Oxaliplatin In Colon Cancer Cells Research Articles

Dihydroartemisinin synergistically enhances the cytotoxic effects of oxaliplatin in colon cancer by targeting the PHB2-RCHY1 mediated signaling pathway.

Dihydroartemisinin (DHA) has recently attracted increasing attention for its low toxicity and high antitumor activity. DHA has been reported to have synergistic anticancer effects with a variety of drugs in the clinic; however, the molecular mechanism by which DHA inhibits tumorigenesis and improves oxaliplatin cytotoxicity in colon cancer cells is still not well understood. In this study, we found that DHA can inhibit cell proliferation and colony formation in a dose-dependent manner. Prohibitin 2 (PHB2) is a potential target by which DHA exerts its antitumor and cytotoxic effects. The function and molecular mechanism of PHB2 in colon cancer tumorigenesis were fully studied to determine the regulatory mechanism between DHA and PHB2. We found that PHB2, a mitochondrial inner membrane scaffold protein, has a higher expression level in colon cancer tissues than in adjacent nontumor tissues and is mainly localized in mitochondria. Overexpression of PHB2 can promote cell proliferation and colony formation in vitro and accelerate tumor growth in vivo. We also found that the expression level of PHB2 was inversely related to the cytotoxicity of DHA and oxaliplatin in colon cancer cells. The molecular mechanism of PHB2 in tumorigenesis and cancer therapy was further studied. The results showed that 20 μM DHA can downregulate PHB2 expression in a ubiquitylation-dependent manner and subsequently block PHB2-induced RCHY1 upregulation and p53 and p21 downregulation. In this process, RCHY1 is necessary for PHB2 to play a tumor-promoting role. Thus, PHB2 and RCHY1 are effective targets for colon cancer therapy, and DHA has synergistic anticancer effects with oxaliplatin via promoting PHB2 degradation in colon cancer cells.

Metformin Enhancement of Therapeutic Effects of 5-Fluorouracil and Oxaliplatin in Colon Cancer Cells and Nude Mice.

Studies have demonstrated that metformin has antitumor effects in addition to therapeutic effects on hyperglycemia; however, few studies have explored the effects of metformin in chemotherapy. Therefore, we hypothesized that the administration of metformin would enhance the therapeutic effects of 5-fluorouracil and oxaliplatin (FuOx) to inhibit the growth of colorectal cancer (CRC) cells in vitro and in vivo. The results of our in vitro experiments demonstrated that metformin significantly increased the effects of FuOx with respect to cell proliferation (p < 0.05), colony formation (p < 0.05), and migration (p < 0.01) and induced cell cycle arrest in the G0/G1 phase in HT29 cells and the S phase in SW480 and SW620 cells (p < 0.05). Flow cytometry analysis revealed that metformin combined with FuOx induced late apoptosis (p < 0.05) by mediating mitochondria-related Mcl-1 and Bim protein expression. Furthermore, in vivo, metformin combined with FuOx more notably reduced tumor volume than FuOx or metformin alone did in BALB/c mice (p < 0.05). These findings demonstrate that metformin may act as an adjunctive agent to enhance the chemosensitivity of CRC cells to FuOx. However, further clinical trials are warranted to validate the clinical implications of the findings.

Dihydroartemisinin enhances the anti-tumor activity of oxaliplatin in colorectal cancer cells by altering PRDX2-reactive oxygen species-mediated multiple signaling pathways

BackgroundGlobally, colorectal cancer (CRC) is one of the leading causes of cancer-related deaths. Oxaliplatin based treatments are frequently used as chemotherapeutic methods for CRC, however, associated side effects and drug resistance often limit their clinical application. Dihydroartemisinin (DHA) induces apoptosis in various cancer cells by increasing reactive oxygen species (ROS) production. However, the direct target of DHA and underlying molecular mechanisms in oxaliplatin-mediated anti-tumor activities against CRC are unclear. MethodsWe used 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT), flow cytometry, and colony formation assays to investigate cell phenotype alterations and ROS generation. We also used quantitative Real-Time PCR (qRT-PCR) and western blotting to measure relative gene and protein expression. Finally, an in vivo mouse xenograft model was used to assess the anti-tumor activity of oxaliplatin and DHA alone, and combinations. ResultsDHA synergistically enhanced the anti-tumor activity of oxaliplatin in colon cancer cells by regulating ROS-mediated ER stress, signal transducer and activator of transcription 3 (STAT3), C-Jun-amino-terminal kinase (JNK), and p38 signaling pathways. Mechanistically, DHA increased ROS levels by inhibiting peroxiredoxin 2 (PRDX2) expression, and PRDX2 knockdown sensitized DHA-mediated cell growth inhibition and ROS production in CRC cells. A mouse xenograft model showed strong anti-tumor effects from combination treatments when compared with single agents. ConclusionsWe demonstrated an improved therapeutic strategy for CRC patients by combining DHA and oxaliplatin treatments.

