CCK Receptor Inhibition Reduces Pancreatic Tumor Fibrosis and Promotes Nanoparticle Delivery.

The poor prognosis for pancreatic ductal adenocarcinoma (PDAC) patients is due in part to the highly fibrotic nature of the tumors that impedes delivery of therapeutics, including nanoparticles (NPs). Our prior studies demonstrated that proglumide, a cholecystokinin receptor (CCKR) antagonist, reduced fibrosis pervading PanIN lesions in mice. Here, we further detail how the reduced fibrosis elicited by proglumide achieves the normalization of the desmoplastic tumor microenvironment (TME) and improves nanoparticle uptake. One week following the orthotopic injection of PDAC cells, mice were randomized to normal or proglumide-treated water for 3-6 weeks. Tumors were analyzed ex vivo for fibrosis, vascularity, stellate cell activation, vascular patency, and nanoparticle distribution. The histological staining and three-dimensional imaging of tumors each indicated a reduction in stromal collagen in proglumide-treated mice. Proglumide treatment increased tumor vascularity and decreased the activation of cancer-associated fibroblasts (CAFs). Additionally, PANC-1 cells with the shRNA-mediated knockdown of the CCK2 receptor showed an even greater reduction in collagen, indicating the CCK2 receptors on tumor cells contribute to the desmoplastic TME. Proglumide-mediated reduction in fibrosis also led to functional changes in the TME as evidenced by the enhanced intra-tumoral distribution of small (<12 nm) Rhodamine-loaded nanoparticles. The documented in vivo, tumor cell-intrinsic anti-fibrotic effects of CCK2R blockade in both an immunocompetent syngeneic murine PDAC model as well as a human PDAC xenograft model demonstrates that CCK2R antagonists, such as proglumide, can improve the delivery of nano-encapsulated therapeutics or imaging agents to pancreatic tumors.

VCAM-1-targeted nanoparticles to diagnose, monitor and treat atherosclerosis.

Vascular cell adhesion molecule-1 (VCAM-1) was identified over 2 decades ago as an endothelial adhesion receptor involved in leukocyte recruitment and cell-based immune responses. In atherosclerosis, a chronic inflammatory disease of the blood vessels that is the leading cause of death in the USA, endothelial VCAM-1 is robustly expressed beginning in the early stages of the disease. The interactions of circulating immune cells with VCAM-1 on the activated endothelial cell surface promote the uptake of monocytes and the progression of atherosclerotic lesions in susceptible vessels. Herein, we review the role of VCAM-1 in atherosclerosis and the use of VCAM-1 binding peptides, antibodies and aptamers as targeting agents for nanoplatforms for early detection and treatment of atherosclerotic disease.

Addition of in situ clay catalysts at different process points in a cascaded hydrothermal carbonization-pyrolysis process for agro-industrial waste valorization.

Agro-industrial wastes can be thermochemically converted to sustainable fuels and upcycled carbon products. However, processing such feedstocks through pyrolysis or hydrothermal carbonization (HTC) alone yields fuels that require significant downstream upgrading. In this work, apple pomace was treated via a cascaded HTC-pyrolysis process using inexpensive and abundant clay catalysts, montmorillonite and attapulgite. Clays were added pre-HTC to raw biomass or to hydrochar pre-pyrolysis to examine the effect of addition as a function of process insertion point. Both clays produce similar bio-oils when they are added at the same process point. However, bio-oil was affected by the point in which clay was added to the process (before or after HTC). When clay was added pre-HTC, the bio-oil had an average hydrocarbon content twice that when clay was added to the hydrochar after HTC, prior to pyrolysis.

Reactive Oxygen Species-Triggered Hydrogen Sulfide Release and Cancer-Selective Antiproliferative Effect of Anethole Dithiolethione-Containing Polymeric Micelles.

