Design and characterization of a prototype tertiary device for proton beam stereotactic radiosurgery.

Though potentially beneficial, proton beam stereotactic radiosurgery has not been adopted widely secondary to the technical challenge of safely delivering multiple focused beams of proton radiation. In this study, we describe the design and characterization of a proton beam stereotactic radiosurgery system that can be adopted by existing passive scattering systems. This system utilizes a helmet-like device in which patient-specific brass apertures required for final beam collimation are positioned on a scaffold that is separate from the treatment gantry. The proton snout is then fitted with a generic aperture to focus the primary proton beam onto the patient specific apertures that are in the helmet-like device. The patient-specific apertures can all be placed at the start of the treatment, thus treatment with multiple beams can be accomplished without the delay of switching the apertures. In this report we describe a prototype design of this collimation system and dosimetric testing to verify efficacy. Subsequently, we describe a custom 3D printing of a prototype device and report on overall localization accuracy using Winston-Lutz tests. Our results show that it is possible to develop an add-on device for proton beam radiosurgery that is safe and efficient and capable of wide adoption on existing proton delivery systems.

Proton vs Hyperarc™ radiosurgery: A planning comparison.

For many patients, stereotactic radiosurgery (SRS) offers a minimally invasive, curative option when surgical techniques are not possible. To date, the literature supporting the efficacy and safety of SRS treatment techniques uses photon beams. However, with the number of proton therapy facilities exponentially growing and the favorable physical properties of proton beam radiation therapy, there is an opportunity to develop proton therapy techniques for SRS. The goal of this paper is to determine the ability of clinical proton treatment planning systems to model small field dosimetry accurately and to compare various planning metrics used to evaluate photon SRS to determine the optimum beam configurations and settings for proton SRS (PSRS) treatment plans. Once established, these plan settings were used to perform a planning comparison on a variety of different SRS cases and compare SRS metrics between the PSRS plans and HyperArc™ (VMAT) SRS plans.

The prospective Hemophilia Inhibitor PUP Study reveals distinct antibody signatures prior to FVIII inhibitor development.

Preventing factor VIII (FVIII) inhibitors following replacement therapies with FVIII products in patients with hemophilia A remains an unmet medical need. Better understanding of the early events of evolving FVIII inhibitors is essential for risk identification and the design of novel strategies to prevent inhibitor development. The Hemophilia Inhibitor Previously Untreated Patients (PUPs) Study (HIPS; www.clinicaltrials.gov #NCT01652027) is the first prospective cohort study to evaluate comprehensive changes in the immune system during the first 50 exposure days (EDs) to FVIII in patients with severe hemophilia A. HIPS participants were enrolled prior to their first exposure to FVIII or blood products ("true PUPs") and were evaluated for different immunological and clinical parameters at specified time points during their first 50 EDs to a single source of recombinant FVIII. Longitudinal antibody data resulting from this study indicate that there are 4 subgroups of patients expressing distinct signatures of FVIII-binding antibodies. Subgroup 1 did not develop any detectable FVIII-binding immunoglobulin G (IgG) antibodies. Subgroup 2 developed nonneutralizing, FVIII-binding IgG1 antibodies, but other FVIII-binding IgG subclasses were not observed. Subgroup 3 developed transient FVIII inhibitors associated with FVIII-binding IgG1 antibodies, similar to subgroup 2. Subgroup 4 developed persistent FVIII inhibitors associated with an initial development of high-affinity, FVIII-binding IgG1 antibodies, followed by IgG3 and IgG4 antibodies. Appearance of FVIII-binding IgG3 was always associated with persistent FVIII inhibitors and the subsequent development of FVIII-binding IgG4. Some of the antibody signatures identified in HIPS could serve as candidates for early biomarkers of FVIII inhibitor development.

Anti-C1 domain antibodies that accelerate factor VIII clearance contribute to antibody pathogenicity in a murine hemophilia A model.