The POU2F1-ALDOA axis promotes the proliferation and chemoresistance of colon cancer cells by enhancing glycolysis and the pentose phosphate pathway activity

Cancer metabolic reprogramming enhances its malignant behaviors and drug resistance, which is regulated by POU domain transcription factors. This study explored the effect of POU domain class 2 transcription factor 1 (POU2F1) on metabolic reprogramming in colon cancer. The POU2F1 expression was analyzed in GEO dataset, TCGA cohorts and human colon cancer tissues by bioinformatics and immunohistochemistry. The effects of altered POU2F1 expression on proliferation, glucose metabolism and oxaliplatin sensitivity of colon cancer cells were tested. The impacts of POU2F1 on aldolase A (ALDOA) expression and malignant behaviors of colon cancer cells were examined. We found that up-regulated POU2F1 expression was associated with worse prognosis and oxaliplatin resistance in colon cancer. POU2F1 enhanced the proliferation, aerobic glycolysis and the pentose phosphate pathway (PPP) activity, but reduced oxidative stress and apoptosis in colon cancer cells, dependent on up-regulating ALDOA expression. Mechanistically, POU2F1 directly bound to the ALDOA promoter to enhance the ALDOA promoter activity in colon cancer cells. Moreover, activation of the POU2F1-ALDOA axis decreased the sensitivity to oxaliplatin in colon cancer cells. These data indicate that the POU2F1-ALDOA axis promotes the progression and oxaliplatin resistance by enhancing metabolic reprogramming in colon cancer. Our findings suggest that the POU2F1-ALDOA axis may be new therapeutic targets to overcome oxaliplatin resistance in colon cancer.

Perifosine enhances the potential antitumor effect of 5-fluorourasil and oxaliplatin in colon cancer cells harboring the PIK3CA mutation

Phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA) mutation in colon cancer contributes to the poor prognosis of the disease and chemoresistance of tumors. New therapies are needed; however, the lack of knowledge of the mechanism of chemoresistance has hindered progress. In this study, we investigated the mechanism of the reduced sensitivity of colon cancer cells to 5-fluorouracil (5-FU) and oxaliplatin (L-OHP), and the effects of perifosine, an Akt inhibitor that enhances the cytotoxicity of 5-FU and L-OHP in colon cancer cells harboring the PIK3CA mutation. The use of 5-FU or L-OHP alone or in combination induced significant death of Caco-2 cells (PIK3CA wild type), but only weakly decreased the viability of DLD-1 and SW948 cells harboring the PIK3CA mutation. The use of 5-FU and L-OHP, either alone or in combination, strongly suppressed Akt activation, Survivin, Bcl-2, and Bcl-xL expression, and enhanced Puma, phospho-p53, and p53 expression in Caco-2 cells than in DLD-1 cells. In addition, perifosine enhanced the cytotoxicity of the 5-FU and L-OHP combination, inhibited Akt activation and the expression of Survivin, Bcl-2, and Bcl-xL, and increased the expression of Puma, phospho-p53, and p53 in DLD-1 cells. These results indicate that PIK3CA mutation contributes to reduced sensitivity to 5-FU and L-OHP via Akt activation in colon cancer cells. Perifosine increases the efficacy of 5-FU and L-OHP by suppressing Akt activation. Thus, the use of an Akt inhibitor in combination with 5-FU and L-OHP may be beneficial in colon cancer with cells harboring the PIK3CA mutation.

