Tumor microenvironment reprogramming improves nanomedicine-based chemo-immunotherapy in sarcomas.

BACKGROUND/INTRODUCTION: Sarcomas are a heterogenous group of rare cancers that originate in soft tissues or bones. Their complexity and tendency for metastases makes treatment challenging, highlighting the need for new therapeutic approaches to improve patient survival. The difficulties in treating these cancers primarily stem from abnormalities within the tumor microenvironment (TME), which lead to reduced blood flow and oxygen levels in tumors. Consequently, this hampers the effective delivery of drugs to tumors and diminishes treatment efficacy despite higher, toxic doses of chemotherapy. Here, we tested the mechanotherapeutic ketotifen combined with either pegylated-liposomal doxorubicin (PLD) or pegylated-liposomal co-encapsulated alendronate-doxorubicin (PLAD) plus anti-PD-1 antibody in mouse models of fibrosarcoma and osteosarcoma. RESULTS: We found that ketotifen successfully reprogrammed the TME by reducing tumor stiffness and increasing perfusion, proven by changes measured by shear-wave-elastography (SWE) and contrast-enhanced-ultrasound (CEUS) respectively, and enhanced the therapeutic efficacy of our nanomedicine-based chemo-immunotherapy protocols. An additional observation was a trend to improved antitumor response when nano-chemotherapy is given alongside anti-PD1 and when the immunomodulator alendronate was present in the treatment. We next investigated the mechanisms of action of this combination. Ketotifen combined with nanomedicine-based chemo-immunotherapy, increased T-cell infiltration, specifically cytotoxic CD8+ T cells and CD4+ T helper-cell and decreased the number of regulatory-T-cells. In addition, the combination also altered the polarization of tumor associated macrophages, favouring the M1 immune-supportive phenotype over the M2 immuno-suppressive phenotype. CONCLUSION: Collectively, our findings provide evidence that ketotifen-induced TME reprograming can improve the efficacy of nanomedicine-based chemoimmunotherapy in sarcomas.

Mechanisms and Barriers in Nanomedicine: Progress in the Field and Future Directions.

In recent years, steady progress has been made in synthesizing and characterizing engineered nanoparticles, resulting in several approved drugs and multiple promising candidates in clinical trials. Regulatory agencies such as the Food and Drug Administration and the European Medicines Agency released important guidance documents facilitating nanoparticle-based drug product development, particularly in the context of liposomes and lipid-based carriers. Even with the progress achieved, it is clear that many barriers must still be overcome to accelerate translation into the clinic. At the recent conference workshop "Mechanisms and Barriers in Nanomedicine" in May 2023 in Colorado, U.S.A., leading experts discussed the formulation, physiological, immunological, regulatory, clinical, and educational barriers. This position paper invites open, unrestricted, nonproprietary discussion among senior faculty, young investigators, and students to trigger ideas and concepts to move the field forward.

The impact of germline BRCA pathogenic variants in locally advanced, triple negative breast cancer treated with platinum-based neoadjuvant chemotherapy.

BACKGROUND: Whether germline BRCA (gBRCA) pathogenic variants (PV) affect prognosis of women with triple negative breast cancer (TNBC) and whether it has implications for treatment decisions in the neoadjuvant setting is unclear. METHODS: This is a retrospective two-center cohort study comprising all women with early stage TNBC who have completed genetic testing and were treated with neoadjuvant dose-dense doxorubicin and cyclophosphamide followed by paclitaxel and carboplatin. All eligible patients treated between 10.2014 and 3.2020 were included. Data on clinico-pathological, pathological response, overall survival (OS) and disease-free survival (DFS) were evaluated. Differences in clinico-pathological features and outcomes were analyzed according to gBRCA status. RESULTS: Sixty-four women were included in the final analysis, of which 31 had gBRCA PV (gBRCA carriers) and 33 were gBRCA wild-type. Clinico-pathological characteristics were similar between both groups. The odds for pathological complete response (pCR) were significantly higher in gBRCA carriers (74.2%) compared to BRCA wild-type women (48.5%), p = 0.035. At a median follow-up of 30 months, gBRCA carriers had significantly favorable OS (HR = 8.64, 95% CI 1.08-69.21, p = 0.042). The difference in DFS did not reach statistical significance (HR = 7.4, 95% CI 0.91-60.27, p = 0.062). The favorable OS for gBRCA carriers remained significant in multivariate analysis (p = 0.029) and was noted regardless of pathological response (p = 0.018). CONCLUSION: Compared to wild-type, gBRCA carriers with locally advanced TNBC treated with neoadjuvant chemotherapy containing carboplatin had a higher pCR rate and better outcomes. These results strengthen the contention that gBRCA status should be considered when tailoring treatment decisions in women with locally advanced TNBC.