Essentials Inhibitor formation remains a challenging complication of hemophilia A care. The Bethesda assay is the primary method used for determining bleeding risk and management. Antibodies that block factor VIII binding to von Willebrand factor can increase FVIII clearance. Antibodies that increase clearance contribute to antibody pathogenicity. SUMMARY: Background The development of neutralizing anti-factor VIII (FVIII) antibodies remains a challenging complication of modern hemophilia A care. In vitro assays are the primary method used for quantifying inhibitor titers, predicting bleeding risk, and determining bleeding management. However, other mechanisms of inhibition are not accounted for in these assays, which may result in discrepancies between the inhibitor titer and clinical bleeding symptoms. Objectives To evaluate FVIII clearance in vivo as a potential mechanism for antibody pathogenicity and to determine whether increased FVIII dosing regimens correct the associated bleeding phenotype. Methods FVIII-/- or FVIII-/- /von Willebrand factor (VWF)-/- mice were infused with anti-FVIII mAbs directed against the FVIII C1, C2 or A2 domains, followed by infusion of FVIII. Blood loss via the tail snip bleeding model, FVIII activity and FVIII antigen levels were subsequently measured. Results Pathogenic anti-C1 mAbs that compete with VWF for FVIII binding increased the clearance of FVIII-mAb complexes in FVIII-/- mice but not in FVIII-/- /VWF-/- mice. Additionally, pathogenic anti-C2 mAbs that inhibit FVIII binding to VWF increased FVIII clearance in FVIII-/- mice. Anti-C1, anti-C2 and anti-A2 mAbs that do not inhibit VWF binding did not accelerate FVIII clearance. Infusion of increased doses of FVIII in the presence of anti-C1 mAbs partially corrected blood loss in FVIII-/- mice. Conclusions A subset of antibodies that inhibit VWF binding to FVIII increase the clearance of FVIII-mAb complexes, which contributes to antibody pathogenicity. This may explain differences in the bleeding phenotype observed despite factor replacement in some patients with hemophilia A and low-titer inhibitors.

Innovating immune tolerance induction for haemophilia.

INTRODUCTION: Haemophilia A is an X-linked bleeding disorder characterized by a deficiency of coagulation protein factor VIII (FVIII). A challenging complication of therapeutic FVIII infusions is the formation of neutralizing alloantibodies against the FVIII protein defined as inhibitors. The development of FVIII inhibitors drastically alters the quality of life of the patients and is associated with tremendous increases in morbidity as well as treatment costs. AIM: Current clinical immune tolerance induction protocols to reverse inhibitors are lengthy, costly and not effective in all patients. Prophylactic protocols to prevent inhibitor formation have not yet been developed in the clinical setting. However, there has been ample progress towards this goal in recent years in preclinical studies using animal models of haemophilia. METHODS: Here, we review the mechanisms that lead to inhibitor formation against FVIII and two promising new strategies for antigen-specific tolerance induction. RESULTS: CD4+ T cells play an important role in the FVIII-specific B cell response. Immune tolerance can be induced based on transplacental delivery of FVIII domains fused to Fc or on oral delivery of leaf cells from chloroplast transgenic crop plants. CONCLUSIONS: Recent literature suggests that prophylactic tolerance induction protocols for FVIII may be feasible in haemophilia A patients.

Characterization of a genetically engineered mouse model of hemophilia A with complete deletion of the F8 gene.

UNLABELLED: ESSENTIALS: Anti-factor VIII (FVIII) inhibitory antibody formation is a severe complication in hemophilia A therapy. We genetically engineered and characterized a mouse model with complete deletion of the F8 coding region. F8(TKO) mice exhibit severe hemophilia, express no detectable F8 mRNA, and produce FVIII inhibitors. The defined background and lack of FVIII in F8(TKO) mice will aid in studying FVIII inhibitor formation. BACKGROUND: The most important complication in hemophilia A treatment is the development of inhibitory anti-Factor VIII (FVIII) antibodies in patients after FVIII therapy. Patients with severe hemophilia who express no endogenous FVIII (i.e. cross-reacting material, CRM) have the greatest incidence of inhibitor formation. However, current mouse models of severe hemophilia A produce low levels of truncated FVIII. The lack of a corresponding mouse model hampers the study of inhibitor formation in the complete absence of FVIII protein. OBJECTIVES: We aimed to generate and characterize a novel mouse model of severe hemophilia A (designated the F8(TKO) strain) lacking the complete coding sequence of F8 and any FVIII CRM. METHODS: Mice were created on a C57BL/6 background using Cre-Lox recombination and characterized using in vivo bleeding assays, measurement of FVIII activity by coagulation and chromogenic assays, and anti-FVIII antibody production using ELISA. RESULTS: All F8 exonic coding regions were deleted from the genome and no F8 mRNA was detected in F8(TKO) mice. The bleeding phenotype of F8(TKO) mice was comparable to E16 mice by measurements of factor activity and tail snip assay. Similar levels of anti-FVIII antibody titers after recombinant FVIII injections were observed between F8(TKO) and E16 mice. CONCLUSIONS: We describe a new C57BL/6 mouse model for severe hemophilia A patients lacking CRM. These mice can be directly bred to the many C57BL/6 strains of genetically engineered mice, which is valuable for studying the impact of a wide variety of genes on FVIII inhibitor formation on a defined genetic background.