GW4064 enhances the chemosensitivity of colorectal cancer to oxaliplatin by inducing pyroptosis

Repeated and long-term oxaliplatin therapy leads to drug resistance and severe adverse events, which limit its clinical use. These difficulties highlight the importance of identifying potent and specific drug combinations to enhance the antitumor effects of oxaliplatin. The farnesoid X receptor (FXR) deficiency in colorectal cancer (CRC) suggests that restoring FXR function might be a promising strategy for CRC treatment. A drug combination study showed that the GW4064 acted synergistically with oxaliplatin in colon cancer cells. The combination of oxaliplatin plus GW4064 inhibited cell growth and colony formation, induced apoptosis and pyroptosis in vitro, and slowed tumor growth in vivo. Mechanistically, GW4064 enhanced the chemosensitivity of cells to oxaliplatin by inducing BAX/caspase-3/GSDME-mediated pyroptosis. Furthermore, the combination of oxaliplatin and GW4064 synergistically inhibited STAT3 signaling by restoring SHP expression. Our study revealed that GW4064 could enhance the antitumor effects of oxaliplatin against CRC, which provides a novel therapeutic strategy based on a combinational approach for CRC treatment.

Abstract 3824: Increased production of endogenous hydrogen sulfide (H2S) induced resistance to 5-FU and Oxaliplatin in colon cancer cells

Abstract The combination of 5-FU and Oxaliplatin has been widely adopted as one of the first line chemotherapeutic regimens for colon cancer patients. However, the development of chemoresistance can lead to early recurrence and metastasis to distant organs, thus remaining a leading cause of death from colon cancer. In the present study, the increased production of hydrogen sulfide (H2S) in colon cancer tissues was validated and the effect of inhibition of production of endogenous H2S on the responses of colon cancer cells to 5-FU and Oxaliplatin was further analyzed. The results suggested that inhibition of the enzymatic activity of cystathionine-β-synthase (CBS), a major producer of H2S in colon cancer cells, significantly decreased the IC50s of both 5-FU and Oxaliplatin. Inhibition of CBS leads to decreased PI3K-Akt signaling and consequently decreased expression of thymidylate synthetase (TYMS), which is the target of 5-FU. Inhibition of CBS utilizing aminooxyacetic acid (AOAA) leads to increased production of reactive oxygen species (ROS) and exaggerated mitochondrial damages in colon cancer cells. Analysis performed on CompuSyn confirmed the synergetic effect of AOAA with both 5-FU and Oxaliplatin. In vivo imaging of tumorigenesis in nude mice further confirmed the effect of inhibition of CBS on the therapeutic effect of both 5-FU and Oxaliplatin. Together, these results suggested that inhibition of production of endogenous H2S might be a promising target for chemoresistance in colon cancer. Citation Format: Shanwen Chen, Shuai Zuo. Increased production of endogenous hydrogen sulfide (H2S) induced resistance to 5-FU and Oxaliplatin in colon cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3824.

Genetic variants in RPA1 associated with the response to oxaliplatin-based chemotherapy in colorectal cancer.

Oxaliplatin (L-OHP) is a commonly used first-line chemotherapy for colorectal cancer. Genetic variants in nucleotide excision repair (NER) pathway genes may alter genomic integrity and the efficacy of oxaliplatin-based chemotherapy in colorectal cancer. We investigated the association between genetic variants in 19 NER pathway genes and the disease control rate (DCR) and progression-free survival (PFS) among 166 colorectal cancer patients who received oxaliplatin-based chemotherapy. Expression quantitative trait loci (eQTL) analysis was performed using the Genotype-Tissue Expression (GTEx) portal. Gene harboring significant SNP was overexpressed or knocked down to demonstrate the effect on cell phenotypes with or without oxaliplatin treatment. We found that rs5030740, located in the 3'-untranslated region (3'-UTR) of RPA1, was associated with DCR [OR = 2.99 (1.33-5.69), P = 4.00 × 10-3] and PFS [HR = 1.86 (1.30-2.68), P = 7.39 × 10-4]. The C allele was significantly associated with higher RPA1 mRNA expression levels according to eQTL analysis (P = 0.010 for sigmoid colonand P = 0.004 for transversecolon). The C allele of rs5030740 disrupted let-7e-5p binding to enhance RPA1 expression. Functionally, RPA1 knockdown inhibited cell proliferation and promoted cell apoptosis, whereas RPA1 overexpression promoted proliferation and suppressed apoptosis. Furthermore, low RPA1 expression increased sensitivity to oxaliplatin in colon cancer cells and inhibited proliferation after oxaliplatin treatment. Our findings demonstrate an association between rs5030740 and the DCR and PFS of colorectal cancer patients. RPA1 functions as a putative oncogene in tumorigenesis by reducing sensitivity to oxaliplatin and could serve as a potential prognostic biomarker in colorectal cancer.