Harnessing Nanomedicine to Potentiate the Chemo-Immunotherapeutic Effects of Doxorubicin and Alendronate Co-Encapsulated in Pegylated Liposomes.

Encapsulation of Doxorubicin (Dox), a potent cytotoxic agent and immunogenic cell death inducer, in pegylated (Stealth) liposomes, is well known to have major pharmacologic advantages over treatment with free Dox. Reformulation of alendronate (Ald), a potent amino-bisphosphonate, by encapsulation in pegylated liposomes, results in significant immune modulatory effects through interaction with tumor-associated macrophages and activation of a subset of gamma-delta T lymphocytes. We present here recent findings of our research work with a formulation of Dox and Ald co-encapsulated in pegylated liposomes (PLAD) and discuss its pharmacological properties vis-à-vis free Dox and the current clinical formulation of pegylated liposomal Dox. PLAD is a robust formulation with high and reproducible remote loading of Dox and high stability in plasma. Results of biodistribution studies, imaging with radionuclide-labeled liposomes, and therapeutic studies as a single agent and in combination with immune checkpoint inhibitors or gamma-delta T lymphocytes suggest that PLAD is a unique product with distinct tumor microenvironmental interactions and distinct pharmacologic properties when compared with free Dox and the clinical formulation of pegylated liposomal Dox. These results underscore the potential added value of PLAD for chemo-immunotherapy of cancer and the relevance of the co-encapsulation approach in nanomedicine.

Pegylated Liposomal Alendronate Biodistribution, Immune Modulation, and Tumor Growth Inhibition in a Murine Melanoma Model.

While tumor-associated macrophages (TAM) have pro-tumoral activity, the ablation of macrophages in cancer may be undesirable since they also have anti-tumoral functions, including T cell priming and activation against tumor antigens. Alendronate is a potent amino-bisphosphonate that modulates the function of macrophages in vitro, with potential as an immunotherapy if its low systemic bioavailability can be addressed. We repurposed alendronate in a non-leaky and long-circulating liposomal carrier similar to that of the clinically approved pegylated liposomal doxorubicin to facilitate rapid clinical translation. Here, we tested liposomal alendronate (PLA) as an immunotherapeutic agent for cancer in comparison with a standard of care immunotherapy, a PD-1 immune checkpoint inhibitor. We showed that the PLA induced bone marrow-derived murine non-activated macrophages and M2-macrophages to polarize towards an M1-functionality, as evidenced by gene expression, cytokine secretion, and lipidomic profiles. Free alendronate had negligible effects, indicating that liposome encapsulation is necessary for the modulation of macrophage activity. In vivo, the PLA showed significant accumulation in tumor and tumor-draining lymph nodes, sites of tumor immunosuppression that are targets of immunotherapy. The PLA remodeled the tumor microenvironment towards a less immunosuppressive milieu, as indicated by a decrease in TAM and helper T cells, and inhibited the growth of established tumors in the B16-OVA melanoma model. The improved bioavailability and the beneficial effects of PLA on macrophages suggest its potential application as immunotherapy that could synergize with T-cell-targeted therapies and chemotherapies to induce immunogenic cell death. PLA warrants further clinical development, and these clinical trials should incorporate tumor and blood biomarkers or immunophenotyping studies to verify the anti-immunosuppressive effect of PLA in humans.

