Pep5-based antitumor peptides containing multifunctional fragments with enhanced activity and synergistic effect

Carrier Cell–mediated Delivery of a Replication-competent Adenovirus for Cancer Gene Therapy

Synergistic effects of a combined treatment of PI3K/mTOR dual inhibitor LY3023414 and carboplatin on human endometrial carcinoma

Targeting Hypoxia-inducible Factor-1α With Tf–PEI–shRNA Complex via Transferrin Receptor–mediated Endocytosis Inhibits Melanoma Growth

Background: MIV-818, a nucleotide prodrug of troxacitabine-monophosphate, has been designed to deliver high levels of the chain-terminating nucleotide troxacitabine-triphosphate (TRX-TP) to the liver after oral dosing while minimizing systemic exposure. Sorafenib is a multikinase inhibitor with antiangiogenic and antiproliferative effects that is approved for the treatment of advanced hepatocellular carcinoma (HCC). Hypoxia is induced as a result of the antiangiogenic effects of sorafenib. Since hypoxic conditions have been shown to increase cytotoxicity of TRX via increased conversion of TRX-diphosphate to TRX-TP, we investigated the effects of combining MIV-818 or TRX with sorafenib in cell lines and in xenograft mouse models of HCC. Methods: Synergy of MIV-818 and sorafenib was evaluated in vitro using Bliss Independence combination analysis. In vivo effects were evaluated in nude mice with subcutaneous Hep3B or HepG2 xenografts. Due to instability of MIV-818 in mouse blood, treatment with TRX was also included. MIV-818 (30 or 100 mg/kg PO) or TRX (2.5 mg/kg IP) was given twice daily for 5 days alone or in combination with sorafenib (30 mg/kg PO) once daily for 21 days. Quantitative immuno-fluorescence was used to assess DNA damage (pH2AX), proliferation (BrdU), and hypoxia (pimonidazole) in the tumors. Results: MIV-818 shows strong synergistic anti-proliferative activity with sorafenib in several HCC cell lines in vitro. In the Hep3B xenograft model, treatment with TRX or sorafenib alone resulted in tumor growth inhibition (TGI) of 32% and 52%, respectively. Combination of TRX and sorafenib was substantially more active than either agent alone, reaching a TGI of 90%. Exposures of TRX and sorafenib in plasma and tumor were similar in the combination and single agent groups, suggesting no pharmacokinetic interactions. TRX treatment resulted in significant inhibition of proliferation (by 80%) and induction of DNA damage (15-fold) in vivo. Sorafenib treatment resulted in a small but significant inhibition of proliferation (by 27%) and 3-fold induction in hypoxia, but no induction of DNA damage. The data are consistent with the expected mechanisms of action of each agent. Combination of TRX and sorafenib resulted in 92% inhibition of proliferation and induction of DNA damage (15-fold). Notably, clear responses were seen even in hypoxic regions of the tumor, which can be resistant to therapy, indicating effective delivery of TRX-TP to regions far from blood vessels. Conclusions: Additive and synergistic anti-tumor effects were observed in nonclinical HCC models after combination of MIV-818 or TRX with sorafenib. The results suggest that add-on of MIV-818 to sorafenib may be beneficial for the treatment of HCC. MIV-818 is currently in nonclinical development in preparation for clinical trials in patients with advanced HCC and other liver cancers. Citation Format: Biljana Rizoska, Johan Bylund, Sveinn Briem, Alastair Kyle, Andrew Minchinton, Fredrik Öberg, Karin Göhlin, Mark Albertella. Synergistic and additive anti-tumor effects of MIV-818 in combination with sorafenib in nonclinical hepatocellular carcinoma models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2930.

