Inhaled treprostinil in patients with pulmonary hypertension associated with interstitial lung disease with less severe haemodynamics: a post hoc analysis of the INCREASE study.

BACKGROUND: Inhaled treprostinil (iTre) is the only treatment approved for pulmonary hypertension due to interstitial lung disease (PH-ILD) to improve exercise capacity. This post hoc analysis evaluated clinical worsening and PH-ILD exacerbations from the 16-week INCREASE study and change in 6-minute walking distance (6MWD) in the INCREASE open-label extension (OLE) in patients with less severe haemodynamics. METHODS: Patients were stratified by baseline pulmonary vascular resistance (PVR) of <4 Wood units (WU) versus ≥4 WU and <5 WU versus ≥5 WU. Exacerbations of underlying lung disease, clinical worsening and change in N-terminal prohormone of brain natriuretic peptide (NT-proBNP) in INCREASE were evaluated. For the OLE, patients previously assigned to placebo were considered to have a 16-week treatment delay. 6MWD and clinical events in the OLE were evaluated by PVR subgroup. RESULTS: Of the 326 patients enrolled in INCREASE, patients with less severe haemodynamics receiving iTre had fewer exacerbations of underlying lung disease and clinical worsening events. This was supported by the Bayesian analysis of the risk of disease progression (HR<1), and significant decreases in NT-proBNP levels. In the OLE, patients without a treatment delay had improved exercise capacity after 1-year compared with those with a 16-week treatment delay (22.1 m vs -10.3 m). Patients with a PVR of ≤5 WU without a treatment delay had a change of 5.5 m compared with -8.2 m for those with a treatment delay. Patients without a treatment delay had a prolonged time to hospitalisation, lung disease exacerbation and death. CONCLUSION: Treatment with iTre led to consistent benefits in clinical outcomes in patients with PH-ILD and less severe haemodynamics. Earlier treatment in less severe PH-ILD may lead to better exercise capacity long-term, however, the subgroup analyses in this post hoc study were underpowered and confirmation of these findings is needed.

Nano-trapping CXCL13 reduces regulatory B cells in tumor microenvironment and inhibits tumor growth.

Interactions between different cell types in the tumor microenvironment (TME) affect tumor growth. Tumor-associated fibroblasts produce C-X-C motif chemokine ligand 13 (CXCL13) which recruits B cells to the TME. B-cells in the TME differentiate into regulatory B cells (Bregs) (IL-10+CD1d+CD5+CD138+CD19+). We highlight these Breg cells as a new important factor in the modulation of the immunosuppressive TME in different desmoplastic murine tumor models. In addition, CXCL13 also stimulates epithelial-mesenchymal transition (EMT) of the tumor cells. The tumorigenic roles of CXCL13 led us to explore an innovative anti-cancer strategy based on delivering plasmid DNA encoding a CXCL13 trap to reduce Bregs differentiation and normalize EMT, thereby suppressing tumor growth. CXCL13 trap suppressed tumor growth in pancreatic cancer, BRAF-mutant melanoma, and triple-negative breast cancer. In this study, following treatment, the affected tumor remained dormant resulting in prolonged progression-free survival of the host.

Tumor neoantigen heterogeneity impacts bystander immune inhibition of pancreatic cancer growth.

The immunogenic clonal-fraction threshold in heterogeneous solid-tumor required to induce effective bystander-killing of non-immunogenic subclones is unknown. Pancreatic cancer poses crucial challenges for immune therapeutic interventions due to low mutational-burden and consequent lack of neoantigens. Here, we designed a model to incorporate artificial-neoantigens into genes-of -interest in cancer-cells and to test their potential to actuate bystander-killing. By precisely controlling a neoantigen's abundance in the tumor, we studied the impact of neoantigen frequency on immune-response and immune-escape. Our results showed single, strong, widely-expressed neoantigen could lead to robust antitumor response when over 80% of cancer cells express the neoantigen. Further, immunological assays demonstrated T-cell responses against non-target self-antigen on KRAS-oncoprotein, when we inoculated animals with a high frequency of tumor-cells expressing test-neoantigen. Using nanoparticle-based gene-therapy, we successfully altered tumor-microenvironment by perturbing interleukin-12 and interleukin-10 gene-expression. The subsequent microenvironment-remodeling reduced the neoantigen frequency threshold at which bioluminescent signal intensity for tumor-burden decreased 1.5-log-fold, marking robust tumor-growth inhibition, from 83% to 29%. Our results thus suggest bystander killing is inefficient in immunologically-cold tumors like pancreatic-cancer and requires high neoantigen abundance. However, bystander killing mediated antitumor response can be rescued by adjuvant-immune therapy.