Pegylated Liposomal Mitomycin C Lipidic Prodrug in Combination With External Beam Radiation Therapy in Patients With Advanced Cancer: A Phase 1B Study.

PURPOSE: The aim of this study was to evaluate a formulation of pegylated liposomal mitomycin C lipidic prodrug (PL-MLP) in patients concomitantly undergoing external beam radiation therapy (RT). METHODS AND MATERIALS: Patients with metastatic disease or inoperable primary solid tumors requiring RT for disease control or symptom relief were treated with 2 courses of PL-MLP (1.25, 1.5, or 1.8 mg/kg) at 21-day intervals, along with 10 fractions of conventional RT or 5 stereotactic body RT fractions initiated 1 to 3 days after the first PL-MLP dose and completed within 2 weeks. Treatment safety was monitored for 6 weeks, and disease status was re-evaluated at 6-week intervals thereafter. MLP levels were analyzed 1 hour and 24 hours after each PL-MLP infusion. RESULTS: Overall, 19 patients with metastatic (18) or inoperable (1) disease received combination treatment, with 18 completing the full protocol. Most patients (16) had diagnoses of advanced gastrointestinal tract cancer. One grade 4 neutropenia event possibly related to study treatment was reported; other adverse events were mild or moderate. Of the 18 evaluable patients, 16 were free of RT target lesion progression at first re-evaluation. Median survival of the entire patient population was 63.3 weeks. Serum MLP level correlated with dose increases and similar long circulating profiles were observed before and after RT. CONCLUSIONS: PL-MLP up to 1.8 mg/kg in combination with RT treatment is safe, with a high rate of tumor control. Drug clearance is not affected by radiation. PL-MLP is potentially an attractive option for chemoradiation therapy that warrants further evaluation in randomized studies in the palliative and curative settings.

Comparative effects of free doxorubicin, liposome encapsulated doxorubicin and liposome co-encapsulated alendronate and doxorubicin (PLAD) on the tumor immunologic milieu in a mouse fibrosarcoma model.

Background: We have previously shown that alendronate, an amino-bisphosphonate, when reformulated in liposomes, can significantly enhance the efficacy of cytotoxic chemotherapies and help remodel the immunosuppressive tumor microenvironment towards an immune-permissive milieu resulting in increased anticancer efficacy. In addition, we have previously shown that the strong metal-chelating properties of alendronate can be exploited for nuclear imaging of liposomal biodistribution. To further improve anticancer efficacy, a pegylated liposome formulation co-encapsulating alendronate and doxorubicin (PLAD) has been developed. In this study, we examined the effects of PLAD on the tumor immunologic milieu in a mouse fibrosarcoma model in which the tumor microenvironment is heavily infiltrated with tumor-associated macrophages (TAM) that are associated with poor prognosis and treatment resistance. Methods: Doxorubicin biodistribution, characterization of the tumor immunologic milieu, cellular doxorubicin uptake, and tumor growth studies were performed in Balb/c mice bearing subcutaneously implanted WEHI-164 fibrosarcoma cells treated intravenously with PLAD, pegylated liposomal doxorubicin (PLD), free doxorubicin, or vehicle. Results: PLAD delivery resulted in a high level of tumor doxorubicin that was 20 to 30-fold greater than in free doxorubicin treated mice, and non-significantly higher than in PLD treated mice. PLAD also resulted in increased uptake in spleen and slightly lower plasma levels as compared to PLD. Importantly, our results showed that PLAD, and to a lesser extent PLD, shifted cellular drug uptake to TAM and to monocytic myeloid-derived suppressor cells (MDSC), while there was no drug uptake in neutrophilic MDSC or lymphoid cells. Free doxorubicin cellular drug uptake was below detectable levels. PLAD, and to a lesser extent PLD, also induced significant changes in number and functionality of tumor-infiltrating TAM, MDSC, Treg, NKT, and NK cells that are consistent with enhanced antitumor immune responses in the tumor microenvironment. In contrast, free doxorubicin induced moderate changes in the tumor microenvironment that could promote (decreased Treg) or be detrimental to antitumor immune responses (decreased M1 TAM and NK cells). These immune modulatory effects are reflected in the therapeutic study which showed that PLAD and PLD inhibited tumor growth and significantly prolonged survival, while free doxorubicin showed little or no anticancer activity. Conclusion: We show that liposomal delivery of doxorubicin not only alters pharmacokinetics, but also dramatically changes the immune modulatory activity of the drug cargo. In addition, our data support that the PLAD nanotheranostic platform further enhances some immune changes that may act in synergy with its cytotoxic chemotherapy effects.