US Guidelines for immune tolerance induction in patients with haemophilia a and inhibitors.

INTRODUCTION: The development of anti-factor VIII (FVIII) antibodies (inhibitors) is the most serious treatment-related complication in patients with hemophilia A, rendering standard replacement therapy ineffective, heightening the risk for uncontrollable bleeding and morbidity, decreasing quality of life, and increasing healthcare costs. AIM: Formulate evidence-based guidelines for optimizing immune tolerance induction (ITI) in patients with hemophilia A and inhibitors. METHODS: Results from the International ITI study and other available evidence were used to develop guidelines for ITI. RESULTS: Predictors of ITI success were identified and recommendations made for ITI with regard to candidates, timing, product, regimen, monitoring, defining success, concurrent immunomodulation, duration of treatment, and bleed management before and during ITI. CONCLUSION: Evidence-based recommendations to guide treatment decisions may increase the likelihood of successful inhibitor eradication and the induction of FVIII tolerance in patients with hemophilia A who develop inhibitory antibodies.

Expanding the ortholog approach for hemophilia treatment complicated by factor VIII inhibitors.

BACKGROUND: The formation of neutralizing antibodies (inhibitors) directed against human coagulation factor VIII (hFVIII) is a life-threatening pathogenic response that occurs in 20-30% of severe congenital hemophilia A patients and 0.00015% of the remaining population (i.e. acquired hemophilia A). Interspecies amino acid sequence disparity among FVIII orthologs represents a promising strategy to mask FVIII from existing inhibitors while retaining procoagulant function. Evidence for the effectiveness of this approach exists in clinical data obtained for porcine FVIII (pFVIII) products, which have demonstrated efficacy in the setting of congenital and acquired hemophilia. OBJECTIVES: In the current study, recombinant (r) ovine FVIII (oFVIII) was evaluated for antigenicity and procoagulant activity in the context of human patient-derived and murine model-generated FVIII inhibitors. METHODS: The antigenicity of roFVIII was assessed using (i) inhibitor patient plasma samples, (ii) murine anti-FVIII MAbs, (iii) immunized murine hemophilia A plasmas and (iv) an in vivo model of acquired hemophilia A. RESULTS: Overall, roFVIII demonstrated reduced reactivity to, and inhibition by, anti-hFVIII immunoglobulin in patient plasmas. Additionally, several hFVIII epitopes were predicted and empirically shown not to exist within roFVIII. In a murine hemophilia A model designed to mimic clinical inhibitor formation, it was demonstrated that inhibitor titers to roFVIII were significantly reduced when compared with the orthologous immunogens, rhFVIII or rpFVIII. Furthermore, in a murine model of acquired hemophilia A, roFVIII administration conferred protection from bleeding following tail transection. CONCLUSION: These data support the investigation of FVIII orthologs as treatment modalities in both the congenital and acquired FVIII inhibitor settings.

Epitope mapping of inhibitory antibodies targeting the C2 domain of coagulation factor VIII by hydrogen-deuterium exchange mass spectrometry.

