Synergetic Pyroptosis with Apoptosis Improving Phototherapy of Mitochondria-Targeted Cyanines with Superior Photostability.

Pyroptosis has been reported to improve the antitumor effect by evoking a more intense immune response and a therapeutic effect. For phototherapy, several photosensitizers have been found to initiate pyroptosis. However, the effect of pyroptosis associated with apoptosis in enhancing the antitumor therapy needs sufficient characterization, especially under long-term treatment. As a NIR photosensitizer, heptamethine cyanines have been discovered for anticancer phototherapy for deep tissue penetration and inherent tumor-targeted capability. However, they are not quite stable for long-term performance. To investigate the effect of pyroptosis along with apoptosis on the anticancer immune responses and phototherapy, here, we chemically modulate the cyanine IR780 to regulate hydrophobicity, stability, and intracellular targeting. Two photosensitizers, T780T-TPP and T780T-TPP-C12, were finally optimized and showed excellent photostability with high photothermal conversion efficiency. Although the cellular uptake of the two molecules was both mediated by OATP transporters, T780T-TPP induced tumor cell death via pyroptosis and apoptosis and accumulated in tumor accumulation, while T780T-TPP-C12 was prone to accumulate in the liver. Ultimately, via one injection-multiple irradiation treatment protocol, T780T-TPP displayed a significant antitumor effect, even against the growth of large tumors (200 mm3).

Mucus-Penetrating Nanoassembly as Potential Oral Phototherapeutic Formulation against Multi-Drug Resistant Helicobacter pylori Infection.

Helicobacter pylori (H. pylori) infection presents increasing challenges to antibiotic therapies in limited penetration through gastric mucus, multi-drug resistance (MDR), biofilm formation, and intestinal microflora dysbiosis. To address these problems, herein, a mucus-penetrating phototherapeutic nanomedicine (RLs@T780TG) against MDR H. pylori infection is engineered. The RLs@T780TG is assembled with a near-infrared photosensitizer T780T-Gu and an anionic component rhamnolipids (RLs) for deep mucus penetration and light-induced anti-H. pylori performances. With optimized suitable size, hydrophilicity and weak negative surface, the RLs@T780TG can effectively penetrate through the gastric mucus layer and target the inflammatory site. Subsequently, under irradiation, the structure of RLs@T780TG is disrupted and facilitates the T780T-Gu releasing to target the H. pylori surface and ablate multi-drug resistant (MDR) H. pylori. In vivo, RLs@T780TG phototherapy exhibits impressive eradication against H. pylori. The gastric lesions are significantly alleviated and intestinal bacteria balance is less affected than antibiotic treatment. Summarily, this work provides a potential nanomedicine design to facilitate in vivo phototherapy in treatment of H. pylori infection.

Mitochondria-Mediated HSP Inhibition Strategy for Enhanced Low-Temperature Photothermal Therapy.

Low-temperature photothermal therapy (PTT) has the advantage of causing less damage to normal tissues and has attracted great attention in recent years. However, the efficacy of low-temperature PTT is restricted by the overexpression of heat shock proteins (HSPs), specifically HSP70 and HSP90. Inhibiting the function of these HSPs is a major strategy used in the development of new cancer therapies. Herein, we designed four T780T-containing thermosensitive nanoparticles to interrupt the energy supply for HSP expression using their TPP-based mitochondrial targeting action. The reversal behavior of the nanoparticles on the gambogic acid (GA)-induced compensatory increase of HSP70 was investigated in vitro by Western blot and in vivo by immunohistochemistry. The in vivo anticancer efficacy of the low-temperature PTT based on these thermosensitive nanoparticles was also systematically examined. The design proposes for the first time to utilize and elucidate the mechanism of the mitochondrial targeting of T780T-containing NPs in synergy with the HSP90 inhibition of GA to achieve an effective low-temperature PTT. This work not only provides a novel pathway for the dual inhibition of HSP70 and HSP90 but also opens up a new approach for low-temperature PTT of tumors.

