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.

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.

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.

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.

Targeting of folate-conjugated liposomes with co-entrapped drugs to prostate cancer cells via prostate-specific membrane antigen (PSMA).

Folate-targeted liposomes (FTL) were tested as drug delivery vehicles to PSMA-positive cancer cells. We used FL with co-entrapped mitomycin C lipophilic prodrug (MLP) and doxorubicin (DOX), and the LNCaP prostate cancer cell line which expresses PSMA but is negative for folate receptor. A major increase in cell drug levels was observed when LNCaP cells were incubated with FTL as compared to non-targeted liposomes (NTL). MLP was activated to mitomycin C, and intracellular and nuclear fluorescence of DOX was detected, indicating FTL processing and drug bioavailability. PMPA (2-(phosphonomethyl)-pentanedioic acid), a specific inhibitor of PSMA, blocked the uptake of FTL into LNCaP cells, but did not affect the uptake of FTL into PSMA-deficient and folate receptor-positive KB cells. The cytotoxic activity of drug-loaded FTL was found significantly enhanced when compared to NTL in LNCaP cells. FTL may provide a new tool for targeted therapy of cancers that over-express the PSMA receptor.

Characterization of Pegylated Liposomal Mitomycin C Lipid-Based Prodrug (Promitil) by High Sensitivity Differential Scanning Calorimetry and Cryogenic Transmission Electron Microscopy.

The effect of a lipidated prodrug of mitomycin C (MLP) on the membrane of a pegylated liposome formulation (PL-MLP), also known as Promitil, was characterized through high-sensitivity differential scanning calorimetry (DSC) and cryo-TEM. The thermodynamic analysis demonstrated that MLP led to the formation of heterogeneous domains in the membrane plane of PL-MLP. MLP concentrated in prodrug-rich domains, arranged in high-ordered crystal-like structures, as suggested by the sharp and high enthalpy endotherm in the first heating scanning. After thiolytic cleavage of mitomycin C from MLP by dithiothreitol (DTT) treatment, the crystal-like prodrug domain disappears and a homogeneous membrane with stronger lipid interactions and higher phase transition temperature compared with the blank (MLP-free) liposomes is observed by DSC. In parallel, the rod-like discoid liposomes and the "kissing liposomes" seen by cryo-TEM in the PL-MLP formulation disappear, and liposome mean size and polydispersity increase after DTT treatment. Both MLP and the residual postcleavage lipophilic moiety of the prodrug increased the rigidity of the liposome membrane as indicated by DSC. These results confirm that MLP is inserted in the PL-MLP liposome membrane via its lipophilic anchor, and its mitomycin C moiety located mainly at the region of the phospholipid glycerol backbone and polar headgroup. We hypothesize that π-π stacking between the planar aromatic rings of the mitomycin C moieties leads to the formation of prodrug-rich domains with highly ordered structure on the PL-MLP liposome membrane. This thermodynamically stable conformation may explain the high stability of the PL-MLP formulation. These results also provide us with an interesting example of the application of high sensitivity DSC in understanding the composition-structure-behavior dynamics of liposomal nanocarriers having a lipid-based drug as pharmaceutical ingredient.

Dexrazoxane added to doxorubicin-based adjuvant chemotherapy of breast cancer: a retrospective cohort study with a comparative analysis of toxicity and survival.