BACKGROUND: The development of anti-factor VIII antibodies (inhibitors) is a significant complication in the management of patients with hemophilia A, leading to significant increases in morbidity and treatment cost. Using a panel of mAbs against different epitopes on FVIII, we have recently shown that epitope specificity, inhibitor kinetics and time to maximum inhibition are more important than inhibitor titer in predicting responses to FVIII and the combination of FVIII and recombinant FVIIa. In particular, a subset of high-titer inhibitors responded to high-dose FVIII, which would not be predicted on the basis of their inhibitor titer alone. Thus, the ability to quickly map the epitope spectrum of patient plasma with a clinically feasible assay may fundamentally change how clinicians approach the treatment of high-titer inhibitor patients. OBJECTIVES: To map the epitopes of anti-FVIII mAbs, three of which are classic inhibitors and one of which is a non-classic inhibitor, by the use of hydrogen-deuterium exchange coupled with mass spectrometry (HDX-MS). METHODS: The binding epitopes of four mAbs targeting the FVIII C2 domain were mapped with HDX-MS. RESULTS: The epitopes determined with HDX-MS are consistent with those obtained earlier through structural characterization and antibody competition assays. In addition, classic and non-classic inhibitor epitopes could be distinguished by the use of a limited subset of C2 domain-derived peptic fragments. CONCLUSION: Our results demonstrate the effectiveness and robustness of the HDX-MS method for epitope mapping, and suggest a potential role of rapid mapping of FVIII inhibitor epitopes in facilitating individualized treatment of inhibitor patients.

Late immune tolerance induction in haemophilia A patients.

The incidence of inhibitor development in patients with severe haemophilia A is approximately 30%. Immune tolerance induction (ITI) is commonly utilized to eradicate these antibodies and is successful in 63-100% of cases. Potential predictors of a poor outcome in ITI include a high preinduction titre, high historical peak titre, older age at start of ITI and prolonged interval from diagnosis to start of ITI. The goal of this study was to characterize the outcomes of patients from our centre who have undergone late ITI, many of whom had poor prognostic features. Medical records of patients in our centre with severe/moderately severe haemophilia A (<2% FVIII activity) and history of inhibitor were reviewed. Data were abstracted from all patients who attempted late ITI. Nine patients underwent late ITI between January 1999 and December 2011. Within this cohort, 7 (78%) patients were black, 6 (67%) were <21 years old and 4 (44%) had a family history of inhibitor. Three patients had previously received ITI unsuccessfully. To date, 4 (44%) patients are tolerized (persistently negative inhibitor titre, FVIII recovery >66% and successfully treated with FVIII products ± FVIII t(½) of >6 h). Three patients are partially tolerized (have low responding inhibitor, variable FVIII recovery and successfully treated with FVIII products). Two patients are not tolerized. Some patients with haemophilia A and long-standing inhibitors may benefit from ITI.

Medical radiation exposure in children with bleeding disorders: an institutional experience.

Patients with bleeding disorders may be exposed to ionizing radiation during medical care. We hypothesized that children with severe haemophilia may have higher radiation exposure than those with mild bleeding disorders (MBDs). To compare medical radiation exposure rates between children with severe haemophilia and MBDs. Charts of 35 pediatric patients with severe haemophilia were randomly selected from a database of active male patients followed in our bleeding disorders clinic from 2000 to 2010. Case patients were age and sex matched with two control patients with MBDs [Type 1 von Willebrand disease (VWD) or mild platelet function defect (PFD)]. By retrospective review, data on radiation exposure in millisieverts (mSv) was collected from radiological studies performed within Emory/CHOA. The rates of exposure between cohorts were compared using the Mann-Whitney Test. Case patients had a mean of 11.3 (median 8, IQR = 29) radiographic studies compared with 1.8 (median 1, IQR = 11) for controls (P < 0.001). The mean effective dose of radiation per patient per year of study was two mSv for case patients (median 0.4, IQR = 3) and 0.4 mSv for control patients (median 0.01, IQR = 0.3) (P < 0.001). Overall, 1.4% of controls and 31.4% of cases accumulated high to very high levels of exposure ( > 20 mSv). Case patients with severe hemophilia accumulated significantly more medical radiation exposure than controls. While the use of ionizing radiation is often necessary for management of these patients, avoidance of unnecessary exposure along with exploration of alternative imaging techniques and low dose protocols should be considered whenever possible.

Direct response to proton beam linear energy transfer (LET) in a novel polymer gel dosimeter formulation.

