Indium-111-labeled CD166-targeted peptide as a potential nuclear imaging agent for detecting colorectal cancer stem-like cells in a xenograft mouse model.

BACKGROUND: Cancer stem cells (CSCs) are involved in drug resistance, metastasis, and relapse of cancers, which can significantly affect tumor therapy. Hence, to develop specifically therapeutic target probe at CSCs for improvement of survival and quality of life of cancer patients is urgently needed. The CD166 protein has been suggested to be involved in colorectal cancer (CRC) tumorigenesis and to be considered a marker for colorectal CSCs (CRCSCs) detection. In this study, therefore, we attend to apply a nuclear imaging agent probe, Glycine18-Cystine-linked CD166-targeted peptides (CD166tp-G18C), to detect the changes of CD166 level in a CRC xenograft mouse model. RESULTS: We isolated the CD166-positive cells from the HCT15 CRC cell line (CD166+HCT15) and evaluated their morphology and ability of clone formation, migration, protein expression, and drug resistance. The CD166-positive HCT15 cells display the CSCs characteristics. We discovered and designed a CD166-targeted peptide (CD166tp-G18C) as a targeted probe of CRC stem-like cell for cell binding assay. The CD166tp-G18C confirmed the CD166 protein targeting ability in CD166+HCT15 cells. The diethylenetriaminopentaacetic acid (DTPA)-conjugated CD166tp-G18C further was labeled with indium-111 (111In-DTPA-CD166tp-G18C) as nuclear imaging agent for imaging and bio-distribution analysis in vivo. Finally, we observed that the 111In-DTPA-CD166tp-G18C was significantly enhanced in tumor tissues of CD166+HCT15 xenograft mice as compared to the non-CD166tp-G18C control. CONCLUSIONS: Our results indicated that the indium-111-labeled CD166tp-G18C may be served as a powerful tool for colorectal CSCs nuclear imaging in the CRC patients.

The new ophthalmic formulation for infection control by combining collagen/gelatin/alginate biomaterial with liposomal chloramphenicol.

Eye drops are a conventional method of drug delivery to the eye, accounting for 90% of currently accessible ophthalmic formulations. The major problem with eye drop treatments is rapid pre-corneal drug loss. Furthermore, the need for frequent administration of eye drops can profoundly affect the quality of life of ophthalmological patients. In the current study, we developed a liposomal nanoparticle encapsulated with chloramphenicol mixed with biodegradable materials against ophthalmological disease. We first established a protocol for chloramphenicol (CAP) loaded into liposomal nanoparticle (LipoCAP). We also established the collagen/gelatin/sodium alginate (CGA) as the component of biodegradable polymers and calibrated the novel drug-releasing formulation. Finally, we combined LipoCAP with CGA to generate an 8-h degradable ophthalmic chloramphenicol gel, CGA-LipoCAP-8. CGA-LipoCAP-8 reached the effective working concentration in 75 min and prolonged the drug-releasing time for at least 12 h. In addition, CGA-LipoCAP-8 could stably and continuously inhibit E. coli proliferation. The inhibiting phenomenon was more pronounced over time. Furthermore, there were no significant toxicities observed when CGA-LipoCAP-8 co-cultured with ocular epithelial cells. In conclusion, CGA-LipoCAP-8 achieved effective CAP dose concentrations in a short time and sustained CAP release for a prolonged period. Our results provide an innovative concept in relation to novel drug-release formulations, with safety and efficiency supporting use in future treatments for ophthalmological diseases.

Evaluation Efficacy of Rhenium-188-Loaded Micro-particles for Radiotherapy in a Mouse Model of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is one of the leading causes of mortality worldwide. The aim of the present study was to evaluate the distribution and the therapeutic effect of 188Re-Tin-colloid micro-particles in subcutaneous HCC-bearing mice. The synthesis and characterization of micro-particles labeled with the 188Re isotope were performed. The micro-particles were injected into the tumor site subcutaneously in the BNL HCC-bearing mice with three treatment groups, normal saline, 188Re micro-particles, and 188Re-Tin-colloid micro-particles. The results of biodistribution showed that major radioactivity (188Re) of 188Re-Tin-colloid micro-particles (18.69 ± 4.28 %ID/g) remained at the tumor sites, compared with 188Re micro-particles (0.21 ± 0.12 %ID/g), 24 h post injection. Following the injection of 188Re-Tin-colloid micro-particles for 14 days, all BNL tumors in mice were regressed during the observation period. By contrast, all of the mice treated with normal saline or 188Re micro-particles had died by 24 and 28 days, respectively. The 188Re-Tin-colloid micro-particles demonstrated high accumulation and therapeutic potential in the subcutaneous HCC-bearing mice.

