPQ and the long and short isoforms. Evidence has shown that the short isoform exists primarily in cancers. Furthermore, immunocytometry studies and IHC analysis have revealed that SFPQ levels are consistent with ELISA results. In addition, enhanced DNA methylation in the SFPQ gene may facilitate the SFPQ expression differences between control and cancer cells. Considering this, elevated SFPQ level and the isoform location could serve as a cancer diagnostic and prognostic marker.

4Ei-10 interdiction of oncogenic cap-mediated translation as therapy for non-small cell lung cancer.

In order to suppress 5' cap-mediated translation a highly available inhibitor of the interaction between the 5' mRNA cap and the eIF4E complex has been developed. 4Ei-10 is a member of the class of ProTide compounds and has elevated membrane permeability and is a strong active chemical antagonist for eIF4E. Once taken up by cells it is converted by anchimeric activation of the lipophilic 2-(methylthio) ethyl protecting group and after that Hint1 P-N bond cleavage to N7-(p-chlorophenoxyethyl) guanosine 5'-monophosphate (7-Cl-Ph-Ethyl-GMP). Using this powerful interaction, it has been demonstrated that 4Ei-10 inhibits non-small cell lung cancer (NSCLC) cell growth. In addition, treatment of NSCLC cells with 4Ei-10 results in suppression of translation and diminished expression of a cohort of cellular proteins important to maintaining the malignant phenotype and resisting apoptosis such as Bcl-2, survivin, and ornithine decarboxylase (ODC). Finally, as a result of targeting the translation of anti-apoptotic proteins, NSCLC cells are synergized to be more sensitive to the existing anti-neoplastic treatment gemcitabine currently used in NSCLC therapy.

Synchronous breast carcinoma and peritoneal mesothelioma.

Breast cancer may be associated with other primary cancers via germline mutations; however, sporadic occurrences of other malignancies are rare. With increased use of advanced breast cancer imaging, including MRI and PET/CT, other incidental synchronous cancers are increasingly identified. Such cases can represent unique diagnostic and treatment challenges. Here, we present a case of a young woman diagnosed with primary breast cancer who underwent imaging studies identifying an incidental primary peritoneal mesothelioma.

Survival of Lung Cancer Patients Dependent on the LOH Status for DMP1, ARF, and p53.

Lung cancer is the leading cause of cancer deaths in the world, and accounts for more solid tumor deaths than any other carcinomas. The prognostic values of DMP1, ARF, and p53-loss are unknown in lung cancer. We have conducted survival analyses of non-small cell lung cancer (NSCLC) patients from the University of Minnesota VA hospital and those from the Wake Forest University Hospital. Loss of Heterozygosity (LOH) for hDMP1 was found in 26 of 70 cases (37.1%), that of the ARF/INK4a locus was found in 33 of 70 (47.1%), and that of the p53 locus in 43 cases (61.4%) in the University of Minnesota samples. LOH for hDMP1 was associated with favorable prognosis while that of p53 predicted worse prognosis. The survival was much shorter for ARF-loss than INK4a-loss, emphasizing the importance of ARF in human NSCLC. The adverse effect of p53 LOH on NSCLC patients' survival was neutralized by simultaneous loss of the hDMP1 locus in NSCLC and breast cancer, suggesting the possible therapy of epithelial cancers with metastatic ability.

4EGI-1 represses cap-dependent translation and regulates genome-wide translation in malignant pleural mesothelioma.

