Assessment of Clinical Response to V937 Oncolytic Virus After Intravenous or Intratumoral Administration Using Physiologically-Based Modeling.

Oncolytic viruses (OVs) represent a potential therapeutic strategy in cancer treatment. However, there is currently a lack of comprehensive quantitative models characterizing clinical OV kinetics and distribution to the tumor. In this work, we present a mechanistic modeling framework for V937 OV, after intratumoral (i.t.) or intravascular (i.v.) administration in patients with cancer. A minimal physiologically-based pharmacokinetic model was built to characterize biodistribution of OVs in humans. Viral dynamics was incorporated at the i.t. cellular level and linked to tumor response, enabling the characterization of a direct OV killing triggered by the death of infected tumor cells and an indirect killing induced by the immune response. The model provided an adequate description of changes in V937 mRNA levels and tumor size obtained from phase I/II clinical trials after V937 administration. The model showed prominent role of viral clearance from systemic circulation and infectivity in addition to known tumor aggressiveness on clinical response. After i.v. administration, i.t. exposure of V937 was predicted to be several orders of magnitude lower compared with i.t. administration. These differences could be overcome if there is high virus infectivity and/or replication. Unfortunately, the latter process could not be identified at the current clinical setting. This work provides insights on selecting optimal OV considering replication rate and infectivity.

Stealth nanoparticles in oncology: Facing the PEG dilemma.

Nanoparticles (Nps) have revolutionized the landscape of many treatments, by modifying not only pharmacokinetic properties of the encapsulated agent, but also providing a significant protection of the drug from non-desired interactions, and reducing side-effects of the enclosed therapeutic, enabling co-encapsulation of possibly synergistic compounds or activities, allowing a controlled release of content and improving the therapeutic effect. Nevertheless, in systemic circulation, Nps suffer a rapid removal by opsonisation and the action of Mononuclear phagocyte system (MPS). To overcome this problem, different polymers, in particular Polyethyleneglycol (PEG), have been used to cover the surface of these nanocarriers forming a hydrophilic layer that allows the delay of the removal. These advantages contrast with some drawbacks such as the difficulty to interact with cell membranes and the development of immunological reactions, conforming the known, "PEG dilemma". To address and minimize this phenomenon, different strategies have been applied. Therefore, this review aims to summarize the state of the art of Pegylation strategies, comment in depth on the principal characteristics of PEG and describe the main alternatives, which are the use of cleavable PEG, addition of different polymers or even use other derivatives of cell membranes to camouflage Nps.

Semi-Mechanistic Model for the Antitumor Response of a Combination Cocktail of Immuno-Modulators in Non-Inflamed (Cold) Tumors.

Immune checkpoint inhibitors, administered as single agents, have demonstrated clinical efficacy. However, when treating cold tumors, different combination strategies are needed. This work aims to develop a semi-mechanistic model describing the antitumor efficacy of immunotherapy combinations in cold tumors. Tumor size of mice treated with TC-1/A9 non-inflamed tumors and the drug effects of an antigen, a toll-like receptor-3 agonist (PIC), and an immune checkpoint inhibitor (anti-programmed cell death 1 antibody) were modeled using Monolix and following a middle-out strategy. Tumor growth was best characterized by an exponential model with an estimated initial tumor size of 19.5 mm3 and a doubling time of 3.6 days. In the treatment groups, contrary to the lack of response observed in monotherapy, combinations including the antigen were able to induce an antitumor response. The final model successfully captured the 23% increase in the probability of cure from bi-therapy to triple-therapy. Moreover, our work supports that CD8+ T lymphocytes and resistance mechanisms are strongly related to the clinical outcome. The activation of antigen-presenting cells might be needed to achieve an antitumor response in reduced immunogenic tumors when combined with other immunotherapies. These models can be used as a platform to evaluate different immuno-oncology combinations in preclinical and clinical scenarios.

Mechanistic Modeling of a Novel Oncolytic Virus, V937, to Describe Viral Kinetic and Dynamic Processes Following Intratumoral and Intravenous Administration.

