Size discrimination in rat and mouse gastric emptying.

OBJECTIVES: To investigate the relationship between particle size and gastric emptying in rodents using radiolabeled insoluble polymethyl methacrylate (PMMA) microcapsules/beads. METHODS: PMMA microcapsules (50-500 µm) and beads (0.5-3 mm) loaded with technetium-99 m diethylenetriamine pentaacetic acid ((99m) Tc-DTPA) were administered to ICR mice or Sprague Dawley (SD) rats by oral gavage. Gamma scintiscans were acquired initially following administration and then at hourly intervals to 4 hours. RESULTS: Scintiscans revealed that the smallest PMMA microcapsules (50-100 µm) or beads (0.5-1 mm) were impeded in the stomach and emptied slower than large particles in both rodent species. In mice, no significant difference in gastric emptying was found with microcapsules between 100 and 300 µm in diameter (p = 0.25) and particles more than 300 µm could not be administered. In rats, capsules containing 0.5-3 mm beads were stuck to the esophagus (up to 1 hour), this was a limitation of dosing beads of this size because they cannot be suspended in a liquid media for oral gavage purposes. Beads with diameters of 2-3 mm stayed in the stomach for up to 4 hours. CONCLUSIONS: The cut-off emptying size in ICR mice could not be determined, due to the limitation of current available dosing methods. The cut-off emptying size in SD rats was between 1.5 and 2 mm. Therefore, particles with a diameter greater than 2 mm should not be used for gastric emptying studies of intact particles in SD rats, as their emptying is retarded in the stomach.

Liposomal formulations of poorly soluble camptothecin: drug retention and biodistribution.

Camptothecin (CPT) represents a potent anticancer drug. However, its therapeutic use is impaired by both drug solubility, hydrolysis, and protein interactions in vivo. Use of liposomes as a drug-formulation approach could overcome some of these challenges. The aim of this study was to perform a mechanistic study of the incorporation and retention of the lipophilic parent CPT compound in different liposome formulations using radiolabeled CPT and thus to be able to identify promising CPT delivery systems. In this context, we also wanted to establish an appropriate mouse tumor model, in vivo scintigraphic imaging, and biodistribution methodology for testing the most promising formulation. CPT retention in various liposome formulations after incubation in buffer and serum was determined. The HT-29 mouse tumor model, (111)In-labeled liposomes, as well as (3)H-labeled CPT were used to investigate the biodistribution of liposomes and drug. The ability of different liposome formulations to retain CPT in buffer was influenced by lipid concentration and drug/lipid ratio, rather than lipid composition. The tested formulations were cleared from the blood in the following order: CPT solution > CPT liposomes > (111)In-labeled liposomes, and liposomes mainly accumulated in the liver. Lipid composition did not influence CPT retention to the same extent as earlier observed from incorporation studies. The set-up for the biodistribution study works well and is suited for future in vivo studies on CPT liposomes. The biodistribution study showed that liposomes circulated longer than free drug, but premature release of drug from liposomes occurred. Further studies to develop formulations with higher retention potential and prolonged circulation are desired.

Modulation of oxidative stability of haemoglobin inside liposome-encapsulated haemoglobin.

The major hurdle in the formulation of liposome-encapsulated haemoglobin (LEH) is the oxidation of haemoglobin (Hb) into methaemoglobin during storage and after administration. In order to reduce this oxidative degradation, we tested various reducing conditions in the presence of catalase. We found that at 37°C more than 50% of Hb oxidized to methaemoglobin within 24 h, whereas in presence of catalase, the oxidation was significantly reduced. The effect of catalase was further enhanced by a reduction mixture containing ß-NAD, d-glucose, adenine, inosine, MgCl2, KCl, KH2PO4 and Na2HPO4; only 14% methaemoglobin was generated in the presence of catalase and reduction mixture. Contrary to the expectation, glutathione, deferoxamine and homocysteine enhanced Hb oxidation. The presence of CRM inside liposomes (250 nm) significantly decreased Hb oxidation. The results suggest that catalase and a well-defined mixture of co-factors may help control Hb oxidation for improvement in the functional life of LEH.

Feasibility of eradication of breast cancer cells remaining in postlumpectomy cavity and draining lymph nodes following intracavitary injection of radioactive immunoliposomes.