Sesquiterpenes α-humulene and β-caryophyllene oxide enhance the efficacy of 5-fluorouracil and oxaliplatin in colon cancer cells.

The present study is designed to find out if sesquiterpenes, α-humulene (HUM), valencene (VAL), β-caryphyllene-oxide (CAO) and trans-nerolidol (NER), are able to improve the antiproliferative effect of classical cytostatic drugs, 5-fluorouracil (FU) and oxaliplatin (1,2-diaminocyclohexaneoxalato-platinum, OxPt), in colon cancer cell lines Caco-2 and SW-620. In addition, the possible mechanisms of sesquiterpene action are studied. The results show significant ability of HUM and especially of CAO to enhance the anti-proliferative effects of FU and OxPt in cancer cell lines Caco-2 and SW-620. On the other hand, VAL and NER are ineffective. The action of CAO could be partly based on its ability to disrupt the mitochondrial membrane potential and to activate initiator caspases, but other mechanisms are probably also involved. Based on these results, CAO seems to have the potential for combination therapy of colon cancers and deserves further study.

Rosmarinic Acid Mitigates Mitochondrial Dysfunction and Spinal Glial Activation in Oxaliplatin-induced Peripheral Neuropathy.

Chemotherapy-induced peripheral neuropathy (CIPN) is a dose-limiting complication which develops as a consequence of treatment with chemotherapeutic agents like oxaliplatin and is a mainstay of therapy for colorectal cancer. Ever since CIPN was identified, understanding its exact pathomechanisms remains a clinical challenge. The role of mitochondrial dysfunction and glial cell activation has surfaced in the etiology of CIPN. Rosmarinic acid (RA), a known mitoprotectant exerts neuroprotection against the oxidative stress and neuroinflammation in various disease conditions. Hence, in the present study, we investigated the effect using rosmarinic acid (25 and 50mg/kg, po) in the experimental model of oxaliplatin-induced peripheral neuropathy (OIPN) in rats. Results showed that RA significantly (p < 0.001) prevented the functional deficits, reversed oxaliplatin-induced mechanical allodynia and cold hyperalgesia in rats. It reduced the oxidative stress, improved the mitochondrial function, and prevented the oxaliplatin-induced loss of ATP levels. RA significantly (p < 0.01) inhibited the spinal glial cell activation and suppressed the expression of inflammatory markers. RA treatment also resulted in the activation of adenosine monophosphate-activated protein kinase (AMPK) in the peripheral nerves and dorsal root ganglion (DRG) which also might have contributed to its neuroprotective actions. In vitro screening also revealed that RA did not compromise the anti-cancer activity of oxaliplatin in colon cancer cells (HT-29). Taken together, the above results demonstrate the therapeutic activity of RA against the oxaliplatin-induced mitochondrial dysfunction and neuroinflammation and thus, suggest its potential for the management of OIPN. Graphical Abstract Schematic representation of neuroprotective mechanisms of rosmarinic acid via AMPK activation in oxaliplatin-evoked peripheral neuropathy.

Deficiency of gap junction composed of connexin43 contributes to oxaliplatin resistance in colon cancer cells.