Hydrogen sulfide (H2 S) is a gaseous signaling molecule in the human body and has attracted attention in cancer therapy due to its regulatory roles in cancer cell proliferation and migration. Accumulating evidence suggests that continuous delivery of H2 S to cancer cells for extended periods of time suppresses cancer progression. However, one major challenge in therapeutic applications of H2 S is its controlled delivery. To solve this problem, polymeric micelles are developed containing H2 S donating-anethole dithiolethione (ADT) groups, with H2 S release profiles optimal for suppressing cancer cell proliferation. The micelles release H2 S upon oxidation by reactive oxygens species (ROS) that are present inside the cells. The H2 S release profiles can be controlled by changing the polymer design. Furthermore, the micelles that show a moderate H2 S release rate exert the strongest anti-proliferative effect in human colon cancer cells in in vitro assays as well as the chick chorioallantoic membrane cancer model, while the micelles do not affect proliferation of human umbilical vein endothelial cells. This study shows the importance of fine-tuning H2 S release profiles using a micelle approach for realizing the full therapeutic potential of H2 S in cancer treatment.

Enzyme aggregation and fragmentation induced by catalysis relevant species.

It is usually assumed that enzymes retain their native structure during catalysis. However, the aggregation and fragmentation of proteins can be difficult to detect and sometimes conclusions are drawn based on the assumption that the protein is in its native form. We have examined three model enzymes, alkaline phosphatase (AkP), hexokinase (HK) and glucose oxidase (GOx). We find that these enzymes aggregate or fragment after addition of chemical species directly related to their catalysis. We used several independent techniques to study this behavior. Specifically, we found that glucose oxidase and hexokinase fragment in the presence of D-glucose but not L-glucose, while hexokinase aggregates in the presence of Mg2+ ion and either ATP or ADP at low pH. Alkaline phosphatase aggregates in the presence of Zn2+ ion and inorganic phosphate. The aggregation of hexokinase and alkaline phosphatase does not appear to attenuate their catalytic activity. Our study indicates that specific multimeric structures of native enzymes may not be retained during catalysis and suggests pathways for different enzymes to associate or separate over the course of substrate turnover.

Aptamer-Targeted Calcium Phosphosilicate Nanoparticles for Effective Imaging of Pancreatic and Prostate Cancer.

PURPOSE: Accurate tumor identification and staging can be difficult. Aptamer-targeted indocyanine green (ICG)-nanoparticles can enhance near-infrared fluorescent imaging of pancreatic and prostate tumors and could improve early cancer detection. This project explored whether calcium-phosphosilicate nanoparticles, also known as NanoJackets (NJs), that were bioconjugated with a tumor-specific targeting DNA aptamer could improve the non-invasive detection of pancreatic and prostate tumors. METHODS: Using in vivo near-infrared optical imaging and ex vivo fluorescence analysis, DNA aptamer-targeted ICG-loaded NJs were compared to untargeted NJs for detection of tumors. RESULTS: Nanoparticles were bioconjugated with the DNA aptamer AP1153, which binds to the CCK-B receptor (CCKBR). Aptamer bioconjugated NJs were not significantly increased in size compared with unconjugated nanoparticles. AP1153-ICG-NJ accumulation in orthotopic pancreatic tumors peaked at 18 h post-injection and the ICG signal was cleared by 36 h with no evidence on uptake by non-tumor tissues. Ex vivo tumor imaging confirmed the aptamer-targeted NJs accumulated to higher levels than untargeted NJs, were not taken up by normal pancreas, exited from the tumor vasculature, and were well-dispersed throughout pancreatic and prostate tumors despite extensive fibrosis. Specificity for AP1153-NJ binding to the CCK-B receptor on pancreatic tumor cells was confirmed by pre-treating tumor-bearing mice with the CCK receptor antagonist proglumide. Proglumide pre-treatment reduced the in vivo tumoral accumulation of AP1153-NJs to levels comparable to that of untargeted NJs. CONCLUSION: Through specific interactions with CCK-B receptors, tumor-targeted nanoparticles containing either ICG or rhodamine WT were well distributed throughout the matrix of both pancreatic and prostate tumors. Tumor-targeted NJs carrying various imaging agents can enhance tumor detection.

Preferential uptake of antibody targeted calcium phosphosilicate nanoparticles by metastatic triple negative breast cancer cells in co-cultures of human metastatic breast cancer cells plus bone osteoblasts.

