Purpose Sonodynamic therapy (SDT) is emerging as a cancer treatment alternative with significant advantages over conventional therapies, including its minimally invasive and site-specific nature, its radical antitumour efficacy with minimal side effects, and its capacity to raise an antitumour immune response. The study explores the efficacy of SDT in combination with nanotechnology against pancreatic ductal adenocarcinoma. Methods A nanoparticulate formulation (HPNP) based on a cathepsin B-degradable glutamate-tyrosine co-polymer that carries hematoporphyrin was used in this study for the SDT-based treatment of PDAC. Cathepsin B levels in BxPC-3 and PANC-1 cells were correlated to cellular uptake of HPNP. The HPNP efficiency to induce a sonodynamic effect at varying ultrasound parameters, and at different oxygenation and pH conditions, was investigated. The biodistribution, tumour accumulation profile, and antitumour efficacy of HPNP in SDT were examined in immunocompetent mice carrying bilateral ectopic murine pancreatic tumours. The immune response profile of excised tumour tissues was also examined. Results The HPNP formulation significantly improved cellular uptake of hematoporphyrin for both BxPC-3 and PANC-1 cells, while increase of cellular uptake was positively correlated in PANC-1 cells. There was a clear SDT-induced cytotoxicity at the ultrasound conditions tested, and the treatment impaired the capacity of both BxPC-3 and PANC-1 cells to form colonies. The overall acoustic energy and pulse length, rather than the power density, were key in eliciting the effects observed in vitro. The SDT treatment in combination with HPNP resulted in 21% and 27% reduction of the target and off-target tumour volumes, respectively, within 24 h. A single SDT treatment elicited an antitumour effect that was characterized by an SDT-induced decrease in immunosuppressive T cell phenotypes. Conclusion SDT has significant potential to serve as a monotherapy or adjunctive treatment for inoperable or borderline resectable PDAC.

Abstract Background The emergence of immune checkpoint inhibitors (ICI’s) in the past decade has proven transformative in the area of immuno-oncology. The PD-1 / PD-L1 axis has been particularly well studied and monoclonal antibodies developed to block either the receptor (anti PD-1) or its associated ligand (anti PD-L1) can generate potent anti-tumour immunity in certain tumour models. However, many “immune cold” tumours remain unresponsive to ICI’s. Sonodynamic therapy (SDT) is a targeted anti-cancer treatment that uses ultrasound to activate a sensitiser with the resulting generation of reactive oxygen species (ROS) causing direct cell death. SDT has also been shown to stimulate the adaptive immune system in a pre-clinical cancer model. We investigate the ability of combining ICI and microbubble mediated SDT at controlling tumour growth in a bilateral pancreatic cancer model. Methods Preparation of O2MB-RB are shown below (scheme 1). Cytotoxicity of SDT and immunotherapy in-vivo are illustrated below (Figure 1). Figure 1 highlights that T110299 cells were subcutaneously implanted in the right and left dorsum of C57 mice. Group 1 received an IP injection of anti-mouse PD-L1 antibody (10mg/kg). After 2 hours, mice in this group received an IV injection of O2MB-RB suspension while receiving ultrasound applied to the right-hand-side (target) tumour. Group 2 received the same as Group 1 but no anti PD-L1 antibody; Group 3 received anti-PD-L1 antibody alone and Group 4 remained untreated. Flow cytometry analysis were carried out to investigate tumour infiltrating CD4+ and CD8+ T-lymphocytes. Results The results demonstrated a significant 287% decrease in tumour volume when compared to untreated animals 11 days following the initial treatment with SDT, which reduced further to 369% when SDT was combined with anti-PD-L1 ICI treatment. Analysis of residual tumour tissues remaining after treatment revealed increased levels of infiltrating CD4+ and CD8+ T-lymphocytes (respectively 4.65 and 3.16-fold more) in the off-target tumours of animals where the target tumour was treated with SDT and anti-PD-L1, when compared to untreated tumours. These results suggest that SDT treatment elicits an adaptive immune response that is potentiated by the anti-PD-L1 ICI in this particular model of pancreatic cancer. Conclusions In conclusion, microbubble mediated SDT treatment of a target tumour in a bilateral tumour model of pancreatic cancer, enables growth control at both the target and off-target tumours which is further enhanced when combined with anti-PD-L1 ICI treatment. Combining SDT with anti-PD-L1 ICI treatment, which is also well tolerated, could provide an attractive treatment option for pancreatic cancer, particularly for patients with advanced disease who may not be physically capable of undertaking a toxic chemotherapy regimen.