Deregulation of cap-dependent translation has been implicated in the malignant transformation of numerous human tissues. 4EGI-1, a novel small-molecule inhibitor of cap-dependent translation, disrupts formation of the eukaryotic initiation factor 4F (eIF4F) complex. The effects of 4EGI-1-mediated inhibition of translation initiation in malignant pleural mesothelioma (MPM) were examined. 4EGI-1 preferentially inhibited cell viability and induced apoptosis in MPM cells compared to normal mesothelial (LP9) cells. This effect was associated with hypophosphorylation of 4E-binding protein 1 (4E-BP1) and decreased protein levels of the cancer-related genes, c-myc and osteopontin. 4EGI-1 showed enhanced cytotoxicity in combination with pemetrexed or gemcitabine. Translatome-wide polysome microarray analysis revealed a large cohort of genes that were translationally regulated upon treatment with 4EGI-1. The 4EGI-1-regulated translatome was negatively correlated to a previously published translatome regulated by eIF4E overexpression in human mammary epithelial cells, which is in agreement with the notion that 4EGI-1 inhibits the eIF4F complex. These data indicate that inhibition of the eIF4F complex by 4EGI-1 or similar translation inhibitors could be a strategy for treating mesothelioma. Genome wide translational profiling identified a large cohort of promising target genes that should be further evaluated for their potential significance in the treatment of MPM.

Small-molecule inhibition of oncogenic eukaryotic protein translation in mesothelioma cells.

Deranged cap-mediated translation is implicated in the genesis, maintenance and progression of many human cancers including mesothelioma. In this study, disrupting the eIF4F complex by antagonizing the eIF4E-mRNA-cap interaction is assessed as a therapy for mesothelioma. Mesothelioma cells were treated with 4Ei-1, a membrane permeable prodrug that when converted to the active drug, 7-benzyl guanosine monophosphate (7Bn-GMP) displaces capped mRNAs from the eIF4F complex. Colony formation was measured in mesothelioma treated with 4Ei-1 alone or combined with pemetrexed. Proliferation was examined in cells treated with 4Ei-1. Binding to a synthetic cap-analogue was used to study the strength of eIF4F complex activation in lysates exposed to 4Ei-1. 4Ei-1 treatment resulted in a dose dependent decrease in colony formation and cell viability. Combination therapy of 4Ei-1 with pemetrexed further reduced colony number. Formation of eIF4F cap-complex decreased in response to 4Ei-1 exposure. 4Ei-1 is a novel prodrug that reduces proliferation, represses colony formation, diminishes association of eIF4F with the mRNA cap, and sensitizes mesothelioma cells to pemetrexed.

Cancer sensitizing effect of deazaflavin analogs is associated with increased intracellular drug accumulation.

As part of our efforts geared towards developing mechanism-based cancer sensitizing agents, we have previously synthesized and characterized novel deazaflavin analogs as potent tyrosyl DNA phosphodiesterase 2 (TDP2) inhibitors for combination treatments with topoisomerase II (TOP2) poisons. Interestingly, the sensitizing effect of a few analogs toward TOP2 poison etoposide (ETP) was associated with a significant increase in intracellular drug accumulation, which could be an alternative mechanism to boost the clinical efficacy of ETP in cancer chemotherapies. Hence, we evaluated more deazaflavin TDP2 inhibitors for their impact on drug retention in cancer cells. We found that all but one tested TDP2 inhibitors substantially increased the ETP retention in DT40 cells. Particularly, we identified an exceptionally potent analog, ZW-1226, which at 3 nM increased the intracellular ETP by 13-fold. Significantly, ZW-1226 also stimulated cellular accumulation of two other anticancer drugs, TOP2 poison teniposide and antifolate pemetrexed, and produced an effect more pronounced than those of ABC transporter inhibitors verapamil and elacridar in human leukemic CCRF-CEM cells toward ETP. Lastly, ZW-1226 potentiated the action of ETP in the sensitive human CCRF-CEM cells and a few resistant non-small-cell lung cancer (NSCLC) cells, including H460 and H838 cells. Collectively, the results of this study strongly suggest that deazaflavin analog ZW-1226 could be an effective cancer sensitizing agent which warrants further investigation.

A tri-specific killer engager against mesothelin targets NK cells towards lung cancer.