Phototherapy and Mechanism Exploration of Biofilm and Multidrug-Resistant Helicobacter pylori by Bacteria-Targeted NIR Photosensitizer.

Helicobacter pylori (H. pylori) infection has been the leading cause of gastric cancer development. In recent years, the resistance of H. pylori against antibiotic treatment has been a great challenge for most countries worldwide. Since biofilm formation is one of the reasons for the antibiotic resistance of H. pylori, and phototherapy has emerged as a promisingly alternative antibacterial treatment, herein the bacteria-targeted near-infrared (NIR) photosensitizer (T780T-Gu) by combining positively-charged guanidinium (Gu) with an efficient phototherapeutic agent T780T is developed. The proposed molecule T780T-Gu exhibits synergistic photothermal therapy/photodynamic therapy effect against both H. pylori biofilms and multidrug-resistant (MDR) clinical strains. More importantly, the phototherapy mechanism of T780T-Gu acquired by the RNA-seq analysis indicates that structural deficiency as well as a decrease in metabolism and defense activity are the possible reasons for the efficient H. pylori phototherapy.

Synergistic Release of Photothermal Molecules from Nanocarriers Induced by Light and Hyperthermia Benefits Efficient Anticancer Phototherapy.

Recently, nanoformulations have been widely applied in the delivery of organic photothermal agents (OPTAs) for cancer therapy to prolong blood circulation or improve tumor-targeting capacity. However, the systematic evaluations of their effects on the photothermal behavior of OPTAs are limited, especially for different types of nanoparticle systems. Herein, we prepared two kinds of nanoparticles (BSA and PEG nanoparticles (NPs)) to load an OPTA, a cyanine photosensitizer (IR780-O-TPE), and investigated their photothermal response, organelle targeting, and in vivo therapeutic efficacy. Due to different assembly forms, the two NPs showed distinct morphological changes after exposure to laser or hyperthermia. Under laser irradiation at 808 nm, BSA NPs could release IR780-O-TPE more efficiently than PEG NPs. We speculate that this phenomenon is probably caused by dual-responsive release of IR780-O-TPE from BSA NPs against light and hyperthermia. Moreover, IR780-O-TPE/BSA NPs were highly mitochondria-targeting and therefore displayed significant inhibition of cell viability. In contrast, IR780-O-TPE/PEG NPs were "shell-core" nanostructures and more stable under laser stimulation. As a consequence, the mitochondria-targeting and anticancer photothermal therapy by IR780-O-TPE/PEG NPs was less obvious. This study revealed the significance of nanocarrier design for OPTA delivery and demonstrated that BSA NPs could release IR780-O-TPE more effectively for efficient photothermal therapy. We also believe that the dual-responsive release of OPTAs from NPs can provide an effective strategy to promote anticancer photothermal treatment.

A Tumor-Targeting Near-Infrared Heptamethine Cyanine Photosensitizer with Twisted Molecular Structure for Enhanced Imaging-Guided Cancer Phototherapy.

In recent years, cancer phototherapy has been extensively studied as noninvasive cancer treatment. To present efficient recognition toward cancer cells, most photosensitizers (PSs) are required to couple with tumor-targeted ligands. Interestingly, the heptamethine cyanine IR780 displays an intrinsic tumor-targeted feature even without modification. However, the photothermal efficacy and photostability of IR780 are not sufficient enough for clinical use. Herein, we involve a twisted structure of tetraphenylethene (TPE) between two molecules of IR780 to improve the photothermal conversion efficiency (PCE). The obtained molecule T780T shows strong near-infrared (NIR) fluorescence and improved PCE (38.5%) in the dispersed state. Also, the photothermal stability and ROS generation capability of T780T at the NIR range (808 nm) are both promoted. In the aqueous phase, the T780T was formulated into uniform nanoaggregates (∼200 nm) with extremely low fluorescence and PTT response, which would reduce in vivo imaging background and side effect of PTT response in normal tissues. After intravenous injection into tumor-bearing mice, the T780T nanoaggregates display high tumor accumulation and thus remarkably inhibit the tumor growth. Moreover, the enhanced photostability of the T780T allows for twice irradiation after one injection and leads to more significant tumor inhibition. In summary, our study presents a tumor-targeted small-molecule PS for efficient cancer therapy and brings a new design of heptamethine cyanine PS for potential clinical applications.