Dexrazoxane is indicated as a cardioprotective agent for patients receiving doxorubicin who are at increased risk for cardiotoxicity. Concerns have been raised on the use of dexrazoxane, particularly in adjuvant therapy, because of the risk of interference with the antitumor effect of doxorubicin. Two meta-analyses in metastatic breast cancer have rejected this hypothesis, but have shown an apparent increase in the severity of myelosuppression when dexrazoxane is used. Here, we analyzed retrospectively a cohort of our institute database to assess whether the addition of dexrazoxane causes more bone marrow suppression in breast cancer patients receiving doxorubicin-based adjuvant therapy. The secondary objectives were assessment of the incidence of febrile neutropenia, dose-schedule modifications, recorded cardiac events or cardiac test abnormalities, and overall survival. Eight hundred and twenty-two female patients who received adjuvant (or neoadjuvant) doxorubicin and cyclophosphamide for breast cancer between 2001 and 2013 were included. One hundred and four of these patients also received dexrazoxane concurrently with the adjuvant treatment. Hospital records and, when accessible, community clinic records were reviewed. The median follow-up duration was 7 years for patients receiving dexrazoxane and 7.5 years for patients not receiving dexrazoxane. 85.6% of patients were alive at data lock. Compared with the nondexrazoxane group, patients who received dexrazoxane were older (median age at diagnosis 59 vs. 52 years) and more likely to receive dose-dense AC therapy (73 vs. 59%) and adjuvant trastuzumab treatment (29 vs. 15%). Compared with the nondexrazoxane group, dexrazoxane treatment was associated with a higher rate of hematological side effects: leukopenia (48 vs. 39%), neutropenia (45 vs. 31%, P=0.003), anemia (86 vs. 73%, P=0.005), and thrombocytopenia (37 vs. 22%, P=0.001). There were more febrile neutropenia hospitalizations (20 vs. 10%, P=0.001) and dose reductions (22 vs. 8%, P<0.001) in the dexrazoxane group, but no significant difference in the incidence of treatment delays or cancellations. The incidence of cardiac events was the same in both treatment groups with and without dexrazoxane. There was a nonsignificantly lower mortality rate in the dexrazoxane group (9.6%) compared with the nondexrazoxane group (15.0%) at data lock. Adding dexrazoxane to doxorubicin in adjuvant therapy patients leads to higher rates of bone marrow suppression in all blood components, as well as more febrile neutropenia events, and dose reductions. No differences in events defined as cardiac toxicities were detected. Dexrazoxane had no detrimental effect on survival, despite the higher hematological toxicity, the older median age, and the higher prevalence of HER2-positive disease in the dexrazoxane group.

New insights and evolving role of pegylated liposomal doxorubicin in cancer therapy.

We herein review various pharmacological and clinical aspects of pegylated liposomal doxorubicin (PLD), the first nanomedicine to be approved for cancer therapy, and discuss the gap between its potent antitumor activity in preclinical studies and its comparatively modest achievements in clinical studies and limited use in clinical practice. PLD is a complex formulation of doxorubicin based on pharmaceutical nanotechnology with unique pharmacokinetic and pharmacodynamic properties. Its long circulation time with stable retention of the payload and its accumulation in tumors with high vascular permeability both result in important advantages over conventional chemotherapy. The ability of PLD to buffer a number of undesirable side effects of doxorubicin, including a major risk reduction in cardiac toxicity, is now well-established and confers a major added value in a number of disease conditions. PLD is approved for the treatment of ovarian cancer, breast cancer, multiple myeloma, and Kaposi sarcoma. In addition, clinically significant antitumor activity of PLD has been reported in a number of other cancer types, including lymphomas and soft tissue sarcomas. In spite of this, PLD has not replaced conventional doxorubicin in common applications such as the adjuvant and neoadjuvant treatment of breast cancer, and its use in the clinic has not become as widespread as one may have predicted. Exploiting the unique pharmacology of PLD, analyzing its selective biodistribution and homing to tumors in cancer patients with proper theranostic tools, and harnessing its complex interaction with the immune system, will lead to a more selective and rational use of PLD that may have great impact on future clinical results and may help realize its largely untapped potential.

MEK inhibitor treatment is effective in a patient with metastatic carcinoma of the ampulla of Vater with BRAF and NRAS mutations shown by next-generation sequencing.

Here, we present a case of an 84-year-old woman who developed obstructive jaundice and was diagnosed with nonoperable adenocarcinoma originating from the ampulla of Vater, a lethal disease with a median overall survival of less than a year. Her tumor was examined by next-generation sequencing, which showed BRAF and NRAS mutations. To target these mutations, a MEK inhibitor was chosen for treatment. The patient has been treated with a MEK inhibitor for the last 12 months since diagnosis, with clinical and laboratory improvement and manageable side effects. PET-computed tomography imaging has shown stable disease or improvement in the primary and metastatic lesions. This is the first case report of an ampulla of a Vater cancer patient with NRAS and BRAF mutations, identified in next-generation sequencing, and treated successfully with a MEK inhibitor.

Pharmacologic Studies of a Prodrug of Mitomycin C in Pegylated Liposomes (Promitil(®)): High Stability in Plasma and Rapid Thiolytic Prodrug Activation in Tissues.