Calcium phosphosilicate nanoparticles (CPSNPs) are bioresorbable nanoparticles that can be bioconjugated with targeting molecules and encapsulate active agents and deliver them to tumor cells without causing damage to adjacent healthy tissue. Data obtained in this study demonstrated that an anti-CD71 antibody on CPSNPs targets these nanoparticles and enhances their internalization by triple negative breast cancer cells in-vitro. Caspase 3,7 activation, DNA damage, and fluorescent microscopy confirmed the apoptotic breast cancer response caused by targeted anti-CD71-CPSNPs encapsulated with gemcitabine monophosphate, the active metabolite of the chemotherapeutic gemcitabine used to treat cancers including breast and ovarian. Targeted anti-CD71-CPSNPs encapsulated with the fluorophore, Rhodamine WT, were preferentially internalized by breast cancer cells in co-cultures with osteoblasts. While osteoblasts partially internalized anti-CD71-GemMP-CPSNPs, their cell growth was not affected. These results suggest that CPSNPs may be used as imaging tools and selective drug delivery systems for breast cancer that has metastasized to bone.

The use of nanoparticulates to treat breast cancer.

Breast cancer is a major ongoing public health issue among women in both developing and developed countries. Significant progress has been made to improve the breast cancer treatment in the past decades. However, the current clinical approaches are invasive, of low specificity and can generate severe side effects. As a rapidly developing field, nanotechnology brings promising opportunities to human cancer diagnosis and treatment. The use of nanoparticulate-based platforms overcomes biological barriers and allows prolonged blood circulation time, simultaneous tumor targeting and enhanced accumulation of drugs in tumors. Currently available and clinically applicable innovative nanoparticulate-based systems for breast cancer nanotherapies are discussed in this review.

Effective encapsulation and biological activity of phosphorylated chemotherapeutics in calcium phosphosilicate nanoparticles for the treatment of pancreatic cancer.

Drug resistant cancers like pancreatic ductal adenocarcinoma (PDAC) are difficult to treat, and nanoparticle drug delivery systems can overcome some of the limitations of conventional systemic chemotherapy. In this study, we demonstrate that FdUMP and dFdCMP, the bioactive, phosphorylated metabolites of the chemotherapy drugs 5-FU and gemcitabine, can be encapsulated into calcium phosphosilicate nanoparticles (CPSNPs). The non-phosphorylated drug analogs were not well encapsulated by CPSNPs, suggesting the phosphate modification is essential for effective encapsulation. In vitro proliferation assays, cell cycle analyses and/or thymidylate synthase inhibition assays verified that CPSNP-encapsulated phospho-drugs retained biological activity. Analysis of orthotopic tumors from mice treated systemically with tumor-targeted FdUMP-CPSNPs confirmed the in vivo up take of these particles by PDAC tumor cells and release of active drug cargos intracellularly. These findings demonstrate a novel methodology to efficiently encapsulate chemotherapeutic agents into the CPSNPs and to effectively deliver them to pancreatic tumor cells.

A Cholecystokinin B Receptor-Specific DNA Aptamer for Targeting Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinomas (PDACs) constitutively express the G-protein-coupled cholecystokinin B receptor (CCKBR). In this study, we identified DNA aptamers (APs) that bind to the CCKBR and describe their characterization and targeting efficacy. Using dual SELEX selection against "exposed" CCKBR peptides and CCKBR-expressing PDAC cells, a pool of DNA APs was identified. Further downselection was based on predicted structures and properties, and we selected eight APs for initial characterizations. The APs bound specifically to the CCKBR, and we showed not only that they did not stimulate proliferation of PDAC cell lines but rather inhibited their proliferation. We chose one AP, termed AP1153, for further binding and localization studies. We found that AP1153 did not activate CCKBR signaling pathways, and three-dimensional Confocal microscopy showed that AP1153 was internalized by PDAC cells in a receptor-mediated manner. AP1153 showed a binding affinity of 15 pM. Bioconjugation of AP1153 to the surface of fluorescent NPs greatly facilitated delivery of NPs to PDAC tumors in vivo. The selectivity of this AP-targeted NP delivery system holds promise for enhanced early detection of PDAC lesions as well as improved chemotherapeutic treatments for PDAC patients.