Nano-delivery of Gemcitabine Derivative as a Therapeutic Strategy in a Desmoplastic KRAS Mutant Pancreatic Cancer.

Pancreatic ductal adenocarcinoma remains one of the challenging malignancies to treat, and chemotherapy is the primary treatment strategy available to most patients. Gemcitabine, one of the oldest chemotherapeutic drugs approved for pancreatic cancer, has limited efficacy, due to low drug distribution to the tumor and chemoresistance following therapy. In this study, we delivered gemcitabine monophosphate using lipid calcium phosphate nanoparticles, to desmoplastic pancreatic tumors. Monophosphorylation is a critical, rate-limiting step following cellular uptake of gemcitabine and precursor of the pharmacologically active gemcitabine triphosphate. Our drug delivery strategy enabled us to achieve robust tumor regression with a low parenteral dose in a clinically relevant, KRAS mutant, syngeneic orthotopic allograft, lentivirus-transfected KPC cell line-derived model of pancreatic cancer. Treatment with gemcitabine monophosphate significantly increased apoptosis of cancer cells, enabled reduction in the proportion of immunosuppressive tumor-associated macrophages and myeloid-derived suppressor cells, and did not increase expression of cancer stem cell markers. Overall, we could trigger a strong antitumor response in a treatment refractory PDAC model, while bypassing critical hallmarks of gemcitabine chemoresistance.

Nano Codelivery of Oxaliplatin and Folinic Acid Achieves Synergistic Chemo-Immunotherapy with 5-Fluorouracil for Colorectal Cancer and Liver Metastasis.

FOLFOX, the combinational strategy of folinic acid (FnA), 5-fluorouracil (5-Fu), and oxaliplatin (OxP), has been used as standard treatment of colorectal cancer (CRC) for decades. Despite the improved survival, patients still suffer from drawbacks such as low efficacy, high toxicity, and long course of treatment. New strategies to address these issues are needed to further clinical benefits. In this study, a nanoprecipitate (C26H35N9O7Pt) was formed by the active form of OxP ([Pt(DACH)(H2O)2]2+) and FnA, which was formulated into an aminoethyl anisamide targeted PEGylated lipid nanoparticle within microemulsions using nanoprecipitation technique. The resultant formulation (namely Nano-Folox) significantly promoted the blood circulation and tumor accumulation of platinum drug and FnA in an orthotopic CRC mouse model. Emerging evidence indicates that OxP can not only provide anticancer cytotoxic effects but also induce immunogenic cell death (a type of apoptosis that primes anticancer immune responses). Consequently, Nano-Folox demonstrated favorable chemo-immunotherapeutic activities in orthotopic CRC mice. In addition, when compared to FOLFOX the significantly stronger chemo-immunotherapeutic responses were achieved by the combination of Nano-Folox and 5-Fu without showing toxicity. Moreover, the anti-PD-L1 monoclonal antibody enhanced Nano-Folox/5-Fu for decreased liver metastases in mice. These results indicate the potential of Nano-Folox-based combination strategy for the treatment of CRC.

Nanotherapeutics for Immuno-Oncology: A Crossroad for New Paradigms.

With the rapid increase in the use of nanotechnology and immunotherapy for cancer management in the recent past, there are great implications for using nanotechnology in immuno-oncology. However, to deliver clinical success, the scientific and clinical rationale must be critically evaluated when applying nanotechnology to immuno-oncology challenges. This opinion article distinguishes between designing nanotherapeutics for immunotherapy and the past focus on the placement of chemotherapy agents in nanoparticles. We believe the integration of nanotechnology with cancer immunotherapy for nano-immunotherapeutics provides unique opportunities for both fields, paving the way for entirely new therapeutic paradigms. As a particular focus in our article, we envision the necessities and challenges of nanotechnology in the development of in situ cancer vaccines, immune checkpoint inhibitors, adoptive cell transfer, and bispecific antibody therapy.