To evaluate the inhibitory effect of imrecoxib combined with lobaplatin on tumor growth and lymph node metastasis of human lung adenocarcinoma xenografts in nude mice, and to explore its possible mechanisms. Human lung cancer A549 cells were injected into Bal B/c nude mice subcutaneously. Twenty-eight healthy male nude mice were randomly divided into 4 groups: the control group, imrecoxib group, lobaplatin group and imrecoxib combined with lobaplatin group. Each group was treated with appropriate drugs and the tumor size was measured every five days. The expression of ezrin and E-cadherin protein was detected by immunohistochemistry and flow cytometry. Ezrin and E-cadherin mRNA were detected by real-time PCR. The tumor inhibition rates of imrecoxib group, lobaplatin group and combination group were 36.7%, 54.6% and 69.2%, respectively. The tumor volumes of imrecoxib group [(905.33±113.31) mm(3)] and combination group [(507.74±77.50) mm(3)] were significantly lower than that of the control group (1355.33±189.04) mm(3) (P<0.05), and the tumor weights were significantly reduced [(1.13±0.14) g, (0.63±0.10) g respectively] vs. (1.69±0.24) g (P<0.05). The expressions of ezrin protein and mRNA in the imrecoxib group and combined treatment group were significantly lower than that of the control group (136.53±35.52, 74.72±19.48 vs. 175.62±21.16 for protein expression level; 0.54±0.03, 0.36±0.03 vs. 1.02±0.02 for mRNA expression level, respectively, P<0.05 for both), while the expression of E-cadherin protein and mRNA in the imrecoxib group and combined treatment group was significantly higher than that of the control group (253.78±38.87, 308.94±24.67 vs. 213.66±30.31 for protein expression level; 2.19±0.02, 3.02±0.02 vs. 1.05±0.03 for mRNA expression level, respectively, P<0.05 for both). There was a significant negative correlation between ezrin protein and E-cadherin protein (r=-0.737, P<0.01), as well as between ezrin mRNA and E-cadherin mRNA (r=-0.977, P<0.01). Administration of imrecoxib combined with lobaphatin has inhibitory effects on the growth of non-small cell lung cancer xenografts and lymph node metastasis via down-regulated ezrin and upregulated E-cadherin. Imrecoxib and lobaplatin have a synergistic antitumor effect.

A synergistic antitumor effect of polyphyllin I and formosanin C on hepatocarcinoma cells

Synergistic antitumor effects of HER2/neu antisense oligodeoxynucleotides and conventional chemotherapeutic agents

Immunogenic cell death (ICD) involves the activation of cytotoxic T lymphocyte-driven adaptive immunity with long-term immunological memory. OBI-3424 is a highly selective prodrug that is activated by AKR1C3 to release a potent bis-alkylating agent. This study aims to investigate whether OBI-3424 can induce ICD and create a tumor microenvironment that benefits the combination therapy of OBI-3424 with immune checkpoint inhibitors. OBI-3424 induced ICD was examined in vitro by incubation of the prodrug with AKR1C3 positive cells followed by the detection of damaging-associated molecular patterns (DAMPs). The ICD-related immunity was assessed in vivo using advanced severe immunodeficient mice that were engrafted with human peripheral blood mononuclear cells (PBMCs). Anti-tumor effect of OBI-3424 in combination with pembrolizumab was evaluated in a xenograft model using PBMC-humanized mice. Incubation of OBI-3424 with AKR1C3 positive cells H460 and HepG2 induced the release of DAMPs including calreticulin, HMGB1, and ATP, in dose- and time-dependent manners. The detection of the DAMPs indicated that OBI-3424 induced ICD in vitro. The OBI-3424-induced ICD and its related immunity were also assessed in vivo. PBMC-humanized mice were immunized with OBI-3424- or PBS-treated HepG2 cells and then challenged with live HepG2 cells. No tumor growth was noted in mice that were immunized with OBI-3424-treated cells, indicating that the dying HepG2 cells induced by OBI-3424 elicited an adaptive, tumor-specific immune response. Furthermore, OBI-3424 showed a synergistic anti-tumor effect in combination with pembrolizumab in HepG2 xenograft model using PBMC-humanized mice. OBI-3424 plus pembrolizumab exhibited significantly stronger inhibition on tumor growth (TGI 77.2%) when compared with the treatment of OBI-3424 (TGI 27.8%) or pembrolizumab (TGI -15.3%) alone. Analysis of tumor-infiltrating lymphocytes (TILs) showed that OBI-3424 treatment induced the populations of activated cytotoxic CD8 T-cells (CD45+/CD8+/CD69+ and CD45+/CD8+/Granzyme), activated helper CD4 T-cells (CD45+/CD4+/CD69+), and mature dendritic cells (CD11b+/CD86+). In addition, OBI-3424 treatment also increased PD-1 expression on CD8 and CD4 cells, which in turn potentiated the anti-tumor effect of pembrolizumab. We demonstrated that OBI-3424 was able to induce ICD as shown by the release of DAMPs in vitro and tumor-specific immunity in vivo. OBI-3424 also created a tumor microenvironment that enhances the function of pembrolizumab, supported by the synergistic effect in animals with the cotreatment of the two drugs. The results suggest that a combination therapy of OBI-3424 and anti-PD-1 in human clinical study is warranted. OBI-3424 is currently in Phase 1/2 clinical trials for solid tumor and acute lymphoblastic leukemia (NCT03592264 and NCT04315324). Citation Format: Chun-Chung Wang, Wan-Fen Li, Chih-Chan Lee, Lu-Tzu Chen, Jhih-Jie Yang, Jiann-Shiun Lai, Ming-Tain Lai. OBI-3424, an AKR1C3-activated prodrug, exhibits in vivo synergistic anti-tumor effect in combination with pembrolizumab by induction of immunogenic cell death [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 6111.