PURPOSE: Pegylated liposomal (PL) mitomycin C lipid-based prodrug (MLP) has recently entered clinical testing. We studied here the preclinical pharmacology of PL-MLP. METHODS: The stability, pharmacokinetics, biodistribution, and other pharmacologic parameters of PL-MLP were examined. Thiolytic cleavage of MLP and release of active mitomycin C (MMC) were studied using dithiothreitol (DTT), and by incubation with tissue homogenates. RESULTS: MLP was incorporated in the bilayer at 10% molar ratio with nearly 100% entrapment efficiency, resulting in a formulation with high plasma stability. In vitro, DTT induced cleavage of MLP with predictable kinetics, generating MMC and enhancing pharmacological activity. A long circulation half-life of MLP (10-15 h) was observed in rodents and minipigs. Free MMC is either extremely low or undetectable in plasma. However, urine from PL-MLP injected rats revealed delayed but significant excretion of MMC indicating in vivo activation of MLP. Studies in mice injected with H3-cholesterol radiolabeled PL-MLP demonstrated relatively greater tissue levels of H3-cholesterol than MLP. MLP levels were highest in tumor and spleen, and very low or undetectable in liver and lung. Rapid cleavage of MLP in various tissues, particularly in liver, was shown in ex-vivo experiments of PL-MLP with tissue homogenates. PL-MLP was less toxic in vivo than equivalent doses of MMC. Therapeutic studies in C26 mouse tumor models demonstrated dose-dependent improved efficacy of PL-MLP over MMC. CONCLUSIONS: Thiolytic activation of PL-MLP occurs in tissues but not in plasma. Liposomal delivery of MLP confers a favorable pharmacological profile and greater therapeutic index than MMC.

Adoptive immunotherapy of epithelial ovarian cancer with Vγ9Vδ2 T cells, potentiated by liposomal alendronic acid.

Adoptive immunotherapy using γδ T cells harnesses their natural role in tumor immunosurveillance. The efficacy of this approach is enhanced by aminobisphosphonates such as zoledronic acid and alendronic acid, both of which promote the accumulation of stimulatory phosphoantigens in target cells. However, the inefficient and nonselective uptake of these agents by tumor cells compromises the effective clinical exploitation of this principle. To overcome this, we have encapsulated aminobisphosphonates within liposomes. Expanded Vγ9Vδ2 T cells from patients and healthy donors displayed similar phenotype and destroyed autologous and immortalized ovarian tumor cells, following earlier pulsing with either free or liposome-encapsulated aminobisphosphonates. However, liposomal zoledronic acid proved highly toxic to SCID Beige mice. By contrast, the maximum tolerated dose of liposomal alendronic acid was 150-fold higher, rendering it much more suited to in vivo use. When injected into the peritoneal cavity, free and liposomal alendronic acid were both highly effective as sensitizing agents, enabling infused γδ T cells to promote the regression of established ovarian tumors by over one order of magnitude. Importantly however, liposomal alendronic acid proved markedly superior compared with free drug following i.v. delivery, exploiting the "enhanced permeability and retention effect" to render advanced tumors susceptible to γδ T cell-mediated shrinkage. Although folate targeting of liposomes enhanced the sensitization of folate receptor-α(+) ovarian tumor cells in vitro, this did not confer further therapeutic advantage in vivo. These findings support the development of an immunotherapeutic approach for ovarian and other tumors in which adoptively infused γδ T cells are targeted using liposomal alendronic acid.

Emerging delivery systems to reduce doxorubicin cardiotoxicity and improve therapeutic index: focus on liposomes.

Anthracyclines are powerful anticancer agents and among the most important tools in the chemotherapy armamentarium of medical oncologists. They are approved for use in the treatment of a broad variety of solid and hematologic neoplasms. However, the usefulness of these agents, particularly doxorubicin, the most widely used anthracycline, is limited by considerable toxicity, especially damage to the cardiac muscle, which is cumulative and mostly irreversible, restricting extended use of this drug. In the last 30 years, extensive research with a variety of drug-delivery systems has attempted to overcome this limitation, with clinical success mostly confined to liposome formulations. Liposomal doxorubicin, and particularly pegylated liposomal doxorubicin, has shown significant pharmacologic advantages and an added clinical value over doxorubicin. Here, we review the mechanisms of action and toxicity of doxorubicin, and ways to reduce toxicity, with a focus on liposome-based drug-delivery systems.

Pharmacokinetics of low molecular weight heparin in patients with malignant tumors.