Most diagnosed early stage breast cancer cases are treated by lumpectomy and adjuvant radiation therapy, which significantly decreases the locoregional recurrence but causes inevitable toxicity to normal tissue. By using a technique of preparing liposomes carrying technetium-99m ((99m)Tc), rhenium-186 ((186)Re), or rhenium-188 ((188)Re) radionuclides, as well as chemotherapeutic agents, or their combination, for cancer therapy with real time image-monitoring of pharmacokinetics and prediction of therapy effect, this study investigated the potential of a novel targeted focal radiotherapy with low systemic toxicity using radioactive immunoliposomes to treat both the surgical cavity and draining lymph nodes in a rat breast cancer xenograft positive surgical margin model. Immunoliposomes modified with either panitumumab (anti-EGFR) or bevacizumab (anti-VEGF) were remote loaded with (99m)Tc diagnostic radionuclide, and injected into the surgical cavity of female nude rats with positive margins postlumpectomy. Locoregional retention and systemic distribution of (99m)Tc-immunoliposomes were investigated by nuclear imaging, stereofluorescent microscopic imaging, and gamma counting. Histopathological examination of excised draining lymph nodes was performed. The locoregional retention of (99m)Tc-immunoliposomes in each animal was influenced by the physiological characteristics of the surgical site of individual animals. Panitumumab- and bevacizumab-liposome groups had higher intracavitary retention compared with the control liposome groups. Draining lymph node uptake was influenced by both the intracavitary radioactivity retention level and metastasis status. The panitumumab-liposome group had higher accumulation on the residual tumor surface and in the metastatic lymph nodes. Radioactive liposomes that were cleared from the cavity were metabolized quickly and accumulated at low levels in vital organs. Therapeutic radionuclide-carrying specifically targeted panitumumab- and bevacizumab-liposomes have increased potential compared to non-antibody targeted liposomes for postlumpectomy focal therapy to eradicate remaining breast cancer cells inside the cavity and draining lymph nodes with low systemic toxicity.

Post-lumpectomy intracavitary retention and lymph node targeting of (99m)Tc-encapsulated liposomes in nude rats with breast cancer xenograft.

Liposomes are recognized drug delivery systems with tumor-targeting capability. In addition, therapeutic or diagnostic radionuclides can be efficiently loaded into liposomes. This study investigated the feasibility of utilizing radiotherapeutic liposomes as a new post-lumpectomy radiotherapy for early-stage breast cancer by determining the locoregional retention and systemic distribution of liposomes radiolabeled with technetium-99m ((99m)Tc) in an orthotopic MDA-MB-231 breast cancer xenograft nude rat model. To test this new brachytherapy approach, a positive surgical margin lumpectomy model was set up by surgically removing the xenograft and deliberately leaving a small tumor remnant in the surgical cavity. Neutral, anionic, and cationic surface-charged fluorescent liposomes of 100 and 400 nm diameter were manufactured and labeled with (99m)Tc-BMEDA. Locoregional retention and systemic distribution of (99m)Tc-liposomes injected into the post-lumpectomy cavity were determined using non-invasive nuclear imaging, ex vivo tissue gamma counting and fluorescent stereomicroscopic imaging. The results indicated that (99)Tc-liposomes were effectively retained in the surgical cavity (average retention was 55.7 ± 24.2% of injected dose for all rats at 44 h post-injection) and also accumulated in the tumor remnant (66.9 ± 100.4%/g for all rats). The majority of cleared (99m)Tc was metabolized quickly and excreted into feces and urine, exerting low radiation burden on vital organs. In certain animals (99m)Tc-liposomes significantly accumulated in the peripheral lymph nodes, especially 100 nm liposomes with anionic surface charge. The results suggest that post-lumpectomy intracavitary administration of therapeutic radionuclides delivered by 100-nm anionic liposome carrier is a potential therapy for the simultaneous treatment of the surgical cavity and the draining lymph nodes of early-stage breast cancer.

Chemoradionuclide therapy with 186Re-labeled liposomal doxorubicin in combination with radiofrequency ablation for effective treatment of head and neck cancer in a nude rat tumor xenograft model.