Characterization of a Mesoporous Silica Nanoparticle Formulation Loaded with Mitomycin C Lipidic Prodrug (MLP) and In Vitro Comparison with a Clinical-Stage Liposomal Formulation of MLP.

Nanomedicines have revolutionized the treatment of certain types of cancer, as is the case of doxil, liposomal formulation with doxorubicin encapsulated, in the treatment of certain types of ovarian cancer, AIDS-related Kaposi sarcoma, and multiple myeloma. These nanomedicines can improve the performance of conventional chemotherapeutic treatments, with fewer side effects and better efficiency against cancer. Although liposomes have been used in some formulations, different nanocarriers with better features in terms of stability and adsorption capabilities are being explored. Among the available nanoparticles in the field, mesoporous silica nanoparticles (MSNP) have attracted great attention as drug delivery platforms for the treatment of different diseases. Here, a novel formulation based on MSNP loaded with a potent antitumor prodrug that works in vitro as well as in a clinically evaluated liposomal formulation has been developed. This novel formulation shows excellent prodrug encapsulation efficiency and effective release of the anticancer drug only under certain stimuli typical of tumor environments. This behavior is of capital importance for translating this nanocarrier to the clinic in the near future.

Ex-vivo activation of a liposomal prodrug of mitomycin C by human tumors.

PURPOSE: To examine the ex- vivo ability of explanted human tumors and normal tissue to activate liposomal mitomycin C lipidic prodrug (MLP) by releasing the active free drug form, mitomycin C (MMC). METHODS: We tested conversion of MLP to MMC in an ex vivo assay using explanted tissues obtained during routine surgery to remove primary tumors or metastases. Tumor and adjacent normal tissue were obtained from freshly explanted tumors and were immediately deep frozen at - 70 °C. On test day, the fragments were thawed, homogenized and incubated in the presence of a fixed amount of liposomal MLP at 37 °C for 1 h. We measured MLP and its rate of conversion to MMC by HPLC. Controls included plasma, malignant effusions, red blood cells, tumor cell lines, mouse liver, and buffer with dithiothreitol, a potent reducing agent. RESULTS: Most patients tested (16/20) were diagnosed with colo-rectal carcinoma. The average fraction of MLP cleaved per 100-mg tumor tissue (21.1%, SEM = 1.8) was greater than per 100-mg normal tissue (16.6%, SEM = 1.3). When the tumor and normal tissue samples were paired by patient, the difference was statistically significant (p = 0.022, paired t test). Biological fluids did not activate liposomal MLP, while normal liver tissue strongly does. Interestingly, the omental fatty tissue also greatly activated MLP. CONCLUSIONS: Tumor tissue homogenates activate MLP with greater efficiency than the surrounding normal tissues, but far less than liver and adipose tissue. These observations demonstrate the bioavailability of liposomal MLP in human tumors, and its pharmacologic potential in cancer therapy.

Liposome co-encapsulation of anti-cancer agents for pharmacological optimization of nanomedicine-based combination chemotherapy.