V937 is an investigational novel oncolytic non-genetically modified Kuykendall strain of Coxsackievirus A21 which is in clinical development for the treatment of advanced solid tumor malignancies. V937 infects and lyses tumor cells expressing the intercellular adhesion molecule I (ICAM-I) receptor. We integrated in vitro and in vivo data from six different preclinical studies to build a mechanistic model that allowed a quantitative analysis of the biological processes of V937 viral kinetics and dynamics, viral distribution to tumor, and anti-tumor response elicited by V937 in human xenograft models in immunodeficient mice following intratumoral and intravenous administration. Estimates of viral infection and replication which were calculated from in vitro experiments were successfully used to describe the tumor response in vivo under various experimental conditions. Despite the predicted high clearance rate of V937 in systemic circulation (t1/2 = 4.3 min), high viral replication was observed in immunodeficient mice which resulted in tumor shrinkage with both intratumoral and intravenous administration. The described framework represents a step towards the quantitative characterization of viral distribution, replication, and oncolytic effect of a novel oncolytic virus following intratumoral and intravenous administrations in the absence of an immune response. This model may further be expanded to integrate the role of the immune system on viral and tumor dynamics to support the clinical development of oncolytic viruses.

Drug transport kinetics of intravascular triggered drug delivery systems.

Intravascular triggered drug delivery systems (IV-DDS) for local drug delivery include various stimuli-responsive nanoparticles that release the associated agent in response to internal (e.g., pH, enzymes) or external stimuli (e.g., temperature, light, ultrasound, electromagnetic fields, X-rays). We developed a computational model to simulate IV-DDS drug delivery, for which we quantified all model parameters in vivo in rodent tumors. The model was validated via quantitative intravital microscopy studies with unencapsulated fluorescent dye, and with two formulations of temperature-sensitive liposomes (slow, and fast release) encapsulating a fluorescent dye as example IV-DDS. Tumor intra- and extravascular dye concentration dynamics were extracted from the intravital microscopy data by quantitative image processing, and were compared to computer model results. Via this computer model we explain IV-DDS delivery kinetics and identify parameters of IV-DDS, of drug, and of target tissue for optimal delivery. Two parameter ratios were identified that exclusively dictate how much drug can be delivered with IV-DDS, indicating the importance of IV-DDS with fast drug release (~sec) and choice of a drug with rapid tissue uptake (i.e., high first-pass extraction fraction). The computational model thus enables engineering of improved future IV-DDS based on tissue parameters that can be quantified by imaging.

Dual activity of PD-L1 targeted Doxorubicin immunoliposomes promoted an enhanced efficacy of the antitumor immune response in melanoma murine model.

BACKGROUND: The immunomodulation of the antitumor response driven by immunocheckpoint inhibitors (ICIs) such as PD-L1 (Programmed Death Ligand-1) monoclonal antibody (α-PD-L1) have shown relevant clinical outcomes in a subset of patients. This fact has led to the search for rational combinations with other therapeutic agents such as Doxorubicin (Dox), which cytotoxicity involves an immune activation that may enhance ICI response. Therefore, this study aims to evaluate the combination of chemotherapy and ICI by developing Dox Immunoliposomes functionalized with monovalent-variable fragments (Fab') of α-PD-L1. RESULTS: Immunoliposomes were assayed in vitro and in vivo in a B16 OVA melanoma murine cell line over-expressing PD-L1. Here, immune system activation in tumor, spleen and lymph nodes, together with the antitumor efficacy were evaluated. Results showed that immunoliposomes bound specifically to PD-L1+ cells, yielding higher cell interaction and Dox internalization, and decreasing up to 30-fold the IC50, compared to conventional liposomes. This mechanism supported a higher in vivo response. Indeed, immunoliposomes promoted full tumor regression in 20% of mice and increased in 1 month the survival rate. This formulation was the only treatment able to induce significant (p < 0.01) increase of activated tumor specific cytotoxic T lymphocytes at the tumor site. CONCLUSION: PD-L1 targeted liposomes encapsulating Dox have proved to be a rational combination able to enhance the modulation of the immune system by blocking PD-L1 and selectively internalizing Dox, thus successfully providing a dual activity offered by both, chemo and immune therapeutic strategies.

Modulation of intratumoural myeloid cells, the hallmark of the anti-tumour efficacy induced by a triple combination: tumour-associated peptide, TLR-3 ligand and α-PD-1.