PURPOSE: To determine the therapeutic efficacy of rhenium 186 ((186)Re)-labeled PEGylated liposomal doxorubicin ((186)Re-liposomal doxorubicin) in combination with radiofrequency (RF) ablation of human head and neck squamous cell carcinoma (HNSCC) xenograft in nude rats. MATERIALS AND METHODS: This investigation was approved by the animal care committee. Sixty nude rats with subcutaneously implanted HNSCC xenografts (six per group) were treated with (a) RF ablation (70 °C for 5 minutes), (b) PEGylated liposomes, (c) liposomal doxorubicin, (d) (186)Re-PEGylated liposomes (1295 MBq/kg), (e) (186)Re-liposomal doxorubicin (555 MBq/kg), (f) PEGylated liposomes plus RF ablation, (g) liposomal doxorubicin plus RF ablation, (h) (186)Re-PEGylated liposomes plus RF ablation, or (i) (186)Re-liposomal doxorubicin plus RF ablation. Six rats did not receive any treatment (control group). Tumor uptake in (186)Re therapy groups was monitored with small-animal single photon emission computed tomography for 5 days. Therapeutic efficacy was monitored for 6 weeks with measurement of tumor volume, calculation of the percentage injected dose of fluorine 18 fluorodeoxyglucose (FDG) in tumor from small-animal positron emission tomography (PET) images, and determination of viable tumor volume at histopathologic examination. Significant differences between groups were determined with analysis of variance. RESULTS: The average tumor volume (± standard deviation) on the day of therapy was 1.32 cm(3) ± 0.17. At 6 weeks after therapy, control of tumor growth was better with (186)Re-liposomal doxorubicin than with liposomal doxorubicin alone (tumor volume, 2.26 cm(3) ± 0.89 vs 5.43 cm(3) ± 0.93, respectively; P < .01). The use of RF ablation with liposomal doxorubicin and (186)Re-liposomal doxorubicin further improved tumor control (tumor volume, 2.05 cm(3) ± 1.36 and 1.49 cm(3) ± 1.47, respectively). The tumor growth trend correlated with change in percentage of injected dose of FDG in tumor for all groups (R(2) = 0.85, P < .001). Viable tumor volume was significantly decreased in the group treated with (186)Re-liposomal doxorubicin plus RF ablation (0.54 cm(3) ± 0.38; P < .001 vs all groups except (186)Re-liposomal doxorubicin alone). CONCLUSION: Triple and dual therapies had an observable trend ((186)Re-liposomal doxorubicin plus RF ablation > (186)Re-liposomal doxorubicin > liposomal doxorubicin plus RF ablation > liposomal doxorubicin) of improved tumor growth control and decreased viable tumor compared with other therapies. FDG PET could be used as a noninvasive surrogate marker for tumor growth and viability in this tumor model.

Novel asparagine-derived lipid enhances distearoylphosphatidylcholine bilayer resistance to acidic conditions.

A novel asparagine-derived lipid analogue (ALA(11,17)) bearing a tetrahydropyrimidinone headgroup and two fatty chains (11 and 17 indicate the lengths of linear alkyl groups) was synthesized in high yield and purity. The thin film hydration of formulations containing 5 mol % or greater ALA(11,17) in distearoylphosphatidylcholine (DSPC) generated multilamellar vesicles (MLVs) that remained unaggregated according to optical microscopy, while those formed from DSPC only were highly clustered. The MLVs were processed into unilamellar liposomes via extrusion and were characterized by dynamic light scattering (DLS), zeta potential, turbidity, and scanning electron microscopy (SEM) analysis. Results show that the presence of ALA(11,17) in DSPC liposomes significantly alters the morphology, colloidal stability, and retention of encapsulated materials in both acidic and neutral conditions. The ability of ALA(11,17)-hybrid liposomes to encapsulate and retain inclusions under neutral and acidic conditions (pH < 2) was demonstrated by calcein dequenching experiments. DLS and SEM confirmed that ALA(11,17)/DSPC liposomes remained intact under these conditions. The bilayer integrity observed under neutral and acidic conditions and the likely biocompatibility of these fatty amino acid analogues suggest that ALA(11,17) is a promising additive for modulating phosphatidylcholine lipid bilayer properties.

Remote-loading labeling of liposomes with (99m)Tc-BMEDA and its stability evaluation: effects of lipid formulation and pH/chemical gradient.