Anti-angiogenic treatment (Bevacizumab) improves the responsiveness of photodynamic therapy in colorectal cancer.

Photodynamic therapy (PDT) is a treatment utilizing the combined action of photosensitizers and light for the treatment of various cancers. The mechanisms for tumor destruction after PDT include direct tumor cell kill by singlet oxygen species (OS), indirect cell kill via vascular damage, and an elicited immune response. However, it has been reported that many cellular activators, including vascular endothelial growth factor (VEGF), are produced by tumor cells after PDT. In this study, we demonstrate that meta-tetra(hydroxyphenyl) chlorin (mTHPC)-based photodynamic therapy combined with bevacizumab (Avastin™), an anti-VEGF neutralizing monoclonal antibody that blocks the binding of VEGF to its receptor, can enhance the effectiveness of each treatment modality. We evaluated the efficacy of bevacizumab-based anti-angiogenesis in combination with PDT as well as the resulting VEGF levels and microvessel density (MVD) in a mouse model of human colon cancer. Enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC) were performed to assess VEGF concentrations and microvessel density in the various treatment groups, and confocal imaging and high performance liquid chromatography (HPLC) analyses were used to measure the distribution and concentration of mTHPC in tumors. Our results demonstrate that combination of PDT followed by bevacizumab significantly elicits a greater tumor response whereas bevacizumab treatment prior to PDT led to a reduced tumor response. Immunostaining and ELISA analyses revealed a lower expression of VEGF in tumors treated with combination therapy of PDT followed by bevacizumab. However, bevacizumab treatment decreased the accumulation of mTHPC in tumors 24 h after administration, which complemented the results of decreased anti-tumor efficacy of bevacizumab followed by PDT.

Polyethylene glycol-conjugated HER2-targeted peptides as a nuclear imaging probe for HER2-overexpressed gastric cancer detection in vivo.

BACKGROUND: The human epidermal growth factor receptor 2 (HER2) involved proliferation, angiogenesis, and reduced apoptosis in gastric cancer (GC), which is a common target for tumor therapy. HER2 is usually overexpressed in more than 15% GC patients, developing a reliable diagnostic tool for tumor HER2 detection is important. In this study, we attend to use polyethylene glycol (PEG) linked anti-HER2/neu peptide (AHNP-PEG) as a nuclear imaging agent probe for HER2 detection in GC xenograft animal model. METHODS: The HER2 expression of human sera and tissues were detected in GC patients and normal subjects. GC cell lines NCI-N87 (high HER2 levels) and MKN45 (low HER2 levels) were treated with AHNP-PEG to assess the cell viability and HER2 binding ability. The NCI-N87 was treated with AHNP-PEG to observe the level and phosphorylation of HER2. The MKN45 and NCI-N87-induced xenograft mice were intravenous injection with fluorescence labeled AHNP-PEG for detecting in vivo fluorescence imaging properties and biodistribution. The AHNP-PEG was conjugated with diethylenetriaminopentaacetic acid (DTPA) for indium-111 labeling (111In-DTPA-AHNP-PEG). The stability of was assessed in vitro. The imaging properties and biodistribution of 111In-DTPA-AHNP-PEG were observed in NCI-N87-induced xenograft mice. RESULTS: The serum HER2 (sHER2) levels in GC patients were significantly higher than the normal subjects. The sHER2 levels were correlated with the tumor HER2 levels in different stages of GC patients. The AHNP-PEG inhibited the cell growth and down-regulated HER2 phosphorylation in HER2-overexpressed human GC cells (NCI-N87) via specific HER2 interaction of cell surface. In addition, the GC tumor tissues from HER2-postive xenograft mice presented higher HER2 fluorescence imaging as compared to HER2-negative group. The HER2 levels in the tumor tissues were also higher than other organs in NCI-N87-induced xenograft mice. Finally, we further observed that the 111In-DTPA-AHNP-PEG was significantly enhanced in tumor tissues of NCI-N87-induced xenograft mice compared to control. CONCLUSIONS: These findings suggest that the sHER2 measurement may be as a potential tool for detecting HER2 expressions in GC patients. The radioisotope-labeled AHNP-PEG may be useful to apply in GC patients for HER2 nuclear medicine imaging.

pH-Responsive Nanophotosensitizer for an Enhanced Photodynamic Therapy of Colorectal Cancer Overexpressing EGFR.

