Gene therapy for aromatic L-amino acid decarboxylase deficiency: Requirements for safe application and knowledge-generating follow-up.

The autosomal recessive defect of aromatic L-amino acid decarboxylase (AADC) leads to a severe neurological disorder with manifestation in infancy due to a pronounced, combined deficiency of dopamine, serotonin and catecholamines. The success of conventional drug treatment is very limited, especially in patients with a severe phenotype. The development of an intracerebral AAV2-based gene delivery targeting the putamen or substantia nigra started more than 10 years ago. Recently, the putaminally-delivered construct, Eladocagene exuparvovec has been approved by the European Medicines Agency and by the British Medicines and Healthcare products Regulatory Agency. This now available gene therapy provides for the first time also for AADC deficiency (AADCD) a causal therapy, leading this disorder into a new therapeutic era. By using a standardized Delphi approach members of the International Working Group on Neurotransmitter related Disorders (iNTD) developed structural requirements and recommendations for the preparation, management and follow-up of AADC deficiency patients who undergo gene therapy. This statement underlines the necessity of a framework for a quality-assured application of AADCD gene therapy including Eladocagene exuparvovec. Treatment requires prehospital, inpatient and posthospital care by a multidisciplinary team in a specialized and qualified therapy center. Due to lack of data on long-term outcomes and the comparative efficacy of alternative stereotactic procedures and brain target sites, a structured follow-up plan and systematic documentation of outcomes in a suitable, industry-independent registry study are necessary.

Prevalence of DDC genotypes in patients with aromatic L-amino acid decarboxylase (AADC) deficiency and in silico prediction of structural protein changes.

Aromatic L-amino acid decarboxylase (AADC) deficiency is a rare autosomal recessive genetic disorder affecting the biosynthesis of dopamine, a precursor of both norepinephrine and epinephrine, and serotonin. Diagnosis is based on the analysis of CSF or plasma metabolites, AADC activity in plasma and genetic testing for variants in the DDC gene. The exact prevalence of AADC deficiency, the number of patients, and the variant and genotype prevalence are not known. Here, we present the DDC variant (n = 143) and genotype (n = 151) prevalence of 348 patients with AADC deficiency, 121 of whom were previously not reported. In addition, we report 26 new DDC variants, classify them according to the ACMG/AMP/ACGS recommendations for pathogenicity and score them based on the predicted structural effect. The splice variant c.714+4A>T, with a founder effect in Taiwan and China, was the most common variant (allele frequency = 32.4%), and c.[714+4A>T];[714+4A>T] was the most common genotype (genotype frequency = 21.3%). Approximately 90% of genotypes had variants classified as pathogenic or likely pathogenic, while 7% had one VUS allele and 3% had two VUS alleles. Only one benign variant was reported. Homozygous and compound heterozygous genotypes were interpreted in terms of AADC protein and categorized as: i) devoid of full-length AADC, ii) bearing one type of AADC homodimeric variant or iii) producing an AADC protein population composed of two homodimeric and one heterodimeric variant. Based on structural features, a score was attributed for all homodimers, and a tentative prediction was advanced for the heterodimer. Almost all AADC protein variants were pathogenic or likely pathogenic.

AADC deficiency from infancy to adulthood: Symptoms and developmental outcome in an international cohort of 63 patients.

Aromatic l-amino acid decarboxylase deficiency (AADCD) is a rare, autosomal recessive neurodevelopmental disorder characterized by impaired synthesis of dopamine, noradrenaline, adrenaline and serotonin, leading to a complex syndrome of motor, behavioral, and autonomic symptoms. This retrospective study assessed the symptoms and developmental outcome of a large international cohort of patients with AADCD via physician and/or caregiver responses to a detailed, standardized questionnaire. Sixty-three patients (60% female; ages 6 months-36 years, median 7 years; 58 living) from 23 individual countries participated. Common symptoms at onset (median age 3 months, range 0-12 months) were hypotonia, developmental delay, and/or oculogyric crises. Oculogyric crises were present in 97% of patients aged 2 to 12 years, occurred in the majority of patients in all age groups, and tended to be most severe during early childhood. Prominent non-motor symptoms were sleep disturbance, irritable mood, and feeding difficulties. The majority of subjects (70%) had profound motor impairment characterized by absent head control and minimal voluntary movement, while 17% had mild motor impairment and were able to walk independently. Dopamine agonists were the medications most likely to produce some symptomatic benefit, but were associated with dose-limiting side effects (dyskinesia, insomnia, irritability, vomiting) that led to discontinuation 25% of the time. The age distribution of our cohort (70% of subjects under age 13 years) and the observation of a greater proportion of patients with a more severe disease phenotype in the younger compared to the older patients, both suggest a significant mortality risk during childhood for patients with severe disease.