Efficient, convenient, and stable radiolabeling plays a critical role for the monitoring of liposome behavior via either blood sampling, organ distribution, or noninvasive nuclear imaging. The direct labeling of liposome-carrying drugs without any prior modification undoubtedly is convenient and optimal for liposomal drug testing. In this article, we investigated the effect of various lipid formulations and pH/chemical gradients on the radiolabeling efficiency and entrapment stability of technetium-99m ((99m)Tc) remotely loaded into liposomes, using (99m)Tc-N,N-bis(2-mercaptoethyl)-N',N'-diethyl-ethylenediamine ((99m)Tc-BMEDA) complex. The tested liposomes either contained unsaturated lipid or possessed various surface charges. (99m)Tc could be efficiently loaded into various premanufactured liposomes containing either an ammonium sulfate pH, citrate pH, or glutathione (GSH) chemical gradient. (99m)Tc-entrapment stabilities of these liposomes in phosphate-buffered saline (PBS; pH 7.4) buffer at 25°C were mainly dependent on the pH/chemical gradient, but not lipid formulation. Stability sequence was ammonium sulfate pH-gradient>citrate pH-gradient>GSH-gradient. Stabilities of (99m)Tc-liposomes in 50% fetal bovine serum (FBS)/PBS (pH 7.4) buffer at 37°C are dependent on both lipid formulation and pH/chemical gradient. Specifically, (99m)Tc labeling of the ammonium sulfate pH-gradient liposomes were less stable in 50% FBS/PBS than in PBS, whereas noncationic liposomes with citrate pH- or GSH-gradient displayed higher stability, except that anionic citrate pH-gradient liposomes showed no stability difference in these two media. Cationic liposomes aggregated in 50% FBS/PBS, forming a new discrete fraction with larger particle sizes. These in vitro characterization results have indicated the optimism of using (99m)Tc-BMEDA for labeling pH/GSH gradient liposomes without the requirement of modifying lipid formulation for liposomal therapeutic-agent development.

Interventional therapy of head and neck cancer with lipid nanoparticle-carried rhenium 186 radionuclide.

PURPOSE: Minimally invasive interventional cancer therapy with drug-carrying lipid nanoparticles (ie, liposomes) via convection-enhanced delivery by an infusion pump can increase intratumoral drug concentration and retention while facilitating broad distribution throughout solid tumors. The authors investigated the utility of liposome-carrying beta-emitting radionuclides to treat head and neck cancer by direct intratumoral infusion in nude rats. MATERIALS AND METHODS: Four groups of nude rats were subcutaneously inoculated with human tongue cancer cells. After tumors reached an average size of 1.6 cm(3), the treatment group received an intratumoral infusion of liposomal rhenium-186 ((186)Re) (185 MBq [5 mCi]/cm(3) tumor). Three control groups were intratumorally infused with unlabeled liposomes, unencapsulated (186)Re-perrhenate, or unencapsulated intermediate (186)Re compound ((186)Re-N,N-bis[2-mercaptoethyl]-N',N'-diethyl-ethylenediamine [BMEDA]). In vivo distribution of (186)Re activity was measured by planar gamma-camera imaging. Tumor therapy and toxicity were assessed by tumor size, body weight, and hematology. RESULTS: Average tumor volume in the (186)Re-liposome group on posttreatment day 14 decreased to 87.7% +/- 20.1%, whereas tumor volumes increased to 395.0%-514.4% on average in the other three groups (P< .001 vs (186)Re-liposome). The (186)Re-liposomes provided much higher intratumoral retention of (186)Re activity, resulting in an average tumor radiation absorbed dose of 526.3 Gy +/- 93.3, whereas (186)Re-perrhenate and (186)Re-BMEDA groups had only 3.3 Gy +/- 1.2 and 13.4 Gy +/- 9.2 tumor doses, respectively. No systemic toxicity was observed. CONCLUSIONS: Liposomal (186)Re effectively treated head and neck cancer with minimal side effects after convection-enhanced interventional delivery. These results suggest the potential of liposomal (186)Re for clinical application in interventional therapy of cancer.

Intraoperative 186Re-liposome radionuclide therapy in a head and neck squamous cell carcinoma xenograft positive surgical margin model.