Lung cancer has a poor 5-year survival rate and is the leading cause of cancer-related deaths worldwide. Thus, the development of more efficient therapeutic strategies is urgently needed. Many studies have shown that EGCG, a major polyphenol found in green tea, has potential anticancer effects. The present study aims to investigate the molecular mechanism of EGCG-mediated inhibition of proliferation in lung cancer cells and to explore the effects of combined treatment with EGCG and an NF-κB inhibitor, BAY11-7082, in A549 and H1299 cells both in vitro and in vivo. Our results showed that EGCG inhibits cell proliferation and migration and induces apoptosis in A549 and H1299 cells at relatively high concentrations (IC50=86.4 µM for A549 cells and 80.6 µM for H1299 cells). These effects are partially achieved via inhibition of the NF-κB signaling pathway. Combined treatment with EGCG and BAY11-7082, a potent NF-κB inhibitor, shows significant synergistic effects at relatively low concentrations. The inhibition rate reached approximately 50% in cells treated for 72 h with 20 µM EGCG and 5 µM (A549 cells) or 2.5 µM BAY11-7082 (H1299 cells). This synergistic anti-tumor effect was also observed in a xenograft model. These results indicated that EGCG inhibits lung cancer cell proliferation by suppressing NF-κB signaling. Coadministration of EGCG and BAY11-7082 has a synergistic effect both in vitro and in vivo and may serve as a novel therapeutic strategy for lung cancer.

Synergistic antitumor effect mediated by a paclitaxel-conjugated polymeric micelle-coated oncolytic adenovirus.

Introduction: Gut microbiota has been reported to have a potential to modulate tumor sensitivity to chemo- or immuno-therapies although the underlying mechanism is still unclear. Oncolytic virotherapy is a promising antitumor treatment that selectively induces oncolytic cell death in tumor cells. We have previously developed OBP-702, a telomerase-specific oncolytic adenovirus armed with the p53 tumor suppressor gene, highlighting potent antitumor effects across different cancer types. To explore the potential of gut microbial metabolites, particularly butyrate, in augmenting the antitumor efficacy of oncolytic virotherapy. Materials and Methods: We assessed the synergistic antitumor effects of butyrate and OBP-702 in cytotoxic assay using two human (HCT116, SW48) and two murine (CT26, MC38) colorectal cancer cell lines in vitro, and the in vivo antitumor effects in subcutaneous tumor models using HCT116 and CT26. We explored underlying mechanisms of the synergistic effects, focusing on the influences of butylate on viral infectivity via Coxsackie-Adenovirus Receptor (CAR) and integrins, and antitumor immunity via tumor MHC-I expression and CD8-positive T cells. Results: Butyrate and OBP-702 showed potent synergistic effects in all four cell lines in vitro, and the combination suppressed tumor growth significantly in HCT116 and CT26 subcutaneous tumors compared to each monotherapy. These synergistic effects were produced by butyrate increasing the potential of OBP-702 by enhancing viral infection efficiency via upregulating expressions of CAR and integrins on tumor cells. Butyrate also increased MHC-I expression on tumor cells by activating the cGAS-STING pathway, leading to increased CXCL10 expression, which led to activation of antitumor immunity through recruitment of CD8-positive T cells in tumor tissues. The synergistic effects of butyrate and OBP-702 were proved in an orthotopic colorectal tumor model with liver metastases using CT26 cells expressing luciferase, in which this combination not only reduced tumor growth, but also improved survival by activating antitumor immunity via CD8-positive T cells. Conclusion: Butyrate and OBP-702 showed synergistic antitumor effects via activation of systemic antitumor immunity in addition to direct effects of butyrate on viral cytotoxic potential, which provides valuable insights into the development of innovative cancer treatment strategies. Citation Format: Masaki Sakamoto. Gut microbial metabolite butyrate facilitates antitumor efficacy of telomerase-specific oncolytic adenovirus via MHC-I and cGAS-STING pathway activation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6690.