Photodynamic therapy (PDT) has been shown to kill cancer cells and improve survival and quality of life in cancer patients, and numerous new approaches have been considered for maximizing the efficacy of PDT. In this study, a new multifunctional nanophotosensitizer Ce6/GE11-(pH)micelle was developed to target epidermal growth factor receptor (EGFR) overexpressing colorectal cancer (CRC) cells. This nanophotosensitizer was synthesized using a micelle comprising pH-responsive copolymers (PEGMA-PDPA), biodegradable copolymers (mPEG-PCL), and maleimide-modified biodegradable copolymers (Mal-PEG-PCL) to entrap the potential hydrophobic photosensitizer chlorin e6 (Ce6) and to present EGFR-targeting peptides (GE11) on its surface. In the presence of Ce6/GE11-(pH)micelles, Ce6 uptake by EGFR-overexpressing CRC cells significantly increased due to GE11 specificity. Moreover, Ce6 was released from Ce6/GE11-(pH)micelles in tumor environments, leading to improved elimination of cancer cells in PDT. These results indicate enhanced efficacy of PDT using Ce6/GE11-(pH)micelle, which is a powerful nanophotosensitizer with high potential for application to future PDT for CRC.

Biodegradable and Multifunctional Microspheres for Treatment of Hepatoma through Transarterial Embolization.

To improve the effectiveness of cancer treatment, this study aimed to develop biodegradable microspheres and to combine chemotherapy and radiotherapy via transcatheter arterial embolization/chemoembolization (TAE/TACE) with local radiation therapy for the slow release of chemotherapeutic agents. Microparticles were prepared by double emulsification using a biodegradable and biocompatible polymer Poly(D,l-lactide-co-glycolide) (PLGA). Since the microspheres contain a water-soluble Poly(vinylsulfonic acid) (PVSA) solution, the functional groups of this polymer dissociate into -SO3- in water. The positively charged doxorubicin can be loaded into beads, ensuring slow release. After 188Retin colloids were added into the microspheres, TACE was performed in a rat hepatocellular carcinoma model. Single photon emission computed tomography/computed tomography imaging and biodistribution analyses showed that the microspheres were still in the liver after 72 h. During 4 weeks of observation, ultrasound images showed that the Re/DOX@MS treatment had the most significant inhibitory effect on tumor growth. Biodegradable PLGA microspheres have the advantage of enabling local embolization therapy with reduced adverse effects. In the future, microspheres could serve as a drug delivery system for cancer treatment by combining therapeutic radionuclides and chemotherapeutic drugs, thereby improving treatment effects for hepatocellular carcinoma.

EGFR-targeted micelles containing near-infrared dye for enhanced photothermal therapy in colorectal cancer.

The purpose of this research was to investigate the effectiveness of epidermal growth factor receptor (EGFR) targeted micelles loaded with IR-780 (Cetuximab/IR-780/micelles) for generating tumor targeting, multimodal images, and photothermal therapy (PTT). We initially studied the cellular uptake of these micelles using the HCT-116 and SW-620 cell lines. HCT-116 (high expression of EGFR) and SW-620 (low expression of EGFR) cell lines were used to examine biodistribution and antitumor effects of Cetuximab/IR-780/micelles. Time-lapse near-IR fluorescence (NIRF) images also indicated the highest IR-780 accumulation from Cetuximab/IR-780/micelles in HCT-116 tumors (p<0.05). HCT-116 tumors in tumor-bearing mice exhibited significantly higher accumulations of Cetuximab/IR-780/111In-micelles than SW-620 tumors in Micro-SPECT/CT imaging and biodistribution studies (p<0.05). Dual-radioisotope Nano-SPECT/CT imaging of Cetuximab/131I-IR-780/111In-micelles demonstrated simultaneous high accumulation of both IR-780 and micelles in HCT-116 tumors, but not in SW-620 tumors. Regarding antitumor effects, following the Cetuximab/IR-780/micelles with PPT on day 6, all HCT-116 tumor-bearing mice were cured. In contrast, SW-620 tumors relapsed at 13days after treatment. In summary, we expect that the Cetuximab/IR-780/micelles could enhance the antitumor effects by PTT in EGFR overexpression colorectal cancers through effective drug delivery nanoparticles.