PURPOSE: Positive surgical margins in advanced head and neck squamous cell carcinoma (HNSCC) have a well-documented association with an increased risk of locoregional recurrence and significantly poorer survival. Traditionally, unresectable tumor is treated with postoperative radiotherapy and/or chemotherapy. However, these therapeutic options can delay treatment and increase toxicity. The potential value of intraoperative injection of liposomal therapeutic radionuclides as a locoregional, targeted therapy in unresectable advanced HNSCC was assessed in a nude rat xenograft positive surgical margin model. EXPERIMENTAL DESIGN: The therapeutic effects of beta-emission rhenium-186 (186Re) carried by liposomes into the tumor remnants in a nude rat squamous cell carcinoma xenograft model were studied. Following the partial resection of tumor xenografts, the animals were intratumorally injected with 186Re-labeled or unlabeled (control) neutrally charged or positively charged 100-nm-diameter liposomes. Tumor size, body weight, hematology, and toxicity were monitored for 35 days posttherapy. RESULTS: The neutral (n = 4) and cationic (n = 4) liposome control groups showed an increase in tumor growth of 288.0 +/- 37.3% and 292.2 +/- 133.7%, respectively, by day 15. The 186Re-neutral-liposome group (n = 8) and the 186Re-cationic-liposome group (n = 8) presented with an average final tumor volume of 25.6 +/- 21.8% and 28.5 +/- 32.2%, respectively, at the end of the study (day 35). All groups showed consistent increases in body weight. No significant systemic toxicity was observed in any of the animals. CONCLUSIONS: With excellent tumor suppression and minimal side-effect profile, the intraoperative use of liposomal therapeutic radionuclides may play a role in the management of positive surgical margins in advanced HNSCC.

Selective uptake of surface-modified phospholipid vesicles by bone marrow macrophages in vivo.

An advantage of using vesicles (liposomes) as drug delivery carriers is that their pharmacokinetics can be controlled by surface characteristics, which can permit specific delivery of the encapsulated agents to organs or cells in vivo. Here we report a vesicle formulation which targets the bone marrow after intravenous injection in rabbits. Surface modification of the vesicle with an anionic amphiphile; L-glutamic acid, N-(3-carboxy-1-oxopropyl)-, 1,5-dihexadecyl ester (SA) results in significant targeting of vesicles to bone marrow. Further incorporation of as little as 0.6 mol% of poly(ethylene glycol)-lipid (PEG-DSPE) passively enhanced the distribution of SA-vesicles into bone marrow and inhibited hepatic uptake. In this model, more than 60% of the intravenously injected vesicles were distributed to bone marrow within 6 h after administration of a small dose of lipid (15 mg/kg b.w.). Histological evidence indicates that the targeting was achieved due to uptake by bone marrow macrophages (BMMphi). The efficient delivery of encapsulated scintigraphic and fluorescent imaging agents to BMMphi suggests that vesicles are promising carriers for the specific targeting of BMMphi and may be useful for delivering a wide range of therapeutic agents to bone marrow.

Cerebral oxygen delivery by liposome-encapsulated hemoglobin: a positron-emission tomographic evaluation in a rat model of hemorrhagic shock.

Liposome-encapsulated Hb (LEH) is being developed as an artificially assembled, low-toxicity, and spatially isolated Hb-based oxygen carrier (HBOC). Standard methods of evaluating oxygen carriers are based on surrogate indicators of physiology in animal models of shock. Assessment of actual delivery of oxygen by HBOCs and resultant improvement in oxygen metabolism at the tissue level has been a technical challenge. In this work, we report our findings from 15O-positron emission tomographic (15O-PET) evaluation of LEH in a rat model of 40% hypovolemic shock. In vitro studies showed that PEGylated LEH formulation containing approximately 7.5% Hb and consisting of neutral lipids (distearoylphosphatidylcholine:cholesterol:alpha-tocopherol, 51.4:46.4:2.2) efficiently picks up 15O-labeled oxygen gas. The final preparation of LEH contained 5% human serum albumin to provide oncotic pressure. Cerebral PET images of anesthetized rats inhaling 15O-labeled O2 gas showed efficient oxygen-carrying and delivery capacity of LEH formulation. From the PET images, we determined cerebral metabolic rate of oxygen (CMR(O2)) as a direct indicator of oxygen-carrying capacity of LEH as well as oxygen delivery and metabolism in rat brain. Compared with control fluids [saline and 5% human serum albumin (HSA)], LEH significantly improved CMR(O2) to approximately 80% of baseline level. Saline and HSA resuscitation could not improve hypovolemia-induced decrease in CMR(O2). On the other hand, resuscitation of shed blood was the most efficient in restoring oxygen metabolism. The results suggest that 15O-PET technology can be successfully employed to evaluate potential oxygen carriers and blood substitutes and that LEH resuscitation in hemorrhage enhances oxygen delivery to the cerebral tissue and improves oxygen metabolism in brain.

Use of avidin/biotin-liposome system for enhanced peritoneal drug delivery in an ovarian cancer model.