Liposomal Nanostructures for Drug Delivery in Gastrointestinal Cancers.

Gastrointestinal (GI) cancers like liver, pancreatic, colorectal, and gastric cancer remain some of the most difficult and aggressive cancers. Nanoparticles like liposomes had been approved in the clinic for cancer therapy dating as far back as 1995. Over the years, liposomal formulations have come a long way, facing several roadblocks and failures, and advancing by optimizing formulations and incorporating novel design approaches to navigate therapeutic delivery challenges. The first liposomal formulation for a GI cancer drug was approved recently in 2015, setting the stage for further clinical developments of liposome-based delivery systems for therapies against GI malignancies. This article reviews the design considerations and strategies that can be used to deliver drugs to GI tumors, the wide range of therapeutic agents that have been explored in preclinical as well as clinical studies, and the current therapies that are being investigated in the clinic against GI malignancies.

Nanoparticle Delivery of RIG-I Agonist Enables Effective and Safe Adjuvant Therapy in Pancreatic Cancer.

Local immunomodulation can be a promising strategy to augment the efficacy and decrease off-target toxicities associated with cancer treatment. Pancreatic cancer is resistant to immunotherapies due to the immunosuppressive tumor microenvironment. Herein, we investigated a therapeutic approach involving delivery of a short interfering double-stranded RNA (dsRNA), specific to Bcl2, with 5' triphosphate ends, by lipid calcium phosphate nanoparticles, in an orthotopic allograft KPC model of pancreatic cancer. Retinoic acid-inducible gene I (RIG-I)-like receptors can bind to 5' triphosphate dsRNA (ppp dsRNA), a pathogen-associated molecular pattern, producing type I interferon, while Bcl2 silencing can drive apoptosis of cancer cells. Our approach demonstrated a robust enrichment of tumor tissue with therapeutic nanoparticles and enabled a significant tumor growth inhibition, prolonging median overall survival. Nanoparticles encapsulating dual-therapeutic ppp dsRNA allowed strong induction in levels of pro-inflammatory Th1 cytokines, further increasing proportions of CD8+ T cells over regulatory T cells, M1 over M2 macrophages, and decreased levels of immunosuppressive B regulatory and plasma cells in the tumor microenvironment. Thus, these results provide a new immunotherapy approach for pancreatic cancer.

RNA Interference-Based Cancer Drugs: The Roadblocks, and the "Delivery" of the Promise.

Nucleic acid-based therapeutics like synthetic small interfering RNAs have been exploited to modulate gene function, taking advantage of RNA interference (RNAi), an evolutionally conserved biological process. Recently, the world's first RNAi drug was approved for a rare genetic disorder in the liver. However, there are significant challenges that need to be resolved before RNAi can be translated in other genetic diseases like cancer. Current drug delivery platforms for therapeutic silencing RNAs are tailored to hepatic targets. RNAi therapies for nonhepatic conditions are still at early clinical phases. In this study, we discuss the critical design considerations in anticancer RNAi drug development, insights gained from initial clinical trials, and new strategies that are entering clinical development, shaping the future of RNAi in cancer.

Local Blockade of Interleukin 10 and C-X-C Motif Chemokine Ligand 12 with Nano-Delivery Promotes Antitumor Response in Murine Cancers.

In many cancers, the tumor microenvironment (TME) is largely immune suppressive, blocking the antitumor immunity and resulting in immunotherapy resistance. Interleukin 10 (IL-10) is a major player controlling the immunosuppressive TME in different murine tumor models. Increased IL-10 production suppresses intratumoral dendritic cell production of interleukin 12, thereby limiting antitumor cytotoxic T-cell responses and activation of NK cells during therapy. We engineered, formulated, and delivered genes encoding an IL-10 protein trap to change immunosuppressive TME, which could enhance antitumor immunity. Additionally, to achieve stronger and long-term therapeutic efficacy in a pancreatic cancer model, we targeted C-X-C motif chemokine ligand 12 (CXCL12), a key factor for inhibiting T-cell tumor infiltration, and simultaneously delivered an IL-10 trap. Following three injections of the lipid-protamine-DNA (LPD) nanoparticles loaded with trap genes (IL-10 trap and CXCL12 trap), we found tumor growth reduction and significantly prolonged survival of the host compared to control groups. Furthermore, the combination trap gene treatment significantly reduced immunosuppressive cells, such as M2 macrophages, MDSCs, and PD-L1+ cells, and activated immunosuppressive tolerogenic dendritic cells, NK cells, and macrophages intratumorally. We have also shown that, when effectively delivered to the tumor, the IL-10 trap gene alone can inhibit triple-negative breast cancer growth. This strategy may allow clinicians and researchers to change the immunosuppressive microenvironment in the tumor with either a single therapeutic agent or in combination with other immunotherapies to prime the immune system, preventing cancer invasion and prolonging patient survival.

