Enhancing photodynamic and radionuclide therapy by small interfering RNA (siRNA)-RAD51 transfection via self-emulsifying delivery systems (SNEDDS).

BACKGROUND AIMS: Gene-silencing by small interfering RNA (siRNA) is an attractive therapy to regulate cancer death, tumor recurrence or metastasis. Because siRNAs are easily degraded, it is necessary to develop transport and delivery systems to achieve efficient tumor targeting. Self-nanoemulsifying systems (SNEDDS) have been successfully used for pDNA transport and delivery, so they may be useful for siRNA. The aim of this work is to introduce siRNA-RAD51 into a SNEDDS prepared with Phospholipon-90G, Labrafil-M1944-CS and Cremophor-RH40 and evaluate its efficacy in preventing homologous recombination of DNA double-strand breaks caused by photodynamic therapy (PDT) and ionizing radiation (IR). METHODS: The siRNA-RAD51 was loaded into SNEDDS using chitosan. Transfection capacity was estimated by comparison with Lipofectamine-2000. RESULTS: SNEDDS(siRNA-RAD51) induced gene silencing effect on the therapies evaluated by cell viability and clonogenic assays using T47D breast cancer cells. CONCLUSIONS: SNEDDS(siRNA-RAD51) shown to be an effective siRNA-delivery system to decrease cellular resistance in PDT or IR.

Pharmaceutical Nanoplatforms Based on Self-nanoemulsifying Drug Delivery Systems for Optimal Transport and Co-delivery of siRNAs and Anticancer Drugs.

Small interfering RNAs (siRNAs) have the ability to induce selective gene silencing, although siRNAs are vulnerable to degradation in vivo. Various active pharmaceutical ingredients (APIs) are currently used as effective therapeutics in the treatment of cancer. However, routes of administration are limited due to their physicochemical and biopharmaceutical properties. This research aimed to develop oral pharmaceutical formulations based on self-nanoemulsifying drug delivery systems (SNEDDS) for optimal transport and co-delivery of siRNAs related to cancer and APIs. Formulations were developed using optimal mixing design (Design-Expert 11 software) for SNEDDS loading with siRNA (water/oil emulsion), API (oil/water emulsion), and siRNA-API (multiphase water/oil/water emulsion). The final formulations were characterized physicochemically and biologically. The nanosystems less than 50 nm in size had a drug loading above 48 %. The highest drug release occurred at intestinal pH, allowing drug protection in physiological fluids. SNEDDS-siRNA-APIs showed a twofold toxicity effect than APIs in solution and higher transfection and internalization of siRNA in cancer cells with respect to free siRNAs. In the duodenum, higher permeability was observed with SNEDDS-API than with the API solution, as determined by ex-vivo fluorescence microscopy. The multifunctional formulation based on SNEDDS was successfully prepared, siRNA, hydrophobic chemotherapeutics (doxorubicin, valrubicin and methotrexate) and photosensitizers (rhodamine b and protoporphyrin IX) agents were loaded, using a chitosan-RNA core, and Labrafil® M 1944 CS, Cremophor® RH40, phosphatidylcholine shell, forming stable hybrid SNEDDS as multiphasic emulsion, suitable as co-delivery system with a potent anticancer activity.

Brainstem auditory-evoked responses among children afflicted by severely hypoxic CHD.

MAIN AIM: To electrophysiologically determine the impact of moderate to severe chronic hypoxia (H) resulting from a wide array of CHD (HCHD) conditions on the integrity of brainstem function. MATERIALS AND METHODS: Applying brainstem auditory-evoked response methodology, 30 chronically afflicted HCHD patients, who already had undergone heart surgery, were compared to 28 healthy control children (1-15 yo) matched by age, gender and socioeconomic condition. Blood oxygen saturation was clinically determined and again immediately before brainstem auditory-evoked response testing. RESULTS: Among HCHD children, auditory wave latencies (I, III and V) were significantly longer (medians: I, 2.02 ms; III, 4.12 ms, and; V, 6.30 ms) compared to control (medians: I, 1.67ms; III, 3.72 ms, and; V, 5.65 ms), as well as interpeak intervals (HCHD medians: I-V, 4.25 ms, and; III-V, 2.25ms; control medians: I-V, 3.90 ms and, III-V, 1.80 ms) without significant differences in wave amplitudes between groups. A statistically significant and inverse correlation between average blood oxygen saturation of each group (control, 94%; HCHD, 78%) and their respective wave latencies and interpeak intervals was found. CONCLUSIONS: As determined by brainstem auditory-evoked responses, young HCHD patients manifestly show severely altered neuronal conductivity in the auditory pathway strongly correlated with their hypoxic condition. These observations are strongly supported by different brainstem neurological and image studies showing that alterations, either in microstructure or function, result from the condition of chronic hypoxia in CHD. The non-altered wave amplitudes are indicative of relatively well-preserved neuronal relay nuclei.

