Directed Evolution of AAV Targeting Primate Retina by Intravitreal Injection Identifies R100, a Variant Demonstrating Robust Gene Delivery and Therapeutic Efficacy in Non-Human Primates

Targeted AAV vectors are needed for safe and efficient delivery to and transduction of specific tissue target(s) in patients. Effective intravitreal delivery for retina gene therapy is not feasible with wildtype AAV. We employed directed evolution in nonhuman primates (NHP) to discover an AAV variant (R100) for intravitreal treatment of multiple target cells in the primate retina. R100 demonstrated superior transduction of human retinal cells compared to wildtype AAV. Furthermore, three R100-based gene therapeutics demonstrated safety, delivery, and durable pan-retinal expression of intracellular or secreted transgenes throughout the NHP retina following intravitreal administration. Finally, efficacy of R100 mediated delivery of therapeutic transgenes was demonstrated in patient-derived retinal cells (monogenic diseases) and in an NHP model of pathogenic retinal angiogenesis.

Exploiting Tissue Dielectric Properties to Shape Microwave Thermal Ablation Zones

The dielectric characterization of tissue targets of microwave thermal ablation (MTA) have improved the efficacy and pre-procedural planning of treatment. In some clinical scenarios, the tissue target lies at the interface with an external layer of fat. The aim of this work is to investigate the influence of the dielectric contrast between fat and target tissue on the shape and size of the ablation zone. A 2.45 GHz monopole antenna is placed parallel to an interface modelled by fat and a tissue characterized by higher dielectric properties and powered at 30 and 60 W for 60 s. The performances of MTA are numerically investigated considering different interface scenarios (i.e., different widths of fat layer, shifts in the antenna alignment) and a homogeneous reference scenario. Experiments (N = 10) are conducted on ex vivo porcine tissue to validate the numerical results. Asymmetric heating patterns are obtained in the interface scenario, the ablation zone in the target tissue is two-fold to ten-fold the size of the zone in the adipose tissue, and up to four times larger than the homogenous scenario. The adipose tissue reflects the electromagnetic energy into the adjacent tissue target, reducing the heating in the opposite direction.

Characterisation of Ex Vivo Liver Thermal Properties for Electromagnetic-Based Hyperthermic Therapies

Electromagnetic-based hyperthermic therapies induce a controlled increase of temperature in a specific tissue target in order to increase the tissue perfusion or metabolism, or even to induce cell necrosis. These therapies require accurate knowledge of dielectric and thermal properties to optimise treatment plans. While dielectric properties have been well investigated, only a few studies have been conducted with the aim of understanding the changes of thermal properties as a function of temperature; i.e., thermal conductivity, volumetric heat capacity and thermal diffusivity. In this study, we experimentally investigate the thermal properties of ex vivo ovine liver in the hyperthermic temperature range, from 25 °C to 97 °C. A significant increase in thermal properties is observed only above 90 °C. An analytical model is developed to model the thermal properties as a function of temperature. Thermal properties are also investigated during the natural cooling of the heated tissue. A reversible phenomenon of the thermal properties is observed; during the cooling, thermal properties followed the same behaviour observed in the heating process. Additionally, tissue density and water content are evaluated at different temperatures. Density does not change with temperature; mass and volume losses change proportionally due to water vaporisation. A 30% water loss was observed above 90 °C.

The Neuroimmune Basis of Acupuncture: Correlation of Cutaneous Mast Cell Distribution with Acupuncture Systems in Human

The hypothesis that cutaneous mast cells (MCs) are responsible for skin phenomena in acupuncture was proposed 40 years ago, but very little is known about the correlation of MC distribution with acupuncture systems in human. The aim of this study is to quantify cutaneous mast cells at different body sites and compare them with the distributions of classical acupuncture points and micro-acupuncture systems. Skin biopsies from dermatological practice were evaluated under microscope with H&E or CD117 stains. Dermal MCs were counted and expressed as MCs per high power field. Densities of classical acupuncture points at different body sites were also calculated and expressed as points per dm2. MC densities at special sites of the body were compared with micro-acupuncture systems. After examining 285 skin biopsies, MC enriched special sites (MESS) were found at peripheral parts of the body and around orifices of body surfaces. Comparative mapping showed that patterns of MC distribution are highly correlated with the distributions of classic acupuncture points in 14 classic acupuncture meridians, with the exception of the trunk areas. Mapping also revealed that all micro-acupuncture systems were established at MESS, including ear, scalp, hand, foot, eye, face, and umbilicus. The conclusion is that the densities of cutaneous MCs are highly correlated with classical acupuncture points and micro-acupuncture systems. These findings provide tissue evidence of neuroimmune basis of acupuncture and suggest that MC is a tissue target for acupuncture stimulation and may serve as a tissue marker for acupuncture points.