A novel temperature-responsive micelle for enhancing combination therapy.

A novel thermosensitive polymer p(N-isopropylacrylamide-co-poly[ethylene glycol] methyl ether acrylate)-block-poly(epsilon-caprolactone), p(NIPAAM-co-PEGMEA)-b-PCL, was synthesized and developed as nanomicelles. The hydrophobic heat shock protein 90 inhibitor 17-allylamino-17-demethoxygeldanamycin and the photosensitizer cyanine dye infrared-780 were loaded into the core of the micelles to achieve both chemotherapy and photothermal therapy simultaneously at the tumor site. The release of the drug could be controlled by varying the temperature due to the thermosensitive nature of the micelles. The micelles were less than 200 nm in size, and the drug encapsulation efficiency was >50%. The critical micelle concentrations were small enough to allow micelle stability upon dilution. Data from cell viability and animal experiments indicate that this combination treatment using photothermal therapy with chemotherapy had synergistic effects while decreasing side effects.

Targeting Colorectal Cancer Stem-Like Cells with Anti-CD133 Antibody-Conjugated SN-38 Nanoparticles.

Cancer stem-like cells play a key role in tumor development, and these cells are relevant to the failure of conventional chemotherapy. To achieve favorable therapy for colorectal cancer, PEG-PCL-based nanoparticles, which possess good biological compatibility, were fabricated as nanocarriers for the topoisomerase inhibitor, SN-38. For cancer stem cell therapy, CD133 (prominin-1) is a theoretical cancer stem-like cell (CSLC) marker for colorectal cancer and is a proposed therapeutic target. Cells with CD133 overexpression have demonstrated enhanced tumor-initiating ability and tumor relapse probability. To resolve the problem of chemotherapy failure, SN-38-loaded nanoparticles were conjugated with anti-CD133 antibody to target CD133-positive (CD133(+)) cells. In this study, anti-CD133 antibody-conjugated SN-38-loaded nanoparticles (CD133Ab-NPs-SN-38) efficiently bound to HCT116 cells, which overexpress CD133 glycoprotein. The cytotoxic effect of CD133Ab-NPs-SN-38 was greater than that of nontargeted nanoparticles (NPs-SN-38) in HCT116 cells. Furthermore, CD133Ab-NPs-SN-38 could target CD133(+) cells and inhibit colony formation compared with NPs-SN-38. In vivo studies in an HCT116 xenograft model revealed that CD133Ab-NPs-SN-38 suppressed tumor growth and retarded recurrence. A reduction in CD133 expression in HCT116 cells treated with CD133Ab-NPs-SN-38 was also observed in immunohistochemistry results. Therefore, this CD133-targeting nanoparticle delivery system could eliminate CD133-positive cells and is a potential cancer stem cell targeted therapy.

Blocking heme oxygenase-1 by zinc protoporphyrin reduces tumor hypoxia-mediated VEGF release and inhibits tumor angiogenesis as a potential therapeutic agent against colorectal cancer.