New treatments are required to enhance current therapies for lung cancer. Mesothelin is a surface protein overexpressed in non-small cell lung cancer (NSCLC) that shows promise as an immunotherapeutic target in phase I clinical trials. However, the immunosuppressive environment in NSCLC may limit efficacy of these therapies. We applied time-of-flight mass cytometry to examine the state of circulating mononuclear cells in fourteen patients undergoing treatment for unresectable lung cancer. Six patients had earlier stage NSCLC (I-IVA) and eight had highly advanced NSCLC (IVB). The advanced NSCLC patients relapsed with greater frequency than the earlier stage patients. Before treatment, patients with very advanced NSCLC had a greater proportion of CD14- myeloid cells than patients with earlier NSCLC. These patients also had fewer circulating natural killer (NK) cells bearing an Fc receptor, CD16, which is crucial to antibody-dependent cellular cytotoxicity. We designed a high affinity tri-specific killer engager (TriKE®) to enhance NK cytotoxicity against mesothelin+ targets in this environment. The TriKE consisted of CD16 and mesothelin binding elements linked together by IL-15. TriKE enhanced proliferation of lung cancer patient NK cells in vitro. Lung cancer lines are refractory to NK cell killing, but the TriKE enhanced cytotoxicity and cytokine production by patient NK cells when challenged with tumor. Importantly, TriKE triggered NK cell responses from patients at all stages of disease and treatment, suggesting TriKE can enhance current therapies. These pre-clinical studies suggest mesothelin-targeted TriKE has the potential to overcome the immunosuppressive environment of NSCLC to treat disease.

Exceptional response to afatinib in a patient with persistent G719A EGFR-mutant NSCLC.

We present a patient with metastatic NSCLC harboring a compound EGFR mutation with co-occurring G719A and T790M mutation. T790M mutation was treatment emergent mutation when patient was on early generation tyrosine kinase inhibitors. Initial Guardant 360 showed that G719A was the dominant clone. Following, osimertinib, the patient had only a radiographic disease stabilization and then developed both clinical and radiographic progression. On progression, T790M was undetectable but G719A continued to be the dominant clone. Subsequent administration of afatinib led to a clinical and radiological response. To our knowledge, this is the first case report describing co-occurrence of EGFR G719A and T790M mutations and the clonal evolution during treatment with anti-EGFR therapies.

Morphine-induced epidermal growth factor pathway activation in non-small cell lung cancer.

BACKGROUND: Epidermal growth factor receptor (EGFR) is coactivated by the µ-opioid receptor (MOR), expressed on non-small cell lung cancer (NSCLC) cells and human lung cancer. We hypothesized that clinically used opioid analgesics that are MOR agonists coactivate EGFR, resulting in growth- and survival-promoting signaling. METHODS: We used H2009, a human adenocarcinoma NSCLC cell line, with constitutive EGFR phosphorylation, which showed increased expression of MOR and the δ-opioid receptor by reverse transcriptase polymerase chain reaction. We used Western immunoblotting, magnetic bead-based Bio-Plex cytokine assay, immunofluorescent staining, BrdU incorporation enzyme-linked immunosorbent assay, and BioCoat™ Matrigel™ invasion assay to examine cell signaling, cytokine expression, colocalization of MOR and EGFR in human lung cancer, and cell proliferation and invasion, respectively. RESULTS: Similar to epidermal growth factor (EGF), morphine stimulated phosphorylation of EGFR, Akt/protein kinase B (Akt), and mitogen-activated protein kinase/extracellular signal regulated kinase (MAPK/ERK) signaling in H2009 cells. Opioid receptor (OR) antagonist, naloxone, EGFR tyrosine kinase inhibitor, erlotinib, and silencing of MOR and δ-opioid receptor abrogated morphine- and EGF-induced phosphorylation of signaling, suggestive of OR-mediated coactivation of EGFR. H2009 cells secreted significantly higher levels of cytokines compared with control Beas2B epithelial cells. H2009-conditioned medium stimulated MOR expression in Beas2B cells, suggesting that cytokines secreted by H2009 may be associated with increased OR expression in H2009. We observed colocalization of EGFR and MOR, in human NSCLC tissue. Functionally, morphine- and EGF-induced proliferation and invasion of H2009 cells was ameliorated by naloxone as well as erlotinib. CONCLUSION: Morphine-induced phosphorylation of EGFR occurs via ORs, leading to downstream MAPK/ERK, Akt phosphorylation, cell proliferation, and increased invasion. Notably, ORs are also associated with EGF-induced phosphorylation of EGFR. Increased coexpression of MOR and EGFR in human lung cancer suggests that morphine may have a growth-promoting effect in lung cancer.