BACKGROUND: Anti-programmed cell death 1 (PD-1)/programmed death-ligand 1 (PD-L1) monoclonal antibodies (mAbs) show remarkable clinical anti-tumour efficacy. However, rational combinations are needed to extend the clinical benefit to primary resistant tumours. The design of such combinations requires the identification of the kinetics of critical immune cell populations in the tumour microenvironment. METHODS: In this study, we compared the kinetics of immune cells in the tumour microenvironment upon treatment with immunotherapy combinations with different anti-tumour efficacies in the non-inflamed tumour model TC-1/A9. Tumour-bearing C57BL/6J mice were treated with all possible combinations of a human papillomavirus (HPV) E7 long peptide, polyinosinic-polycytidylic acid (PIC) and anti-PD-1 mAb. Tumour growth and kinetics of the relevant immune cell populations were assessed over time. The involvement of key immune cells was confirmed by depletion with mAbs and immunophenotyping with multiparametric flow cytometry. RESULTS: The maximum anti-tumour efficacy was achieved after intratumoural administration of HPV E7 long peptide and PIC combined with the systemic administration of anti-PD-1 mAb. The intratumoural immune cell kinetics of this combination was characterised by a biphasic immune response. An initial upsurge of proinflammatory myeloid cells led to a further rise in effector CD8+ T lymphocytes at day 8. Depletion of either myeloid cells or CD8+ T lymphocytes diminished the anti-tumour efficacy of the combination. CONCLUSIONS: The anti-tumour efficacy of a successful immunotherapy combination in a non-inflamed tumour model relies on an early inflammatory process that remodels the myeloid cell compartment.

Sensitization of drug resistant sarcoma tumors by membrane modulation via short chain sphingolipid-containing nanoparticles.

Nanoparticles such as liposomes are able to overcome cancer treatment challenges such as multidrug resistance by increasing the bioavailability of the encapsulated drug, bypassing drug pumps or through targeting resistant cells. Here, we merge enhanced drug delivery by nanotechnology with tumor cell membrane modulation combined in a single formulation. This is achieved through the incorporation of Short chain sphingolipids (SCSs) in the liposomal composition, which permeabilizes cell membranes to amphiphilic drugs such as Doxorubicin (Dxr). To study the mechanism and capability of SCS-containing nanodevices to overcome Dxr resistance, a sensitive uterine sarcoma cell line, MES-SA, and a resistant derived cell line, MES-SA/MX2, were used. The mechanism of resistance was explored by lipidomics and flow cytometry, revealing significant differences in lipid composition and in P glycoprotein (Pgp) expression. In vitro assays show that SCS liposomes were able to reverse cell resistance, and importantly, display a higher net effect on resistant than sensitive cells. SCS lipids modulated the cell membrane of MES-SA/MX2 drug resistant cells, while Pgp expression was not affected. Furthermore, SCS-modified liposomes were evaluated in a sarcoma xenograft model on drug accumulation, pharmacokinetics and efficacy. SCS liposomes improved Dxr levels in tumor nuclei of MES-SA/MX2 tumor cells, which was accompanied by a delay in tumor growth of the resistant model. Here we show that Dxr accumulation in tumor cells by SCS-modified liposomes was especially improved in Dxr resistant cells, rendering Dxr as effective as in sensitive cells. Moreover, this phenomenon translated to improved efficacy when Dxr liposomes where modified with SCSs in the drug resistant tumor model, while no benefit was seen in the sensitive tumors.

Quantification of Pharmacokinetic Profiles of PD-1/PD-L1 Antibodies by Validated ELISAs.

Immunotherapy has changed the paradigm of cancer treatments. In this way, several combinatorial strategies based on monoclonal antibodies (mAb) such as anti (a)-PD-1 or anti (a)-PD-L1 are often reported to yield promising clinical benefits. However, the pharmacokinetic (PK) behavior of these mAbs is a critical issue that requires selective analytical techniques. Indeed, few publications report data on a-PD1/a-PD-L1 exposure and its relationship with therapeutic or toxic effects. In this regard, preclinical assays allow the time profiles of antibody plasma concentrations to be characterized rapidly and easily, which may help to increase PK knowledge. In this study, we have developed and validated two in-house ELISAs to quantify a-PD-1 and a-PD-L1 in plasma collected from tumor-bearing mice. The linear range for the a-PD-1 assay was 2.5-125 ng/mL and 0.11-3.125 ng/mL for the a-PD-L1 assay, whereas the intra-and inter-day precision was lower than 20% for both analytes. The PK characterization revealed a significant decrease in drug exposure after administration of multiple doses. Plasma half-life for a-PD-1 was slightly shorter (22.3 h) than for a-PD-L1 (46.7 h). To our knowledge, this is the first reported preclinical ELISA for these immune checkpoint inhibitors, which is sufficiently robust to be used in different preclinical models. These methods can help to understand the PK behavior of these antibodies under different scenarios and the relationship with response, thus guiding the choice of optimal doses in clinical settings.

