Jonathan P. Dyke

Treatment of Late Infantile Neuronal Ceroid Lipofuscinosis by CNS Administration of a Serotype 2 Adeno-Associated Virus Expressing CLN2 cDNA

Late infantile neuronal ceroid lipofuscinosis (LINCL) is an autosomal recessive, neurodegenerative lysosomal storage disease affecting the CNS and is fatal by age 8 to 12 years. A total average dose of 2.5 10(12) particle units of an adeno-associated virus (AAV) serotype 2 vector expressing the human CLN2 cDNA (AAV2 CU h-CLN2) was administered to 12 locations in the CNS of 10 children with LINCL. In addition to safety parameters, a neurological rating scale (primary variable) and three quantitative magnetic resonance imaging (MRI) parameters (secondary variables) were used to compare the rate of neurological decline for 18 months in treated subjects compared with untreated subjects. Although there were no unexpected serious adverse events that were unequivocally attributable to the AAV2 CU hCLN2 vector, there were serious adverse effects, the etiology of which could not be determined under the conditions of the experiment. One subject died 49 days postsurgery after developing status epilepticus on day 14, but with no evidence of CNS inflammation. Four of the 10 subjects developed a mild, mostly transient, humoral response to the vector. Compared with control subjects, the measured rates of decline of all MRI parameters were slower, albeit the numbers were too small for statistical significance. Importantly, assessment of the neurologic rating scale, which was the primary outcome variable, demonstrated a significantly reduced rate of decline compared with control subjects. Although the trial is not matched, randomized, or blinded and lacked a contemporaneous placebo/sham control group, assessment of the primary outcome variable suggests a slowing of progression of LINCL in the treated children. On this basis, we propose that additional studies to assess the safety and efficacy of AAV-mediated gene therapy for LINCL are warranted.

Possible axonal regrowth in late recovery from the minimally conscious state

We used diffusion tensor imaging (DTI) to study 2 patients with traumatic brain injury. The first patient recovered reliable expressive language after 19 years in a minimally conscious state (MCS); the second had remained in MCS for 6 years. Comparison of white matter integrity in the patients and 20 normal subjects using histograms of apparent diffusion constants and diffusion anisotropy identified widespread altered diffusivity and decreased anisotropy in the damaged white matter. These findings remained unchanged over an 18-month interval between 2 studies in the first patient. In addition, in this patient, we identified large, bilateral regions of posterior white matter with significantly increased anisotropy that reduced over 18 months. In contrast, notable increases in anisotropy within the midline cerebellar white matter in the second study correlated with marked clinical improvements in motor functions. This finding was further correlated with an increase in resting metabolism measured by PET in this subregion. Aberrant white matter structures were evident in the second patients DTI images but were not clinically correlated. We propose that axonal regrowth may underlie these findings and provide a biological mechanism for late recovery. Our results are discussed in the context of recent experimental studies that support this inference.

Dissociations between behavioural and functional magnetic resonance imaging-based evaluations of cognitive function after brain injury

Functional neuroimaging methods hold promise for the identification of cognitive function and communication capacity in some severely brain-injured patients who may not retain sufficient motor function to demonstrate their abilities. We studied seven severely brain-injured patients and a control group of 14 subjects using a novel hierarchical functional magnetic resonance imaging assessment utilizing mental imagery responses. Whereas the control group showed consistent and accurate (for communication) blood-oxygen-level-dependent responses without exception, the brain-injured subjects showed a wide variation in the correlation of blood-oxygen-level-dependent responses and overt behavioural responses. Specifically, the brain-injured subjects dissociated bedside and functional magnetic resonance imaging-based command following and communication capabilities. These observations reveal significant challenges in developing validated functional magnetic resonance imaging-based methods for clinical use and raise interesting questions about underlying brain function assayed using these methods in brain-injured subjects.

Treatment planning for prostate implants using magnetic-resonance spectroscopy imaging

PURPOSE Recent studies have demonstrated that magnetic-resonance spectroscopic imaging (MRSI) of the prostate may effectively distinguish between regions of cancer and normal prostatic epithelium. This diagnostic imaging tool takes advantage of the increased choline plus creatine versus citrate ratio found in malignant compared to normal prostate tissue. The purpose of this study is to describe a novel brachytherapy treatment-planning optimization module using an integer programming technique that will utilize biologic-based optimization. A method is described that registers MRSI to intraoperative-obtained ultrasound images and incorporates this information into a treatment-planning system to achieve dose escalation to intraprostatic tumor deposits. METHODS MRSI was obtained for a patient with Gleason 7 clinically localized prostate cancer. The ratios of choline plus creatine to citrate for the prostate were analyzed, and regions of high risk for malignant cells were identified. The ratios representing peaks on the MR spectrum were calculated on a spatial grid covering the prostate tissue. A procedure for mapping points of interest from the MRSI to the ultrasound images is described. An integer-programming technique is described as an optimization module to determine optimal seed distribution for permanent interstitial implantation. MRSI data are incorporated into the treatment-planning system to test the feasibility of dose escalation to positive voxels with relative sparing of surrounding normal tissues. The resultant tumor control probability (TCP) is estimated and compared to TCP for standard brachytherapy-planned implantation. RESULTS The proposed brachytherapy treatment-planning system is able to achieve a minimum dose of 120% of the 144 Gy prescription to the MRS positive voxels using (125)I seeds. The preset dose bounds of 100-150% to the prostate and 100-120% to the urethra were maintained. When compared to a standard plan without MRS-guided optimization, the estimated TCP for the MRS-optimized plan is superior. The enhanced TCP was more pronounced for smaller volumes of intraprostatic tumor deposits compared to estimated TCP values for larger lesions. CONCLUSIONS Using this brachytherapy-optimization system, we could demonstrate the feasibility of MRS-optimized dose distributions for (125)I permanent prostate implants. Based on probability estimates of anticipated improved TCP, this approach may have an impact on the ability to safely escalate dose and potentially improve outcome for patients with organ-confined but aggressive prostatic cancers. The magnitude of the TCP enhancement, and therefore the risks of ignoring the MR data, appear to be more substantial when the tumor is well localized; however, the gain achievable in TCP may depend quite considerably on the MRS tumor-detection efficiency.