BACKGROUND: Hypoxia in tumor niche is one of important factors to start regeneration of blood vessels, leading to increase survival, proliferation, and invasion in cancer cells. Under hypoxia microenvironment, furthermore, steadily increased hypoxia-inducible factor -1α (HIF-1α) is observed, and can increase vascular endothelial growth factor (VEGF) expression and promote angiogenesis. Zinc protoporphyrin (ZnPP), a heme oxygenase-1 (HO-1) inhibitor, is potential to inhibit tumor proliferation and progression. However, the mechanism of ZnPP in inhibition of tumor is not completely clear. We hypothesize that ZnPP may modulate HIF-1α through inhibiting HO-1, and then inhibit angiogenesis and tumor progression. This study aimed to dissect the mechanism of ZnPP in tumor suppression. RESULTS: We observed the amount of VEGF was increased in the sera of the colorectal cancer (CRC) patients (n = 34, p < 0.05). Furthermore, increased VEGF expression was also measured in colorectal cancer cells, HCT-15, culturing under mimicking hypoxic condition. It suggested that hypoxia induced VEGF production from cancer cells. VEGF production was significantly reduced from HCT-15 cells after exposure to HIF-1α inhibitor KC7F2, suggesting that HIF-1α regulated VEGF production. Moreover, we observed that the HO-1 inhibitor ZnPP inhibited the expressions of HIF-1α and VEGF coupled with cell proliferations of HCT-15 cells, suggesting that ZnPP blocked HIF-1α expression, and then inhibited the consequent VEGF production. In the xenograft model, we also observed that the animals exposed to ZnPP displayed much smaller tumor nodules and less degree of angiogenesis with decreased expression of the angiogenesis marker, αvß3 integrin, compared to that in normal control. CONCLUSIONS: This study demonstrated that VEGF level in serum was elevated in the patients with CRC. The HO-1 inhibitor, ZnPP, possessed the properties of anti-tumor agent by decreasing HIF-1α levels, blocking VEGF production, impairing tumor angiogenesis, and inhibiting tumor growth.

Sulfonamide derivative targeting carbonic anhydrase IX as a nuclear imaging probe for colorectal cancer detection in vivo.

Hypoxic microenvironment is a common situation in solid tumors. Carbonic anhydrase IX (CA9) is one of the reliable cellular biomarkers of hypoxia. The role of CA9 in colorectal cancer (CRC) remains to be clarified. CA9 inhibitor such as sulfonamides is known to block CA9 activation and reduce tumor growth consequently. Here, we aimed to investigate the CA9 expression in serum and tumor from different stages of CRC patients and utilize sulfonamide derivative with indium-111 labeling as a probe for CRC nuclear imaging detection in vivo. The serum CA9 was correlated with the tumor CA9 levels in different stages of CRC patients. Hypoxia increased cell viability and CA9 expression in colorectal cancer HCT-15 cells. Sulfonamide derivative 5-(2-aminoethyl)thiophene-2-sulfonamide (ATS) could bind with CA9 in vitro under hypoxia. Moreover, tumor tissues in HCT-15-induced xenograft mice possessed higher hypoxic fluorescence signal as compared with other organs. We also found that the radioisotope signal of indium-111 labeled ATS, which was utilized for CRC detection in HCT-15-induced xenograft mice, was markedly enhanced in tumors as compared with non-ATS control. Taken together, these findings suggest that CA9 is a potential hypoxic CRC biomarker and measurement of serum CA9 can be as a potential tool for diagnosing CA9 expressions in CRC clinical practice. The radioisotope-labeled sulfonamide derivative (ATS) may be useful to apply in CRC patients for nuclear medicine imaging.

111In-cetuximab as a diagnostic agent by accessible epidermal growth factor (EGF) receptor targeting in human metastatic colorectal carcinoma.

Colorectal adenocarcinoma is a common cause death cancer in the whole world. The aim of this study is to define the 111In-cetuximab as a diagnosis tracer of human colorectal adenocarcinoma. In this research, cell uptake, nano-SPECT/CT scintigraphy, autoradiography, biodistribution and immunohitochemical staining of EGF receptor were included. HCT-116 and HT-29 cell expressed a relatively high and moderate level of EGF receptor, respectively. The nano-SPECT/CT image of 111In-cetuximab showed tumor radiation uptake of subcutaneous HCT-116 xenograft tumor was higher than SW-620. Autoradiography image also showed that tumor of HCT-116 had high 111In-cetuximab uptake. Mice that bearing CT-26 in situ xenograft colorectal tumors showed similar high uptake in vivo and ex vivo through nano-SPECT/CT imaging at 72 hours. Metastatic HCT-116/Luc tumors demonstrated the highest uptake at 72 hours after the injection of 111In-cetuximab. Relatively, results of 111In-DTPA showed that metabolism through urinary system, especially in the kidney. The quantitative analysis of biodistribution showed count value of metastatic HCT-116/Luc tumors that treated with 111In-cetuximab had a significant difference (P < 0.05) compared with that treated with 111In-DTPA after injection 72 hours. Result of immunohistologic staining of EGF receptor also showed high EGF receptor expression and uptake in metastatic colorectal tumors. In summary, we suggested that 111In-cetuximab will be a potential tool for detecting EGF receptor expression in human metastatic colorectal carcinoma.