3D Printing of Personalized Artificial Bone Scaffolds.

Additive manufacturing technologies, including three-dimensional printing (3DP), have unlocked new possibilities for bone tissue engineering. Long-term regeneration of normal anatomic structure, shape, and function is clinically important subsequent to bone trauma, tumor, infection, nonunion after fracture, or congenital abnormality. Due to the great complexity in structure and properties of bone across the population, along with variation in the type of injury or defect, currently available treatments for larger bone defects that support load often fail in replicating the anatomic shape and structure of the lost bone tissue. 3DP could provide the ability to print bone substitute materials with a controlled chemistry, shape, porosity, and topography, thus allowing printing of personalized bone grafts customized to the patient and the specific clinical condition. 3DP and related fabrication approaches of bone grafts may one day revolutionize the way clinicians currently treat bone defects. This article gives a brief overview of the current advances in 3DP and existing materials with an emphasis on ceramics used for 3DP of bone scaffolds. Furthermore, it addresses some of the current limitations of this technique and discusses potential future directions and strategies for improving fabrication of personalized artificial bone constructs.

Novel strategies for managing pancreatic cancer.

With the incidence reports of pancreatic cancer increasing every year, research over the last several decades has been focused on the means to achieve early diagnosis in patients that are at a high risk of developing the malignancy. This review covers current strategies for managing pancreatic cancer and further discusses efforts in understanding the role of early onset symptoms leading to tumor progression. Recent investigations in this discussion include type 3c diabetes, selected biomarkers and pathways related to pancreatic intraepithelial neoplasia lesions, drug resistance, and advances in nanomedicine which may provide significant solutions for improving early detection and treatments in future medicine.

PhotoImmunoNanoTherapy reveals an anticancer role for sphingosine kinase 2 and dihydrosphingosine-1-phosphate.

Tumor-associated inflammation mediates the development of a systemic immunosuppressive milieu that is a major obstacle to effective treatment of cancer. Inflammation has been shown to promote the systemic expansion of immature myeloid cells which have been shown to exert immunosuppressive activity in laboratory models of cancer as well as cancer patients. Consequentially, significant effort is underway toward the development of therapies that decrease tumor-associated inflammation and immunosuppressive cells. The current study demonstrated that a previously described deep tissue imaging modality, which utilized indocyanine green-loaded calcium phosphosilicate nanoparticles (ICG-CPSNPs), could be utilized as an immunoregulatory agent. The theranostic application of ICG-CPSNPs as photosensitizers for photodynamic therapy was shown to block tumor growth in murine models of breast cancer, pancreatic cancer, and metastatic osteosarcoma by decreasing inflammation-expanded immature myeloid cells. Therefore, this therapeutic modality was termed PhotoImmunoNanoTherapy. As phosphorylated sphingolipid metabolites have been shown to have immunomodulatory roles, it was hypothesized that the reduction of immature myeloid cells by PhotoImmunoNanoTherapy was dependent upon bioactive sphingolipids. Mechanistically, PhotoImmunoNanoTherapy induced a sphingosine kinase 2-dependent increase in sphingosine-1-phosphate and dihydrosphingosine-1-phosphate. Furthermore, dihydrosphingosine-1-phosphate was shown to selectively abrogate myeloid lineage cells while concomitantly allowing the expansion of lymphocytes that exerted an antitumor effect. Collectively, these findings revealed that PhotoImmunoNanoTherapy, utilizing the novel nontoxic theranostic agent ICG-CPSNP, can decrease tumor-associated inflammation and immature myeloid cells in a sphingosine kinase 2-dependent manner. These findings further defined a novel myeloid regulatory role for dihydrosphingosine-1-phosphate. PhotoImmunoNanoTherapy holds the potential to be a revolutionary treatment for cancers with inflammatory and immunosuppressive phenotypes.

Calcium phosphosilicate nanoparticles for imaging and photodynamic therapy of cancer.