Engineered rHDL Nanoparticles as a Suitable Platform for Theranostic Applications.

Reconstituted high-density lipoproteins (rHDLs) can transport and specifically release drugs and imaging agents, mediated by the Scavenger Receptor Type B1 (SR-B1) present in a wide variety of tumor cells, providing convenient platforms for developing theranostic systems. Usually, phospholipids or Apo-A1 lipoproteins on the particle surfaces are the motifs used to conjugate molecules for the multifunctional purposes of the rHDL nanoparticles. Cholesterol has been less addressed as a region to bind molecules or functional groups to the rHDL surface. To maximize the efficacy and improve the radiolabeling of rHDL theranostic systems, we synthesized compounds with bifunctional agents covalently linked to cholesterol. This strategy means that the radionuclide was bound to the surface, while the therapeutic agent was encapsulated in the lipophilic core. In this research, HYNIC-S-(CH2)3-S-Cholesterol and DOTA-benzene-p-SC-NH-(CH2)2-NH-Cholesterol derivatives were synthesized to prepare nanoparticles (NPs) of HYNIC-rHDL and DOTA-rHDL, which can subsequently be linked to radionuclides for SPECT/PET imaging or targeted radiotherapy. HYNIC is used to complexing 99mTc and DOTA for labeling molecules with 111, 113mIn, 67, 68Ga, 177Lu, 161Tb, 225Ac, and 64Cu, among others. In vitro studies showed that the NPs of HYNIC-rHDL and DOTA-rHDL maintain specific recognition by SR-B1 and the ability to internalize and release, in the cytosol of cancer cells, the molecules carried in their core. The biodistribution in mice showed a similar behavior between rHDL (without surface modification) and HYNIC-rHDL, while DOTA-rHDL exhibited a different biodistribution pattern due to the significant reduction in the lipophilicity of the modified cholesterol molecule. Both systems demonstrated characteristics for the development of suitable theranostic platforms for personalized cancer treatment.

Photoactivation of Chemotherapeutic Agents with Cerenkov Radiation for Chemo-Photodynamic Therapy.

Cerenkov radiation (CR) can be used as an internal light source in photodynamic therapy (PDT). Methotrexate (MTX) and paclitaxel (PTX), chemotherapeutic agents with wide clinical use, have characteristics of photosensitizers (PS). This work evaluates the possibility of photoexciting MTX and PTX with CR from 18F-FDG to produce reactive oxygen species (ROS) capable of inducing cytotoxicity. PTX did not produce ROS when excited by CR from 18F-FDG, so it is not useful for PDT. In contrast, MTX produces 1O2 (detected by ABMA) in amounts sufficient to significantly decrease the viability of the T47D cells. MTX solutions of 100 nM combined with 18F-FDG activities of 50 (1.85 MBq) and 100 µCi (3.7 MBq) produced a significant decrease in cell viability to (50.09 ± 4.95) and (47.96 ± 11.19)%, respectively, compared to MTX (66.29 ± 5.92)% and 18F-FDG (91.35 ± 7.00% for 50 µCi and 99.43 ± 11.03% for 100 µCi) alone. Using the CellRox Green reagent, the intracellular production of ROS was confirmed as the main mechanism of cytotoxicity. The results confirm the therapeutic potential of photoactivation with CR and the synergy of the combined treatment with chemotherapy + photodynamic therapy (CMT + PDT). The combination of chemotherapeutic agents with PS properties and ß-emitting radiopharmaceuticals, previously approved for clinical use, will make it possible to shorten the evaluation stages of new CMT + PDT systems.