The goal of this study was to determine the distribution of the avidin/biotin-liposome system in an ovarian cancer xenograft model. Optimal avidin/biotin-liposome injection sequence with enhanced liposome accumulation to the peritoneum was determined. Two weeks after NIH:OVCAR-3 cell inoculation, rats were divided into three groups. Group 1 (B-A) (n=4), received an intraperitoneal injection of (99m)Tc-blue-biotin-liposomes 30 min before an intraperitoneal injection of avidin. Group 2 (A-B) (n=4), received an intraperitoneal injection of avidin 30 min before an intraperitoneal injection of (99m)Tc-blue-biotin-liposomes. Group 3 (A-B 2h) (n=5), received an intraperitoneal injection of avidin 2h before an intraperitoneal injection of (99m)Tc-blue-biotin-liposomes. Three additional non-tumor nude rats served as controls in each group, and were subjected to the same injection sequences. Scintigraphic imaging commenced at various times post (99m)Tc-blue-biotin-liposome injection. After imaging, rats were euthanized at 23 h post-liposome injection for tissue biodistribution. Images showed no apparent difference in liposome distribution between control and tumor animals. Regional uptake analysis at 4h for tumor rats showed significantly higher lymphatic channel uptake in the A-B 2h group (p<0.05) and a trend of increased peritoneal uptake in A-B group. By 22 h, peritoneal and lymphatic channel uptake was similar for all groups. At necropsy, most activity was found in blue-stained omentum, diaphragm, mediastinal and abdominal nodes. Bowel activity was minimal. These results correlate with previous normal rat studies, and demonstrate potential use of this avidin/biotin-liposome system for prolonging drug delivery to the peritoneal cavity and associating lymph nodes in this ovarian cancer xenograft model.

Techniques for Loading Technetium-99m and Rhenium-186/188 Radionuclides into Preformed Liposomes for Diagnostic Imaging and Radionuclide Therapy.

Liposomes can serve as carriers of radionuclides for diagnostic imaging and therapeutic applications. Herein, procedures are outlined for radiolabeling liposomes with the gamma-emitting radionuclide, technetium-99m (99mTc), for noninvasive detection of disease and for monitoring the pharmacokinetics and biodistribution of liposomal drugs, and/or with therapeutic beta-emitting radionuclides, rhenium-186/188 (186/188Re), for radionuclide therapy. These efficient and practical liposome radiolabeling methods use a post-labeling mechanism to load 99mTc or 186/188Re into preformed liposomes prepared in advance of the labeling procedure. For all liposome radiolabeling methods described, a lipophilic chelator is used to transport 99mTc or 186/188Re across the lipid bilayer of the preformed liposomes. Once within the liposome interior, the pre-encapsulated glutathione or ammonium sulfate (pH) gradient provides for stable entrapment of the 99mTc and 186/188Re within the liposomes. In the first method, 99mTc is transported across the lipid bilayer by the lipophilic chelator, hexamethylpropyleneamine oxime (HMPAO) and 99mTc-HMPAO becomes trapped by interaction with the pre-encapsulated glutathione within the liposomes. In the second method, 99mTc or 186/188Re is transported across the lipid bilayer by the lipophilic chelator, N,N-bis(2-mercaptoethyl)-N',N'-diethylethylenediamine (BMEDA), and 99mTc-BMEDA or 186/188Re-BMEDA becomes trapped by interaction with pre-encapsulated glutathione within the liposomes. In the third method, an ammonium sulfate (pH) gradient loading technique is employed using liposomes with an extraliposomal pH of 7.4 and an interior pH of 5.1. BMEDA, which is lipophilic at pH 7.4, serves as a lipophilic chelator for 99mTc or 186/188Re to transport the radionuclides across the lipid bilayer. Once within the more acidic liposome interior, 99mTc/186/188Re-BMEDA complex becomes protonated and more hydrophilic, which results in stable entrapment of the 99mTc/186/188Re-BMEDA complex within the liposomes. Since many commercially available liposomal drugs use an ammonium sulfate (pH) gradient for drug loading, these liposomal drugs can be directly radiolabeled with 99mTc-BMEDA for noninvasive monitoring of tissue distribution during treatment or with 186/188Re-BMEDA for combination chemo-radionuclide therapy.

Mediastinal node and diaphragmatic targeting after intracavitary injection of avidin/99mTc-blue-biotin-liposome system.