Improved cancer phototheranostic efficacy of hydrophobic IR780 via parenteral route by association with tetrahedral nanostructured DNA.

As a photosensitizer with effective photothermal (PTT) and photodynamic (PDT) response, IR780 has been widely explored as promising cancer phototheranostic molecule. However, the systematic administration of IR780 usually suffers from poor water solubility and low photostability, so that it cannot be administrated by parenteral route. In this study, we design a tetrahedral DNA (Td)-based nanosystem to load IR780 (IR780@Td) via electrostatic interaction and π-π stacking. After encapsulation, the water solubility and photostability of IR780 have been greatly improved, and the IR780@Td shows an appropriate nanoformulated size (224 nm) to facilitate hyperthermia-mediated tumor targeting by EPR effect. The nanostructure of Td is proved to be crucial for the proper size and good stability of IR780@Td nanoformulation for in vivo application. The in vitro and ex vivo PTT/PDT efficiencies of IR780 are improved in IR780@Td group. In the tumor-bearing mice, the accumulation of IR780 in tumor site is significantly high in IR780@Td group. Under near-infrared laser irradiation, the intravenous administration of IR780@Td promotes the tumor imaging and enhances anti-tumor effect than IR780 treatment. In summary, the proposed strategy shows promising effect in facilitating intravenous injection of IR780 and enhancing the phototheranostic efficacy for cancer treatment.

Effect of an Artificial Intelligence Clinical Decision Support System on Treatment Decisions for Complex Breast Cancer.

PURPOSE: To examine the impact of a clinical decision support system (CDSS) on breast cancer treatment decisions and adherence to National Comprehensive Cancer Center (NCCN) guidelines. PATIENTS AND METHODS: A cross-sectional observational study was conducted involving 1,977 patients at high risk for recurrent or metastatic breast cancer from the Chinese Society of Clinical Oncology. Ten oncologists provided blinded treatment recommendations for an average of 198 patients before and after viewing therapeutic options offered by the CDSS. Univariable and bivariable analyses of treatment changes were performed, and multivariable logistic regressions were estimated to examine the effects of physician experience (years), patient age, and receptor subtype/TNM stage. RESULTS: Treatment decisions changed in 105 (5%) of 1,977 patients and were concentrated in those with hormone receptor (HR)-positive disease or stage IV disease in the first-line therapy setting (73% and 58%, respectively). Logistic regressions showed that decision changes were more likely in those with HR-positive cancer (odds ratio [OR], 1.58; P < .05) and less likely in those with stage IIA (OR, 0.29; P < .05) or IIIA cancer (OR, 0.08; P < .01). Reasons cited for changes included consideration of the CDSS therapeutic options (63% of patients), patient factors highlighted by the tool (23%), and the decision logic of the tool (13%). Patient age and oncologist experience were not associated with decision changes. Adherence to NCCN treatment guidelines increased slightly after using the CDSS (0.5%; P = .003). CONCLUSION: Use of an artificial intelligence-based CDSS had a significant impact on treatment decisions and NCCN guideline adherence in HR-positive breast cancers. Although cases of stage IV disease in the first-line therapy setting were also more likely to be changed, the effect was not statistically significant (P = .22). Additional research on decision impact, patient-physician communication, learning, and clinical outcomes is needed to establish the overall value of the technology.

Artificial Intelligence Treatment Decision Support For Complex Breast Cancer Among Oncologists With Varying Expertise.

