Synergistic Enhancement of Antitumor Effects by Combining Abemaciclib with Desipramine.

Cyclin-dependent kinase 4 and 6 (CDK4/6) inhibitors, including abemaciclib, have been approved for the treatment of hormone receptor-positive, human epidermal growth factor receptor 2 (HER2)-negative advanced, and metastatic breast cancer. Despite the high therapeutic efficacy of CDK4/6 inhibitors, they are associated with various adverse effects, including potentially fatal interstitial lung disease. Therefore, a combination of CDK4/6 inhibitors with letrozole or fulvestrant has been attempted but has demonstrated limitations in reducing adverse effects, highlighting the need to develop new combination therapies. This study proposes a combination strategy using CDK4/6 inhibitors and tricyclic antidepressants to enhance the therapeutic outcomes of these inhibitors while reducing their side effects. The therapeutic efficacies of abemaciclib and desipramine were tested in different cancer cell lines (H460, MCF7, and HCT-116). The antitumor effects of the combined abemaciclib and desipramine treatment were evaluated in a xenograft colon tumor model. In vitro cell studies have shown the synergistic anticancer effects of combination therapy in the HCT-116 cell line. The combination treatment significantly reduced tumor size compared with control or single treatment without causing apparent toxicity to normal tissues. Although additional in vivo studies are necessary, this study suggests that the combination therapy of abemaciclib and desipramine may represent a novel therapeutic approach for treating solid tumors.

Development of Intraoperative Near-Infrared Fluorescence Imaging System Using a Dual-CMOS Single Camera.

We developed a single-camera-based near-infrared (NIR) fluorescence imaging device using indocyanine green (ICG) NIR fluorescence contrast agents for image-induced surgery. In general, a fluorescent imaging system that simultaneously provides color and NIR images uses two cameras, which is disadvantageous because it increases the imaging head of the system. Recently, a single-camera-based NIR optical imaging device with quantum efficiency partially extended to the NIR region was developed to overcome this drawback. The system used RGB_NIR filters for camera sensors to provide color and NIR images simultaneously; however, the sensitivity and resolution of the infrared images are reduced by 1/4, and the exposure time and gain cannot be set individually when acquiring color and NIR images. Thus, to overcome these shortcomings, this study developed a compact fluorescent imaging system that uses a single camera with two complementary metal-oxide semiconductor (CMOS) image sensors. Sensitivity and signal-to-background ratio were measured according to the concentrations of ICG solution, exposure time, and camera gain to evaluate the performance of the imaging system. Consequently, the clinical applicability of the system was confirmed through the toxicity analysis of the light source and in vivo testing.

Mini-Platform for Off-On Near-Infrared Fluorescence Imaging Using Peptide-Targeting Ligands.

Here, we propose a zwitterionic near-infrared (NIR) fluorophore-tryptophan (Trp) conjugate with a cleavable linker as a minimal-sized versatile platform (MP) for the preparation of peptide ligand-based off-on type molecular probes. The zwitterionic NIR fluorophore in MP undergoes fluorescence quenching via a photoinduced electron transfer mechanism when in close proximity to tryptophan, and nonspecific binding with serum proteins is minimized by the zwitterionicity of the fluorophore. The linker can be cleaved inside cancer cells in response to tumor-associated stimuli. As a proof-of-concept experiment, ATTO655 was covalently linked with Trp via a diarginine linker to form an MP. A cyclic peptide consisting of Arg-Gly-Asp-d-Phe-Lys (cRGD) was used as a cancer-targeting ligand and was conjugated to the MP to form cRGD-MP. The NIR fluorescence of cRGD-MP could be selectively turned on inside the target cancer cells, thereby enabling specific fluorescence imaging of integrin αvß3-overexpressing cancer cells in vitro and in vivo.

Targeted, Stimuli-Responsive, and Theranostic 19F Magnetic Resonance Imaging Probes.