Preclinical evaluation of early multi-organ toxicity induced by liposomal doxorubicin using 67Ga-citrate.

Although liposomal doxorubicin (LPD) is widely used for cancer treatment, knowledge concerning the toxicity induced by this drug in healthy organs and tissues is limited. LPD-induced toxicity studies relative to free doxorubicin (DOX) have focused on cardiotoxicity in tumor-bearing animals. On the other hand, the results on DOX-induced cardiotoxicity depending on gender are controversial. One of the manifestations of toxicity is tissue inflammation. 67Ga-citrate has been used for decades to assess inflammation in various pathologies. In this work, the ex vivo biodistribution of 67Ga-citrate is used to evaluate induced multi-organ toxicity in healthy 10-week-old male and female CD1 mice treated for 5 weeks with LPD. Toxicity in males, determined by 67Ga-citrate, was evident only in the target organs of liposomes (spleen, liver, kidneys, and lungs); the average weight loss was 11% and mortality was 14%. In female mice, 67Ga-citrate revealed a cytotoxic effect in practically all organs, the average weight loss was 37%, and the mortality after the last dose of LPD was 66%. These results confirm the usefulness of 67Ga-citrate and the importance of stratifying by sex in the toxicological evaluation of drugs.

225Ac-rHDL Nanoparticles: A Potential Agent for Targeted Alpha-Particle Therapy of Tumors Overexpressing SR-BI Proteins.

Actinium-225 and other alpha-particle-emitting radionuclides have shown high potential for cancer treatment. Reconstituted high-density lipoproteins (rHDL) specifically recognize the scavenger receptor B type I (SR-BI) overexpressed in several types of cancer cells. Furthermore, after rHDL-SR-BI recognition, the rHDL content is injected into the cell cytoplasm. This research aimed to prepare a targeted 225Ac-delivering nanosystem by encapsulating the radionuclide into rHDL nanoparticles. The synthesis of rHDL was performed in two steps using the microfluidic synthesis method for the subsequent encapsulation of 225Ac, previously complexed to a lipophilic molecule (225Ac-DOTA-benzene-p-SCN, CLog P = 3.42). The nanosystem (13 nm particle size) showed a radiochemical purity higher than 99% and stability in human serum. In vitro studies in HEP-G2 and PC-3 cancer cells (SR-BI positive) demonstrated that 225Ac was successfully internalized into the cytoplasm of cells, delivering high radiation doses to cell nuclei (107 Gy to PC-3 and 161 Gy to HEP-G2 nuclei at 24 h), resulting in a significant decrease in cell viability down to 3.22 ± 0.72% for the PC-3 and to 1.79 ± 0.23% for HEP-G2 at 192 h after 225Ac-rHDL treatment. After intratumoral 225Ac-rHDL administration in mice bearing HEP-G2 tumors, the biokinetic profile showed significant retention of radioactivity in the tumor masses (90.16 ± 2.52% of the injected activity), which generated ablative radiation doses (649 Gy/MBq). The results demonstrated adequate properties of rHDL as a stable carrier for selective deposition of 225Ac within cancer cells overexpressing SR-BI. The results obtained in this research justify further preclinical studies, designed to evaluate the therapeutic efficacy of the 225Ac-rHDL system for targeted alpha-particle therapy of tumors that overexpress the SR-BI receptor.

Targeted photodynamic therapy using reconstituted high-density lipoproteins as rhodamine transporters.