Aim: Co-encapsulation of anti-cancer agents in pegylated liposomes may provide an effective tool to maximize efficacy of combined drug therapy by taking advantage of the long circulation time, passive targeting, and reduced toxicity of liposome formulations. Methods: We have developed several liposome formulations of co-encapsulated drugs using various permutations of three active agents: doxorubicin (Dox), mitomycin-C lipidic prodrug (MLP), and alendronate (Ald). Dox and MLP are available in single drug liposomal formulations: pegylated liposomal Dox (PLD, Doxil®), clinically approved, and pegylated liposomal MLP (PL-MLP, Promitil®), in phase 1-2 clinical testing. We have previously shown that co-encapsulation of Dox and Ald in pegylated liposomes (PLAD) results in a formulation with valuable immuno-pharmacologic properties and superior antitumor properties over PLD in immunocompetent animal models. Building on the PLAD and PL-MLP platforms, we developed a new pegylated liposomal formulation of co-entrapped Dox and MLP (PLAD-MLP), with the former localized in the liposome water phase via remote loading with an ammonium alendronate and the latter passively loaded into the liposome lipid bilayer. An alternative formulation of co-entrapped MLP and Dox in which ammonium Ald was replaced with ammonium sulfate (PLD-MLP) was also tested for comparative purposes. Results: PLAD-MLP displays high loading efficiency of Dox and MLP nearing 100%, and a mean vesicle diameter of 110 nm. Cryo-transmission electron microscopy (cryo-TEM) of PLAD-MLP reveals round vesicles with an intra-vesicle Dox-alendronate precipitate. PLAD-MLP was tested in an in vitro MLP activation assay with the reducing agent dithiothreitol and found to be significantly less susceptible to thiolytic activation than PL-MLP. Alongside thiolytic activation of MLP, a significant fraction of encapsulated Dox was released from liposomes. PLAD-MLP is stable upon in vitro incubation in human plasma with nearly 100% drug retention. In mouse pharmacokinetic studies, PLAD-MLP extended MLP half-life in circulation when compared to that of MLP delivered as PL-MLP. In addition, the MLP levels in tissues were greater than those obtained with PL-MLP, indicating that PLAD-MLP slows down the cleavage of the prodrug MLP to MMC, thus resulting in a more sustained and prolonged exposure. The circulation half-life of Dox in PLAD-MLP was similar to the PLD Dox half-life. The pattern of tissue distribution was similar for the co-encapsulated drugs, although Dox levels were generally higher than those of MLP, as expected from cleavage of MLP to its active metabolite MMC. In mouse tumor models, the therapeutic activity of PLAD-MLP was superior to PL-MLP and PLD with a convenient safety dose window. The Ald-free formulation, PLD-MLP, displayed similar pharmacokinetic properties to PLAD-MLP, but its therapeutic activity was lower. Conclusion: PLAD-MLP is a novel multi-drug liposome formulation with attractive pharmacological properties and powerful antitumor activity and is a promising therapeutic tool for combination cancer chemotherapy.

Repurposing amino-bisphosphonates by liposome formulation for a new role in cancer treatment.

Amino-bisphosphonates (N-BPs) have been commercially available for over four decades and are used for the treatment of osteoporosis, Paget's disease, hypercalcemia of malignancy, and bone metastases derived from various cancer types. Zoledronate and alendronate, two of the most potent N-BPs, have demonstrated direct tumoricidal activity on tumor cells and immune modulatory effects on myeloid cells and T cells in vitro and in animal models of cancer. However, the rapid renal clearance and sequestration in mineral bone of these drugs in free form severely limit their systemic exposure and applications in cancer patients. Reformulation of N-BPs by encapsulation in liposomal nanoparticles addresses these pharmacokinetic barriers, and liposomal zoledronate and alendronate formulations have been found to increase the anticancer efficacy of cytotoxic chemotherapies and adoptive T cell immunotherapies in murine cancer models. Herein, we review the differences in pharmacology between N-BPs versus non-N-BPs (e.g., clodronate), free versus liposomal N-BP formulations, and targeted versus non-targeted liposomal N-BPs, and the clinical and preclinical evidence supporting a role for liposomal N-BPs in the treatment of cancer. We propose that pegylated liposomal alendronate (PLA) has the most potential for clinical translation based on favorable therapeutic index, ability to passively target and accumulate in tumors, proven biocompatibility of the liposome carrier, and preclinical anticancer efficacy.

Development of Promitil®, a lipidic prodrug of mitomycin c in PEGylated liposomes: From bench to bedside.