The emergence of immune checkpoint inhibitors (ICI's) in the past decade has proven transformative in the area of immuno-oncology. The PD-1/PD-L1 axis has been particularly well studied and monoclonal antibodies developed to block either the receptor (anti PD-1) or its associated ligand (anti PD-L1) can generate potent anti-tumour immunity in certain tumour models. However, many "immune cold" tumours remain unresponsive to ICI's and strategies to stimulate the adaptive immune system and make these tumours more susceptible to ICI treatment are currently under investigation. Sonodynamic therapy (SDT) is a targeted anti-cancer treatment that uses ultrasound to activate a sensitiser with the resulting generation of reactive oxygen species (ROS) causing direct cell death by apoptosis and necrosis. SDT has also been shown to stimulate the adaptive immune system in a pre-clinical model of colorectal cancer. In this manuscript, we investigate the ability of microbubble mediated SDT to control tumour growth in a bilateral tumour mouse model of pancreatic cancer by treating the target tumour with SDT and observing the effects at the off-target untreated tumour. The results demonstrated a significant 287% decrease in tumour volume when compared to untreated animals 11 days following the initial treatment with SDT, which reduced further to 369% when SDT was combined with anti-PD-L1 ICI treatment. Analysis of residual tumour tissues remaining after treatment revealed increased levels of infiltrating CD4+ and CD8+ T-lymphocytes (respectively 4.65 and 3.16-fold more) in the off-target tumours of animals where the target tumour was treated with SDT and anti-PD-L1, when compared to untreated tumours. These results suggest that SDT treatment elicits an adaptive immune response that is potentiated by the anti-PD-L1 ICI in this particular model of pancreatic cancer.

Sonodynamic therapy (SDT) is an emerging stimulus-responsive approach for the targeted treatment of solid tumours. However, its ability to generate stimulus-responsive cytotoxic reactive oxygen species (ROS), is compromised by tumour hypoxia. Here we describe a robust means of preparing a pH-sensitive polymethacrylate-coated CaO2 nanoparticle that is capable of transiently alleviating tumour hypoxia. Systemic administration of particles to animals bearing human xenograft BxPC3 pancreatic tumours increases oxygen partial pressures (PO2) to 20?50 mmHg for over 40 min. RT-qPCR analysis of expression of selected tumour marker genes in treated animals suggests that the transient production of oxygen is sufficient to elicit effects at a molecular genetic level. Using particles labelled with the near infra-red (nIR) fluorescent dye, indocyanine green, selective uptake of particles by tumours was observed. Systemic administration of particles containing Rose Bengal (RB) at concentrations of 0.1 mg/mg of particles are capable of eliciting nanoparticle-induced, SDTmediated antitumour effects using the BxPC3 human pancreatic tumour model in immuno-compromised mice. Additionally, a potent abscopal effect was observed in off-target tumours in a syngeneic murine bilateral tumour model for pancreatic cancer and an increase in tumour cytotoxic T cells (CD8+) and a decrease in immunosuppressive tumour regulatory T cells [Treg (CD4+, FoxP3+)] was observed in both target and off-target tumours in SDT treated animals. We suggest that this approach offers significant potential in the treatment of both focal and disseminated (metastatic) pancreatic cancer.

Abstract Background Pancreatic cancer remains a significant therapeutic challenge and its poor prognosis has remained relatively unchanged for the past 40 years. Pancreatic tumours are highly desmoplastic and impenetrable lesions in which both gas and mass transfer is severely compromised. This leads to the development of hypoxia within the tumour and this compromises therapeutic approaches that rely on cytotoxic reactive oxygen species, e.g. photodynamic therapy, sonodynamic therapy and radiotherapy. Hypoxia also results in a relatively low pH within the tumour microenvironment. Here we describe a pH sensitive nanoparticle that can generate oxygen in the tumour and enhance ROS generating therapeutic approaches. Methods CaO2 NPs were generated by exposing to low frequency ultrasound and subsequently coated using a polymethacrylate polymer that becomes soluble at pH 6.4. For some studies, the sonosensitiser, Rose Bengal was attached to the particles. Oxygen generation in tumours (BxPC3) was demonstrated by inserting a dissolved oxygen probe into tumours following IV administration of particles. Particles were also employed together with photodynamic therapy (PDT) and sonodynamic therapy (SDT) using human xenograft and syngeneic pancreatic tumour models. In some cases, tumour tissues were recovered and analysed for tumour infiltrating immune cells using flow cytometry. Results Conclusions Coating CaO2 nanoparticles with a pH sensitive polymer provides in situ oxygen generation in tumours. Transient provision of oxygen enhances therapies that depend on the generation of cytotoxic reactive oxygen species. When used with SDT, and using a bilateral syngeneic pancreatic tumour model, a powerful abscopal effect was observed and this was shown to be immune-mediated. The above data suggest that the particles may be exploited to enhance other therapies that depend on the generation of ROS, e.g. radiotherapy, and further suggest that the approach can be used to treat either local or disseminated forms of pancreatic cancer.