Reconstituted high-density lipoprotein (rHDL) nanoparticles are excellent transporters of molecules and very useful for targeted therapy as they specifically recognize the scavenger receptor, class B1 (SR-B1) that is present on the surface of a wide range of tumor cells. However, they have rarely been employed to transport photosensitizers (PS) for photodynamic therapy (PDT). Rhodamine (R) compounds have been dismissed as useful PSs for PDT due to their low 1O2 production, excitation wavelengths with little tissue penetration, and poor selectivity for tumor cells. It was recently demonstrated that when irradiating at 532 nm or with Cerenkov radiation (CR) from a ß-emitting radionuclide, R123, R6G, and RB undergo electron transfer reactions (type I reaction) with folic acid. R6G also produces type I reactions with O2. In this work, the photodynamic effects of the rHDL-R system were evaluated in vitro. rHDL nanoparticles loaded with R123, R6G, and RB were synthesized, and the PS was internalized into T47D tumor cells. When cells were irradiated with a 532-nm laser in the presence of an rHDL-R systems, a cytotoxic photodynamic effect was obtained in the order R6G > R123 > RB. In the presence of CR from a 177Lu source, cytotoxicity showed the order R6G > RB > R123. The higher cytotoxicity induced by R6G in both cases corresponds to higher cellular internalization and larger production of type I and II reactions. Thus, in this work, it is proposed that rHDL-R/177Lu system can be applied in theragnostics as a multimodal radiotherapy-PDT-imaging system (imaging by SPECT or Cerenkov) and in hypoxic solid tumors in which external radiation is not effective and 177Lu-CR acts as light source.

A Multimodal Theranostic System Prepared from High-Density Lipoprotein Carrier of Doxorubicin and 177Lu.

Recently, it was demonstrated that doxorubicin (Dox.HCl), a chemotherapeutic agent, could be photoactivated by Cerenkov radiation (CR). The objective of the present work was to develop a multimodal chemotherapy-radiotherapy-photodynamic therapeutic system based on reconstituted high-density lipoprotein (rHDL) loaded with Dox.HCl and 177Lu-DOTA. 177Lu acts as a therapeutic radionuclide and CR source. The system can be visualized by nuclear imaging. Fluorescence microscopy showed that rHDL-Dox specifically recognized cancer cells (T47D) that are positive for SR-B1 receptors. Encapsulated Dox.HCl was released into the cells and produced reactive oxygen species when irradiated with a 450-nm laser (photodynamic effect). The same effect occurred when Dox.HCl was irradiated by 177Lu CR. Through in vitro experiments, it was confirmed that the addition of 177Lu-DOTA to the rHDL-Dox nanosystem did not affect the specific recognition of SR-B1 receptors expressed in cells, or the cellular internalization of 177Lu-DOTA. The toxicity induced by the rHDL-Dox/177Lu nanosystem in cell lines with high (T47D and PC3), poor (H9C2) and almost-zero (human fibroblasts (FB)) expression of SR-B1 was evaluated in vitro and confirmed the synergy of the combined chemotherapy-radiotherapy-photodynamic therapeutic effect; this induced toxicity was proportional to the expression of the SR-B1 receptor on the surface of the cells used. The HDL-Dox/177Lu nanosystem experienced uptake by tumor cells and the liver-both tissues with high expression of SR-B1 receptors-but not by the heart. 177Lu CR offered the possibility of imparting photodynamic therapy where laser light could not reach.

Genotoxicity of Mercury and Its Derivatives Demonstrated In Vitro and In Vivo in Human Populations Studies. Systematic Review.

Beside partial coverage in three reviews so far (1994, 2009, 2019), there is no review on genotoxic studies dealing with mercury (Hg) and human exposure using the most usual genotoxic assays: sister chromatid exchanges (SCE), chromosomal aberrations (CA), cytochalasin B blocked micronucleus assay (CBMN), and single-cell gel electrophoresis (SCGE or alkaline comet assay). Fifty years from the first Hg genotoxicity study and with the Minamata Convention in force, the genotoxic potential of Hg and its derivatives is still controversial. Considering these antecedents, we present this first systematic literature overview of genotoxic studies dealing with Hg and human exposure that used the standard genotoxic assays. To date, there is not sufficient evidence for Hg human carcinogen classification, so the new data collections can be of great help. A review was made of the studies available (those published before the end of October 2021 on PubMed or Web of Science in English or Spanish language) in the scientific literature dealing with genotoxic assays and human sample exposure ex vivo, in vivo, and in vitro. Results from a total of 66 articles selected are presented. Organic (o)Hg compounds were more toxic than inorganic and/or elemental ones, without ruling out that all represent a risk. The most studied inorganic (i)Hg compounds in populations exposed accidentally, occupationally, or iatrogenically, and/or in human cells, were Hg chloride and Hg nitrate and of the organic compounds, were methylmercury, thimerosal, methylmercury chloride, phenylmercuric acetate, and methylmercury hydroxide.