Unlike conventional 1H magnetic resonance imaging (MRI), 19F MRI features unambiguous detection of fluorine spins due to negligible background signals. Therefore, it is considered a promising noninvasive and selective imaging method for the diagnosis of cancers and other diseases. For 19F MRI, fluorine-rich molecules such as perfluorocarbons (PFC) have been formulated into nanoemulsions and used as its tracer agent. Along with advancements in other types of nanoparticles as targeted theranostics and stimuli-triggered probes and combined with the advantages of 19F MRI, PFC nanoemulsions are being empowered with these additional functionalities and becoming a promising theranostic platform. In this Review, we provide an overview of fluorine-based materials for sensitive 19F MRI of biological and pathological conditions. In particular, we describe designs and applications of recently reported stimuli-responsive and theranostic 19F MRI probes. Finally, challenges and future perspectives regarding the further development of 19F MRI probes for their clinical applications are described.

Multivalent Mannose-Decorated NIR Nanoprobes for Targeting Pan Lymph Nodes.

Lymphadenectomy is a prerequisite for most malignancies to define the precise staging of cancer, as well as resect the possible metastases completely. While it improves prognosis, lymphadenectomy often causes postoperative edema or bleeding because of unclear surgical margins. In this study, we synthesized near-infrared (NIR) fluorescent nanoprobes with conjugating various mannose moieties on the surface to target macrophages in the lymph node. Armed with these NIR nanoprobes, we demonstrated the feasibility of intraoperative pan lymph nodes (PLN) mapping and real-time optical imaging under the NIR fluorescence imaging system. We found that even single mannose-conjugated ZW800-1 showed specific uptake in lymph nodes within 4 h, and multiple mannose-employed polyrotaxanes highlighted PLN efficiently with low background signals in major organs. This technology can help surgeons perform lymphadenectomy with ease and safety by identifying all regional lymph nodes proficiently after a single intravenous injection of NIR nanoprobes.

Indocyanine green-loaded hollow mesoporous silica nanoparticles as an activatable theranostic agent.

Here we report indocyanine green (ICG)-loaded hollow mesoporous silica nanoparticles (ICG@HMSNP) as an activatable theranostic platform. Near-infrared fluorescence and singlet oxygen generation of ICG@HMSNP was effectively quenched (i.e. turned off) in its native state because of the fluorescence resonance energy transfer between ICG molecules. Therefore, ICG@HMSNP was nonfluorescent and nonphototoxic in the extracellular region. After the nanoparticles entered the cancer cells via endocytosis, they became highly fluorescent and phototoxic. In addition, intracellular uptake of ICG@HMSNP was 2.75 times higher than that of free ICG, resulting in an enhanced phototherapy of cancer.

Polypyrrole-based nanotheranostics for activatable fluorescence imaging and chemo/photothermal dual therapy of triple-negative breast cancer.

Here, we fabricated polypyrrole nanoparticles (PPys) (termed HA10-PPy, HA20-PPy, and HA40-PPy) doped with different average molecular weight hyaluronic acids (HAs) (10, 20, and 40 kDa, respectively), and evaluated the effect of molecular weight of doped HA on photothermal induction, fluorescence quenching, and drug loading efficiencies. Doxorubicin-loaded HA-doped PPys (DOX@HA-PPys) could be used for imaging and therapy of triple-negative breast cancer (TNBC). Fluorescence turn-on, stimuli-responsive drug release, and photo-induced heating of DOX@HA-PPys enabled not only activatable fluorescence imaging but also subsequent chemo/photothermal dual therapy for TNBC. In particular, we illustrated the potential usefulness of the photothermal effect of the nanoparticles for overcoming chemoresistance in TNBC.

A novel endoscopic fluorescent band ligation method for tumor localization.