The major unmet need and crucial challenge hampering the exciting potential of RNAi therapeutics in ophthalmology is to find an effective, safe and non-invasive means of delivering siRNA to the cornea. Although all tissues of the eye are accessible by injection, topical application is preferable for the frequent treatment regimen that would be necessary for siRNA-induced gene silencing. However, the ocular surface is one of the more complex biological barriers for drug delivery due to the combined effect of short contact time, tear dilution and poor corneal cell penetration. Using nanotechnology to overcome the challenges, we developed a unique silicon-based delivery platform for ocular delivery of siRNA. This biocompatible hybrid of porous silicon nanoparticles and lipids has demonstrated an ability to bind nucleic acid and deliver functional siRNA to corneal cells both in vitro and in vivo. Potent transfection of human corneal epithelial cells with siRNA-ProSilic® formulation was followed by a successful downregulation of reporter protein expression. Moreover, siRNA complexed with this silicon-based hybrid and applied in vivo topically to mice eyes penetrated across all cornea layers and resulted in a significant reduction of the targeted protein expression in corneal epithelium. In terms of siRNA loading capacity, system versatility, and potency of action, ProSilic provides unique attributes as a biodegradable delivery platform for therapeutic oligonucleotides.

Pancreatic cancer remains one of the most lethal forms of cancer with a 10-year survival of <1%. With little improvement in survival rates observed in the past 40 years, there is a significant need for new treatments or more effective strategies to deliver existing treatments. The antimetabolite gemcitabine (Gem) is the most widely used form of chemotherapy for pancreatic cancer treatment, but is known to produce significant side effects when administered systemically. We have previously demonstrated the benefit of combined chemo-sonodynamic therapy (SDT), delivered using oxygen carrying microbubbles (O2MB), as a targeted treatment for pancreatic cancer in a murine model of the disease. In this manuscript, we report the preparation of a biotin functionalised Gem ligand for attachment to O2MBs (O2MB-Gem). We demonstrate the effectiveness of chemo-sonodynamic therapy following ultrasound-targeted-microbubble-destruction (UTMD) of the O2MB-Gem and a Rose Bengal loaded O2MB (O2MB-RB) as a targeted treatment for pancreatic cancer. Specifically, UTMD using the O2MB-Gem and O2MB-RB conjugates reduced the viability of MIA PaCa-2, PANC-1, BxPC3 and T110299 pancreatic cancer cells by >60% (p < 0.001) and provided significant tumour growth delay (>80%, p < 0.001) compared to untreated animals when human xenograft MIA PaCa-2 tumours were treated in SCID mice. The toxicity of the O2MB-Gem conjugate was also determined in healthy non-tumour bearing MF1 mice and revealed no evidence of renal or hepatic damage. Therefore, the results presented in this manuscript suggest that chemo-sonodynamic therapy using the O2MB-Gem and O2MB-RB conjugates, is potentially an effective targeted and safe treatment modality for pancreatic cancer.

Background: Epidemiologic evidence suggests that a diet rich in (poly)phenols has beneficial effects on many chronic diseases. Brown seaweed is a rich source of (poly)phenols. Objective: The aim of this study was to investigate the bioavailability and effect of a brown seaweed (Ascophyllum nodosum) (poly)phenol extract on DNA damage, oxidative stress, and inflammation in vivo. Design: A randomized, double-blind, placebo-controlled crossover trial was conducted in 80 participants aged 30-65 y with a body mass index (in kg/m(2)) >= 25. The participants consumed either a 400-mg capsule containing 100 mg seaweed (poly)phenol and 300 mg maltodextrin or a 400-mg maltodextrin placebo control capsule daily for an 8-wk period. Bioactivity was assessed with a panel of blood-based markers including lymphocyte DNA damage, plasma oxidant capacity, C-reactive protein (CRP), and inflammatory cytokines. To explore the bioavailability of seaweed phenolics, an untargeted metabolomics analysis of urine and plasma samples after seaweed consumption was determined by ultra-high-performance liquid chromatography-high-resolution mass spectrometry. Results: Consumption of the seaweed (poly)phenols resulted in a modest decrease in DNA damage but only in a subset of the total population who were obese. There were no significant changes in CRP, antioxidant status, or inflammatory cytokines. We identified phlorotannin metabolites that are considered potential biomarkers of seaweed consumption including pyrogallol/phloroglucinol-sulfate, hydroxytrifurahol A-glucuronide, dioxinodehydroeckol-glucuronide, diphlorethol sulfates, C-O-C dimer of phloroglucinol sulfate, and C-O-C dimer of phloroglucinol. Conclusions: To the best of our knowledge, this work represents the first comprehensive study investigating the bioactivity and bioavailability of seaweed (poly)phenolics in human participants. We identified several potential biomarkers of seaweed consumption. Intriguingly, the modest improvements in DNA damage were observed only in the obese subset of the total population. The subgroup analysis should be considered exploratory because it was not preplanned; therefore, it was not powered adequately. Elucidation of the biology underpinning this observation will require participant stratification according to weight in future studies.