In vitro irradiation of doxorubicin with 18F-FDG Cerenkov radiation and its potential application as a theragnostic system.

Doxorubicin (DOX), an effective chemotherapeutic agent, has a wide excitation band centred at 480 nm. Cerenkov radiation (CR) is considered an internal light source in photodynamic therapy (PDT). DOX could be photoactivated by CR and thus, enhancing its cytotoxicity. In this work, 18F-FDG was used to evaluate the effect of Cerenkov radiation on DOX, in comparison to irradiation with a 450-nm laser beam, in terms of ROS production. The production of 1O2 and O2⁎- reactive species during DOX irradiation was detected indirectly by ABMA and DCPIP bleaching, respectively. The cytotoxic effect of the DOX / 18F-FDG CR system was evaluated in the T47D breast cancer cell line. The irradiation of DOX produced 1O2 and O2⁎- species using both 18F-FDG CR and a 450-nm laser beam. The majority reactive species produced in both cases was 1O2; a favourable result, given the greater cytotoxicity of this species. The viability of T47D cells in presence of DOX (5 nM), 18F-FDG (37.5 µCi) and DOX (5 nM)/18F-FDG (37.5 µCi) was (86 ± 9)%, (84 ± 8)% and (64 ± 5)%, respectively; these results suggest a synergistic cytotoxic effect derived from the cytotoxic activity of DOX and its photoactivation by 18F-FDG CR. It is worth noting that the system could be optimized in terms of DOX concentration and 18F-FDG activity for better results. Due to the fact that 18F-FDG is widely used in nuclear imaging, the DOX/18F-FDG system also possesses theragnostic characteristics. Thus, in this work, it is demonstrated that DOX can be used in a dual therapy system based on chemotherapy-PDT when 18F-FDG CR is used as a DOX excitation source.

Preparation and in vitro evaluation of radiolabeled HA-PLGA nanoparticles as novel MTX delivery system for local treatment of rheumatoid arthritis.

Radiosynovectomy is a technique used to decrease inflammation of the synovial tissue by intraarticular injection of a ß-emitting radionuclide, such as 177Lu, which is suitable for radiotherapy due to its decay characteristics. Drug-encapsulating nanoparticles based on poly lactic­co­glycolic acid (PLGA) polymer are a suitable option to treat several arthritic diseases, used as anti-inflammatory drugs transporters of such as methotrexate (MTX), which has been widely used in the arthritis treatment (RA), and hyaluronic acid (HA), which specifically binds the CD44 and hyaluronan receptors overexpressed on the inflamed synovial tissue cells. The 1,4,7,10­Tetraazacyclododecane­1,4,7,10­tetraacetic acid (DOTA) was used as complexing agent of Lutetium-177 for radiotherapy porpoises. The aim of this research was to synthesize 177Lu-DOTA-HA-PLGA(MTX) as a novel, smart drug delivery system with target-specific recognition, potentially useful in radiosynovectomy for local treatment of rheumatoid arthritis. The polymeric nanoparticle system was prepared and chemically characterized. The MTX encapsulation and radiolabelling were performed with suitable characteristics for its in vitro evaluation. The HA-PLGA(MTX) nanoparticle mean diameter was 167.6 nm ±â€¯57.4 with a monomodal and narrow distribution. Spectroscopic techniques demonstrated the effective conjugation of HA and chelating agent DOTA to the polymeric nanosystem. The MTX encapsulation was 95.2% and the loading efficiency was 6%. The radiochemical purity was 96 ±â€¯2%, determined by ITLC. Conclusion: 177Lu-DOTA-HA-PLGA(MTX) was prepared as a biocompatible polymeric PLGA nanoparticle conjugated to HA for specific targeting. The therapeutic nanosystem is based on bi-modal mechanisms using MTX as a disease-modifying antirheumatic drug (DMARD) and 177Lu as a radiotherapeutic component. The 177Lu-DOTA-HA-PLGA(MTX) nanoparticles showed properties suitable for radiosynovectomy and further specific targeted anti-rheumatic therapy.

