In Silico Docking of Alkylphosphocholine Analogs to Human Serum Albumin Predicts Partitioning and Pharmacokinetics.

Alkylphosphocholine (APC) analogs are a novel class of broad-spectrum tumor-targeting agents that can be used for both diagnosis and treatment of cancer. The potential for clinical translation for APC analogs will strongly depend on their pharmacokinetic (PK) profiles. The aim of this work was to understand how the chemical structures of various APC analogs impact binding and PK. To achieve this aim, we performed in silico docking analysis, in vitro and in vivo partitioning experiments, and in vivo PK studies. Our results have identified 7 potential high-affinity binding sites of these compounds on human serum albumin (HSA) and suggest that the size of the functional group directly influences the albumin binding, partitioning, and PK. Namely, the bulkier the functional groups, the weaker the agent binds to albumin, the more the agent partitions onto lipoproteins, and the less time the agent spends in circulation. The results of these experiments provide novel molecular insights into the binding, partitioning, and PK of this class of compounds and similar molecules as well as suggest pharmacological strategies to alter their PK profiles. Importantly, our methodology may provide a way to design better drugs by better characterizing the PK profile for lead compound optimization.

Analysis of Cancer-Targeting Alkylphosphocholine Analogue Permeability Characteristics Using a Human Induced Pluripotent Stem Cell Blood-Brain Barrier Model.

Cancer-targeting alkylphosphocholine (APC) analogues are being clinically developed for diagnostic imaging, intraoperative visualization, and therapeutic applications. These APC analogues derived from chemically synthesized phospholipid ethers were identified and optimized for cancer-targeting specificity using extensive structure-activity studies. While they strongly label human brain cancers associated with disrupted blood-brain barriers (BBB), APC permeability across intact BBB remains unknown. Three of our APC analogues, CLR1404 (PET radiotracer), CLR1501 (green fluorescence), and CLR1502 (near-infrared fluorescence), were tested for permeability across a BBB model composed of human induced pluripotent stem cell-derived brain microvascular endothelial cells (iPSC-derived BMECs). This in vitro BBB system has reproducibly consistent high barrier integrity marked by high transendothelial electrical resistance (TEER > 1500 Ω-cm(2)) and functional expression of drug efflux transporters. The radioiodinated and fluorescent APC analogues demonstrated fairly low permeability across the iPSC-BMEC (35 ± 5.7 (CLR1404), 54 ± 3.2 (CLR1501), and 26 ± 4.9 (CLR1502) × 10(-5) cm/min) compared with BBB-impermeable sucrose (13 ± 2.5) and BBB-permeable diazepam (170 ± 29). Only the fluorescent APC analogues (CLR1501, CLR1502) underwent BCRP and MRP polarized drug efflux transport in the brain-to-blood direction of the BBB model, and this efflux can be specifically blocked with pharmacological inhibition. None of the tested APC analogues appeared to undergo substantial P-gp transport. Limited permeability of the APC analogues across an intact BBB into normal brain likely contributes to the high tumor to background ratios observed in initial human trials. Moreover, addition of fluorescent moieties to APCs resulted in greater BMEC efflux via MRP and BCRP, and may affect fluorescence-guided applications. Overall, the characterization of APC analogue permeability across human BBB is significant for advancing future brain tumor-targeted applications of these agents.

Is wax equivalent to tissue in electron conformal therapy planning? A Monte Carlo study of material approximation introduced dose difference.

With CT-based Monte Carlo (MC) dose calculations, material composition is often assigned based on the standard Hounsfield unit ranges. This is known as the density threshold method. In bolus electron conformal therapy (BolusECT), the bolus material, machineable wax, would be assigned as soft tissue and the electron density is assumed equivalent to soft tissue based on its Hounsfield unit. This study investigates the dose errors introduced by this material assignment. BEAMnrc was used to simulate electron beams from a Trilogy accelerator. SPRRZnrc was used to calculate stopping power ratios (SPR) of tissue to wax, SPR (tissue) (wax), and tissue to water, SPR(tissue) (water), for 6, 9, 12, 15, and 18 MeV electron beams, of which 12 and 15MeV beams are the most commonly used energies in BolusECT. DOSXYZnrc was applied in dose distribution calculations in a tissue phantom with either flat wax slabs of various thicknesses or a wedge-shaped bolus on top. Dose distribution for two clinical cases, a chest wall and a head and neck, were compared with the bolus material treated as wax or tissue. The SPR(tissue) (wax) values for 12 and 15MeV beams are between 0.935 and 0.945, while the SPR(tissue) (water) values are between 0.990 and 0.991. For a 12 MeV beam, the dose in tissue immediately under the bolus is overestimated by 2.5% for a 3 cm bolus thickness if the wax bolus is treated as tissue. For 15 MeV beams, the error is 1.4%. However, in both clinical cases the differences in the PTV DVH is negligible. Due to stopping power differences, dose differences of up to 2.5% are observed in MC simulations if the bolus material is misassigned as tissue in BolusECT dose calculations. However, for boluses thinner than 2 cm that are more likely encountered in practice, the error is within clinical tolerance.