Photodynamic therapy (PDT) has emerged as an alternative modality for cancer treatment. PDT works by initiating damaging oxidation or redox-sensitive pathways to trigger cell death. PDT can also regulate tumor angiogenesis and modulate systemic antitumor immunity. The drawbacks to PDT--photosensitizer toxicity, a lack of selectivity and efficacy of photosensitizers, and a limited penetrance of light through deep tissues--are the same pitfalls associated with diagnostic imaging. Developments in the field of nanotechnology have generated novel platforms for optimizing the advantages while minimizing the disadvantages of PDT. Calcium phosphosilicate nanoparticles (CPSNPs) represent an optimal nano-system for both diagnostic imaging and PDT. In this review, we will discuss how CPSNPs can enhance optical agents and serve as selective, non-toxic, and functionally stable photosensitizers for PDT. We will also examine novel applications of CPSNPs and PDT for the treatment of leukemia to illustrate their potential utility in cancer therapeutics.

Calcium phosphate-based composite nanoparticles in bioimaging and therapeutic delivery applications.

Bioimaging and therapeutic delivery applications are areas of biomedicine where nanoparticles have had significant impact, but the use of a nanomaterial in these applications can be limited by its physicochemical properties. Calcium phosphate-based composite nanoparticles are nontoxic and biodegradable, and are therefore considered attractive candidates for bioimaging and therapeutic drug delivery applications. Also, the pH-dependent solubility profiles of calcium phosphate materials make this class of nanoparticles especially useful for in vitro and in vivo delivery of dyes, oligonucleotides, and drugs. In this article, we discuss how calcium phosphate-based composite nanoparticles fulfill some of the requirements typically made for nanoparticles in biomedical applications. We also highlight recent studies in bioimaging and therapeutic delivery applications focusing on how these studies have addressed some of the challenges associated with using these nanoparticles in bioimaging and delivery of therapeutics.

Single-incision laparoscopic sigmoidectomy and rectopexy case series.

Single-incision laparoscopic surgery has recently been investigated as a novel approach to colorectal pathology. This article describes 3 cases of single-incision laparoscopic sigmoidectomy with rectopexy for the treatment of rectal prolapse. We demonstrate our surgical approach and results from these initial patients treated with this novel technique.

Targeted indocyanine-green-loaded calcium phosphosilicate nanoparticles for in vivo photodynamic therapy of leukemia.

Leukemia is one of the most common and aggressive adult cancers, as well as the most prevalent childhood cancer. Leukemia is a cancer of the hematological system and can be divided into a diversity of unique malignancies based on the onset of the disease as well as the specific cell lineages involved. Cancer stem cells, including recently identified leukemia stem cells (LSCs), are hypothesized to be responsible for cancer development, relapse, and resistance to treatment, and new therapeutics targeting these cellular populations are urgently needed. Nontoxic and nonaggregating calcium phosphosilicate nanoparticles (CPSNPs) encapsulating the near-infrared fluoroprobe indocyanine green (ICG) were recently developed for diagnostic imaging and drug delivery as well as for photodynamic therapy (PDT) of solid tumors. Prior studies revealed that specific targeting of CPSNPs allowed for enhanced accumulation within breast cancer tumors, via CD71 targeting, or pancreatic cancer tumors, via gastrin receptor targeting. In the present study, ICG-loaded CPSNPs were evaluated as photosensitizers for PDT of leukemia. Using a novel bioconjugation approach to specifically target CD117 or CD96, surface features enhanced on leukemia stem cells, in vitro ICG-CPSNP PDT of a murine leukemia cell line and human leukemia samples were dramatically improved. Furthermore, the in vivo efficacy of PDT was dramatically enhanced in a murine leukemia model by utilizing CD117-targeted ICG-CPSNPs, resulting in 29% disease-free survival. Altogether, this study demonstrates that leukemia-targeted ICG-loaded CPSNPs offer the promise to effectively treat relapsing and multidrug-resistant leukemia and to improve the life of leukemia patients.

The colloidal stability of fluorescent calcium phosphosilicate nanoparticles: the effects of evaporation and redispersion on particle size distribution.