PURPOSE: The aim of the current study was to assess treatment concordance and adherence to National Comprehensive Cancer Network breast cancer treatment guidelines between oncologists and an artificial intelligence advisory tool. PATIENTS AND METHODS: Study cases of patients (N = 1,977) who were at high risk for recurrence or who had metastatic disease and cell types for which the advisory tool was trained were obtained from the Chinese Society for Clinical Oncology cancer database (2012 to 2017). A cross-sectional observational study was performed to examine treatment concordance and guideline adherence among an artificial intelligence advisory tool and 10 oncologists with varying expertise-three fellows, four attending physicians, and three chief physicians. In a blinded fashion, each oncologist provided treatment advice on an average of 198 cases and the advisory tool on all cases (N = 1,977). Results are reported as rates and logistic regression odds ratios. RESULTS: Concordance for the recommended treatment was 0.56 for all physicians and higher for fellows compared with chief and attending physicians (0.68 v 0.54; 0.49; P = .001). Concordance differed by hormone receptor subtype-TNM stage, with the lowest for hormone receptor-positive human epidermal growth factor receptor 2/neu-positive cancers (0.48) and highest for triple-negative breast cancers (0.71) across most TNM stages. Adherence to National Comprehensive Cancer Network guidelines was higher for oncologists compared with the advisory tool (0.96 v 0.82; P < .003) and lower for fellows compared with attending physicians (0.93 v 0.98; 0.96; P = .04). CONCLUSION: Study findings reflect a complex breast cancer case mix, the limits of medical knowledge regarding optimum treatment, clinician practice patterns, and use of a tool that reflects expertise from one cancer center. Additional research in different practice settings is needed to understand the tool's scalability and its impact on treatment decisions and clinical and health services outcomes.

A colorimetric ATP assay based on the use of a magnesium(II)-dependent DNAzyme.

A colorimetric assay for ATP is described that uses a strategy that combines the concept of split Mg(II)-dependent DNAzyme, split aptamer, and hybridization-induced aggregation of gold nanoparticles (AuNPs). Both ATP aptamer and Mg(II)-dependent DNAzyme are split into two fragments which are allocated to two well-designed DNA probes. The probes also possess mutually complementary stem sequences and spacer sequences. In the presence of ATP, the separated DNAzyme sequences in the two probes assemble via the synchronous recognition of ATP with two fragments of the aptamer. Then, the activated DNAzyme catalyzes multiple cycles of the cleavage of its substrate DNA sequence. The latter acts as a linker and induces the aggregation of two types of ssDNA-modified AuNP through the hybridization between the complementary sequences. Thus, the color of the AuNP solution remains red. However, in the absence of ATP, the detached aptamer cannot induce the assembly of DNAzyme to cleave the linker DNA. This results in the aggregation of AuNP and a concomitant color transition from red to purple. This ATP assay, performed at a wavelength of 530 nm, has a linear detection range that extends from 10 pM to 100 nM, with a detection limit of 5.3 pM. It was applied to the detection of ATP in human serum. Conceivably, the strategy has a wide scope in that it may be applied to the colorimetric detection of various other analytes through the split aptamer configuration. Graphical abstract Schematic presentation of colorimetric assay for adenosine triphosphate (ATP) based on the use of a split Mg(II)-dependent DNAzyme, a split aptamer, and by exploiting the hybridization-induced aggregation of gold nanoparticles that leads to a color change from red to purple.

1,25(OH)2D3 deficiency increases TM40D tumor growth in bone and accelerates tumor-induced bone destruction in a breast cancer bone metastasis model.

Breast cancer is one of the most common malignancies and bone is the commonest site of distant metastases. Evidences indicate that adequate supply of vitamin D will decrease the morbidity and mortality of breast cancer. However, the main role of vitamin D deficiency in breast cancer bone metastases remains unclear. In this study, the relationship between vitamin D and breast cancer bone metastases were evaluated. Results showed that 1,25(OH)2D3 can not only inhibit the proliferation, migration and invasion of breast cancer cell TM40D in vitro, but also attenuate the breast cancer cell TM40D-induced bone destruction in vivo, whose underlying mechanism was at least partially through decreasing the number of the osteoclasts. To our knowledge, this is the first to use 1-alpha-hydroxylase [1α(OH)ase] knockout mice which characterized vitamin D deficiency to establish the breast cancer bone metastases model. Based on this model, we also found that vitamin D deficiency will accelerate the osteolytic lesions, and 1,25(OH)2D3 supplement will restrain osteolytic lesions. Therefore, these findings suggest that vitamin D has the potential capacity to be a therapeutic agent for the breast cancer bone metastases.