Next-Generation Cancer Magnetic Resonance Imaging With Tumor-Targeted Alkylphosphocholine Metal Analogs.

OBJECTIVES: In an effort to exploit the elevated need for phospholipids displayed by cancer cells relative to normal cells, we have developed tumor-targeted alkylphosphocholines (APCs) as broad-spectrum cancer imaging and therapy agents. Radioactive APC analogs have exhibited selective uptake and prolonged tumor retention in over 50 cancer types in preclinical models, as well as over 15 cancer types in over a dozen clinical trials. To push the structural limits of this platform, we recently added a chelating moiety capable of binding gadolinium and many other metals for cancer-targeted magnetic resonance imaging (MRI), positron emission tomography imaging, and targeted radionuclide therapy. The aim of this work was to synthesize, characterize, and validate the tumor selectivity of a new broad-spectrum, tumor-targeted, macrocyclic MRI chelate, Gd-NM600, in xenograft and orthotopic tumor models. A secondary aim was to identify and track the in vivo chemical speciation and spatial localization of this new chelate Gd-NM600 in order to assess its Gd deposition properties. MATERIALS AND METHODS: T1 relaxivities of Gd-NM600 were characterized in water and plasma at 1.5 T and 3.0 T. Tumor uptake and subcellular localization studies were performed using transmission electron microscopy. We imaged 8 different preclinical models of human cancer over time and compared the T1-weighted imaging results to that of a commercial macrocyclic Gd chelate, Gd-DOTA. Finally, matrix-assisted laser desorption and ionization-mass spectrometry imaging was used to characterize and map the tissue distribution of the chemical species of Gd-NM600. RESULTS: Gd-NM600 exhibits high T1 relaxivity (approximately 16.4 s-1/mM at 1.5 T), excellent tumor uptake (3.95 %ID/g at 48 hours), prolonged tumor retention (7 days), and MRI conspicuity. Moreover, minimal tumor uptake saturability of Gd-NM600 was observed. Broad-spectrum tumor-specific uptake was demonstrated in 8 different human cancer models. Cancer cell uptake of Gd-NM600 via endosomal internalization and processing was revealed with transmission electron microscopy. Importantly, tissue mass spectrometry imaging successfully interrogated the spatial localization and chemical speciation of Gd compounds and also identified breakdown products of Gd species. CONCLUSIONS: We have introduced a new macrocyclic cancer-targeted Gd chelate that achieves broad-spectrum tumor uptake and prolonged retention. Furthermore, we have demonstrated in vivo stability of Gd-NM600 by ultrahigh resolution MS tissue imaging. A tumor-targeted contrast agent coupled with the enhanced imaging resolution of MRI relative to positron emission tomography may transform oncologic imaging.

Beyond the margins: real-time detection of cancer using targeted fluorophores.

Over the past two decades, synergistic innovations in imaging technology have resulted in a revolution in which a range of biomedical applications are now benefiting from fluorescence imaging. Specifically, advances in fluorophore chemistry and imaging hardware, and the identification of targetable biomarkers have now positioned intraoperative fluorescence as a highly specific real-time detection modality for surgeons in oncology. In particular, the deeper tissue penetration and limited autofluorescence of near-infrared (NIR) fluorescence imaging improves the translational potential of this modality over visible-light fluorescence imaging. Rapid developments in fluorophores with improved characteristics, detection instrumentation, and targeting strategies led to the clinical testing in the early 2010s of the first targeted NIR fluorophores for intraoperative cancer detection. The foundations for the advances that underline this technology continue to be nurtured by the multidisciplinary collaboration of chemists, biologists, engineers, and clinicians. In this Review, we highlight the latest developments in NIR fluorophores, cancer-targeting strategies, and detection instrumentation for intraoperative cancer detection, and consider the unique challenges associated with their effective application in clinical settings.

Diapeutic cancer-targeting alkylphosphocholine analogs may advance management of brain malignancies.