Theoretical and experimental characterization of emission and transmission spectra of Cerenkov radiation generated by 177Lu in tissue.

Cerenkov radiation (CR) is the emission of UV-vis light generated by the de-excitation of the molecules in the medium, after being polarized by an excited particle traveling faster than the speed of light. When ß particles travel through tissue with energies greater than 219 keV, CR occurs. Tissues possess a spectral optical window of 600 to 1100 nm. The CR within this range can be useful for quantitative preclinical studies using optical imaging and for the in-vivo evaluation of Lu177-radiopharmaceuticals (ß-particle emitters). The objective of our research was to determine the experimental emission light spectrum of Lu177-CR and evaluate its transmission properties in tissue as well as the feasibility to applying CR imaging in the preclinical studies of Lu177-radiopharmaceuticals. The theoretical and experimental characterizations of the emission and transmission spectra of Lu177-CR in tissue, in the vis-NIR region (350 to 900 nm), were performed using Monte Carlo simulation and UV-vis spectroscopy. Mice Lu177-CR images were acquired using a charge-coupled detector camera and were quantitatively analyzed. The results demonstrated good agreement between the theoretical and the experimental Lu177-CR emission spectra. Preclinical CR imaging demonstrated that the biokinetics of Lu177-radiopharmaceuticals in the main organs of mice can be acquired.

Synthesis and preclinical evaluation of the 177Lu-DOTA-PSMA(inhibitor)-Lys3-bombesin heterodimer designed as a radiotheranostic probe for prostate cancer.

BACKROUND: Human tumors show intrinsic heterogeneity and changes in phenotype during disease progression, which implies different expression levels of cell surface receptors. The research on new heterodimeric lutetium-177 (Lu)-radiopharmaceuticals interacting with two different targets on tumor cells is a strategy for improvement of radiotheranostic performance. This study aimed to synthesize and characterize the Lu-DOTA-PSMA(inhibitor)-Lys-bombesin (Lu-DOTA-iPSMA-Lys-BN) heterodimer and to evaluate its potential to target prostate-specific membrane antigen (PSMA) and gastrin-releasing peptide receptor (GRPr) overexpressed in prostate cancer. METHODS: The heterodimeric conjugate was synthesized and characterized by infrarred, mass, and H-NMR spectroscopies. The ligand was labeled with Lu and the radiochemical purity was assessed by radio-high-performance liquid chromatography. PSMA/GRPr affinity and the heterobivalent effect on cell viability were evaluated in LNCaP and PC3 prostate cancer cell lines. The biodistribution profile (3 and 96 h) was assessed in athymic mice with induced prostate tumors. Using pulmonary LNCaP (PSMA-positive) and PC3 (GRPr-negative) micrometastasis models, the influence of heterobivalency and affinity on tumor uptake was quantified (micro-SPECT/CT). RESULTS: Lu-iPSMA-BN (radiochemical purity>98%) showed specific recognition for PSMA and GRPr (IC50=5.62 and 3.49 nmol/l, respectively) with a significant decrease in cell viability (10.15% of cell viability in LNCaP and 40.10% in PC3 at 48 h), as well as high LNCaP and PC3 tumor uptake (5.21 and 3.21% ID/g at 96 h, respectively). Micro-SPECT/CT imaging showed the heterodimer ability to target the tumors (SUVmax of 1.93±0.30 and 1.76±0.10 in LNCaP and PC3, respectively), possibly influenced by the heterobivalent effect. Lu-DOTA-iPSMA-Lys-BN showed suitable affinity for PSMA and GRPr. CONCLUSION: The results warrant further preclinical studies to establish the Lu-radiotracer theranostic efficacy.

[99mTc-HYNIC-N-dodecylamide]: a new hydrophobic tracer for labelling reconstituted high-density lipoproteins (rHDL) for radioimaging.