Liposomes targeted to MHC-restricted antigen improve drug delivery and antimelanoma response.

PURPOSE: Melanoma is the most aggressive form of skin cancer. Chemotherapy at a late stage fails due to low accumulation in tumors, indicating the need for targeted therapy. MATERIALS AND METHODS: To increase drug uptake by tumor cells, we have targeted doxorubicin-containing liposomes using a T-cell receptor (TCR)-like antibody (scFv G8 and Hyb3) directed against melanoma antigen A1 (MAGE-A1) presented by human leukocyte antigen A1 (M1/A1). With the use of flow cytometry and confocal microscopy, we have tested our formulation in vitro. In vivo pharmacokinetics was done in tumor-free nu/nu mice, while biodistribution and efficacy study was done in nu/nu mice xenograft. RESULTS: We demonstrated two to five times higher binding and internalization of these immunoliposomes by M1+/A1+ melanoma cells in vitro in comparison with nontargeted liposomes. Cytotoxicity assay showed significant tumor cell kill at 10 µM doxorubicin (DXR) for targeted vs nontargeted liposomes. In vivo pharmacokinetics of nontargeted and targeted liposomes were similar, while accumulation of targeted liposomes was 2- to 2.5-fold and 6.6-fold enhanced when compared with nontargeted liposomes and free drug, respectively. Notably, we showed a superior antitumor activity of MAGE-A1-targeted DXR liposomes toward M1+/A1+ expressing tumors in mice compared with the treatment of M1-/A1+ tumors. Our results indicate that targeted liposomes showed better cytotoxicity in vitro and pharmacokinetics in vivo. CONCLUSION: Liposomes decorated with TCR-mimicking scFv antibodies effectively and selectively target antigen-positive melanoma. We showed that DXR-loaded liposomes coupled to anti-M1/-A1 scFv inflict a significant antitumor response. Targeting tumor cells specifically promotes internalization of drug-containing nanoparticles and may improve drug delivery and ultimately antitumor efficacy. Our data argue that targeting MAGE in A1 context, by nanosized carriers decorated with TCR-like antibodies mimicking scFv, can be used as a theragnostic platform for drug delivery, immunotherapy, and potentially imaging, and diagnosis of melanoma.

A new immune-nanoplatform for promoting adaptive antitumor immune response.

Immunoliposomes (ILs), obtained with monoclonal antibodies (mAbs) decorating the liposome surface, are used for cancer treatment. These mAbs provide the recognition of molecules upregulated in cancer cells, like Programmed Death-Ligand 1 (PD-L1), an immune-checkpoint involved in tumor resistance, forming a complex that blocks this molecule and thereby, induces antitumor immune response. This mechanism introduces a new concept for ILs. ILs coupled to anti-PD-L1 or its Fab' fragment have been developed and in vitro/in vivo characterized. Factors such as coupling methods, PEG density and ligand size were optimized. In vitro data showed that Fab'-ILs displayed the highest PD-L1 cell interaction, correlating with a higher in vivo tumor accumulation and an increase of effector cytotoxic CD8+ T cells, providing tumor shrinkage and total regression in 20% of mice. Therefore, a novel immune-nanoplatform able to modulate the immune system has been developed, allowing the encapsulation of several agents for combinatorial therapies.

Immunoliposomes in clinical oncology: State of the art and future perspectives.