A method for delivering drugs to sites of disease extension in mediastinal nodes is described. Mediastinal node and lymphatic distributions were determined after intracavitary injection of the avidin/biotin-liposome system in normal rats. The effect of the injected dose on lymphatic targeting of liposomes after intraperitoneal injection of (99m)Tc-blue-biotin-liposomes and intrapleural injection of avidin, and vice versa, is presented. Scintigraphic imaging was used to follow the movement of (99m)Tc-blue-biotin-liposomes to determine the pharmacokinetics and organ uptake. Tissue biodistribution studies were performed 22 h after injection of the (99m)Tc-blue-biotin-liposomes. Results indicated that independent of the cavity in which each agent was injected, a dose of 5.0 mg of each agent results in higher mediastinal node targeting (8%-10% ID/Organ) as compared with the injection of a 0.5 mg dose (2%-5% ID/Organ, p < 0.05). Targeting of diaphragm and associated lymphatics was observed when (99m)Tc-blue-biotin-liposomes were injected in peritoneum and avidin in pleural space. In contrast, pleural, and pericardial lymphatic targeting was observed when (99m)Tc-blue-biotin-liposomes were injected in pleural space and avidin in peritoneum. Intracavitary injection of the avidin/biotin-liposome system could potentially be used for the delivery of prophylactic drugs that could reduce tumor metastasis and infection spread to mediastinal nodes.

Potential use of drug carried-liposomes for cancer therapy via direct intratumoral injection.

Liposomes have recognized advantages as nano-particle drug carriers for tumor therapy. In this study, the pharmacokinetics and distribution of intratumorally administered liposomes were investigated as drug carriers for treating solid tumors via direct intratumoral administration. 99mTc-liposomes were administered intratumorally to nude rats bearing human head and neck squamous cell carcinoma xenografts. Planar gamma camera images were analyzed to evaluate the local retention of the intratumorally administered liposomes. Co-registered pinhole micro-single photon emission computed tomography (SPECT)/computed tomography (CT) images were acquired of the whole animal as well as the dissected tumors to determine intratumoral distribution of the 99mTc-liposomes. For 99mTc-liposomes, there was an initial retention of 47.4 +/- 11.0% (n = 4) in tumors and surrounding tissues. At 20 h, 39.2 +/- 10.6% (n = 4) of 99mTc-activity still remained in the tumor. In contrast, only 18.7 +/- 3.3% (n = 3) of the intratumoral 99mTc-activity remained for unencapsulated 99mTc-complex at 20 h. Pinhole micro-SPECT images demonstrated that 99mTc-liposomes also have a superior intratumoral 99mTc-activity diffusion compared with unencapsulated 99mTc-complex. Higher intratumoral retention of 99mTc-liposomes accompanied by an improved intratumoral diffusion suggests that intratumorally administered liposomal drugs are potentially promising agents for solid tumor local therapy.

Strategies for improving the intratumoral distribution of liposomal drugs in cancer therapy.

INTRODUCTION: A major limitation of current liposomal cancer therapies is the inability of liposome therapeutics to penetrate throughout the entire tumor mass. This inhomogeneous distribution of liposome therapeutics within the tumor has been linked to treatment failure and drug resistance. Both liposome particle transport properties and tumor microenvironment characteristics contribute to this challenge in cancer therapy. This limitation is relevant to both intravenously and intratumorally administered liposome therapeutics. AREAS COVERED: Strategies to improve the intratumoral distribution of liposome therapeutics are described. Combination therapies of intravenous liposome therapeutics with pharmacologic agents modulating abnormal tumor vasculature, interstitial fluid pressure, extracellular matrix components, and tumor associated macrophages are discussed. Combination therapies using external stimuli (hyperthermia, radiofrequency ablation, magnetic field, radiation, and ultrasound) with intravenous liposome therapeutics are discussed. Intratumoral convection-enhanced delivery (CED) of liposomal therapeutics is reviewed. EXPERT OPINION: Optimization of the combination therapies and drug delivery protocols are necessary. Further research should be conducted in appropriate cancer types with consideration of physiochemical features of liposomes and their timing sequence. More investigation of the role of tumor associated macrophages in intratumoral distribution is warranted. Intratumoral infusion of liposomes using CED is a promising approach to improve their distribution within the tumor mass.

186Re-liposome labeling using 186Re-SNS/S complexes: in vitro stability, imaging, and biodistribution in rats.