Although comprehensive strategies in the treatment of colorectal cancer have been developed for a number of years, the five-year survival rate of metastatic colon cancer remains less than 10%. Oxaliplatin, a commonly used chemotherapeutic agent for metastatic colon cancer, improves the response rate of patients and prolongs patients' progression-free survival. However, the generation of resistance limits the clinical application of oxaliplatin, and the mechanisms of this remain unclear. The present study mainly investigated the effect of the gap junction (GJ) composed of connexin43 (Cx43) on oxaliplatin cytotoxicity in colon cancer cells. Three different methods with distinct mechanisms were used to change the function of Cx43 GJs, including cell culture at different densities, pretreatment with a specific inhibitor or enhancer, and special gene knockdown, to observe the cytotoxicity of oxaliplatin and the level of reactive oxygen species (ROS) mediated by Cx43 GJs. The results revealed that the cytotoxicity of oxaliplatin and the level of ROS were decreased with the downregulation of Cx43 GJ function, but exacerbated with the upregulation of Cx43 GJ function. Moreover, ROS scavenging with N-acetyl-L-cysteine and apocynin decreased the cytotoxicity of oxaliplatin. We concluded that the loss of GJ composed of Cx43 contributed to the resistance of oxaliplatin in colon cancer cells, and the mechanism was associated with intracellular ROS alternation.

Grape seed extracts modify the outcome of oxaliplatin in colon cancer cells by interfering with cellular mechanisms of drug cytotoxicity.

Grape seed extracts are commonly utilized as dietary supplements for their antioxidant properties, even from cancer patients. However, whether these natural extracts interfere with chemotherapeutics utilized in colon cancer treatment is still poorly investigated. The cytotoxicity of extracts from Italia and Palieri cultivars either alone or in combination with oxaliplatin was evaluated in colon cancer cells. Grape seed extracts displayed anti-proliferative activity depending on the concentration utilized through apoptosis induction. In combination, they affected the activation of Erk1/2 and counteracted the intrinsic and the extrinsic pathway of apoptosis, the DNA damage and the generation of ROS induced by oxaliplatin. Noteworthy grape seed extracts strongly enhanced the uptake of oxaliplatin into all cells, by affecting the cell transport system of platinum. The addition of these natural extracts to oxaliplatin strongly reduced the cellular response to oxaliplatin and allowed a huge accumulation of platinum into cells. Here, we shed light on the chemical biology underlying the combination of grape seed extracts and oxaliplatin, demonstrating that they might be detrimental to oxaliplatin effectiveness in colon cancer therapy.

Repulsive Guidance Molecule b (Dragon) Induces Resistance to Oxaliplatin in Colon Cancer Cells

Repulsive Guidance Molecule b (Dragon) Induces Resistance to Oxaliplatin in Colon Cancer Cells

Abstract 2177: Rutin enhances the antiproliferative effect of 5-FU and oxaliplatin in colon cancer cells

Abstract Background: Rutin is a strong antioxidant molecule and it has advantageous over other flavonoids due to it is a nontoxic and nonoxidizable molecule. The concept of dual therapy of anti-cancer drugs with natural compounds has become a very promising approach in new strategy to treatment. The aim of this study was to evaluate the antiproliferative and apoptotic effects of rutin flavonoids and its efficacy in enhancing the anticancer effects of 5-FU (5-fluorouracil) and oxaliplatin against colon cancer cells. Material and Methods: Caco-2 human colon cancer cells were treated with rutin and/or anticancer drugs (5-FU and oxaliplatin), cell viability and apoptotic parameters were examined. Cell viability was determined by MTT assay. Apoptotic markers (cleaved caspase 3, cleaved PARP and phosphor-Bad) were determined using specific enzyme-linked immunosorbent assay kits. Caspase-8 and caspase-9 were analyzed by colorimetric activity assay kit. DNA fragmentation was analyzed by agarose gel electrophoresis. Results: The exposure Caco-2 human colon cancer cells to rutin, 5-FU and oxaliplatin resulted growth inhibition in a dose- and time-dependent manner. Cell viability assay shows that rutin inhibiting growth of Caco-2 cells at high concentrations (over 1000 μM). Combination with rutin markedly enhanced 5-FU and oxaliplatin growth inhibiting effects on Caco-2 cells. Results suggested that rutin had significant anticancer abilities; such rutin were capable of causing multifold decreases in the half maximal inhibitory concentration IC50 value of 5-FU and oxaliplatin. This flavonoid increased the levels of proteins associated with apoptotic cell death (phospho-Bad, cleaved caspase 3 and cleaved PARP) alone and combination. The activities of caspase 8 and caspase 9 were stimulated by the combination treatment of rutin and oxaliplatin then alone. DNA fragmentation was observed only on combination treatment. Conclusions: Combined treatment with rutin and anticancer drugs (5-FU and/or oxaliplatin) is more effective than the individual treatments of drugs at inhibiting growth of Caco-2 cells. The use of lower 5-FU and oxaliplatin doses, with similar effects, could be also useful to reduce possible adverse effects of these drugs. However, further studies at molecular level are required to elucidate chemopreventive and chemotherapeutic effects of rutin on colon cancer. Citation Format: Farnoud Nasiri, Gorkem Kismali, Merve Alpay, Funda Kosova, Dilek Ulker Cakir, Tevhide Sel. Rutin enhances the antiproliferative effect of 5-FU and oxaliplatin in colon cancer cells. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2177.