BACKGROUND: Accurate tumor localization is essential for minimally invasive surgery. This study describes the development of a novel endoscopic fluorescent band ligation method for the rapid and accurate identification of tumor sites during surgery. METHODS AND MATERIALS: The method utilized a fluorescent rubber band, made of indocyanine green (ICG) and a liquid rubber solution mixture, as well as a near-infrared fluorescence laparoscopic system with a dual light source using a high-powered light-emitting diode (LED) and a 785-nm laser diode. The fluorescent rubber bands were endoscopically placed on the mucosae of porcine stomachs and colons. During subsequent conventional laparoscopic stomach and colon surgery, the fluorescent bands were assayed using the near-infrared fluorescence laparoscopy system. RESULTS: The locations of the fluorescent clips were clearly identified on the fluorescence images in real time. The system was able to distinguish the two or three bands marked on the mucosal surfaces of the stomach and colon. Resection margins around the fluorescent bands were sufficient in the resected specimens obtained during stomach and colon surgery. CONCLUSION: These novel endoscopic fluorescent bands could be rapidly and accurately localized during stomach and colon surgery. Use of these bands may make possible the excision of exact target sites during minimally invasive gastrointestinal surgery.

A New Strategy for Fluorogenic Esterase Probes Displaying Low Levels of Non-specific Hydrolysis.

A new design for fluorescence probes of esterase activity that features a carboxylate-side pro-fluorophore is demonstrated with boron dipyrromethene (BODIPY)-based probes 1 a and 1 b. Because the design relies on the enzyme-catalyzed hydrolysis of an ester group that is not electronically activated, these probes exhibit a stability to background hydrolysis that is far superior to classical alcohol-side profluorophore-based probes, large signal-to-noise ratios, reduced sensitivity to pH variations, and high enzymatic reactivity. The utility of probe 1 a was established with a real-time fluorescence imaging experiment of endogenous esterase activity that does not require washing of the extracellular medium.

Electroactive polypyrrole nanowire arrays: synergistic effect of cancer treatment by on-demand drug release and photothermal therapy.

An electroresponsive drug release system based on polypyrrole (Ppy) nanowires was developed to induce the local delivery of anticancer drug, doxorubicin (DOX), according to the applied electric field. DOX-conjugated Ppy nanowire (NW) (DOX/Ppy NW) array was initially prepared by electrochemical deposition of a mixture of pyrrole monomers and biotin as dopants in the anodic alumina oxide membrane as a sacrificial template. Morphological observation by scanning electron microscopy revealed free-standing and 3D nanotopographical features with large surface area and high density. In addition, we investigated the antitumor efficacy of DOX released from DOX/Ppy NW array in response to the external electric field using two kinds of cancer cell lines, human oral squamous carcinoma cells (KB cells) and human breast cancer cells (MCF7 cells). Meanwhile, strong photothermal effect as a result of a near-infrared absorbing ability of Ppy synergistically maximizes the chemotherapeutic efficacy. Our results suggested that the proposed multifunctional Ppy platform possessing several beneficial features is very promising for many therapeutic applications including cancer.

Photodynamic therapy using a protease-mediated theranostic agent reduces cathepsin-B activity in mouse atheromata in vivo.

OBJECTIVE: To investigate whether an intravenously injected cathepsin-B activatable theranostic agent (L-SR15) would be cleaved in and release a fluorescent agent (chlorin-e6) in mouse atheromata, allowing both the diagnostic visualization and therapeutic application of these fluorophores as photosensitizers during photodynamic therapy to attenuate plaque-destabilizing cathepsin-B activity by selectively eliminating macrophages. APPROACH AND RESULTS: Thirty-week-old apolipoprotein E knock-out mice (n=15) received intravenous injection of L-SR15 theranostic agent, control agent D-SR16, or saline 3× (D0, D7, D14). Twenty-four hours after each injection, the bilateral carotid arteries were exposed, and Cy5.5 near-infrared fluorescent imaging was performed. Fluorescent signal progressively accumulated in the atheromata of the L-SR15 group animals only, indicating that photosensitizers had been released from the theranostic agent and were accumulating in the plaque. After each imaging session, photodynamic therapy was applied with a continuous-wave diode-laser. Additional near-infrared fluorescent imaging at a longer wavelength (Cy7) with a cathepsin-B-sensing activatable molecular imaging agent showed attenuation of cathepsin-B-related signal in the L-SR15 group. Histological studies demonstrated that L-SR15-based photodynamic therapy decreased macrophage infiltration by inducing apoptosis without significantly affecting plaque size or smooth muscle cell numbers. Toxicity studies (n=24) showed that marked erythematous skin lesion was generated in C57/BL6 mice at 24 hours after intravenous injection of free chlorin-e6 and ultraviolet light irradiation; however, L-SR15 or saline did not cause cutaneous phototoxicity beyond that expected of ultraviolet irradiation alone, neither did we observe systemic toxicity or neurobehavioral changes. CONCLUSIONS: This is the first study showing that macrophage-secreted cathepsin-B activity in atheromata could be attenuated by photodynamic therapy using a protease-mediated theranostic agent.