Several liposome products have been approved for the treatment of cancer. In all of them, the active agents are encapsulated in the liposome water phase passively or by transmembrane ion gradients. An alternative approach in liposomal drug delivery consists of chemically modifying drugs to form lipophilic prodrugs with strong association to the liposomal bilayer. Based on this approach, we synthesized a mitomycin c-derived lipidic prodrug (MLP) which is entrapped in the bilayer of PEGylated liposomes (PL-MLP, Promitil®), and activated by thiolytic cleavage. PL-MLP is stable in plasma with thiolytic activation of MLP occurring exclusively in tissues and is more effective and less toxic than conventional chemotherapy in various tumor models. PL-MLP has completed phase I clinical development where it has shown a favorable safety profile and a 3-fold reduction in toxicity as compared to free mitomycin c. Clinical and pharmacokinetic studies in patients with advanced colo-rectal carcinoma have indicated a significant rate of disease stabilization (39%) in this chemo-refractory population and significant prolongation of median survival in patients attaining stable disease (13.9 months) versus progressive disease patients (6.35 months). The pharmacokinetics of MLP was typically stealth with long T½ (~1 day), slow clearance and small volume of distribution. Interestingly, a longer T½, and slower clearance were both correlated with disease stabilization and longer survival. This association of pharmacokinetic parameters with patient outcome suggests that arrest of tumor growth is related to the enhanced tumor localization of long-circulating liposomes and highlights the importance of personalized pharmacokinetic evaluation in the clinical use of nanomedicines. Another important area where PL-MLP may have an added value is in chemoradiotherapy, where it has shown a strong radiosensitizing effect in animal models based on a unique mechanism of enhanced prodrug activation and encouraging results in early human testing.

Complement Activation: A Potential Threat on the Safety of Poly(ethylene glycol)-Coated Nanomedicines.

In this issue of ACS Nano, Chen et al. provide in vitro and in vivo evidence for monoclonal anti-poly(ethylene glycol) (anti-PEG) antibody-triggered, complement terminal complex-mediated damage to PEGylated liposomal doxorubicin, entailing the release of the encapsulated drug from the vesicles. These results reveal a new dimension of the potential damage of anti-PEG antibody-mediated complement activation on PEGylated nanomedicines in addition to previous observations on infusion hypersensitivity reactions and the accelerated blood clearance effect. The possibility of a destructive attack of the complement system on the liposome drug carrier may have safety implications in patients displaying high levels of preformed anti-PEG antibodies. In this Perspective, we summarize the experimental and clinical data highlighting the relationships among the above adverse immune phenomena and the options available for reducing the risk of immune damage caused by PEGylated nanomedicines.

Translational considerations in nanomedicine: The oncology perspective.

Nanoparticles can provide effective control of the release rate and tissue distribution of their drug payload, leading to major pharmacokinetic and pharmacodynamic changes vis-à-vis the conventional administration of free drugs. In the last two decades, we have witnessed major progress in the synthesis and characterization of engineered nanoparticles for imaging and treatment of cancers, resulting in the approval for clinical use of several products and in new and promising approaches. Despite these advances, clinical applications of nanoparticle-based therapeutic and imaging agents remain limited due to biological, immunological, and translational barriers. There is a need to make high impact advances toward translation. In this review, we address biological, toxicological, immunological, and translational aspects of nanomedicine and discuss approaches to move the field forward productively. Overcoming these barriers may dramatically improve the development potential and role of nanomedicines in the oncology field and help meet the high expectations.

Pharmacokinetics of mitomycin-c lipidic prodrug entrapped in liposomes and clinical correlations in metastatic colorectal cancer patients.