Anti-proliferative effects of simocyclinone D8 (SD8), a novel catalytic inhibitor of topoisomerase II.

INTRODUCTION: Simocyclinone D-8 (SD8), a semi-synthetic compound derived from yeast, has been shown to decrease the proliferation of MCF-7 breast cancer cells. It has been shown to be a potent bacterial DNA gyrase inhibitor, a homologue of human topoisomerase II (hTopoII). We tested SD8 activity alone and in combination with cisplatin against malignant mesothelioma (MM) and non-small cell cancer (NSCLC) cell lines. METHODS: Inhibition of hTopoII supercoiling function by SD8 and a known hTopoII poison, etoposide, were done by in vitro assay using purified hTopoII and kinetoplast DNA as the substrate. The DNA products were analyzed by agarose gel electrophoresis after treatment with increasing concentrations of each drug. Mesothelioma cell lines (H2373, H2461 and H2596) and NSCLC cell lines (H2030, H460, and H2009) grown in RPMI with 10% calf serum were used. Non-malignant mesothelial cells, LP9, were grown in 1:1 ratio of MCDB:199E medium supplemented with 15% calf serum, 0.4 microg/mL hydrocortisone, and 15 ng/mL epidermal grown factor. Cell proliferation assays were performed in 96-well plates using the CCK-8 kit (Dojindo inc.). Cells were treated for 72 h with various SD8 concentrations and controls containing equal volume of the vehicle, DMSO. Treated cells were assayed for the induction of apoptosis with poly ADP-ribose polymerase-1 (PARP) cleavage assay. RESULTS: Biochemical assays revealed that the IC(50) for hTopoII inhibition was 100 microM for SD8 and 400 microM for etoposide. SD8 inhibited hTopoII function without inducing DNA cleavage events. SD8 inhibited the growth of NSCLC and Mesothelioma cells with IC(50) ranging from 75-125 microM. Furthermore, SD8 was not toxic to non-transformed primary mesothelial cell line, LP9 at the IC(50) doses. SD8 induced apoptosis in all cell lines tested. CONCLUSIONS: SD8 inhibits hTopoII in vitro without inducing DNA strands breaks and has significant activity against NSCLC and MM cell lines. While doses required for SD8 anticancer activity are unlikely to be achieved in vivo, chemical modifications to SD8 to increase its potency could lead to improved therapies for these diseases.

p16(INK4a) Peptide mimetics identified via virtual screening.

The transition from G1 to S phase in the cell cycle is highly regulated by Cdk4 and Cdk6, which in turn is inhibited by the tumor suppressor p16(INK4a). Replacement of lost p16(INK4a) activity in cancer cells via gene therapy has worked in vivo to decrease tumor progression; however, practical issues limit gene therapy applications at this time. Here, we report the discovery of compounds that inhibit Cdk4 and Cdk6 activity. The NMR structure of a peptide that exhibits p16(INK4a) activity was solved and combined with known functional data to generate a pharmacophore that was used to mine the NCI chemical database. The hits were filtered utilizing the program Qikprop. Four compounds were subsequently shown to inhibit Cdk4 and/or Cdk6 with IC(50) in the muM range. These compounds form lead compounds upon which further cell cycle inhibitors can be developed.