Consensus guideline for the diagnosis and treatment of aromatic l-amino acid decarboxylase (AADC) deficiency.

Aromatic L-amino acid decarboxylase deficiency (AADCD) is a rare, autosomal recessive neurometabolic disorder that leads to a severe combined deficiency of serotonin, dopamine, norepinephrine and epinephrine. Onset is early in life, and key clinical symptoms are hypotonia, movement disorders (oculogyric crisis, dystonia, and hypokinesia), developmental delay, and autonomic symptoms.In this consensus guideline, representatives of the International Working Group on Neurotransmitter Related Disorders (iNTD) and patient representatives evaluated all available evidence for diagnosis and treatment of AADCD and made recommendations using SIGN and GRADE methodology. In the face of limited definitive evidence, we constructed practical recommendations on clinical diagnosis, laboratory diagnosis, imaging and electroencephalograpy, medical treatments and non-medical treatments. Furthermore, we identified topics for further research. We believe this guideline will improve the care for AADCD patients around the world whilst promoting general awareness of this rare disease.

'Letting them loose!' A different approach to a biochemistry post-exam training session.

This feature looks at changing the format of a well-established information skills session for first-year biochemistry students at UCL, to improve its design, delivery and the overall learning experience for the students. After reviewing current literature, active learning methods were explored and an idea of a research-based quiz based on famous scientists was devised. After a brief introduction to the library resources available, the students were 'let loose' to research their particular scientist and answer a series of questions. Feedback was gained and evaluated and concluded that the session was well received by students. The feature also raises the discussion of generic verses subject-based information skills training. H.S.

An experimental paradigm for the prediction of Post-Operative Pain (PPOP).

Many women undergo cesarean delivery without problems, however some experience significant pain after cesarean section. Pain is associated with negative short-term and long-term effects on the mother. Prior to women undergoing surgery, can we predict who is at risk for developing significant postoperative pain and potentially prevent or minimize its negative consequences? These are the fundamental questions that a team from the University of Washington, Stanford University, the Catholic University in Brussels, Belgium, Santa Joana Women's Hospital in São Paulo, Brazil, and Rambam Medical Center in Israel is currently evaluating in an international research collaboration. The ultimate goal of this project is to provide optimal pain relief during and after cesarean section by offering individualized anesthetic care to women who appear to be more 'susceptible' to pain after surgery. A significant number of women experience moderate or severe acute post-partum pain after vaginal and cesarean deliveries. (1) Furthermore, 10-15% of women suffer chronic persistent pain after cesarean section. (2) With constant increase in cesarean rates in the US (3) and the already high rate in Brazil, this is bound to create a significant public health problem. When questioning women's fears and expectations from cesarean section, pain during and after it is their greatest concern. (4) Individual variability in severity of pain after vaginal or operative delivery is influenced by multiple factors including sensitivity to pain, psychological factors, age, and genetics. The unique birth experience leads to unpredictable requirements for analgesics, from 'none at all' to 'very high' doses of pain medication. Pain after cesarean section is an excellent model to study post-operative pain because it is performed on otherwise young and healthy women. Therefore, it is recommended to attenuate the pain during the acute phase because this may lead to chronic pain disorders. The impact of developing persistent pain is immense, since it may impair not only the ability of women to care for their child in the immediate postpartum period, but also their own well being for a long period of time. In a series of projects, an international research network is currently investigating the effect of pregnancy on pain modulation and ways to predict who will suffer acute severe pain and potentially chronic pain, by using simple pain tests and questionnaires in combination with genetic analysis. A relatively recent approach to investigate pain modulation is via the psychophysical measure of Diffuse Noxious Inhibitory Control (DNIC). This pain-modulating process is the neurophysiological basis for the well-known phenomenon of 'pain inhibits pain' from remote areas of the body. The DNIC paradigm has evolved recently into a clinical tool and simple test and has been shown to be a predictor of post-operative pain.(5) Since pregnancy is associated with decreased pain sensitivity and/or enhanced processes of pain modulation, using tests that investigate pain modulation should provide a better understanding of the pathways involved with pregnancy-induced analgesia and may help predict pain outcomes during labor and delivery. For those women delivering by cesarean section, a DNIC test performed prior to surgery along with psychosocial questionnaires and genetic tests should enable one to identify women prone to suffer severe post-cesarean pain and persistent pain. These clinical tests should allow anesthesiologists to offer not only personalized medicine to women with the promise to improve well-being and satisfaction, but also a reduction in the overall cost of perioperative and long term care due to pain and suffering. On a larger scale, these tests that explore pain modulation may become bedside screening tests to predict the development of pain disorders following surgery.