Different Effects of Epigenetic Modifiers on the Cytotoxicity Induced by 5-Fluorouracil, Irinotecan or Oxaliplatin in Colon Cancer Cells

We investigated the effects of epigenetic modifiers such as DNA methyltransferase (DNMT) or histone deacetylase (HDAC) inhibitors on the cytotoxicity induced by 3 anticancer drugs (5-fluorouracil (5-FU), irinotecan (CPT-11) or its active form SN38, and oxaliplatin (L-OHP)) in human colorectal cancer (CRC) cells. Cytotoxicity in 4 CRC cell lines (HT29, SW480, SW48 and HCT116) was examined by colorimetric assay after drug treatment for 72 h. The effects of drug combinations were analyzed by an isobologram method. SW480 cells showed the lowest sensitivity to cytotoxicity induced by the anticancer drugs among the 4 CRC cell lines. In SW480 cells, DNMT inhibitors, such as decitabine (DAC), azacytidine and zebularine (Zeb), showed synergic effects on the cytotoxicity induced by anticancer drugs except for SN-38 plus Zeb, while HDAC inhibitors, trichostatin A, suberoylanilide hydroxamic acid and valproic acid, showed antagonistic effects. DAC showed the most potent synergic effects among the epigenetic modifiers studied. Thus, we examined whether the synergic effect of DAC is observed in other different CRC cell lines, HT29, SW48 and HCT116 cells. In all 4 CRC cell lines, the cytotoxicity of L-OHP was enhanced in a synergic manner by co-treatment with DAC. However, synergic effects of DAC with 5-FU or CPT-11 (SN-38) were not observed in 4 CRC cell lines.

Abstract C46: The pharmacokinetics and pharmacodynamics study of honokiol in Sprague-Dawley rats and colorectal tumor-bearing mice

Abstract Honokiol, a small-molecule pharmacological bioactive constituent of the bark of Magnolia plants, has shown anti-tumor effect in a variety of human cancer xenograft models through the induction of apoptosis. Additionally, honokiol could enhance the cytotoxicity in combination of oxaliplatin in colon cancer cells. To investigate the anti-tumor effect and the pharmacokinetics (PK) of honokiol, honokiol was tested for anti-tumor activity in a colorectal cancer xenograft mice model and oral-bioavailability of honokiol was assayed in Sprague-Dawley rats. The pharmacokinetic results of 5.0 mg/kg honokiol after intravenous injection showed that the elimination half time was about 0.79 ± 0.03 hours, the Cmax was about 752.67 ± 58.36 ng/mL, and the mean residence time (MRT) was about 0.85 ± 0.05 hours. In vivo study for testing anti-cancer activity of honokiol is underwent and expected that honokiol would inhibit the tumor growth in colorectal tumor-bearing mice. In conclusion, honokiol may be developed be as therapeutic agent for cancer treatment in combination with PK and pharmacodynamics (PD) data. Citation Format: Yung-Jen Tsai, Pei-Yi Tsai, Le-Mei Hung, Mei-Yin Su, Min-Hui Lin, Lung-Yu Kuan, Yih-Chiao Tsai, Hui-Ling Chen, Jia-Ming Chang. The pharmacokinetics and pharmacodynamics study of honokiol in Sprague-Dawley rats and colorectal tumor-bearing mice. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr C46.