Despite the widespread use of nanotechnology in radio-imaging applications, lipoprotein based delivery systems have received only limited attention so far. These studies involve the synthesis of a novel hydrophobic radio-imaging tracer consisting of a hydrazinonicotinic acid (HYNIC)-N-dodecylamide and 99mTc conjugate that can be encapsulated into rHDL nanoparticles (NPs). These rHDL NPs can selectively target the Scavenger Receptor type B1 (SR-B1) that is overexpressed on most cancer cells due to excess demand for cholesterol for membrane biogenesis and thus can target tumors in vivo. We provide details of the tracer synthesis, characterization of the rHDL/tracer complex, in vitro uptake, stability studies and in vivo application of this new radio-imaging approach.

Fluorescent, Plasmonic, and Radiotherapeutic Properties of the 177Lu-Dendrimer-AuNP-Folate-Bombesin Nanoprobe Located Inside Cancer Cells.

The integration of fluorescence and plasmonic properties into one molecule is of importance in developing multifunctional imaging and therapy nanoprobes. The aim of this research was to evaluate the fluorescent properties and the plasmonic-photothermal, therapeutic, and radiotherapeutic potential of 177Lu-dendrimer conjugated to folate and bombesin with gold nanoparticles in the dendritic cavity (177Lu-DenAuNP-folate-bombesin) when it is internalized in T47D breast cancer cells. The intense near-Infrared (NIR) fluorescence emitted at 825 nm from the conjugate inside cells corroborated the usefulness of DenAuNP-folate-bombesin for optical imaging. After laser irradiation, the presence of the nanosystem in cells caused a significant increase in the temperature of the medium (46.8°C, compared to 39.1°C without DenAuNP-folate-bombesin, P < 0.05), resulting in a significant decrease in cell viability (down to 16.51% ± 1.52%) due to the 177Lu-DenAuNP-folate-bombesin plasmonic properties. After treatment with 177Lu-DenAuNP-folate-bombesin, the T47D cell viability decreased 90% because of the radiation-absorbed dose (63.16 ± 4.20 Gy) delivered inside the cells. The 177Lu-DenAuNP-folate-bombesin nanoprobe internalized in cancer cells exhibited properties suitable for optical imaging, plasmonic-photothermal therapy, and targeted radiotherapy.

Haloalkalitolerant Actinobacteria with capacity for anthracene degradation isolated from soils close to areas with oil activity in the State of Veracruz, Mexico.

The use of native strains of microorganisms from soils is an excellent option for bioremediation. To our knowledge, until now there has been no other group working on the isolation of Actinobacteria from contaminated soils in Mexico. In this study, samples of soils close to areas with oil activity in the State of Veracruz, Mexico, were inoculated for the isolation of Actinobacteria. The strains isolated were characterized morphologically, and the concentrations of NaCl and pH were determined for optimal growth. Strain selection was performed by the detection of a phylogenetic marker for Actinobacteria located at the 23S rRNA gene, followed by species identification by sequencing the 16S rRNA gene. Several haloalkalitolerant Actinobacteria were isolated and identified as: Kocuria rosea, K. palustris, Microbacterium testaceum, Nocardia farcinica and Cellulomonas denverensis. Except for C. denverensis, the biomass of all strains increased in the presence of anthracene. The strains capacity to metabolize anthracene (at 48 h), determined by fluorescence emission, was in the range of 46-54%. During this time, dihydroxy aromatic compounds formed, characterized by attenuated total reflectance Fourier transform infrared spectroscopy bands of 1205 cm-1 and 1217 cm-1. Those Actinobacteria are potentially useful for the bioremediation of saline and alkaline environments contaminated with polycyclic aromatic hydrocarbon compounds. [Int Microbiol 2016; 19(1):15-26].

Synthesis and evaluation of Lys¹(α,γ-Folate)Lys³(¹77Lu-DOTA)-Bombesin(1-14) as a potential theranostic radiopharmaceutical for breast cancer.

The aim of this work was to synthesize Lys(1)(α,γ-Folate)-Lys(3)((177)Lu-DOTA)-Bombesin (1-14) ((177)Lu-Folate-BN), as well as to assess its potential for molecular imaging and targeted radiotherapy of breast tumors expressing folate receptors (FR) and gastrin-releasing peptide receptors (GRPR). Radiation absorbed doses of (177)Lu-Folate-BN (74 MBq, i.v.) estimated in athymic mice with T47D-induced breast tumors (positive to FR and GRPR), showed tumor doses of 23.9±2.1 Gy. T47D-tumors were clearly visible (Micro-SPECT/CT images). (177)Lu-Folate-BN demonstrated properties suitable as a theranostic radiopharmaceutical.