Long-term erythropoietin gene expression from transduced cells in bioisolator devices.

Recombinant erythropoietin (EPO) is widely administered for long-term treatment of anemia associated with renal failure and other chronic diseases. The ability to deliver EPO by gene therapy would have clinical and economic benefit. We compared autologous and allogeneic transduced primary vascular smooth muscle cells for their ability to provide sustained EPO gene expression when encapsulated in TheraCyte devices implanted subcutaneously (SQ) or intraperitoneally (IP) in rats. Cells were transduced with retrovirus vector LrEpSN encoding rat EPO cDNA. Rats that received either autologous or allogeneic transduced cells showed elevated hematocrits (HCTs) ranging from 50 to 79% that were sustained for more than 12 months. The HCT of control rats remained at baseline (45.8%). Rats that received second SQ implants of either autologous or allogeneic cells showed elevations in hematocrit that were sustained for up to 12 months, suggesting the absence of immunological responses to transduced cells or implant material. All experimental groups had statistically significant elevated HCT (p < 0.001) when compared with controls. Both SQ and IP implantation were equally effective in delivering EPO long term. There were no significant differences in white blood cell (WBC) or platelet (PLT) values between treated and control animals. Implantation of TheraCyte devices was well tolerated and histological evaluation of the devices up to 12 months after surgery revealed a high degree of vascularization and no evidence of host immune response. TheraCyte devices offer a simple and safe gene delivery system that provides sustained therapeutic gene expression, permit removal and implantation of new devices, and do not require immunosuppression of the host.

Treatment of canine cyclic neutropenia by lentivirus-mediated G-CSF delivery.

Cyclic neutropenia is a rare disease that occurs both in humans and gray collie dogs and is characterized by recurrent severe neutropenia leading to bacterial infections and shortened life expectancy. Daily injections of recombinant granulocyte colony-stimulating factor (rG-CSF) are effective in shortening the period of severe neutropenia and reducing infections. After demonstrating that rG-CSF induced elevated neutrophil production in an affected dog, cytokine administration was stopped and 109 infectious units (IUs) of a lentivirus pseudotyped with vesicular stomatitis virus G protein (VSV-G) encoding canine G-CSF cDNA was administered intramuscularly. Serial blood cell counts showed elevated neutrophil production for longer than 17 months. Although neutrophil counts continued to cycle, the range at nadirs was from 3710 to 5300 cells/microL, well above the nadirs before lentivirus administration. After the injection of lentivirus, mean neutrophil counts +/- SD were 12 460 +/- 4240 cells/microL, significantly increased over both pretreatment values of 3040 +/- 2540 cells/microL(P <.0001) and neutrophil counts during G-CSF administration of 10 290 +/- 4860 cells/microL(P <.007). The changes in blood counts from lentivirus injection were associated with absence of clinical signs of infection and fever. The gray collie continued to gain weight and was no longer housed in a pathogen-free environment. Genomic DNA from muscle at injection sites was positive for provirus, whereas gonad, lung, spleen, heart, liver, kidney, leukocytes, and noninjected muscle samples were all negative for provirus. Thus, intramuscular administration of lentivirus encoding G-CSF provided sustained therapeutic levels of neutrophils, suggesting this approach may be applied for long-term treatment of patients with cyclic and other neutropenias.