Liposomal formulations entrapping a vast number of molecules have improved cancer therapies overcoming certain pharmacokinetic (PK) and pharmacodynamic limitations, many of which are associated with tumor characteristics. In this context, immunoliposomes represent a new strategy that has been widely investigated in preclinical cancer models with promising results, although few have reached the stage of clinical trials. This contrasts with the emerging clinical application of monoclonal antibodies (mAbs). This formulation allows the conjugation of different mAbs or antibody derivatives, such as monovalent variable fragments Fab', to the polymers covering the surface of liposomes. The combination of this targeting strategy together with drug encapsulation in a single formulation may contribute to enhance the efficacy of these associated agents, reducing their toxicities. In this paper we will consider how factors such as particle size, lipid composition and charge, lipid-polymer conjugation, method of production and type of ligand for liposome coupling influence the efficacy of these formulations. Furthermore, the high inter-individual variability in the tumor microenvironment, as well as the poor experimental designs for the PK characterization of liposomes, make the establishment of the relationship between plasma or tumor concentrations and efficacy difficult. Thus, adequate dosing regimens and patient stratification regarding the target expression may contribute to enhance the possibility of incorporating these immunoliposomes into the therapeutic arsenal for cancer treatments. All these issues will be briefly dealt with here, together with a section showing the state of the art of those targeted liposomes that are coming up for testing in clinical trials. Finally, some insights into future developments such as the combination of specificity and controlled release, based on the application of different stimuli, for the manipulation of stability and cargo release, will be offered. This has been included in order to highlight the new opportunities for targeted liposomes, including immunoliposomes.

Cell membrane modulation as adjuvant in cancer therapy.

Cancer is a complex disease involving numerous biological processes, which can exist in parallel, can be complementary, or are engaged when needed and as such can replace each other. This redundancy in possibilities cancer cells have, are fundamental to failure of therapy. However, intrinsic features of tumor cells and tumors as a whole provide also opportunities for therapy. Here we discuss the unique and specific makeup and arrangement of cell membranes of tumor cells and how these may help treatment. Interestingly, knowledge on cell membranes and associated structures is present already for decades, while application of membrane modification and manipulation as part of cancer therapy is lagging. Recent developments of scientific tools concerning lipids and lipid metabolism, opened new and previously unknown aspects of tumor cells and indicate possible differences in lipid composition and membrane function of tumor cells compared to healthy cells. This field, coined Lipidomics, demonstrates the importance of lipid components in cell membrane in several illnesses. Important alterations in cancer, and specially in resistant cancer cells compared to normal cells, opened the door to new therapeutic strategies. Moreover, the ability to modulate membrane components and/or properties has become a reality. Here, developments in cancer-related Lipidomics and strategies to interfere specifically with cancer cell membranes and how these affect cancer treatment are discussed. We hypothesize that combination of lipid or membrane targeted strategies with available care to improve chemotherapy, radiotherapy and immunotherapy will bring the much needed change in treatment in the years to come.

EGFR targeted thermosensitive liposomes: A novel multifunctional platform for simultaneous tumor targeted and stimulus responsive drug delivery.

The epidermal growth factor receptor (EGFR) is a promising target for anti-cancer therapy. The aim of this study was to design thermosensitive liposomes (TSL), functionalized with anti-EGFR ligands for targeted delivery and localized triggered release of chemotherapy. For targeting, EGFR specific peptide (GE11) and Fab' fragments of cetuximab were used and the effect of ligand density on in vitro tumor targeting was investigated. Ligand conjugation did not significantly change the physicochemical characteristics of liposomes. Fab'-decorated TSL (Fab'-TSL) can specifically and more efficiently bind to the EGFR overexpressed cancer cells as compared to GE11 modified TSL. Calcein labeled Fab'-TSL showed adequate stability at 37°C in serum (<4% calcein released after 1h) and a temperature dependent release at above 40°C. FACS analysis and live cell imaging showed efficient and EGFR mediated cellular association as well as dramatic intracellular cargo release upon hyperthermia. Fab'-conjugation and hyperthermia induced enhanced tumor cell cytotoxicity of doxorubicin loaded TSL. The relative cytotoxicity of Fab'-TSL was also correlated to EGFR density on the tumor cells. These results suggest that Fab'-TSL showed great potential for combinational targeted and triggered release drug delivery.

Targeting melanoma with immunoliposomes coupled to anti-MAGE A1 TCR-like single-chain antibody.