UNLABELLED: Liposomes are important carriers for controlling the spatial and temporal distribution of drug molecules or other bioactive molecules. Radiolabeled liposomes have potential applications in diagnostic imaging and radionuclide therapy. The purpose of this study was to develop a practical method for labeling liposomes with therapeutic rhenium radionuclides, using (186)Re as an example. METHODS: An SNS pattern ligand, N,N-bis(2-mercaptoethyl)-N',N'-diethylethylenediamine (BMEDA), and an S pattern ligand, benzene thiol (BT), were used to make 2 kinds of (186)Re-SNS/S complexes, (186)Re-BMEDA and (186)Re-BMEDA + BT. These (186)Re-SNS/S complexes were mixed with neutral liposomes encapsulating cysteine or (NH(4))(2)SO(4) to prepare (186)Re-liposomes. The in vitro labeling stability of (186)Re-liposomes was investigated by incubation in 50% fetal bovine serum/50% phosphate-buffered saline, pH 7.4, at 37 degrees C. Rat distribution studies of (186)Re-liposomes after intravenous injection were also performed. RESULTS: The labeling efficiencies of (186)Re-liposomes were 52.9%-81.3% depending on the (186)Re-SNS/S complex chosen and whether cysteine- or (NH(4))(2)SO(4)-encapsulated liposomes were used. (186)Re-(NH(4))(2)SO(4) liposomes labeled with (186)Re-BMEDA had the best in vitro labeling stability in serum with 89.8% +/- 3.1% of the radioactivity associated with liposomes at 24 h and 76.2% +/- 5.1% at 96 h. A specific activity of 1.85 GBq (50 mCi) of (186)Re per 50 mg of phospholipid could be achieved with good labeling stability. Biodistributions were followed for 72 h and showed good in vivo stability for (186)Re-liposomes that was characterized by a slow blood clearance and a gradually increasing spleen accumulation. (186)Re-BMEDA alone had fast blood clearance and no accumulation in spleen. CONCLUSION: A practical method for labeling liposomes with (186)Re using (186)Re-SNS/S complexes is described. The labeled (186)Re-liposomes were stable in serum and in vivo and could potentially be useful for radionuclide therapy.

Pharmacokinetics and biodistribution of [111In]-avidin and [99mTc]-biotin-liposomes injected in the pleural space for the targeting of mediastinal nodes.

Pharmacokinetics and mediastinal node uptake of [111In]-avidin and [99mTc]-biotin-liposomes following either intrapleural (pleural) or intraperitoneal (ip) injection were determined using scintigraphic imaging. Biodistribution results of [111In]-avidin at 44 h showed 3.3% uptake in mediastinal nodes by pleural injection vs 1.3% with ip injection. Mediastinal node accumulation with [99mTc]-biotin-liposomes was not different between injections (0.6% ip vs 0.5% pleural). This study demonstrates the potential of the pleural route as a technique for mediastinal node targeting using the avidin/biotin-liposome system.

Avidin/biotin-liposome system injected in the pleural space for drug delivery to mediastinal lymph nodes.

The objective of this study was to develop a more effective liposome-based method for delivering drugs to mediastinal nodes. Nodal uptake was determined after intrapleural injection of the avidin/biotin-liposome system in normal rats. The effect of injection sequence (avidin injected 2 h before biotin-liposomes and vice versa), volume injected, and administered dose of the agents is described. Pharmacokinetics of the avidin/biotin-liposome system was monitored with scintigraphic imaging by labeling the biotin-liposomes with technetium-99m ((99m)Tc). To identify the nodes during the biodistribution studies, patent blue dye was encapsulated in the biotin-liposomes. Tissue biodistribution studies were performed 22 h after injection of the (99m)Tc-blue-biotin-liposomes. When avidin was injected before (99m)Tc-blue-biotin-liposomes, better mediastinal node targeting (15.7%; p < 0.05) was achieved than when biotin-liposomes were injected first (8.3%) or when only biotin-liposomes were injected (1.0%). Injection of a small dose of liposomes (0.5 mg phospholipid) and avidin (0.5 mg) resulted in the most favorable drug delivery to mediastinal nodes and other organs. Intrapleural injection of the avidin/biotin-liposome system could potentially be used for drug delivery to disease processes such as lung cancer, anthrax, and tuberculosis that invade mediastinal nodes and use them as centers of incubation and dissemination.