The following is a special report on alkylphosphocholine analogs as targeted imaging and therapy agents for cancer, and their potential role in diagnosis and treatment in glioblastoma and brain metastases. These novel cancer-targeting agents display impressive tumor avidity with low background in the normal brain, and multimodal diagnostic imaging and therapy capabilities. The use of these agents may significantly improve diagnosis, treatment and post-treatment follow-up in patients with brain malignancies.

Fluorescent cancer-selective alkylphosphocholine analogs for intraoperative glioma detection.

BACKGROUND: 5-Aminolevulinic acid (5-ALA)-induced tumor fluorescence aids brain tumor resections but is not approved for routine use in the United States. We developed and describe testing of 2 novel fluorescent, cancer-selective alkylphosphocholine analogs, CLR1501 (green) and CLR1502 (near infrared), in a proof-of-principle study for fluorescence-guided glioma surgery. OBJECTIVE: To demonstrate that CLR1501 and CLR1502 are cancer cell-selective fluorescence agents in glioblastoma models and to compare tumor-to-normal brain (T:N) fluorescence ratios with 5-ALA. METHODS: CLR1501, CLR1502, and 5-ALA were administered to mice with magnetic resonance imaging-verified orthotopic U251 glioblastoma multiforme- and glioblastoma stem cell-derived xenografts. Harvested brains were imaged with confocal microscopy (CLR1501), the IVIS Spectrum imaging system (CLR1501, CLR1502, and 5-ALA), or the Fluobeam near-infrared fluorescence imaging system (CLR1502). Imaging and quantitative analysis of T:N fluorescence ratios were performed. RESULTS: Excitation/emission peaks are 500/517 nm for CLR1501 and 760/778 nm for CLR1502. The observed T:N ratio for CLR1502 (9.28±1.08) was significantly higher (P<.01) than for CLR1501 (3.51±0.44 on confocal imaging; 7.23±1.63 on IVIS imaging) and 5-ALA (4.81±0.92). Near-infrared Fluobeam CLR1502 imaging in a mouse xenograft model demonstrated high- contrast tumor visualization compatible with surgical applications. CONCLUSION: CLR1501 (green) and CLR1502 (near infrared) are novel tumor-selective fluorescent agents for discriminating tumor from normal brain. CLR1501 exhibits a tumor-to-brain fluorescence ratio similar to that of 5-ALA, whereas CLR1502 has a superior tumor-to-brain fluorescence ratio. This study demonstrates the potential use of CLR1501 and CLR1502 in fluorescence-guided tumor surgery.

Monte Carlo comparison of superficial dose between flattening filter free and flattened beams.

This study investigates the superficial dose from FFF beams in comparison with the conventional flattened ones using a Monte Carlo (MC) method. Published phase-space files which incorporated real geometry of a TrueBeam accelerator were used for the dose calculation in phantom and clinical cases. The photon fluence on the central axis is 3 times that of a flattened beam for a 6 MV FFF beam and 5 times for a 10 MV beam. The mean energy across the field in air at the phantom surface is 0.92-0.95 MeV for the 6 MV FFF beam and 1.18-1.30 MeV for the corresponding flattened beam. At 10 MV, the values are 1.52-1.72 and 2.15-2.87 MeV for the FFF and flattened beams, respectively. The phantom dose at the depth of 1 mm in the 6 MV FFF beam is 6% ± 2.5% (of the maximum dose) higher compared to the flattened beam for a 25 × 25 cm(2) field and 14.6% ± 1.9% for the 2 × 2 cm(2) field. For the 10 MV beam, the corresponding differences are 3.4% ± 1.5% and 10.7% ± 0.6%. The skin dose difference at selected points on the patient's surface between the plans using FFF and flattened beams in the head-and-neck case was 6.5% ± 2.3% (1SD), and for the breast case it was 6.4% ± 2.3%. The Monte Carlo simulations showed that due to the lower mean energy in the FFF beam, the clinical superficial dose is higher without the flattening filter compared to the flattened beam.

Dosimetric analysis of modulated and hybrid arcs in stereotactic radiosurgery.