Understanding the colloidal stability of nanoparticles is important for biological applications, such as bio-imaging and drug delivery. This work combines theoretical calculations with experimental data to elucidate the mechanism of stabilization for calcium phosphosilicate nanoparticles containing Cy3 with both citrate and poly(ethylene glycol) (PEG) surface conjugation. The citrate surface is shown to provide electrosteric dispersion in water-ethanol mixtures as well as the ability to redisperse after evaporating the solvent. Improved colloidal stability is afforded with the addition of PEG with respect to redispersion after drying. Changes in average agglomeration number (AAN) are tracked and explained by DLVO and the Napper electrosteric and steric theories for dispersion, respectively.

Validation of a novel laparoscopic adjustable gastric band simulator.

BACKGROUND: Morbid obesity accounts for more than 90,000 deaths per year in the United States. Laparoscopic adjustable gastric banding (LAGB) is the second most common weight loss procedure performed in the US and the most common in Europe and Australia. Simulation in surgical training is a rapidly advancing field that has been adopted by many to prepare surgeons for surgical techniques and procedures. The aim of our study was to determine face, construct, and content validity for a novel virtual reality laparoscopic adjustable gastric band simulator. METHODS: Twenty-eight subjects were categorized into two groups (expert and novice), determined by their skill level in laparoscopic surgery. Experts consisted of subjects who had at least 4 years of laparoscopic training and operative experience. Novices consisted of subjects with medical training but with less than 4 years of laparoscopic training. The subjects used the virtual reality laparoscopic adjustable band surgery simulator. They were automatically scored according to various tasks. The subjects then completed a questionnaire to evaluate face and content validity. RESULTS: On a 5-point Likert scale (1 = lowest score, 5 = highest score), the mean score for visual realism was 4.00 ± 0.67 and the mean score for realism of the interface and tool movements was 4.07 ± 0.77 (face validity). There were significant differences in the performances of the two subject groups (expert and novice) based on total scores (p < 0.001) (construct validity). Mean score for utility of the simulator, as addressed by the expert group, was 4.50 ± 0.71 (content validity). CONCLUSION: We created a virtual reality laparoscopic adjustable gastric band simulator. Our initial results demonstrate excellent face, construct, and content validity findings. To our knowledge, this is the first virtual reality simulator with haptic feedback for training residents and surgeons in the laparoscopic adjustable gastric banding procedure.

Single-incision laparoscopic right colectomy: experience with 17 consecutive cases and comparison with multiport laparoscopic right colectomy.

BACKGROUND: Recently, single-incision laparoscopic surgery has begun to develop as an extension of standard laparoscopic minimally invasive procedures. However, there have been a limited number of reports of single-incision procedures in colorectal disease. PURPOSE: The aim of this study is to describe our initial experience with single-incision laparoscopic right colectomy and to make comparisons with the current standard of care, multiport laparoscopic right colectomy. METHODS: Data from consecutive patients undergoing single-incision laparoscopic right colectomy were analyzed and compared with case-matched multiport laparoscopic right colectomies. Indications for surgery, type of port used, operative time, number of nodes harvested, length of hospital stay, and complications were the outcomes measured. RESULTS: During the study period, 17 patients underwent single-incision laparoscopic colectomy. Of the planned single-incision laparoscopic cases, 15 (88%) were completed with a single incision, whereas 2 required an additional port placement. There were no conversions to open surgery during any of the cases. Indications for surgery were similar between the 2 groups. Operative time was not significantly different in single-incision laparoscopic right colectomy compared with multiport laparoscopic right colectomy (139 min vs 134 min, respectively; P = .61). Length of stay and number of nodes harvested also had no significant differences between the 2 groups. There was one death after discharge to home secondary to pulmonary embolism and one delayed thermal injury in the single-incision laparoscopic group. CONCLUSION: Single-incision laparoscopic right colectomy is feasible, and appears to have results similar to standard multiport right colectomy in our initial comparisons. Ongoing development in instrumentation may help to further shorten operative time and minimize complications, and may make this an equivalent or preferred method for minimally invasive colorectal surgery. Large, prospective, randomized, controlled trials should be conducted to further compare the safety and efficacy of this approach.