Therapy of melanoma using T-cells with genetically introduced T-cell receptors (TCRs) directed against a tumor-selective cancer testis antigen (CTA) NY-ESO1 demonstrated clear antitumor responses in patients without side effects. Here, we exploited the concept of TCR-mediated targeting through introduction of single-chain variable fragment (scFv) antibodies that mimic TCRs in binding major histocompatibility complex-restricted CTA. We produced scFv antibodies directed against Melanoma AntiGEn A1 (MAGE A1) presented by human leukocyte antigen A1 (HLA-A1), in short M1/A1, and coupled these TCR-like antibodies to liposomes to achieve specific melanoma targeting. Two anti-M1/A1 antibodies with different ligand-binding affinities were derived from a phage-display library and reformatted into scFvs with an added cysteine at their carboxyl termini. Protein production conditions, ie, bacterial strain, temperature, time, and compartments, were optimized, and following production, scFv proteins were purified by immobilized metal ion affinity chromatography. Batches of pure scFvs were validated for specific binding to M1/A1-positive B-cells by flow cytometry. Coupling of scFvs to liposomes was conducted by employing different conditions, and an optimized procedure was achieved. In vitro experiments with immunoliposomes demonstrated binding of M1/A1-positive B-cells as well as M1/A1-positive melanoma cells and internalization by these cells using flow cytometry and confocal microscopy. Notably, the scFv with nonenhanced affinity of M1/A1, but not the one with enhanced affinity, was exclusively bound to and internalized by melanoma tumor cells expressing M1/A1. Taken together, antigen-mediated targeting of tumor cells as well as promoting internalization of nanoparticles by these tumor cells is mediated by TCR-like scFv and can contribute to melanoma-specific targeting.

EGF-liposomes promote efficient EGFR targeting in xenograft colocarcinoma model.

AIM: Development of EGF-liposomes (LP-EGF) for selective molecules delivery in tumors expressing EGFR. MATERIAL & METHODS: In vitro cellular interaction of EGF-LP and nontargeted liposomes (LP-N) was assayed at 37 and 4 °C in cells expressing different EGFR levels. Receptor-mediated uptake was investigated by competition with a monoclonal antibody anti-EGFR. Selective intracellular drug delivery and efficacy was tested by oxaliplatin encapsulation. In vivo biodistribution of LP-N and LP-EGF was done in xenograft model. RESULTS: LP-EGF was internalized by an active and selective mechanism through EGFR without receptor activation. Oxaliplatin LP-EGF decreased IC50 between 48 and 13% in cell EGFR+. LP-EGF was accumulated in tumor over 72 h postdosing, while LP-N in spleen. CONCLUSION: LP-EGF represents an attractive nanosystem for cancer therapy or diagnosis.

Cetuximab-oxaliplatin-liposomes for epidermal growth factor receptor targeted chemotherapy of colorectal cancer.

Oxaliplatin (L-OH), a platinum derivative with good tolerability is currently combined with Cetuximab (CTX), a monoclonal antibody (mAb), for the treatment of certain (wild-type KRAS) metastatic colorectal cancer (CRC) expressing epidermal growth factor receptor (EGFR). Improvement of L-OH pharmacokinetics (PK) can be provided by its encapsulation into liposomes, allowing a more selective accumulation and delivery to the tumor. Here, we aim to associate both agents in a novel liposomal targeted therapy by linking CTX to the drug-loaded liposomes. These EGFR-targeted liposomes potentially combine the therapeutic activity and selectivity of CTX with tumor-cell delivery of L-OH in a single therapeutic approach. L-OH liposomes carrying whole CTX or CTX-Fab' fragments on their surface were designed and characterized. Their functionality was tested in vitro using four human CRC cell lines, expressing different levels of EGFR to investigate the role of CTX-EGFR interactions in the cellular binding and uptake of the nanocarriers and encapsulated drug. Next, those formulations were evaluated in vivo in a colorectal cancer xenograft model with regard to tumor drug accumulation, toxicity and therapeutic activity. In EGFR-overexpressing cell lines, intracellular drug delivery by targeted liposomes increased with receptor density reaching up to 3-fold higher levels than with non-targeted liposomes. Receptor specific uptake was demonstrated by competition with free CTX, which reduced internalization to levels similar to non-targeted liposomes. In a CRC xenograft model, drug delivery was strongly enhanced upon treatment with targeted formulations. Liposomes conjugated with monovalent CTX-Fab' fragments showed superior drug accumulation in tumor tissue (2916.0±507.84ng/g) compared to CTX liposomes (1546.02±362.41ng/g) or non-targeted liposomes (891.06±155.1ng/g). Concomitantly, CTX-Fab' targeted L-OH liposomes outperformed CTX-liposomes, which on its turn was still more efficacious than non-targeted liposomes and free drug treatment in CRC bearing mice. These results show that site-directed conjugation of monovalent CTX-Fab' provides targeted L-OH liposomes that display an increased tumor drug delivery and efficacy over a formulation with CTX and non-targeted liposomes.