Maximizing Visualization of the Needle During Ultrasound Procedures

Safe and successful ultrasound-guided interventional procedures depend on the ability to visualize both anatomical structures of interest and the advancing needle. This chapter describes various strategies for optimizing needle visualization and tip localization during freehand guidance of a needle to a tissue target using ultrasound. Challenges to needle visualization include the poor echogenicity of standard needles at insertion angles steeper than 30–45 degrees to the horizontal, the difficulty of aligning the needle with the ultrasound beam, and manipulation in three dimensions based on two-dimensional visual information. Attention to ergonomics improves probe and needle control and facilitates alignment. Needle echogenicity can be improved by using a shallower angle, where possible, and echogenic needle technology. Hand movements of probe and needle should be kept small and controlled. Indirect cues of needle tip location are also extremely useful and should be utilized routinely.

Once-nightly (QD) dual CYP17-Lyase (L) inhibitor / androgen receptor (AR) antagonist VT-464 in patients with CRPC.

343 Background: VT-464 is an oral, CYP17-L inhibitor and non-clinically an antagonist of the AR and its variants associated with clinical resistance to enzalutamide (ENZ) and abiraterone (AA). The safety, tolerability and initial PD effects (tumor responses, PSA and testosterone (T)-decreases) of QD VT-464 were evaluated in this Phase (Ph) 1/2 study (INO-VT-464-CL-004; NCT02361086). Methods: Chemotherapy-naïve (n = 20) M0 (n = 1) and M1 (n = 19) patients with CRPC prior to (TN, treatment-naïve; n = 9), or after AA (n = 6),ENZ (n = 3) or both (n = 2) were enrolled in sequential dose-cohorts of 600 mg QD with dose titration (DT; 300 mg QD q2w), and 600 and 750 mg QD without DT in 28-day continuous dosing cycles. All doses except the first were given at night with dinner. Patient results through the 750 mg QD cohort are reported (data cut off 17 Aug 2015). Results: Patients received VT-464 without steroids at 600 mg QD+DT (n = 3), 600 mg QD (n = 8) or 750 mg QD (n = 9). Eight patients are on study in cycles 3-12. Most adverse events (AEs) were Grade (Gr) 1 or 2. Fourteen Gr 3 AEs were reported with 7 considered unrelated and 7 ≥ possibly related; 5 of the latter were in the 600 mg QD+DT cohort (bradycardia, hypotension, syncope, hypertension and hyponatremia) with 2 in non-DT cohorts (hypotension, bilateral lower leg weakness). No AEs ≥ Gr 4 were reported. One DLT occurred in the 600 mg QD+DT cohort (syncope). No mineralocorticoid excess syndrome, ACTH response changes, or increased LFTs were observed. Plasma T was below the LOQ (0.03nM) in 67% of patients, with progesterone and cortisol unchanged. Cmax was similar in all cohorts to 450 mg bid (de Bono et al, GU ASCO 2015) but AUC was decreased. All 4 TN patients had PSA declines at 600 mg QD including one 30% and one 50% response; there was one 50% response in the 600 mg QD+DT cohort. Both TN patients treated at 750 mg QD had > 50% response. Of 8 patients with ≥ 1 soft tissue target lesion and ≥ 1 post-baseline scan, 5 had stable disease and 1 had a partial response at Cycle 5. Conclusions: The VT-464 750 mg QD regimen demonstrated potent and selective CYP17 L inhibition and has been selected for all Ph 2 clinical studies based upon improved tolerability, PK and efficacy compared to bid+DT dosing (de Bono et al, GU ASCO 2015). Clinical trial information: NCT02361086.

Development of targeted drug delivery system: synthesis of conjugates of address fragment (ra-cooh) with ligand (r-nh2)

One of the main ways to increase the effectiveness of well-known medical formulations well-established in clinical medicine – development of delivery systems using new technological approaches and nanomaterials. Currently, much attention is given to targeted delivery systems. At the same time drug carrier has in addition to medication the so-called vector/address with a high affinity for binding to specific receptors on cells/tissue target. In this paper it is described the method for producing of address conjugates to over-expressed receptors on the tumor cells. As address fragment it was folic acid and as a linker was dodecylamine, causing inclusion the conjugate into lipid nanoparticles.