Improved radiopharmaceutical based on 99mTc-Bombesin-folate for breast tumour imaging.

BACKGROUND: Clinical studies in women using technetium-99m (Tc)-Bombesin have shown successful radionuclide imaging of breast tumours overexpressing gastrin-releasing peptide receptors (GRPRs). Recent studies have demonstrated that most breast tumours overexpress folate receptors (FRα). AIM: The aim of this work was to synthesize the Lys(α,γ-Folate)-Lys(Tc-EDDA/HYNIC)-Bombesin (1-14) conjugate (Tc-Bombesin-Folate), as well as to assess the in-vitro and in-vivo potential of the radiopharmaceutical to target FRα and GRPR. METHODS: LysLys(HYNIC)-Bombesin (1-14) was conjugated to folic acid and the product was purified by size-exclusion high-performance liquid chromatography. Ultraviolet-visible spectroscopy, Fourier transform infrared spectroscopy and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry were used for chemical characterization. Tc labelling was performed using ethylenediamine-N,N'-diacetic acid/tricine as coligands. In-vitro binding studies were carried out in T47D breast cancer cells (positive for FRα and GRPR). Biodistribution studies and micro-single-photon emission computed tomography/computed tomography imaging were carried out on athymic mice with T47D-induced tumours. RESULTS: High-performance liquid chromatography analyses indicated that the radioconjugate was obtained with high radiochemical purity (96±2.1%). In-vitro and in-vivo results showed significant uptake of the radiopharmaceutical in T47D cells and tumours (5.43% ID/g), which was significantly inhibited by preincubation with cold folic acid or cold Bombesin. CONCLUSION: The Tc-Bombesin-folate heterobivalent radiopharmaceutical significantly enhances in-vivo tumour uptake because of the concomitant interaction with FRα and GRPR.

Comparative Effect Between Laser and Radiofrequency Heating of RGD-Gold Nanospheres on MCF7 Cell Viability.

Gold nanoparticles conjugated to cyclo-[Arg-Gly-Asp-D-Phe-Lys(Cys)] peptides (AuNP-c[RGDfK(C)]) have been reported as systems with specific cell internalization in breast cancer cells. AuNPs have also been proposed as localized heat sources for cancer treatment using laser irradiation or radiofrequency (RF). The aim of this research was to analyze, based on the Mie theory, the AuNP-c[RGDfK(C)] absorption cross-sections (C(abs)) of low-frequency electromagnetic waves (13.56 MHz, λ = 22 m) and optical frequency waves (laser at λ = 532 nm) and to compare their effect on MCF7 cell viability as thermal conversion sources in AuNPs (20 nm) located inside cells. Cell viability was assessed in MCF7 cells treated with AuNP-c[RGDfK(C)] or water after exposure to the RF field (200 W, 100 V/cm) or laser irradiation (Irradiance 0.65 W/cm2). In both cases (RF and laser) the presence of nanoparticles in cells caused a significant increase in the temperature of the medium (RF: AT = 29.9 ± 1.7 degrees C for AuNP compared to ΔT = 13.0 ± 1.4 degrees C for water; laser: ΔT = 13.5 ± 0.7 degrees C for AuNP compared to 3.3 ± 0.5 degrees C for water). Although RF induced a higher increase in the temperature of the medium with nanoparticles, the largest effect on the cell viability was produced by laser when nanoparticles were located inside the cells (8.7?0.7% for laser compared to 19.4 ± 0.9% for RF). The differences obtained in C(abs) values (laser: 3.7 x 10- (16) m2; RF: 7.9 x 10-(23) m2) and the observed effect on MFC7 cell viability support two mechanisms previously proposed "wave energy absorption by AuNPs" when laser is used as a thermal conversion source, and "attenuation of the wave passing through the AuNP suspension" when RF is applied. The AuNP-c[RGDfK(C)] nanosystem shows suitable properties to improve hyperthermia treatments under laser irradiation due to a larger heat release inside cells.