Targeted drug delivery to melanoma.

Melanoma derived from melanocytes is the most aggressive genre of skin cancer. Although the considerable advancement in the study of human cancer biology and drug discovery, most advanced melanoma patients are inevitably unable to be cured. With the emergence of nanotechnology, the use of nano-carriers is widely expected to alter the landscape of melanoma treatment. In this review, we will discuss melanoma biology, current treatment options, mechanisms behind drug resistance, and nano-based solutions for effective anti-cancer therapy, followed by challenges and perspectives in both pre-clinical and clinical settings.

Transient and Local Expression of Chemokine and Immune Checkpoint Traps To Treat Pancreatic Cancer.

Pancreatic tumors are known to be resistant to immunotherapy due to the extensive immune suppressive tumor microenvironment (TME). We hypothesized that CXCL12 and PD-L1 are two key molecules controlling the immunosuppressive TME. Fusion proteins, called traps, designed to bind with these two molecules with high affinity (Kd = 4.1 and 0.22 nM, respectively) were manufactured and tested for specific binding with the targets. Plasmid DNA encoding for each trap was formulated in nanoparticles and intravenously injected to mice bearing orthotopic pancreatic cancer. Expression of traps was mainly seen in the tumor, and secondarily, accumulations were primarily in the liver. Combination trap therapy shrunk the tumor and significantly prolonged the host survival. Either trap alone only brought in a partial therapeutic effect. We also found that CXCL12 trap allowed T-cell penetration into the tumor, and PD-L1 trap allowed the infiltrated T-cells to kill the tumor cells. Combo trap therapy also significantly reduced metastasis of the tumor cells to other organs. We conclude that the trap therapy significantly modified the immunosuppressive TME to allow the host immune system to kill the tumor cells. This can be an effective therapy in clinical settings.

Chitosan derivatives cross-linked with iodinated 2,5-dimethoxy-2,5-dihydrofuran for non-invasive imaging.

Radiopaque polymer derivatives were successfully prepared through surface diffusion mediated cross-linking of chitosan with iodinated 2,5-dimethoxy-2,5-dihydrofuran. The incorporation of iodine in 2,5-dimethoxy-2,5-dihydrofuran was validated by (1)H NMR and mass spectroscopy. The cross-linking of the glucosamine moieties of chitosan with the iodinated product was confirmed by (13)C NMR and energy-dispersive X-ray spectroscopy. Radiography analysis proved inherent opacity of the iodinated fibrous sheets and microspheres that were comparable to the X-ray visibility of aluminum hollow rings of equivalent thickness and commercially available radiopaque tape, respectively. Microscopic studies evidenced retention of the fiber/microsphere morphology after the iodination/cross-linking reactions. The effects of iodination/cross-linking on the mechanical and biodegradation properties of fibers were studied by nanoindentation and enzymatic assay, respectively. In vitro and in vivo studies established the nontoxic, biodegradable nature of radiopaque derivatives. Iodinated fiber mesh implanted in a rabbit model was significantly X-ray opaque compared to the uncross-linked fiber mesh and medical grade surgical swabs. Further, opacity of the iodinated mesh was evident even after 60 days, though the intensity was reduced, which indicates the biodegradable nature of the iodinated polymer. The opacity of the iodinated sutures was also established in the computed tomography images. Finally, the sufficient in vivo contrast property of the radiopaque microspheres in the gastrointestinal tract indicates its possible role in clinical diagnostics.