Combination immunotherapy regimens incorporating agents that target more than a single regulatory pathway or immune inhibitory checkpoint can confer marked enhancement of anti-tumor immune responses when combined with active immunotherapy. We have tested the antitumor effect of the IDO pathway inhibitor NLG-919 (currently in Phase I clinical trial) in combination with indoximod (a different IDO pathway inhibitor currently in Phase II clinical trials), chemotherapy, vaccination and/or PD-1/PD-L1/PD-L2 blockade. Here we show that the highly potent, orally-bioavailable IDO pathway inhibitor NLG-919 is synergistic with blockade of the PD-1/PD-L1/PD-L2 pathway in mouse models of large established tumors. The combination of NLG-919 plus PD-1/PD-L1/PD-L2 blockade showed significantly enhanced anti-tumor effect compared to either agent alone. This synergy was particularly evident in the setting of large established tumors treated with normally ineffective immunotherapy regimens (e.g., a single dose of chemotherapy plus a vaccine against a poorly immunogenic shared-self antigen). In vivo, administration of NLG-919 enhanced anti-tumor vaccine responses against B16F10 melanoma treated with hgp100 vaccine plus resting, non-activated pmel-1 T cells NLG-919 also reduced Treg-mediated suppression in tumor-bearing hosts, and enhanced dendritic cell activation in tumors and TDLNs. In combination with chemotherapy, treatment with NLG-919 allowed effector T cell responses against endogenous tumor antigens released by chemotherapy. By each of the preceding readouts, the in vivo biologic effect of NLG-919 was qualitatively identical to that of indoximod, but the same immunologic effects could be achieved at lower plasma concentrations in vivo. In multiple models, a combination of oral NLG-919 with oral indoximod produced synergistic anti-tumor effects, which was further enhanced by blockade of the PD 1/PD L1/PD-L2 pathway. In a more stringent B16F10 melanoma tumor model that did not involve adoptive transfer of pmel-1 cells, a combination of immune checkpoint inhibition involving NLG-919, indoximod and anti-PD1/PD-L1/PD-L2 antibodies with chemotherapy and hgp100 peptide vaccine was able to achieve a significant antitumor effect. The current preclinical studies suggest a mechanistic rationale for a combining NLG919 with indoximod and with agents targeting the PD 1/PD L1/PD-L2 pathway. Citation Format: Mario R. Mautino, Charles J. Link, Nicholas N. Vahanian, James T. Adams, Clarissa Van Allen, Madhav D. Sharma, Theodore S. Johnson, David Munn. Synergistic antitumor effects of combinatorial immune checkpoint inhibition with anti-PD-1/PD-L antibodies and the IDO pathway inhibitors NLG-919 and indoximod in the context of active immunotherapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5023. doi:10.1158/1538-7445.AM2014-5023

To investigate the synergistic cytotoxicity of TRAIL in combination with chemotherapeutic agents in A549 cell lines, we systematically evaluated the cytotoxicity of TRAIL alone and TRAIL in combination with cisplatin, paclitaxel (Taxol) or actinomycin D in A549 cell lines in vitro and in vivo, and whether the sensitivity was correlated with the expression level of TRAIL receptors. We investigated the cytotoxicity of TRAIL alone and the synergistic antitumor effects of TRAIL in combination with chemotherapeutic agents in A549 cells by crystal violet staining and FACS in vitro. The expression levels of DR4, DR5, DcR1 and DcR2 were measured in TRAIL-treated and chemotherapeutic agent-treated A549 cells by Western blotting. The growth inhibition of tumors was evaluated in terms of incidence, volume and weight in a A549-implanted nude mice model. Chemotherapeutic agents cisplatin (5.56 mug/ml), Taxol (10 and 30 mug/ml) or actinomycin D (9.26, 83.3 and 750 ng/ml) augmented the cytotoxicity of TRAIL in A549 cell lines within a range of concentrations of TRAIL (1.98-160 ng/ml) in vitro. The expression levels of DR4 and DR5 were not significantly different and the expression of DcR2 was slightly downregulated, but the expression of DcR1 was not detected in non-treated, TRAIL-treated and chemotherapeutic agent-treated A549 cells. The rates of tumor inhibition following treatment with TRAIL alone (15 mg/kg per day, daily for 10 days) and TRAIL/cisplatin (15 mg/kg per day TRAIL, daily for 10 days; 1.5 mg/kg per day cisplatin, daily for 10 days with 7-day intervals) were 28.3% and 76.8% by tumor weight ( P<0.05 for TRAIL alone versus control, P<0.05 for TRAIL/cisplatin versus cisplatin alone and TRAIL alone) on day 65 in vivo. TRAIL in combination with chemotherapeutic agents cisplatin, Taxol or actinomycin D exerted synergistic antitumor effects in A549 cell lines in vitro and TRAIL/cisplatin demonstrated synergistic antitumor effects in vivo. The expression levels of TRAIL receptors suggested that the synergistic effects of TRAIL in combination with chemotherapeutic agents are not at the receptor level in A549 cell lines.