Blood Outgrowth Endothelial Cells as a Cellular Carrier for Oncolytic Vesicular Stomatitis Virus Expressing Interferon-ß in Preclinical Models of Non-Small Cell Lung Cancer.

Oncolytic viruses have demonstrated efficacy in numerous tumor models including non-small cell lung cancer (NSCLC). One limitation of viral therapy for metastatic lung cancer is that systemic administration can be hindered by complement and antiviral immunity. Thus, we investigated whether ex vivo-infected blood outgrowth endothelial cells (BOECs) could be used to deliver VSV-IFNß in preclinical models of NSCLC. BOECs were obtained from human donors or C57/Bl6 mice. VSV was engineered to produce GFP or IFNß. Human and murine BOECs could be infected by VSV-GFP and VSV-IFNß. Infected BOECs resulted in killing of NSCLC cells in vitro and shielded VSV-IFNß from antibody neutralization. Mouse BOECs localized to lungs of mice bearing syngeneic LM2 lung tumors, and infected murine BOECs reduced tumor burden in this model. In an immune-deficient A549 xenograft model, mice treated with VSV-IFNß-infected human BOECs exhibited superior antitumor activity and survival of mice (n = 10, P < .05 compared to VSV-IFNß alone). We conclude that BOECs can be used as a carrier for delivery of oncolytic VSV-IFNß. This may be an effective strategy for clinical translation of oncolytic virotherapy for patients with metastatic NSCLC.

Repression of oncogenic cap-mediated translation by 4Ei-10 diminishes proliferation, enhances chemosensitivity and alters expression of malignancy-related proteins in mesothelioma.

Activated cap-dependent translation promotes cancer by stimulating translation of mRNAs encoding malignancy-promoting proteins. The nucleoside monophosphate Protide, 4Ei-10, undergoes intracellular uptake and conversion by Hint1 to form 7-Cl-Ph-Ethyl-GMP. 7-Cl-Ph-Ethyl-GMP is an analog of cap and inhibits protein translation by binding and sequestering eIF4E thus blocking eIF4E from binding to the mRNA cap. The effects of inhibiting translation initiation by disruption of the eIF4F complex with 4Ei-10 were examined in malignant mesothelioma (MM). In a cell-free assay system, formation of the eIF4F complex was disabled in response to exposure to 4Ei-10. Treatment of MM with 4Ei-10 resulted in decreased cell proliferation, increased sensitivity to pemetrexed and altered expression of malignancy-related proteins. In light of these findings, suppression of translation initiation by small molecule inhibitors like 4Ei-10 alone or in combination with pemetrexed represents an encouraging strategy meriting further evaluation in the treatment of MM.

Minimally invasive versus open Roux-en-Y gastric bypass: effect on immune effector cells.

BACKGROUND: Minimally invasive surgery (MIS) has several potential benefits compared with the open approach, including potentially less perioperative immunosuppression. Data characterizing the differential stress responses have been limited to serum cytokine analyses and animal studies. We hypothesized that the open approach to Roux-en-Y gastric bypass (RYGB) has a more deleterious, negative, quantifiable effect on the peripheral blood mononuclear cells than does the MIS approach. METHODS: Patients undergoing open and MIS RYGB for morbid obesity had blood samples collected preoperatively and postoperatively on days 1 and 2 and at the first follow-up visit. The peripheral blood mononuclear cells were isolated and analyzed for phenotype using flow cytometry, natural killer cell cytotoxicity using 51-chromium release assay, and gene expression using Affymetrix U133 Plus 2.0 microarray. RESULTS: Patient age and body mass index were similar between the 2 groups. Postoperatively, differences within the open group were seen for CD3+/CD16- (T lymphocytes), CD3-/CD16+ (natural killer cells), CD3+/CD4+ (T-helper lymphocytes), and CD4/CD8 subsets (P<.05). No differences were seen within the open group CD3+/CD8+ (cytotoxic T lymphocytes) or within the MIS subsets. Between the 2 approaches, no phenotypic differences were found, except for the postoperative day 1 CD3+/CD16- (P<.05). Within each group, significant decreases were found in cytotoxicity on days 1 and 2 compared with preoperatively (P<.05). The cytotoxicity seen after MIS had returned to the preoperative levels at the first follow-up visit, but the cytotoxicity after open RYGB had not (P<.05). Between the 2 groups, the open group had greater cytotoxic decreases than did the MIS group at postoperative days 1 and 2 (P<.05). Microarray analysis of the preoperative (n=20) and day 2 (n=20) specimens identified a 20-gene signature that correlated with the surgical approach. CONCLUSION: Open RYGB surgery causes greater inhibition of innate immunity than does MIS. This inhibition was not accounted for by phenotypic changes. Gene expression changes from surgical stress might represent the molecular basis of this differential immune response.