Menin mediates Tat-induced neuronal apoptosis in brain frontal cortex of SIV-infected macaques and in Tat-treated cells.

The molecular mechanisms involved in human immunodeficiency virus (HIV)-associated neurocognitive disorder (HAND) remain poorly understood. It has been recently reported that HIV-1 Tat transactivation requires menin, suggesting that menin may be involved in HAND pathogenesis. But the role of menin is not clear. Here, we found that protein level of menin was increased in simian-human immunodeficiency chimeric virus (SHIV)-SF162.P4 and simian immunodeficiency virus (SIV) sm543-3-infected rhesus macaques compared with the controls by immunohistochemistry (IHC) and western blot. Menin mainly expressed in the frontal cortex neurons of the brain, more importantly, the number of menin-staining cells was positively correlated with cleaved-caspase-3-positive cells while it was negatively correlated with a neuron-specific nuclear protein NeuN-positive cells, suggesting that expression of menin may induce neuronal apoptosis. Further studies showed that menin level was significantly increased during Tat-induced apoptosis, while downregulation of menin by pll3.7-MEN1-shRNA attenuated the Tat-induced cleavage of caspase-3 and caspase-8 in SY5Y cells and primary neuron cultures. Together, our findings reveal a pro-apoptotic role of menin in the brains of the SIV-infected macaques and the cultured neurons, indicating that targeting menin may be potential to block the HIV-1 Tat induced neuronal damage in HAND.

Sensitive colorimetric assays for α-glucosidase activity and inhibitor screening based on unmodified gold nanoparticles.

A colorimetric sensor has been developed in this work to sensitively detect α-glucosidase activity and screen α-glucosidase inhibitors (AGIs) utilizing unmodified gold nanoparticles (AuNPs). The sensing strategy is based on triple-catalytic reaction triggered by α-glucosidase. In the presence of α-glucosidase, aggregation of AuNPs is prohibited due to the oxidation of cysteine to cystine in the system. However, with addition of AGIs, cysteine induced aggregation of AuNPs occurs. Thus, a new method for α-glucosidase activity detection and AGIs screening is developed by measuring the UV-vis absorption or visually distinguishing. A well linear relation is presented in a range of 0.0025-0.05 U mL(-1). The detection limit is found to be 0.001 U mL(-1) for α-glucosidase assay, which is one order of magnitude lower than other reports. The IC50 values of four kinds of inhibitors observed with this method are in accordance with other reports. The using of unmodified AuNPs in this work avoids the complicated and time-consuming modification procedure. This simple and efficient colorimetric method can also be extended to other enzymes assays.

A sensitive chemiluminescent immunoassay for point-of-care testing of repaglinide in natural dietary supplements and serum.

For point-of-care testing of the illegal fortification of repaglinide (Rep) in natural dietary supplements, a competitive chemiluminescent immunoassay (CLIA) was established, using a horseradish peroxidase (HRP)-luminol-H2O2 system for signal amplification. Polyclonal antibodies for Rep were produced via immunization technique. Following optimization of the enzyme reaction time and concentrations of antibody and coating antigen, the method showed a limit of quantification (LOQ) of 1.0 ng/mL in PBS and limit of detection (LOD) of 8.3 ng/mL in serum and 6.0 ng/mL in blank tablets. When applied in natural dietary supplements, the method provided results consistent with those from HPLC, suggesting that the proposed method could be used for rapid screening of Rep in natural dietary supplements and detecting Rep in serum after administration.