Development of polysaccharide-complexed nano-sized rice protein dispersion.

The objective of this study was to improve water solubility of the rice protein (RP) by forming complexes with anionic polysaccharides, such as sodium alginate (SA) and xanthan gum (XG). In addition, utilization of the RP complexes as an emulsifier was evaluated. The prepared RP-SA or RP-XG complexes were analyzed by measuring their particle size, ζ-potential, and water solubility as well as by confocal laser scanning microscopy. The formation of a complex between RP-SA and RP-XG improved the water solubility and dispersibility of RP over a wide range of pH values (3, 5, 7, and 9). Confocal fluorescence images showed that the aggregation of RP molecules was prevented by the formation of complexes between RP and polysaccharides. When soybean oil-in-water emulsions were prepared with complexes, RP-SA (ratio 4:1) and RP-XG(ratio 4:1) complex-stabilized emulsions were stable for 4 weeks of storage.

A ratiometric fluorescent probe based on a BODIPY-DCDHF conjugate for the detection of hypochlorous acid in living cells.

A colorimetric and ratiometric fluorescent probe consisting of a boron-dipyrromethene (BODIPY) dye conjugated with a 2-dicyanomethylene-3-cyano-2,5-dihydrofuran (DCDHF) group has been designed for the selective and sensitive detection of HOCl/OCl(-)via oxidative cleavage of an alkene linker between BODIPY and DCDHF.

Real-Time Monitoring of Colorectal Cancer Location and Lymph Node Metastasis and Photodynamic Therapy Using Fucoidan-Based Therapeutic Nanogel and Near-Infrared Fluorescence Diagnostic-Therapy System.

We report real-time monitoring of colorectal cancer, lymph node metastasis of colorectal cancer cells, and tumor growth inhibition through photodynamic therapy (PDT) using a near-infrared fluorescence diagnostic-therapy system with a light source for PDT and a fucoidan-based theranostic nanogel (CFN-gel) with good accumulation efficiency in cancer cells. To confirm the effect of the fabricated system and developed CFN-gel, in vitro and in vivo experiments were performed. Chlorin e6 (Ce6) and 5-aminolevulinic acid (5-ALA) were used for comparison. We confirmed that CFN-gel has a high accumulation efficiency in cancer cells and high fluorescence signals in near-infrared light for a long period, and only CFN-gel delayed the growth rate of cancer in terms of its size in PDT. In addition, using the near-infrared fluorescence diagnostic-therapy system and CFN-gel prepared for these experiments, the lymph node metastasis of cancer cells was imaged in real time, and the metastasis was confirmed through H&E staining. The possibility of image-guided surgery and identification of lymph node metastasis in colorectal cancer can be confirmed through CFN-gel and a near-infrared fluorescence diagnostic-therapy system that includes various light sources.

FOXM1 Inhibition Enhances the Therapeutic Outcome of Lung Cancer Immunotherapy by Modulating PD-L1 Expression and Cell Proliferation.