Gene silencing for epidermal growth factor receptor variant III induces cell-specific cytotoxicity.

Epidermal growth factor receptor variant III (EGFRvIII) is a constitutively active mutant form of EGFR that is expressed in 40% to 50% of gliomas and several other malignancies. Here, we describe the therapeutic effects of silencing EGFRvIII on glioma cell lines in vitro and in vivo. A small interfering RNA molecule against EGFRvIII was introduced into EGFRvIII-expressing glioma cells (U87Delta) by electroporation resulting in complete inhibition of expression of EGFRvIII as early as 48 h post-treatment. During EGFRvIII silencing, a decrease in the proliferation and invasiveness of U87Delta cells was accompanied by an increase in apoptosis (P < 0.05). Notably, EGFRvIII silencing inhibited the signal transduction machinery downstream of EGFRvIII as evidenced by decreases in the activated levels of Ras and extracellular signal-regulated kinase. A lentivirus capable of expressing anti-EGFRvIII short hairpin RNA was also able to achieve progressive silencing of EGFRvIII in U87Delta cells in addition to inhibiting cell proliferation, invasiveness, and colony formation in a significant manner (P < 0.05). Silencing EGFRvIII in U87Delta cultures with this virus reduced the expression of factors involved in epithelial-mesenchymal transition including N-cadherin, beta-catenin, Snail, Slug, and paxillin but not E-cadherin. The anti-EGFRvIII lentivirus also affected the cell cycle progression of U87Delta cells with a decrease in G(1) and increase in S and G(2) fractions. In an in vivo model, tumor growth was completely inhibited in severe combined immunodeficient mice (n = 10) injected s.c. with U87Delta cells treated with the anti-EGFRvIII lentivirus (P = 0.005). We conclude that gene specific silencing of EGFRvIII is a promising strategy for treating cancers that contain this mutated receptor.

JAK/STAT inhibition with ruxolitinib enhances oncolytic virotherapy in non-small cell lung cancer models.

The concept of using viruses to treat cancer has now shown proof of concept in several recent clinical trials, leading to the first FDA approval of virotherapy for melanoma last year. Vesicular stomatitis virus (VSV) is a promising oncolytic virus that has inhibitory effects on a number of cancer types including non-small cell lung cancer. One of the major mechanisms of resistance to VSV infection is the type I interferon (IFN) response, leading to the development of VSV expressing IFNß which will lead to resistance of viral replication in normal cells which have intact IFN signaling but allow replication in cancer cells with defective IFNß signaling. However, some cancer cells have intact or partially intact IFN signaling pathways leading to resistance to virotherapy. Here we utilized JAK/STAT inhibitor, ruxolitinib, in combination with VSV-IFNß to see if inhibition of JAK/STAT signaling will enhance VSV-IFNß therapy for lung cancer. We used five human and two murine NSCLC cell lines in vitro, and the combination treatment was assayed for cytotoxicity. We performed western blots on treated cells to see the effects of ruxolitinib on JAK/STAT signaling and PDL-1 expression in treated cells. Finally, the combination of VSV-IFNß and ruxolitinib was tested in an immune competent murine model of NSCLC. Ruxolitinib enhanced virotherapy in resistant and sensitive NSCLC cells. The addition of ruxolitinib inhibited STAT1 phosphorylation and to a lesser extent STAT3 phosphorylation. Ruxolitinib treatment prevented NSCLC cells from enhancing PDL-1 expression in response to virotherapy. Combination ruxolitinib and VSV-IFNß therapy resulted in a trend toward improved survival of mice without substantially effecting PDL-1 levels or levels of immune infiltration into the tumor. These results support further clinical evaluation of the combination of JAK/STAT inhibition with virotherapy.