BackgroundDiffuse large B cell lymphoma (DLBCL) is the most common form of lymphoma. Although durable remissions can be achieved in more than half of these patients, DLBCL remains a significant clinical challenge, with approximately 30% of patients not being cured. BCR-associated kinases (SYK, BTK, and PI3K) inhibitors have exhibited encouraging pre-clinical and clinical effects, as reported by many researchers. Early studies demonstrated that protein kinase C-β (PKCβ) inhibitors alter phosphorylation level the Bruton’s tyrosine kinase (BTK), which leads to enhanced BTK signaling. Here, for the first time, we investigate whether the combination of PKCβ inhibitor enzastaurin and BTK inhibitor ibrutinib has synergistic anti-tumor effects in DLBCL.MethodsIn vitro cell proliferation was analyzed using Cell Titer-Glo Luminescent Cell Viability Assay. Induction of apoptosis and cell cycle arrest were measured by flow cytometry. Western Blotting analysis was used to detect the essential regulatory enzymes in related signaling pathways. RNA-seq was conducted to evaluate the whole transcriptome changes brought by co-treatment with low doses of enzastaurin and ibrutinib. The synergistic anti-tumor effects of enzastaurin and ibrutinib were also evaluated in vivo.ResultsCombination of enzastaurin and ibrutinib produced a lasting synergistic effect on the survival and proliferation of DLBCL cells, including reduction of proliferation, promoting apoptosis, inducting G1 phase arrest, preventing cell invasion and migration, and down-regulating activation of downstream signaling. More importantly, whole-transcriptome changes results showed that combination therapy worked synergistically to regulate whole-transcriptome expression compared with enzastaurin and ibrutinib alone. Co-treatment with low doses of enzastaurin and ibrutinib could effectively downregulate BCR, NF-κB, JAK and MAPK related signaling pathway. Furthermore, the mRNA expression analysis further indicated that co-treatment significantly decreased the mRNA levels of NOTCH1. The combination effect in inhibiting proliferation of DLBCL cells probably was realized through suppression of NOTCH1 expression. Finally, the anti-tumor activity of co-treatment also was demonstrated in vivo.ConclusionsCombination of enzastaurin and ibrutinib had synergistic anti-tumor effects in DLBCL, independent of molecular subtype. These results provided a sound foundation for an attractive therapeutic treatment, and the simultaneous suppression of BTK and PKCβ might be a new treatment strategy for DLBCL.

Non-small-cell lung carcinomas do not sufficiently respond to cancer chemotherapeutic drugs. Combination effects of cancer chemotherapy drugs (paclitaxel and carboplatin) with nobiletin or powdered Shiikuwasha extract from Citrus depressa were examined by isobologram and combination index analyses. It was demonstrated that the combination generated a synergistic inhibitory effect against the proliferation of the human non-small-cell lung carcinoma cell lines A549 and H460 and that of the two chemotherapy drugs, paclitaxel was responsible for this synergistic effect. Furthermore, the percentage of apoptotic cells was decreased with increasing rates of nobiletin to paclitaxel and carboplatin. These findings were considered to be attributed to the ability of nobiletin to regulate cells in the G1 phase, which escaped cell death initiated by paclitaxel and carboplatin. An antitumor activity assay showed that this combination significantly suppressed the growth of subcutaneous A549 tumor xenografts in nude mice.