The Medial Prefrontal Cortex and the Emergence of Self-Conscious Emotion in Adolescence

In the present study, we examined the relationship between developmental modulation of socioaffective brain systems and adolescents’ preoccupation with social evaluation. Child, adolescent, and adult participants viewed cues indicating that a camera was alternately off, warming up, or projecting their image to a peer during the acquisition of behavioral-, autonomic-, and neural-response (functional MRI) data. Believing that a peer was actively watching them was sufficient to induce self-conscious emotion that rose in magnitude from childhood to adolescence and partially subsided into adulthood. Autonomic arousal was uniquely heightened in adolescents. These behavioral patterns were paralleled by emergent engagement of the medial prefrontal cortex (MPFC) and striatum-MPFC connectivity during adolescence, which are thought to promote motivated social behavior in adolescence. These findings demonstrate that adolescents’ self-consciousness is related to age-dependent sensitivity of brain systems critical to socioaffective processes. Further, unique interactions between the MPFC and striatum may provide a mechanism by which social-evaluation contexts influence adolescent behavior.

Quantitative Assessment of the Vascularity of the Proximal Part of the Humerus

BACKGROUND The current consensus in the literature is that the anterolateral branch of the anterior humeral circumflex artery provides the main blood supply to the humeral head. While the artery is disrupted in association with 80% of proximal humeral fractures, resultant osteonecrosis is infrequent. This inconsistency suggests a greater role for the posterior humeral circumflex artery than has been previously described. We hypothesized that the posterior humeral circumflex artery provides a greater percentage of perfusion to the humeral head than the anterior humeral circumflex artery does. METHODS In twenty-four fresh-frozen cadaver shoulders (twelve matched pairs), we cannulated the axillary artery proximal to the thoracoacromial branch and ligated the brachial artery in the forearm. In each pair, one shoulder served as a control with intact vasculature and, in the contralateral shoulder, either the anterior humeral circumflex artery or the posterior humeral circumflex artery was ligated. Gadolinium was injected through the cannulated axillary arteries, and magnetic resonance imaging was performed. After imaging, a urethane polymer was injected, and specimens were dissected. For volumetric analysis, the gadolinium uptake on the magnetic resonance imaging was quantified in each quadrant of the humeral head with use of a custom automated program. The gadolinium uptake was compared between the control and ligated sides and between the ligated anterior humeral circumflex artery and ligated posterior humeral circumflex artery groups. RESULTS The posterior humeral circumflex artery provided 64% of the blood supply to the humeral head overall, whereas the anterior humeral circumflex artery supplied 36%. The posterior humeral circumflex artery also provided significantly more of the blood supply in three of the four quadrants of the humeral head. CONCLUSIONS The finding that the posterior humeral circumflex artery provides 64% of the blood supply to the humeral head provides a possible explanation for the relatively low rates of osteonecrosis seen in association with displaced fractures of the proximal part of the humerus. In addition, protecting the posterior humeral circumflex artery during the surgical approach and fracture fixation may minimize loss of the blood supply to the humeral head.

Increased ventricular lactate in chronic fatigue syndrome. III. Relationships to cortical glutathione and clinical symptoms implicate oxidative stress in disorder pathophysiology