Honokiol augments the anti-cancer effects of oxaliplatin in colon cancer cells

Oxaliplatin is an important drug in the chemotherapy of colorectal carcinoma, but its toxicity, especially dose-related neurosensory toxicity, is not well tolerated. In this study, we investigated whether honokiol could augment the anti-tumor effect of oxaliplatin in colon cancer HT-29 cells in vitro and whether honokiol could be used with oxaliplatin to decrease oxaliplatin dose. We used the normal colon cells, human colonic epithelial cells (HCoEpiCs) as control cells. Cell proliferation, apoptosis, prostaglandin E2 (PGE2) and vascular endothelial growth factor (VEGF) levels were also investigated. Expression levels of cyclo-oxygenase 2 (COX-2), VEGF, AKT/p-AKT, extracellular signal-related kinase (ERK)1/2/p-ERK1/2, nuclear factor kappa B (NF-κB) P65/p-P65, and caspase-3 were measured. Honokiol or oxaliplatin suppressed the proliferation of HT-29 cells in a concentration-dependent manner, but only high concentrations of honokiol would suppress the proliferation of HCoEpiCs. HT-29 cells were more sensitive to oxaliplatin treatment in the presence of honokiol. Oxaliplatin combined with honokiol improved the apoptosis rate of HT-29 cell and reduced PGE2 and VEGF secretion levels. Expression levels of COX-2 and VEGF protein and phosphorylation of AKT, ERK1/2, and NF-κB P65 were also inhibited. Caspase-3 levels were upregulated after honokiol treatment. Therefore, honokiol can be used in combination with oxaliplatin in the chemotherapy of colon cancer. This combination allows a reduction in oxaliplatin dose, and thereby reduces its adverse effects. It may also enhance the chemotherapeutic effect of oxaliplatin for this disease.

Abstract 2608: Regulation of response to oxaliplatin by wnt/β-catenin in colon cancer cell lines.

Abstract Background: Oxaliplatin is the standard treatment chemotherapy agent in patients with metastatic colorectal cancer. However, most patients developed the progressive disease after treatment with oxaliplatin due to chemotherapy resistance. Unfortunately the mechanism of resistance of oxaliplatin is unclear. Wnt/β-catenin signaling pathway may play a role in oncogenesis in many cancer and also involve in the control the cancer progression and metastasis throughout the interaction with the tumor microenvironment. In this study, we investigated the response to oxaliplatin regulated by wnt/β-catenin pathway in colon cancer cell line. Method: We used SNU-C2A and HCT116 human colon cancer cell lines. After comparing β-catenin expression in SNU-C2A and HCT116 cells through western blot, we exposed these two cell lines to gradient concentrations of oxaliplatin for 48hrs to examine the sensitivity. The viability was determined by Neutral Red uptake assay (NR assay). To construct a gene silenced cell line, a siRNA against β-catenin was transfected in HCT116 cells. Besides, we stimulated SNU-C2A cells with human recombinant Wnt3a (rhWnt3a) protein to up regulate the β-catenin and estimated the change of sensitivity for oxaliplatin,respectively. We also constructed a HCT116-cis cell line which showed oxaliplatin-resistance. It has been made by long-term exposure to oxaliplatin. Result: β-catenin was highly expressed in HCT116 cell line relatively higher than in SNU-C2A cell line. Through the sensitivity test to oxalipatlin, we confirmed that SNU-C2A cells were more sensitive to oxaliplatin than HCT116 cells. When we transfected β-catenin siRNA into the HCT116 cell line, the cell viability was decreased about 20-50% after oxaliplatin treatment. After rhWnt3a treatment,we confirmed the expression of β-catenin and c-myc was up regulated in SNU-C2A cell line by western blot. After treatment with oxaliplatin, we found that the cell viability was reduced in SNU-C2A cells with β-catenin overexpressing. Moreover,HCT116-cis, oxaliplatin-resistance induced cell line, showed not only the increased protein level of β-catenin but also the upper viability for oxaliplatin compared with HCT116. Conclusion Based on our data, β-catenin is one of the crucial factors to determine the resistance for oxaliplatin in colon cancer cells. Currently we are conducting the analysis with human tissue, and exploring the possible mechanism of resistance developmend to oxaliplatin. Citation Format: Myung Ah Lee, Jin Hee Park. Regulation of response to oxaliplatin by wnt/β-catenin in colon cancer cell lines. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2608. doi:10.1158/1538-7445.AM2013-2608