Traceable Self-Assembly of Laser-Triggered Cyanine-Based Micelle for Synergistic Therapeutic Effect.

To track nanocarriers, many researches adopt nanocarriers labeled with radiotracers or encapsulating near-infrared fluorescence (NIRF) dye. In this study, novel amphiphilic copolymers, methoxy poly(ethylene glycol) (mPEG)-cyanine-poly(ε-caprolactone) (PCL) (mPEG-Cy-PCL) are synthesized. mPEG-Cy-PCL are capable of performing NIRF imaging, photothermal therapy (PTT) on cancer cells and self-assembly nanocarriers. Cy-based micelles can encapsulate doxorubicin (Doxo@Cy-micelle) and achieve NIRF image-guided drug delivery. Doxo@Cy-micelles are nanosized micelles enhancing the accumulation of Doxo in tumor sites and decreasing side effects. Doxo@Cy-micelles exhibit an excellent PTT and synergistic chemotherapy of cancer via laser-triggered release of Doxo from micelles, eventually resulting in decreased cancer recurrence rates. The results show that Cy-based micelles are excellent nanocarriers for NIRF imaging and synergistic photothermal-chemotherapy of cancer.

188Re-HYNIC-trastuzumab enhances the effect of apoptosis induced by trastuzumab in HER2-overexpressing breast cancer cells.

PURPOSE: The development of radioimmunotherapy has provided an impressive alternative approach in improving trastuzumab therapy. However, the mechanisms of trastuzumab and radiation treatment combined to increase therapeutic efficacy are poorly understood. Here, we try to examine the efficacy of cytotoxicity and apoptosis induction for (188)Re-HYNIC-trastuzumab in cancer cell lines with various levels of Her2. MATERIALS AND METHODS: Fluorescence flow cytometry was used to detect the alterations of apoptosis induction after (188)Re-HYNIC-trastuzumab treatment in two breast cancer cell lines with different levels of HER2 (BT-474 and MCF-7) and a colorectal carcinoma cell line (HT-29) for control. RESULTS: Our results indicated that (188)Re-HYNIC-trastuzumab led to cell death of breast cancer cells specifically in HER2 level-dependent and radioactivity dose-dependent fashions. In BT-474 cells, 370 kBq/ml of (188)Re-HYNIC-trastuzumab enhanced the cytotoxicity to a level nearly 100-fold that of trastuzumab-alone treatment. The results also revealed that the mitochondria-dependent pathway attenuated irradiation-induced apoptosis in HER2-expressing breast cancer cells after (188)Re-HYNIC-trastuzumab treatment. In contrast, only after 48 h of (188)Re-HYNIC-trastuzumab treatment, BT-474 cells exhibited typical apoptotic changes, including exposure of phospholipid phosphatidylserine on the cell surface, or fragmented DNA formation, in a radioactivity dose-dependent manner. CONCLUSION: Briefly, our study demonstrates that (188)Re-labeled HYNIC-trastuzumab not only enhances cell death in a radioactivity dose-dependent fashion, but may also prolong the effects of apoptosis involved with the mitochondria-dependent pathway in HER2-overexpressing breast cancer cells. It is possible that the (188)Re-HYNIC-trastuzumab treatment induced a second round of apoptosis to prolong the effects of cell kill in these cancer cells. These data revealed that (188)Re-HYNIC-trastuzumab has the potential for use as a therapeutic radiopharmaceutical agent in HER2-overexpressing breast cancer cell treatment.

Therapeutic and scintigraphic applications of polymeric micelles: combination of chemotherapy and radiotherapy in hepatocellular carcinoma.

This study evaluated a multifunctional micelle simultaneously loaded with doxorubicin (Dox) and labeled with radionuclide rhenium-188 ((188)Re) as a combined radiotherapy and chemotherapy treatment for hepatocellular carcinoma. We investigated the single photon emission computed tomography, biodistribution, antitumor efficacy, and pathology of (188)Re-Dox micelles in a murine orthotopic luciferase-transfected BNL tumor cells hepatocellular carcinoma model. The single photon emission computed tomography and computed tomography images showed high radioactivity in the liver and tumor, which was in agreement with the biodistribution measured by γ-counting. In vivo bioluminescence images showed the smallest size tumor (P<0.05) in mice treated with the combined micelles throughout the experimental period. In addition, the combined (188)Re-Dox micelles group had significantly longer survival compared with the control, (188)ReO4 alone (P<0.005), and Dox micelles alone (P<0.01) groups. Pathohistological analysis revealed that tumors treated with (188)Re-Dox micelles had more necrotic features and decreased cell proliferation. Therefore, (188)Re-Dox micelles may enable combined radiotherapy and chemotherapy to maximize the effectiveness of treatment for hepatocellular carcinoma.