Programmed death-ligand 1 (PD-L1) is a major target to cancer immunotherapy, and anti-PD-L1 and anti-PD-1 antibody-mediated immunotherapy are being increasingly used. However, immune checkpoint inhibitors (ICIs) are ineffective in treating large tumors and cause various immune-related adverse events in nontarget organs, including life-threatening cardiotoxicity. Therefore, the development of new therapeutic strategies to overcome these limitations is crucial. The focus of this study is the forkhead box protein M1 (FOXM1), which is identified as a potential therapeutic target for cancer immunotherapy and is associated with the modulation of PD-L1 expression. Selective small interfering RNA knockdown of FOXM1 or treatment with thiostrepton (TST) significantly reduces PD-L1 expression in non-small-cell lung cancer (NSCLC) cells and inhibits proliferation. Chromatin immunoprecipitation-PCR reveals that FOXM1 selectively upregulates PD-L1 expression by binding directly to the PD-L1 promoter. In vivo animal studies have shown that TST treatment significantly downregulates PD-L1 expression in human NSCLC tumors, while greatly reducing tumor size without side effects on normal tissues. Combined treatment with TST and anti-4-1BB antibody in the LLC-1 syngeneic tumor model induces synergistic therapeutic outcomes against immune resistant lung tumors as well as 2.72-folds higher CD3+ T cells in tumor tissues compared to that in the anti-4-1BB antibody treatment group.

Early On-Treatment Prediction of the Mechanisms of Acquired Resistance to EGFR Tyrosine Kinase Inhibitors.

BACKGROUND: Prediction of resistance mechanisms for epidermal growth factor receptor-tyrosine kinase inhibitors (EGFR-TKIs) remains challenging. Thus, we investigated whether resistant cancer cells that expand shortly after EGFR-TKI treatment would eventually cause the resistant phenotype. METHODS: We generated two EGFR-mutant lung cancer cell lines resistant to gefitinib (PC9GR and HCC827GR). The parent cell lines were exposed to short-term treatment with gefitinib or paclitaxel and then were assessed for EGFR T790M mutation and C-MET expression. These experiments were repeated in vivo and in clinically relevant patient-derived cell (PDC) models. For validation in clinical cases, we measured these gene alterations in plasma circulating tumor DNA (ctDNA) before and 8 weeks after starting EGFR-TKIs in four patients with EGFR-mutant lung cancer. RESULTS: T790M mutation was only detected in the PC9GR cells, whereas C-MET amplification was detected in the HCC827GR cells. The T790M mutation level significantly increased in PC9 cells after short-term treatment with gefitinib but not in the paclitaxel. C-MET mRNA expression was only significantly increased in gefitinib-treated HCC827 cells. We confirmed that the C-MET copy number in HCC827 cells that survived after short-term gefitinib treatment was significantly higher than that in dead HCC827 cells. These findings were reproduced in the in vivo and PDC models. An early on-treatment increase in the plasma ctDNA level of these gene alterations was correlated with the corresponding resistance mechanism to EGFR-TKIs, a finding that was confirmed in post-treatment tumor tissues. CONCLUSIONS: Early on-treatment kinetics in resistance-related gene alterations may predict the final mechanism of EGFR-TKI resistance.

Effects of gold nanorod concentration on the depth-related temperature increase during hyperthermic ablation.

The photothermal properties of gold nanorods (GNRs) provide an opportunity for the clinical application of highly efficient and tumor-specific photothermal therapy. For the effective hyperthermic ablation of tumor tissue using GNRs, it is essential to maintain a homogeneous therapeutic temperature in the target tissue during treatment. This study investigates whether the concentration of GNRs affects the distribution of the temperature increase during hyperthermal therapy. The investigation is conducted using polyacrylamide phantoms containing varying amounts of GNRs. In 0.1, 0.25, and 0.5 nM GNR-suspended phantoms, the change in temperature is relatively uniform along the depth of each phantom during laser irradiation at 2 W cm(-2) . In 1.0, 2.0, and 5.0 nM GNR-suspended phantoms, the rates of temperature increase in the deep regions of the phantoms decrease with increasing GNR concentration. At a laser irradiation of 5 W cm(-2) , the temperature of the GNR-suspended phantoms increases at a faster rate, whereas the range of GNR concentrations for maintaining the homogeneity of the temperature increase is not affected. This suggests that the concentration of GNRs is the major determinant of the depth-related temperature increase during hyperthermic ablation. Therefore, prior to the clinical application of hyperthermic ablation using GNRs, the concentration of GNRs has to be optimized to ensure a homogeneous distribution of therapeutic temperature in the targeted tissue.