Background Pegylated liposomal (PL) mitomycin-c lipidic prodrug MLP) may be a useful agent in patients with metastatic colo-rectal carcinoma (CRC). We report here on the pharmacokinetics and clinical observations in a phase 1A/B study with PL-MLP. Methods Plasma levels of MLP were examined in 53 CRC patients, who received PL-MLP either as single agent or in combination with capecitabine and/or bevacizumab. MLP was determined by an HPLC-UV assay, and its pharmacokinetics was analyzed by noncompartmental methods. The correlation between clinical and pharmacokinetic parameters was statistically analyzed. Results PL-MLP was well tolerated with a good safety profile as previously reported. Stable Disease was reported in 15/36 (42%) of efficacy-evaluable patients. Median survival of stable disease patients (14.4 months) was significantly longer than of progressive disease patients (6.5 months) and non-evaluable patients (2.3 months). MLP pharmacokinetics was stealth-like with long T½ (~1 day), slow clearance, and small volume of distribution (Vd). The addition of capecitabine and/or bevacizumab did not have any apparent effect on the pharmacokinetics of MLP and clinical outcome. High baseline neutrophil count and CEA level were correlated with faster clearance, and larger Vd. Stable disease patients had longer T½ and slower clearance than other patients. T½ and clearance were significantly correlated with survival. Conclusions PL-MLP treatment results in a substantial rate of disease stabilization in metastatic CRC, and prolonged survival in patients achieving stable disease. The correlation of neutrophil count and CEA level with pharmacokinetic parameters of MLP is an unexpected finding that needs further investigation. The association of long T½ of MLP with stable disease and longer survival is consistent with an improved probability of disease control resulting from enhanced tumor localization of long-circulating liposomes and underscores the relevance of personalized pharmacokinetic evaluation in the use of nanomedicines.

2-APB and CBD-Mediated Targeting of Charged Cytotoxic Compounds Into Tumor Cells Suggests the Involvement of TRPV2 Channels.

Targeted delivery of therapeutic compounds to particular cell types such that they only affect the target cells is of great clinical importance since it can minimize undesired side effects. For example, typical chemotherapeutic treatments used in the treatment of neoplastic disorders are cytotoxic not only to cancer cells but also to most normal cells when exposed to a critical concentration of the compound. As such, many chemotherapeutics exhibit severe side effects, often prohibiting their effective use in the treatment of cancer. Here, we describe a new means for facilitated delivery of a clinically used chemotherapy compound' doxorubicin, into hepatocellular carcinoma cell line (BNL1 ME). We demonstrate that these cells express a large pore, cation non-selective transient receptor potential (TRP) channel V2. We utilized this channel to shuttle doxorubicin into BNL1 ME cells. We show that co-application of either cannabidiol (CBD) or 2-APB, the activators of TRPV2 channels, together with doxorubicin leads to significantly higher accumulation of doxorubicin in BNL1 ME cells than in BNL1 ME cells that were exposed to doxorubicin alone. Moreover, we demonstrate that sub-effective doses of doxorubicin when co-applied with either 2-APB or CBD lead to a significant decrease in the number of living BNL1 ME cell and BNL1 ME cell colonies in comparison to application of doxorubicin alone. Finally, we demonstrate that the doxorubicin-mediated cell death is significantly more potent, requiring an order of magnitude lower dose, when co-applied with CBD than with 2-APB. We suggest that CBD may have a dual effect in promoting doxorubicin-mediated cell death by facilitating the entry of doxorubicin via TRPV2 channels and preventing its clearance from the cells by inhibiting P-glycoprotein ATPase transporter. Collectively, these results provide a foundation for the use of large pore cation-non selective channels as "natural" drug delivery systems for targeting specific cell types.

Integrating Nanotechnology into Cancer Care.

Research activity in medical and cancer nanotechnology has grown dramatically over the past 15 years. The field has become a cradle of multidisciplinary investigations bringing together physicists, chemists, and engineers working with clinicians and biologists to address paramount problems in cancer care and treatment. Some have argued that the explosion in the number of research papers has not been followed by sufficient clinical activity in nanomedicine. However, three new nanodrugs have now been approved by the U.S. Food and Drug Administration (FDA) in the past three years, confirming the validity of nanotechnology approaches in cancer. Excitingly, translational pipelines contain several additional intriguing candidates. In this Nano Focus article, we discuss potential barriers inhibiting further incorporation of nanomedicines into patient care, possible strategies to overcome these barriers, and promising new directions in cancer interventions based on nanotechnology. Insights presented herein are outcomes of discussions held at a recent strategic workshop hosted by the National Cancer Institute (NCI), which brought together research, clinical, and commercial leaders of the nanomedicine field.