PURPOSE: Forward-planned conformal arcs (termed dynamic arcs in the BrainLab iPlan system) have been routinely used in cranial stereotactic radiosurgery (SRS) for many yeras. This study compares dosimetric parameters of the newer, inversely-planned volumetric modulated arc therapy (VMAT) and VMAT-conformal hybrid plans with conformal arc plans. In the hybrid plans, various numbers of conformal arcs in a conformal plan are replaced with VMAT arcs. METHODS: Ten brain cancer cases previously treated on a Novalis accelerator for frameless cranial stereotactic radiosurgery using conformal arcs generated by BrainLab iPlan treatment planning system were retrospectively studied with various numbers of VMAT arcs replacing the conformal arcs. Pure VMAT plans of different numbers of arcs were also generated using the same angle and arc length setup for all or part of the conformal arcs and compared with both the original and hybrid plans. Pinnacle version 9.0 was used for treatment plan generation. SmartArc was used for the VMAT planning for a Novalis accelerator. The conformity index (CI), defined as the ratio of the isodose volume to the isodose covered target volume, gradient index (GI), defined as the ratio of the 50% isodose volume (V50) to the 100% isodose volume, volume covered by 12 Gy isodose (V12), mean dose and standard deviation of dose in target were studied. RESULTS: With one of the conformal arcs replaced with a VMAT arc of the same weighting and geometric setup, the average CI with one standard deviation was improved from 1.35 ± 0.18 to 1.29 ± 0.15 (p-value 0.03), while the average GI degraded from 3.8 ± 1.0 to 4.9 ± 1.5 (p-value < 0.01). The degraded GI in the VMAT plans is due to the absence of beam margin limit in SmartArc planning. Pure VMAT plans usually demonstrated better CI values than the hybrid and conformal arc plans when the number of arcs was greater than 2. The GI value was improved with increasing number of arcs in VMAT plans. For target volumes greater than 1 cc, VMAT plans demonstrated improved CI and smaller V50 and V12 than that in conformal arc plans. Better dose fall-off in normal tissue in VMAT plans is accompanied with higher dose heterogeneity in the target volume. CONCLUSION: VMAT and conformal-VMAT hybrid plans usually offer better target conformity. The dose fall-off in normal tissue is also better in VMAT plans when the number of arcs is greater than 3 and target volume is greater than 1 cc, but compromise the dose homogeneity in target volumes. VMAT plans are the best option for targets that are greater than 1 cc for cranial radiosurgery treatments, while for very small targets, conformal arc technique may still be the better choice based on data of dose fall-off in normal tissues.

Volumetric modulated arc planning for lung stereotactic body radiotherapy using conventional and unflattened photon beams: a dosimetric comparison with 3D technique.

PURPOSE: Frequently, three-dimensional (3D) conformal beams are used in lung cancer stereotactic body radiotherapy (SBRT). Recently, volumetric modulated arc therapy (VMAT) was introduced as a new treatment modality. VMAT techniques shorten delivery time, reducing the possibility of intrafraction target motion. However dose distributions can be quite different from standard 3D therapy. This study quantifies those differences, with focus on VMAT plans using unflattened photon beams. METHODS: A total of 15 lung cancer patients previously treated with 3D or VMAT SBRT were randomly selected. For each patient, non-coplanar 3D, coplanar and non-coplanar VMAT and flattening filter free VMAT (FFF-VMAT) plans were generated to meet the same objectives with 50 Gy covering 95% of the PTV. Two dynamic arcs were used in each VMAT plan. The couch was set at ± 5° to the 0° straight position for the two non-coplanar arcs. Pinnacle version 9.0 (Philips Radiation Oncology, Fitchburg WI) treatment planning system with VMAT capabilities was used. We analyzed the conformity index (CI), which is the ratio of the total volume receiving at least the prescription dose to the target volume receiving at least the prescription dose; the conformity number (CN) which is the ratio of the target coverage to CI; and the gradient index (GI) which is the ratio of the volume of 50% of the prescription isodose to the volume of the prescription isodose; as well as the V20, V5, and mean lung dose (MLD). Paired non-parametric analysis of variance tests with post-tests were performed to examine the statistical significance of the differences of the dosimetric indices. RESULTS: Dosimetric indices CI, CN and MLD all show statistically significant improvement for all studied VMAT techniques compared with 3D plans (p < 0.05). V5 and V20 show statistically significant improvement for the FFF-VMAT plans compared with 3D (p < 0.001). GI is improved for the FFF-VMAT and the non-coplanar VMAT plans (p < 0.01 and p < 0.05 respectively) while the coplanar VMAT plans do not show significant difference compared to 3D plans. Dose to the target is typically more homogeneous in FFF-VMAT plans. FFF-VMAT plans require more monitor units than 3D or non-coplanar VMAT ones. CONCLUSION: Besides the advantage of faster delivery times, VMAT plans demonstrated better conformity to target, sharper dose fall-off in normal tissues and lower dose to normal lung than the 3D plans for lung SBRT. More monitor units are often required for FFF-VMAT plans.