Preparation and therapeutic evaluation of (188)Re-thermogelling emulsion in rat model of hepatocellular carcinoma.

Radiolabeled Lipiodol(®) (Guerbet, Villepinte, France) is routinely used in hepatoma therapy. The temperature-sensitive hydrogel polyethylene glycol-b-poly-DL-lactic acid-co-glycolic acid-b-polyethylene glycol triblock copolymer is used as an embolic agent and sustained drug release system. This study attempted to combine the polyethylene glycol-b-poly-DL-lactic acid-co-glycolic acid-b-polyethylene glycol hydrogel and radio-labeled Lipiodol to form a new radio-thermogelling emulsion, rhenium-188-N,N'-1,2-ethanediylbis-L-cysteine diethyl-ester dihydrochloride-Lipiodol/hydrogel ((188)Re-ELH). The therapeutic potential of (188)Re-ELH was evaluated in a rodent hepatoma model. Rhenium-188 chelated with N,N'-1,2-ethanediylbis-L-cysteine diethyl-ester dihydrochloride was extracted with Lipiodol to obtain rhenium-188-N,N'-1,2-ethanediylbis-L-cysteine diethyl-ester dihydrochloride-Lipiodol ((188)Re-EL), which was blended with the hydrogel in equal volumes to develop (188)Re-ELH. The (188)Re-ELH phase stability was evaluated at different temperatures. Biodistribution patterns and micro-single-photon emission computed tomography/computed tomography images in Sprague Dawley rats implanted with the rat hepatoma cell line N1-S1 were observed after in situ tumoral injection of ~3.7 MBq (188)Re-ELH. The therapeutic potential of (188)Re-EL (48.58±3.86 MBq/0.1 mL, n=12) was evaluated in a 2-month survival study using the same animal model. The therapeutic effects of (188)Re-ELH (25.52±4.64 MBq/0.1 mL, n=12) were evaluated and compared with those of (188)Re-EL. The responses were assessed by changes in tumor size and survival rates. The (188)Re-ELH emulsion was stable in the gel form at 25°C-35°C for >52 hours. Biodistribution data and micro-single-photon emission computed tomography/computed tomography images of the (188)Re-ELH group indicated that most activity was selectively observed in hepatomas. Long-term (188)Re-ELH studies have demonstrated protracted reductions in tumor volumes and positive effects on the survival rates (75%) of N1-S1 hepatoma-bearing rats. Conversely, the 2-month survival rate was 13% in the control sham group. Therapeutic responses differed significantly between the two groups (P<0.005). Thus, the hydrogel enhanced the injection stability of (188)Re-EL in an animal hepatoma model. Given the synergistic results, direct (188)Re-ELH intratumoral injection is a potential therapeutic alternative for hepatoma treatment.

Afatinib and its encapsulated polymeric micelles inhibits HER2-overexpressed colorectal tumor cell growth in vitro and in vivo.

Colorectal cancer (CRC) is known as a common malignant neoplasm worldwide. The role of EGFR/HER2 in CRC is unclear. Afatinib is an irreversible EGFR/HER2 inhibitor. There were few studies of afatinib on CRC. Here, we investigated the protein levels/expressions of HER2 in sera and tumors from CRC patients and the therapeutic effect of afatinib on HER2-overexpressed CRC in vitro and in vivo. The increased HER2 levels were detected in the collected sera and tumors of patients with CRC. The serological HER2 levels were correlated with the tumor HER2 expressions in patients. Afatinib also inhibited the HER2-positive tumor cell growth and caused apoptosis in HER2-overexpressed human colorectal cancer HCT-15 cells but not in low HER2 expressed human gastric cancer MKN45 cells. In vivo study showed that afatinib reduced tumor growth in HER2-overexpressed xenografts. Moreover, afatinib-encapsulated micelles displayed higher cytotoxic activity in HCT-15 cells and were more effective for tumor growth suppression in HCT-15-induced tumor xenografts than afatinib performance alone. Taken together, these findings suggest that higher serum HER2 levels reflect the higher HER2 contents in tumors of CRC patients, and the improved afatinib-encapsulated micelles possess high therapeutic efficacy in HER2-overexpressed CRC in vitro and in vivo.