An electrochemical aptasensor for thrombin detection based on the recycling of exonuclease III and double-stranded DNA-templated copper nanoparticles assisted signal amplification.

In this paper, we report an improved electrochemical aptasensor based on exonuclease III and double-stranded DNA (dsDNA)-templated copper nanoparticles (CuNPs) assisted signal amplification. In this sensor, duplex DNA from the hybridization of ligated thrombin-binding aptamer (TBA) subunits and probe DNA can act as an effective template for the formation of CuNPs on the electrode surface, so copper ions released from acid-dissolution of CuNPs may catalyze the oxidation of ο-phenylenediamine to produce an amplified electrochemical response. In the presence of thrombin, a short duplex domain with four complementary base pairs can be stabilized by the binding of TBA subunits with thrombin, in which TBA subunit 2 can be partially digested from 3' terminal with the cycle of exonuclease III, so the ligation of TBA subunits and the subsequent formation of CuNPs can be inhibited. By electrochemical characterization of dsDNA-templated CuNPs on the electrode surface, our aptasensor can display excellent performances for the detection of thrombin in a broad linear range from 100 fM to 1 nM with a low detection limit of 20.3 fM, which can also specially distinguish thrombin in both PBS and serum samples. Therefore, our aptasensor might have great potential for clinical diagnosis of biomarkers in the future.

Application of an enzyme immunoassay for the quantitative determination of azo dye (Orange II) in food products.

This paper reports the preparation of polyclonal antibodies against a synthetic azo dye, Orange II, and the development of an indirect ELISA to detect Orange II in foods. The sulfonic group of Orange II was modified and linked with carrier protein to synthesise an artificial antigen. Based on the checkerboard titration, the method showed excellent sensitivity (IC50 = 0.61 ng g⁻¹) to Orange II in the linear range of 0.05-10 ng g⁻¹. The antibody had little cross-reactivity with Chromotrope FB, Gardenia Yellow, Ponceau 4R, Sunset Yellow and Sudan dyes. The ELISA had limits of detection (LOD) of 0.22, 0.97 and 0.74 ng g⁻¹ in chilli powder, chilli oil and braised pork, respectively. The limits of quantification (LOQ) of the assay were 0.91 ng g⁻¹ in chilli powder, 1.48 ng g⁻¹ in chilli oil and 1.10 ng g⁻¹ in braised pork. For food products fortified with 1-10 ng g⁻¹ Orange II, the inter- and intra-assay variations were all less than 24.0% and 18.0%, respectively. Therefore, the proposed test could be used as a rapid screening method for Orange II detection in food samples.

The regimen of gemcitabine and cisplatin combined with radio frequency hyperthermia for advanced non-small cell lung cancer: a phase II study.

PURPOSE: This phase II randomised trial was designed to evaluate the therapeutic efficacy and feasibility of radio frequency regional hyperthermia in combination with chemotherapy for patients with advanced non-small lung cancer (NSCLC). METHODS: Eighty patients with pathologically proven advanced NSCLC, were enrolled and divided into two groups. Group A patients were treated by radio frequency regional hyperthermia in combination with the regimen of gemcitabine and cisplatin (GP). Group B patients were treated with the GP regimen alone. RESULTS: In group A, one patient achieved a complete response (CR), 18 achieved a partial response (PR), 18 achieved a stable disease and three experienced a progression of the disease. Thirty-three patients had a positive Clinical Benefit Response (CBR). In group B, no patient achieved CR, 17 achieved PR, 19 achieved a stable disease and four experienced a progression of the disease. Nineteen patients had a positive CBR. Significant differences between the two groups were observed for the CBR (P < 0.05), but not for RR. Major toxicities included bone marrow depression, nausea, vomiting, without significant differences between the two groups (P > 0.05). CONCLUSIONS: Radio-frequency regional hyperthermia in combination with chemotherapy (GP) is a safe, well tolerated, and effective therapeutic modality for patients with advanced NSCLC. The addition of hyperthermia improved quality of life.