Effects of medicated serum prepared with Chinese herbal medicine Changweiqing on pharmacokinetics of oxaliplatin in colon cancer cells

To investigate the effects of Changweiqing-medicated serum, which was prepared with a compound traditional Chinese herbal medicine, on the reversal of oxaliplatin (L-OHP) resistance and the relationship between the reversal and cellular accumulation of platinum and proteins associated with copper transporter in HCT116/L-OHP cells. For clarifying the reversal effect of Changweiqing, methyl thiazolyl tetrazolium was applied to determine the L-OHP resistance of HCT116/L-OHP cell line. The relationship between the cellular accumulation of platinum and the L-OHP resistance in HCT116/L-OHP cells, and the effects of drug-medicated serum on intracellular contents of platinum were detected by atomic absorption spectrophotometry. Western blot method was used to determine the expressions of human copper transporter 1 (hCTR1), ATPase Cu(2+) transporting alpha polypeptide (ATP7A), copper-transporting P-type adenosine triphosphatase (ATP7B), glutathione S-transferase-π (GST-π) and multidrug resistance-associated protein 2 (MRP2). The inhibitory concentration 50% values of different pairs of L-OHP-sensitive and -resistant cells were 7.2 and 89.00. The resistance index of HCT116/L-OHP cells was 12.36. The reverse index of drug serum on HCT116/L-OHP cells was 2.74. The platinum content in HCT116/L-OHP cells was decreased compared with HCT116 cells in condition of 7.2 μg/mL L-OHP. After treating by 7.5% Changweiqing-medicated serum, the intracellular platinum contents in L-OHP-sensitive and -resistant cells were increased. It was dose-dependent that drug-medicated serum promoted the uptake of L-OHP by HCT116 or HCT116/L-OHP cells and inhibited the discharge. The 7.5% Changweiqing-medicated serum increased the expression of hCTR1 and decreased the expressions of ATP7A and ATP7B in HCT116/L-OHP cells, but had no effects on GST-π and MRP2 protein expressions. Changweiqing can reverse the L-OHP resistance of HCT116/L-OHP by increasing the cellular platinum-DNA accumulation. Down-regulation of expression of ATP7B and ATP7A, and up-regulation of hCTR1 may cause the increase of intracellular platinum content in HCT116/L-OHP cells.

Nitrogen permease regulator-like 2 enhances sensitivity to oxaliplatin in colon cancer cells.

Colorectal cancer (CRC) is the third most common cancer worldwide. Chemotherapeutic compounds used for the treatment of CRC include oxaliplatin (L-OHP). While L-OHP improves CRC survival, certain patients are resistant. The nitrogen permease regulator like-2 (NPRL2) gene is a candidate tumor suppressor gene that resides in a 120-kb homozygous deletion region on chromosome 3p21.3. In the present study, it was demonstrated that NPRL2 overexpression increases the sensitivity of HCT116 cells to L-OHP. The IC50 of L-OHP was decreased in cells transduced with NPRL2 compared with negative control (NC) cells and the effect of NPRL2 on L-OHP sensitivity was time dependent. Following NPRL2 transduction in HCT116 cells, the cell cycle was arrested in the G1 phase and a partial decrease in the S phase population was observed. Flow cytometric analysis revealed that NPRL2 transduction and L-OHP treatment increased apoptosis compared with NC cells. The mechanism through which NPRL2 overexpression enhances L-OHP sensitivity involves downregulation of the functions of the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin network. Furthermore, L-OHP upregulated caspase-3 and caspase-9 to promote apoptosis in NPRL2-overexpressing cells compared with cells that were transduced with NPRL2 or treated with L-OHP and NC cells (P<0.01). NPRL2 overexpression led to the downregulation of CD24, which could significantly reduce tumor invasiveness and decrease the metastatic capacity of HCT116 cells. These mechanisms are likely active in other types of cancer and may be exploited for the development of novel cancer therapies.