Radioprotective effects of Antrodia cinnamomea are enhanced on immune cells and inhibited on cancer cells.

UNLABELLED: Abstract Purpose: The radioprotective effects of Antrodia cinnamomea (AC) were investigated for understanding the potential usefulness of AC as an adjunct treatment for reducing radiation side-effects. MATERIALS AND METHODS: In this study, we determined the ability of AC extracts (AC539) to reduce radiation side-effects by analyzing cellular viability in normal mouse spleen immune cells and human cancer cells with different radiosensitivity. We further detected the effect of AC on radiation-induced changes in cytokine- and inflammatory-related gene expressions. Furthermore, apoptosis assay was performed to determine whether AC could inhibit radiation-induced cytotoxicity. RESULTS: We found that an AC dose of 100-150 µg/ml in a time-dependent manner was the most effective in blocking radiation-induced cytotoxicity, in vitro. Radiation-induced cytotoxicity was inhibited in spleen immune cells by 37-56%; however, pretreatment of human colorectal cancer cell line HT-29 with AC did not have any effect on radiation-induced cytotoxicity, while pretreatment of radiosensitive human breast cancer cell lines BT-474 with AC caused a moderate enhancement of radiation-induced damage. Furthermore, AC pretreatment differentially regulated the mRNA expression of several important immunomodulatory genes in response to irradiation in normal and cancer cells. CONCLUSIONS: Our data indicate that AC may inhibit important immunoregulatory signaling which could be vital in the avoidance of an over-activated cytotoxic and inflammatory response of the immune system caused by radiation-induced tissue damage. Additionally, AC does not provide a radioprotective effect to tumor cells but instead enhances radiation-induced inflammation and cytotoxicity in cancer.

Neutrophil elastase as a diagnostic marker and therapeutic target in colorectal cancers.

Neutrophil elastase (NE), a serine protease secreted by neutrophils, contributes to the progression of cancers to enhance tumor invasion and metastasis. It has been well reported that the regions surrounding the colorectal cancerous tissues usually are decorated with increased accumulation or aggregation of neutrophils coupled with a higher deposition/expression of NE. Therefore, we hypothesized that an increased expressional level of NE in patients with colorectal cancer (CRC) may represent as one of putative biomarkers for CRC. The aim of this study was to evaluate and assure our hypothesis by measurements of the expressional level of NE in the sera and tissues from CRC patients. Moreover, we also proposed a potential therapeutic strategy by blocking enzymatic activity of NE using sivelestat to inhibit the progression of tumor developments. The infiltrated numbers of neutrophils from specimen tissues of CRC patients, and the secreted forms of NE in the sera were quantitatively measured and compared. To evaluate the serum NE as one of putative biomarkers of CRC patients, the receiver operating characteristic (ROC) curve was made to determine the cut-off value of NE in sera for assurance of CRC diagnosis. To evaluate NE as therapeutic target for CRC, sivelestat, a NE inhibitor, was used and administrated into the CRC xenografts. NE expression level coupled with tumor volume were measured and compared between the control and sivelestat-treated xenografts. We found that more infiltrated neutrophils and an increased NE expression were detected in the cancerous tissues compared to the normal tissues. The serum NE concentration in CRC patients was statistically higher than that in the healthy controls (0.56 ± 0.08 µg/ml vs. 0.22 ± 0.03 µg/ml) (p<0.05), indicating that serum NE can potentially be a putative marker of CRC. To characterize the role of NE in tumorigenesis, the NE activity was detected in HCT-15-xenografts using in vivo imaging system (IVIS). Compare to normal mice, the amounts of active NE in xenografts are significantly higher than normal control animals. In the therapeutic characterizing studies, we found that sivelestat can inhibit tumor growth in the HCT-15-induced xenografts. This study suggests that NE is not only as a putative diagnostic biomarker of CRC, but also a potential therapeutic target for patients suffered with CRC.