Liposomes, a promising strategy for clinical application of platinum derivatives.

INTRODUCTION: Liposomes represent a versatile system for drug delivery in various pathologies. Platinum derivatives have been demonstrated to have therapeutic efficacy against several solid tumors. But their use is limited due to their side effects. Since liposomal formulations are known to reduce the toxicity of some conventional chemotherapeutic drugs, the encapsulation of platinum derivatives in these systems may be useful in reducing toxicity and maintaining an adequate therapeutic response. AREAS COVERED: This review describes the strategies applied to platinum derivatives in order to improve their therapeutic activity, while reducing the incidence of side effects. It also reviews the results found in the literature for the different platinum-drugs liposomal formulations and their current status. EXPERT OPINION: The design of liposomes to achieve effectiveness in antitumor treatment is a goal for platinum derivatives. Liposomes can change the pharmacokinetic parameters of these encapsulated drugs, reducing their side effects. However, few liposomal formulations have demonstrated a significant advantage in therapeutic terms. Lipoplatin, a cisplatin formulation in Phase III, combines a reduction in the toxicity associated with an antitumor activity similar to the free drug. Thermosensitive or targeted liposomes for tumor therapy are also included in this review. Few articles about this strategy applied to platinum drugs can be found in the literature.

Efficient serum-resistant lipopolyplexes targeted to the folate receptor.

In this work, we have developed and evaluated a new targeted lipopolyplex (LPP), by combining polyethylenimine (PEI), 1,2-dioleoyl-3-(trimethylammonium) propane (DOTAP)/Chol liposomes, the plasmids pCMVLuc/pCMVIL-12, and the ligand folic acid (FA), able to transfect HeLa and B16-F10 cells in the presence of very high concentration of serum (60% FBS). These complexes (Fol-LPP) have a net positive surface charge. The combination of folic acid with lipopolyplexes also enhanced significantly the transfection activity of the therapeutic gene interleukin-12 (IL-12), without any significant cytotoxicity. The specificity of the folate receptor (FR)-mediated gene transfer was corroborated by employing a folate receptor deficient cell line (HepG2). This formulation improved gene delivery showed by conventional lipoplexes or polyplexes resulting an efficient, simple, and nontoxic method for gene delivery of therapeutic genes in vitro and very probably in vivo.

Application of different methods to formulate PEG-liposomes of oxaliplatin: evaluation in vitro and in vivo.

In this work, the Film Method (FM), Reverse-Phase Evaporation (REV), and the Heating Method (HM) were applied to prepare PEG-coated liposomes of oxaliplatin with natural neutral and cationic lipids, respectively. The formulations developed with the three methods, showed similar physicochemical characteristics, except in the loading of oxaliplatin, which was statistically lower (P<0.05) using the HM. The incorporation of a semi-synthetic lipid in the formulation developed by FM, provided liposomes with a particle size of 115 nm associated with the lowest polydispersity index and the highest drug loading, 35%, compared with the other two lipids, suggesting an increase in the membrane stability. That stability was also evaluated according to the presence of cholesterol, the impact of the temperature, and the application of different cryoprotectants during the lyophilization. The results indicated long-term stability of the developed formulation, because after its intravenous in vivo administration to HT-29 tumor bearing mice was able to induce an inhibition of tumor growth statistically higher (P<0.05) than the inhibition caused by the free drug. In conclusion, the FM was the simplest method in comparison with REV and HM to develop in vivo stable and efficient PEG-coated liposomes of oxaliplatin with a loading higher than those reported for REV.