In Vivo PET Tracking of 89Zr-Labeled Vγ9Vδ2 T Cells to Mouse Xenograft Breast Tumors Activated with Liposomal Alendronate.

Gammadelta T (γδ-T) cells are strong candidates for adoptive immunotherapy in oncology due to their cytotoxicity, ease of expansion, and favorable safety profile. The development of γδ-T cell therapies would benefit from non-invasive cell-tracking methods and increased targeting to tumor sites. Here we report the use of [89Zr]Zr(oxinate)4 to track Vγ9Vδ2 T cells in vivo by positron emission tomography (PET). In vitro, we showed that 89Zr-labeled Vγ9Vδ2 T cells retained their viability, proliferative capacity, and anti-cancer cytotoxicity with minimal DNA damage for amounts of 89Zr ≤20 mBq/cell. Using a mouse xenograft model of human breast cancer, 89Zr-labeled γδ-T cells were tracked by PET imaging over 1 week. To increase tumor antigen expression, the mice were pre-treated with PEGylated liposomal alendronate. Liposomal alendronate, but not placebo liposomes or non-liposomal alendronate, significantly increased the 89Zr signal in the tumors, suggesting increased homing of γδ-T cells to the tumors. γδ-T cell trafficking to tumors occurred within 48 hr of administration. The presence of γδ-T cells in tumors, liver, and spleen was confirmed by histology. Our results demonstrate the suitability of [89Zr]Zr(oxinate)4 as a cell-labeling agent for therapeutic T cells and the potential benefits of liposomal bisphosphonate treatment before γδ-T cell administration.

Chemo-Radiotherapy of Oligometastases of Colorectal Cancer With Pegylated Liposomal Mitomycin-C Prodrug (Promitil): Mechanistic Basis and Preliminary Clinical Experience.

Hypo-fractionated radiotherapy and stereotactic body radiotherapy are viable options for treatment of oligometastases. A prodrug of mitomycin-C is under clinical testing as a pegylated liposomal formulation (Promitil) with an improved safety profile over mitomycin-C. Promitil was offered to two patients with oligometastases from colorectal cancer as radiosensitizer. Each derived durable clinical benefit from Promitil administered immediately prior to and following irradiation. Transient toxicity to normal tissues of moderate to severe degree was observed. Promitil appears to have potential clinical value in this setting. HIGHLIGHTS - Delivery of radio-sensitizing drugs with pegylated (long-circulating) liposomes is a pharmacologically rational approach which remains largely clinically untested.- A mitomycin-c prodrug delivered by pegylated liposomes (Promitil) is activated by thiol groups, which are produced in excess by radiation-damaged cells, thus potentiating the radio-sensitizing effect of Promitil.- Two durable clinical responses in patient with colorectal oligometastases to Promitil and radiotherapy suggest that this approach may be of value in cancer chemo-radiotherapy.

Folate receptor targeting of radiolabeled liposomes reduces intratumoral liposome accumulation in human KB carcinoma xenografts.

BACKGROUND: Active, ligand-mediated, targeting of functionalized liposomes to folate receptors (FRs) overexpressed on cancer cells could potentially improve drug delivery and specificity. Studies on folate-targeting liposomes (FTLs) have, however, yielded varying results and generally fail to display a clear benefit of FR targeting. METHOD: Tumor accumulating potential of FTLs and NTLs were investigated in a FR overex-pressing xenograft model by positron emission tomography/computed tomography imaging. RESULTS: Tumors displayed significantly lower activity of FTLs than NTLs. Furthermore, FTLs displayed worse circulating properties and increased liver-accumulation than NTLs. CONCLUSION: This study underlines that long-circulating properties of liposomes must be achieved to take advantage of EPR-dependent tumor accumulation which may be lost by functionalization. FR-functionalization negatively affected both tumor accumulation and circulation properties.