Linear energy transfer (LET) of clinical proton beams is an important parameter influencing the biological effects of radiation. This work demonstrates LET-induced response enhancement in novel formulations of polymer gel dosimeters, potentially useful for LET mapping of clinical proton beams. A series of four polymer gel dosimeters (labeled A through D), prepared based on the BANG3-Pro2 formulation, but with varying concentrations of polymerization modifiers, were irradiated by a clinical proton beam with a spread out Bragg peak modulation (SOBP) and read out using the OCTOPUS-IQ optical CT scanner. The evaluation of optical density profiles in the SOBP (constant physical dose) revealed response deviations at the distal end consistent with variations in gel composition. Maximum response deviations were as follows: -3% (under-response) for gel A, and over-response of 2%, 12%, and 17% for gels B, C, and D, respectively, relative to the mean dose in the center of the SOBP. This enhancement in optical response was correlated to LET by analytical calculations. Gels A and B showed no measurable dependence on LET. Gel C responded linearly in the limited range from 1.5 to 3.5 keV/µm. LET response of gel D was linear up to at least 5.5 keV/µm, with the threshold at about 1.3 keV/µm. These results suggest that it may be possible to develop a polymer gel system with direct optical response to LET for mapping of LET distributions for particle therapy beams.

A multi-GPU real-time dose simulation software framework for lung radiotherapy.

PURPOSE: Medical simulation frameworks facilitate both the preoperative and postoperative analysis of the patient's pathophysical condition. Of particular importance is the simulation of radiation dose delivery for real-time radiotherapy monitoring and retrospective analyses of the patient's treatment. METHODS: In this paper, a software framework tailored for the development of simulation-based real-time radiation dose monitoring medical applications is discussed. A multi-GPU-based computational framework coupled with inter-process communication methods is introduced for simulating the radiation dose delivery on a deformable 3D volumetric lung model and its real-time visualization. The model deformation and the corresponding dose calculation are allocated among the GPUs in a task-specific manner and is performed in a pipelined manner. Radiation dose calculations are computed on two different GPU hardware architectures. The integration of this computational framework with a front-end software layer and back-end patient database repository is also discussed. RESULTS: Real-time simulation of the dose delivered is achieved at once every 120 ms using the proposed framework. With a linear increase in the number of GPU cores, the computational time of the simulation was linearly decreased. The inter-process communication time also improved with an increase in the hardware memory. Variations in the delivered dose and computational speedup for variations in the data dimensions are investigated using D70 and D90 as well as gEUD as metrics for a set of 14 patients. Computational speed-up increased with an increase in the beam dimensions when compared with a CPU-based commercial software while the error in the dose calculation was <1%. CONCLUSION: Our analyses show that the framework applied to deformable lung model-based radiotherapy is an effective tool for performing both real-time and retrospective analyses.

A spatial resolution study of a new optical tomography-based polymer gel dosimetry system.

A spatial resolution investigation of the OCTOPUSTM-IQ scanner in combination with the new BANG3-Pro2(r) polymer gel was performed by scanning a high-contrast needle phantom. The phantom contained five thin needles (0.3_mm diameter) embedded in gel positioned in different patterns: needles were inserted (a) at 45° angle from the center of the gel container, and (b) vertically along the gel axis. The non-irradiated needle phantoms were scanned at various slice spacings (0.25-1.0_mm) and for two different laser beam orientations. Optical density profiles and their full width at half maximum (FWHM) were evaluated for resolution limit. The modulation transfer function (MTF) corresponding to measured point spread function (PSF) data was calculated. With high resolution scanning mode and 0.25_mm pixel resolution, the measured PSFs at the center of the gel dosimeter have a FWHM of 0.95_mm. The MTF for the 0.25_mm reconstruction pixel size suggests that the resolution of the system is 0.5_mm or less. We also observed a progressive degradation of the vertical needle images with off-axis distance, attributable to the defocusing of the laser beam. No significant degradation was observed up to the maximum useful reconstructed image radius of 50_mm from the gel dosimeter center axis.

The effect of temporal HU variations on the uncertainty of dose recalculations performed on MVCT images.