Gemcitabine and metabolite pharmacokinetics in advanced NSCLC patients after bronchial artery infusion and intravenous infusion.

PURPOSE: We investigated the safety, pharmacokinetics, and efficacy of gemcitabine administered via bronchial artery infusion (BAI) and IV infusion in advanced NSCLC patients. METHODS: Patients were eligible if they had received at least two prior cytotoxic chemotherapy regimens. Gemcitabine was administered via BAI as 600 mg/m2 on day one of cycle one, followed by IV as 1000 mg/m2 on day eight of cycle one, and IV on days one and eight of all subsequent cycles. Pharmacokinetics for gemcitabine and dFdU metabolite in plasma, and dFdCTP active metabolite in peripheral blood mononuclear cells (PBMC) were evaluated. Intensive pharmacokinetic sampling was performed after BAI and IV infusions during cycle one. RESULTS: Three male patients (age range 59-68 years) were evaluated. All patients responded with stable disease or better. One PR was observed after cycle three, and the remaining had SD. Cmax (mean ± SD) following BAI for gemcitabine, dFdCTP, and dFdU were 7.71 ± 0.13, 66.5 ± 40.6, and 38 ± 6.27 µM and following IV infusion, 17 ± 2.36, 50.8 ± 3.61, and 83.2 ± 12.3 µM, respectively. The AUCinf (mean ± SD) following BAI for gemcitabine, dFdCTP, and dFdU were 6.89 ± 1.2, 791.1 ± 551.2, and 829.9 ± 217.8 µM h and following IV infusion, 12.5 ± 3.13, 584 ± 86.6, and 1394.64 ± 682.2 µM h, respectively. The AUC and Cmax of dFdCTP after BAI were higher than IV. The median OS was 6.27 months. No grade 3 or 4 toxicity was observed. The most common side effects were all grade ≤ 2 involving nausea, vomiting, rigor, thrombocytopenia, and anemia. CONCLUSIONS: Systemic exposure to dFdCTP was higher after BAI than IV in two out of three patients.

Inhibition of oncogenic cap-dependent translation by 4EGI-1 reduces growth, enhances chemosensitivity and alters genome-wide translation in non-small cell lung cancer.

Hyperactivation of eIF4F-mediated translation occurs in many if not all cancers. As a consequence, cancer cells aberrantly enhance expression of malignancy-related proteins that are involved in cell cycle progression, angiogenesis, growth, and proliferation. With this in mind eIF4F is a promising molecular target for therapeutics that counteract pathological eIF4F activity. Here we used 4EGI-1, a small-molecule inhibitor of cap-mediated translation that disrupts formation of the eukaryotic initiation factor 4F (eIF4F) complex to treat non-small cell lung cancer (NSCLC). Treatment of cells with 4EGI-1 reduced cell proliferation, decreased cap-dependent complex formation, induced apoptosis, enhanced sensitivity to gemcitabine, and altered global cellular translation. Suppression of cap-dependent translation by 4EGI-1 resulted in diminished expression of oncogenic proteins c-Myc, Bcl-2, cyclin D1, and survivin, whereas ß-actin expression was left unchanged. In light of these results, small-molecule inhibitors like 4EGI-1 alone or with chemotherapy should be further evaluated in the treatment of NSCLC.