Atorvastatin and clopidogrel interfere with photosensitization in vitro.

Photodynamic therapy (PDT) has been used to eliminate undesired cells by using a combination of photosensitizers and light illumination to generate reactive oxygen species. There is great interest in applying PDT to atherosclerosis; preferential destruction of pro-inflammatory macrophages in atheromata might attenuate plaque growth or rupture-prone vulnerability. Here, we report on a previously unknown interaction between cardiovascular drugs that are commonly prescribed for atherosclerosis patients and the cytolytic effects of photodynamic therapy using Cathepsin B activatable photosensitizer L-SR15 on murine macrophage Raw 264.7 cells in culture. Atorvastatin and clopidogrel significantly interfered with in vitro photosensitization effect while aspirin did this to a lesser extent; these drugs did not change the efficiency of cellular uptake of L-SR15 after in vitro photosensitization. A photosensitization interference effect of atorvastatin and clopidogrel was also observed when using a conventional photosensitizer free Ce6 or NCI-H1299 cancer cells. Considering the clinical implications of PDT, our study merits further investigation in clinical settings as well as in animal models.

A novel endoscopic fluorescent clip for the localization of gastrointestinal tumors.

BACKGROUND: Accurate tumor localization is essential for gastrointestinal surgery, especially in cases of early cancer. This study was designed to develop a novel fluorescent clip for rapid and exact visualization of tumor sites. METHODS: A transparent polymer matrix containing highly bright fluorochromes was coated on the front end of endoscopic clips. The fluorescent clips were placed on the mucosal surface of a porcine colon and stomach, and the operator then attempted to identify the fluorescent clips from the outer serosal side of the colon and stomach. A 532-nm diode laser and filter glass were used for visualizing the fluorescence signals through the colonic tissue. A 650-nm diode laser and a digital charge-coupled device (CCD) camera equipped with a bandpass emission filter were used for the imaging of the fluorescent clips through the thick stomach tissue. RESULTS: When a green light from a 532-nm diode laser (power density=0.35 mW/cm2) was applied on the serosal surface of the porcine colon, we could identify all clips that had been placed endoscopically on the mucosal surface of the inner colonic wall. By using the light from a 650-nm diode laser (power density=0.7 mW/cm2), we identified all fluorescent clips through the stomach wall in real time. Similar results were also obtained with the filtered xenon lamp. CONCLUSION: An endoscopic fluorescent clip can be useful for the rapid and exact localization of tumors, and this technique can also be useful during laparoscopic surgery.

Effects of enzymatically modified starch on the encapsulation efficiency and stability of water-in-oil-in-water emulsions.

The present study was performed to investigate the possibility of using 4-α-glucanotransferase (4αGTase)-treated starch in W/O/W emulsions to increase their encapsulation efficiency (EE) and stability. Emulsions were prepared using soybean oil, polyglycerol polyricinoleate (PGPR), 4αGTase-treated starch and Tween 20. The mean diameter of W/O/W droplets ranged from 4 to 10µm depending on the sonication time. When the dye was loaded in the internal water phase, the emulsion prepared by sonication for 1 and 2min showed a high EE of the dye (>90%). The W/O/W emulsion prepared by sonication for 3min showed an EE of<90%, but this EE was improved by adding 4αGTase-treated starch to the internal water phase. 4αGTase-treated starch was added to the internal water phase of W/O/W emulsions prepared with a low concentration of PGPR, and the PGPR concentration required to maintain an EE>90% was reduced. W/O/W emulsions containing 4αGTase-treated starch also showed better stability against heating and shearing stresses. These results indicated that 4αGTase-treated starch could be used in the preparation of W/O/W emulsions, which would allow the formulation of W/O/W emulsions with a reduced surfactant concentration.