Over the course of radiation therapy, a patient's anatomy may change substantially. The relatively recent addition of frequent in-room imaging to assist with patient localization has provided a database of images that may be used to recalculate dose distributions for adaptive radiotherapy purposes. The TomoTherapy Hi-Art II unit (Accuray Inc., Sunnyvale, CA, USA) uses a helical scanning geometry and a megavoltage (MV) beam to acquire volumetric patient images. This study evaluated the uncertainty of dose calculations performed on megavoltage CT (MVCT) images as a function of temporal Hounsfield Unit (HU) variations observed in the imaging system over three years on two machines. A baseline error between dose calculations performed on kVCT and MVCT images was established using a series of phantoms. This baseline error ranged from -1.4% to 0.6%. Materials of differing densities were imaged and MVCT numbers were measured periodically. The MVCT number of solid water varied from 5 to 103 HU and consistently increased prior to target replacement. Finally, the dosimetric uncertainty of the temporal HU variation was assessed using MVCT images of typical head and neck, lung and prostate cancer patients. Worst-case MVCT recalculation errors could approach 5%, 7% and 10% for the head and neck, lung and prostate images, respectively. However, if a tolerance of ±30 HU were maintained for the MVCT number of solid water, dosimetric errors were limited to ±2.5%, ±3% and ±4%, respectively.

Dosimetric evaluation of a novel polymer gel dosimeter for proton therapy.

PURPOSE: The aim of this study is to evaluate the dosimetric performance of a newly developed proton-sensitive polymer gel formulation for proton therapy dosimetry. METHODS: Using passive scattered modulated and nonmodulated proton beams, the dose response of the gel was assessed. A next-generation optical CT scanner is used as the readout mechanism of the radiation-induced absorbance in the gel medium. Comparison of relative dose profiles in the gel to ion chamber profiles in water is performed. A simple and easily reproducible calibration protocol is established for routine gel batch calibrations. Relative stopping power ratio measurement of the gel medium was performed to ensure accurate water-equivalent depth dose scaling. Measured dose distributions in the gel were compared to treatment planning system for benchmark irradiations and quality of agreement is assessed using clinically relevant gamma index criteria. RESULTS: The dosimetric response of the gel was mapped up to 600 cGy using an electron-based calibration technique. Excellent dosimetric agreement is observed between ion chamber data and gel. The most notable result of this work is the fact that this gel has no observed dose quenching in the Bragg peak region. Quantitative dose distribution comparisons to treatment planning system calculations show that most (> 97%) of the gel dose maps pass the 3%/3 mm gamma criterion. CONCLUSIONS: This study shows that the new proton-sensitive gel dosimeter is capable of reproducing ion chamber dose data for modulated and nonmodulated Bragg peak beams with different clinical beam energies. The findings suggest that the gel dosimeter can be used as QA tool for millimeter range verification of proton beam deliveries in the dosimeter medium.

Non-classical anti-factor VIII C2 domain antibodies are pathogenic in a murine in vivo bleeding model.

OBJECTIVE: The pathogenicity of anti-human factor (F) VIII monoclonal antibodies (MAbs) was tested in a murine bleeding model. METHODS: MAbs were injected into the tail veins of hemophilia A mice to a peak plasma concentration of 60 nm, followed by injection of human B domain-deleted FVIII at 180 U kg(-1), producing peak plasma concentrations of approximately 2 nm. At 2 h, blood loss following a 4-mm tail snip was measured. The following MAbs were tested: (i) 4A4, a type I anti-A2 FVIII inhibitor, (ii) I54 and 1B5, classical type I anti-C2 inhibitors, (iii) 2-77 and B45, non-classical type II anti-C2 inhibitors, and (iv) 2-117, a non-classical anti-C2 MAb with inhibitory activity less than 0.4 Bethesda Units per mg IgG. RESULTS: All MAbs except 2-117 produced similar amounts of blood loss that were significantly greater than control mice injected with FVIII alone. Increasing the dose of FVIII to 360 U kg(-1) overcame the bleeding diathesis produced by the type II MAbs 2-77 and B45, but not the type I antibodies, 4A4, I54, and 1B5. These results were consistent with the in vitro Bethesda assay in which 4A4 completely inhibited both 1 U mL(-1) and 3 U mL(-1) FVIII, while there was 40% residual activity at saturating concentrations of 2-77 at either concentration of FVIII. CONCLUSIONS: For patients with an inhibitor response dominated by non-classical anti-C2 antibodies both the in vivo and in vitro results suggest that treatment with high-dose FVIII rather than bypassing agents may be warranted.