Chronic fatigue syndrome (CFS) is a complex illness, which is often misdiagnosed as a psychiatric illness. In two previous reports, using 1H MRSI, we found significantly higher levels of ventricular cerebrospinal fluid (CSF) lactate in patients with CFS relative to those with generalized anxiety disorder and healthy volunteers (HV), but not relative to those with major depressive disorder (MDD). In this third independent cross‐sectional neuroimaging study, we investigated a pathophysiological model which postulated that elevations of CSF lactate in patients with CFS might be caused by increased oxidative stress, cerebral hypoperfusion and/or secondary mitochondrial dysfunction. Fifteen patients with CFS, 15 with MDD and 13 HVs were studied using the following modalities: (i) 1H MRSI to measure CSF lactate; (ii) single‐voxel 1H MRS to measure levels of cortical glutathione (GSH) as a marker of antioxidant capacity; (iii) arterial spin labeling (ASL) MRI to measure regional cerebral blood flow (rCBF); and (iv) 31P MRSI to measure brain high‐energy phosphates as objective indices of mitochondrial dysfunction. We found elevated ventricular lactate and decreased GSH in patients with CFS and MDD relative to HVs. GSH did not differ significantly between the two patient groups. In addition, we found lower rCBF in the left anterior cingulate cortex and the right lingual gyrus in patients with CFS relative to HVs, but rCBF did not differ between those with CFS and MDD. We found no differences between the three groups in terms of any high‐energy phosphate metabolites. In exploratory correlation analyses, we found that levels of ventricular lactate and cortical GSH were inversely correlated, and significantly associated with several key indices of physical health and disability. Collectively, the results of this third independent study support a pathophysiological model of CFS in which increased oxidative stress may play a key role in CFS etiopathophysiology. Copyright

Imaging therapeutic response in human bone marrow using rapid whole-body MRI†

Whole‐body imaging of therapeutic response in human bone marrow was achieved without introduced contrast agents using diffusion‐weighted echo‐planar magnetic resonance imaging of physiologic water. Bone marrow disease was identified relative to the strong overlying signals from water and lipids in other anatomy through selective excitation of the water resonance and generation of image contrast that was dependent upon differential nuclear relaxation times and self‐diffusion coefficients. Three‐dimensional displays were generated to aid image interpretation. The geometric distortion inherent in echo‐planar imaging techniques was minimized through the acquisition of multiple axial slices at up to 12 anatomic stations over the entire body. Examples presented include the evaluation of therapeutic response in bone marrow during cytotoxic therapy for leukemia and metastatic prostate cancer and during cytokine administration for marrow mobilization prior to stem cell harvest. Magn Reson Med 52:1234–1238, 2004.

Long-Term Expression and Safety of Administration of AAVrh.10hCLN2 to the Brain of Rats and Nonhuman Primates for the Treatment of Late Infantile Neuronal Ceroid Lipofuscinosis

Late infantile neuronal ceroid lipofuscinosis (LINCL), a fatal, lysosomal storage disorder caused by mutations in the CLN2 gene, results in a deficiency of tripeptidyl-peptidase I (TPP-I) activity in neurons. Our prior studies showed that delivery of the human CLN2 cDNA directly to the CNS, using an adeno-associated virus serotype 2 (AAV2) vector, is safe in children with LINCL. As a second-generation strategy, we have demonstrated that AAVrh.10hCLN2, a rhesus-derived AAV vector, mediates wide distribution of TPP-I through the CNS in a murine model. This study tests the hypothesis that direct administration of AAVrh.10hCLN2 to the CNS of rats and nonhuman primates at doses scalable to humans has an acceptable safety profile and mediates significant CLN2 expression in the CNS. A dose of 10(11) genome copies (GC) was administered bilaterally to the striatum of Sprague Dawley rats with sacrifice at 7 and 90 days with no significant impact except for mild vector-related histopathological changes at the site of vector administration. A dose of 1.8×10(12) GC of AAVrh.10hCLN2 was administered to the CNS of 8 African green monkeys. The vector-treated monkeys did not differ from controls in any safety parameter except for mild to moderate white matter edema and inflammation localized to the administration sites of the vector. There were no clinical sequelae to these localized findings. TPP-I activity was >2 SD over background in 31.7±8.1% of brain at 90 days. These findings establish the dose and safety profile for human clinical studies for the treatment of LINCL with AAVrh.10hCLN2.

Perfusion abnormalities in subchondral bone associated with marrow edema, osteoarthritis, and avascular necrosis

Abstract:  Bone marrow edema is seen in osteoarthritis, avascular necrosis, and other clinical conditions including the bone marrow edema syndrome. Bone marrow edema is associated with bone pain and may be related to the pathophysiology of osteoarthritis. Our hypothesis is that bone marrow edema is associated with a reduction in perfusion in subchondral bone, which contributes to focal and segmental bone necrosis and cartilage breakdown. We further hypothesize that altered fluid dynamics in subchondral bone comprise part of the physicochemical environment to which osteocytes are highly sensitive and alter their cytokine expression profile in response to changes in fluid flow, pressure, and oxygen gradients. We have used contrast‐enhanced magnetic resonance imaging with Gd‐DTPA to characterize changes in subchondral bone perfusion in two relevant and related models—the Dunkin–Hartley guinea pig model of osteoarthritis and human bone marrow edema associated with osteoarthritis and avascular necrosis. Pharmacokinetic modeling was used to extract dynamic parameters of perfusion. Representative time‐intensity curves are derived, which characterize normal bone and bone with marrow edema. Dynamic contrast‐enhanced magnetic resonance imaging may be a useful tool for the early diagnosis of bone perfusion abnormalities and may be used to characterize marrow edema associated with a number of clinical conditions. This technique may also shed light on the pathophysiology of subchondral perfusion in osteoarthritis and avascular necrosis.