Cancer patients have an increased risk for venous thromboembolism (VTE). Low molecular weight heparin (LMWH) is the mainstay of VTE treatment in these patients. Heparanase, which degrades heparin and LMWH, is an enzyme secreted from a variety of malignant tumors. The objective of this study was to elucidate the pharmacokinetics of LMWH in patients with locally advanced or metastatic cancer. A total of 10 cancer patients with VTE treated with the LMWH enoxaparin at a standard dose of 1 mg/kg every 12 h were enrolled. Blood samples were obtained before the injection of LMWH and at 1, 2, 3, 4, 6, and 8 h after LMWH administration, and they were tested for anti-factor Xa activity and heparanase levels. Peak anti-Xa activity was achieved 2-8 h after subcutaneous administration of LMWH. Six patients did not reach a therapeutic anti-Xa activity target (0.6-1.2 IU/ml) at 4 h after LMWH administration. Four patients did not reach anti-factor Xa values of 0.6 IU/ml throughout the trial. The median anti-Xa activity before LMWH injection was 0.24 IU/ml (range 0.07-0.7 IU/ml), as opposed to 0.52 IU/ml in historical controls. The median anti-Xa activity 4 h after LMWH injection was 0.58 IU/ml (range 0.22-1.23 IU/ml), as opposed to 1.2 IU/ml in historical controls. The blood level of heparanase in patients with malignancy and VTE was 6.24 ± 4.3 ng/ml, compared with 2.67 ± 1.09 ng/ml in cancer-free, age-matched, normal controls. In this pilot study, a substantial proportion of cancer patients suffering from VTE and treated with LMWH had subtherapeutic anti-Xa activity.

Liposome promotion of tumor growth is associated with angiogenesis and inhibition of antitumor immune responses.

Liposomes have tremendous potential as drug carriers in the treatment of cancer. However, despite enhanced tumor drug delivery and decreased toxicity, patient survival rates have not improved significantly compared to corresponding free drug treatments. Importantly, we found that a liposomal nanoparticle currently used as a drug carrier in cancer patients enhanced tumor growth in an immune competent murine model of cancer. This was associated with increased tumor angiogenesis and suppression of antitumor immune responses as indicated by decreased cytokine production by tumor macrophages and cytotoxic T cells, diminished tumor infiltration of tumor-specific T cells, and decreased number of dendritic cells in tumor draining lymph nodes. These results suggest that carrier-induced immunosuppression and angiogenesis have the potential to reduce the antitumor effects of drugs loaded within. These findings may have significant implications for the current use and future development of anticancer nanoparticles and further investigations are urgently needed. FROM THE CLINICAL EDITOR: This study discusses important implications of nanoliposome-based drug delivery systems in cancer therapy, and demonstrates that nanoliposomes may have immunosuppressive and angiogenetic properties, directly counterbalancing their anti-cancer activity, which may also have important clinical implications related to more widespread applications of such systems.

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.

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 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.

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.

Is renal thrombotic angiopathy an emerging problem in the treatment of ovarian cancer recurrences?

BACKGROUND AND OBJECTIVE: Ovarian cancer is usually diagnosed at an advanced stage, with most patients undergoing surgery followed by platinum- and taxane-based chemotherapy. After initial clinical remission, the majority recur, leading to additional treatments, including not only platinums and taxanes but also pegylated liposomal doxorubicin (PLD), gemcitabine, topotecan, and, more recently, bevacizumab, which may extend survival times. PLD, in particular, has been extensively studied by our group, with encouraging therapeutic results. We, however, observed instances of chronic kidney disease (CKD) developing among patients who received long-term treatment for recurrent ovarian cancer. To document the frequency and contributing factors to the emergence of CKD, we initiated a retrospective review at two institutions. PATIENTS AND METHODS: Fifty-six consecutive patients with recurrent ovarian cancer receiving treatment at New York University Cancer Institute were reviewed for the presence of renal disease in 1997-2010. At Shaare Zedek Medical Center, 73 consecutive patients with ovarian cancer were reviewed in 2002-2010. Patients were diagnosed with CKD if they had an estimated GFR <60 mL/minute per 1.73 m2 for >3 months and were staged according to the National Kidney Foundation guidelines. RESULTS: Thirteen patients (23%) developed stage ≥3 CKD. Three patients had renal biopsies performed that showed thrombotic microangiopathy. CONCLUSIONS: CKD is emerging as a potential long-term consequence of current chemotherapy for recurrent ovarian cancer.