Synthetic certified DNA reference material for analysis of human erythropoietin transgene and transcript in gene doping and gene therapy / A. Baoutina, S. Bhat, M. Zheng, L. Partis, M. Dobeson, I.E. Alexander, K.R. Emslie. - (Gene Therapy 23 (2016) 10 (October; p. 708-717)

• PMID: 27439362
• DOI: 10.1038/gt.2016.47

Abstract

There is a recognised need for standardisation of protocols for vector genome analysis used in vector manufacturing, to establish dosage, in biodistribution studies and to detect gene doping in sport. Analysis of vector genomes and transgene expression is typically performed by qPCR using plasmid-based calibrants incorporating transgenic sequences. These often undergo limited characterisation and differ between manufacturers, potentially leading to inaccurate quantification, inconsistent inter-laboratory results and affecting clinical outcomes. Contamination of negative samples with such calibrants could cause false positive results. We developed a design strategy for synthetic reference materials (RMs) with modified transgenic sequences to prevent false positives due to cross-contamination. When such RM is amplified in transgene-specific assays, the amplicons are distinguishable from transgene's amplicons based on size and sequence. Using human erythropoietin as a model, we produced certified RM according to this strategy and following ISO Guide 35. Using non-viral and viral vectors, we validated the effectiveness of this RM in vector genome analysis in blood in vitro. The developed design strategy could be applied to production of RMs for other transgenes, genes or transcripts. Together with validated PCR assays, such RMs form a measurement tool that facilitates standardised, accurate and reliable genetic analysis in various applications.

Synthetic isoflavones and doping : A novel class of aromatase inhibitors? / Michele Iannone, Francesco Botrè, Nicoletta Cardillo, Xavier de la Torre. - (Drug Testing and Analysis 11 (2019) 2 (February); p. 208-214)

• PMID: 30118172.
• DOI: 10.1002/dta.2482

Abstract

Anectodical information suggests that flavonoids may be widely used among athletes for their multiple biochemical and pharmacological effects. We have evaluated in vitro the effects of two synthetic isoflavones, methoxyisoflavone and ipriflavone, on the catalytic activity of human aromatase (CYP19), the enzyme catalyzing the conversion of androgens (ie, testosterone or androstenedione) to estrogens (ie, estradiol and estrone). The potential inhibitory effect was evaluated by measuring the rate of aromatization of testosterone, monitored by gas chromatography-mass spectrometry (GC-MS), both in the presence and in the absence of methoxyisoflavone or ipriflavone, comparing their effects with those of synthetic aromatase inhibitors (formestane, anastrozole, and aminoglutethimide) presently included in the list of prohibited substances and methods, and of natural flavonoids (chrysin, quercetin, and daidzein), that are known inhibitors of CYP19. The preliminary results of our in vitro study show that methoxyisoflavone and ipriflavone act as competitive inhibitors of aromatase, the degree of inhibition measured in vitro being of the same order of magnitude of that of the aromatase inhibitors commonly used in anti-estrogenic therapies. Our preliminary in vitro results indicate that, in principle, a sufficiently large intake of isoflavones could alter the kinetics of the dynamic equilibria between androgens and estrogens, suggesting their monitoring in doping control routine analysis.

Systematic Review : Nonmedical Use of Prescription Stimulants: Risk Factors, Outcomes, and Risk Reduction Strategies / Stephen V. Faraone, Anthony L. Rostain, C. Brendan Montano, Oren Mason, Kevin M. Antshel, Jeffrey H. Newcorn. - (Journal of the American Academy Child & Adolescent Psychiatry 59 (2020) 1 (January); p. 100-112)

• PMID: 31326580
• DOI: 10.1016/j.jaac.2019.06.012

Abstract

Objective: To review all literature on the nonmedical use (NMU) and diversion of prescription stimulants to better understand the characteristics, risk factors, and outcomes of NMU and to review risk-reduction strategies.
Method: We systematically searched PubMed, PsycINFO, and SCOPUS from inception to May 2018 for studies containing empirical data about NMU and diversion of prescription stimulants. Additional references identified by the authors were also assessed for inclusion.
Results: A total of 111 studies met inclusion criteria. NMU and diversion of stimulants are highly prevalent; self-reported rates among population samples range from 2.1% to 58.7% and from 0.7% to 80.0%, respectively. A variety of terms are used to describe NMU, and most studies have examined college students. Although most NMU is oral, non-oral NMU also occurs. The majority of NMU is associated with no, or minor, medical effects; however, adverse medical outcomes, including death, occur in some individuals, particularly when administered by non-oral routes. Although academic and occupational performance enhancement are the most commonly cited motivations, there is little evidence that academic performance is improved by NMU in individuals without attention-deficit/hyperactivity disorder.
Conclusion: NMU of stimulants is a significant public health problem, especially in college students, but variations in the terms used to describe NMU and inconsistencies in the available data limit a better understanding of this problem. Further research is needed to develop methods to detect NMU, identify individuals at greatest risk, study routes of administration, and devise educational and other interventions to help reduce occurrence of NMU. Colleges should consider including NMU in academic integrity policies.

Systematic review : the effects of growth hormone on athletic performance / Hau Liu, Dena M. Bravata, Ingram Olkin, Anne Friedlander, Vincent Liu, Brian Roberts, Eran Bendavid, Olga Saynina, Shelley R. Salpeter, Alan M. Garber, Andrew R. Hoffman. - (Annals of Internal Medicine 148 (2008) 10 (20 May); p. 747-758)

• PMID: 18347346
• DOI: 10.7326/0003-4819-148-10-200805200-00215

Abstract

Background: Human growth hormone is reportedly used to enhance athletic performance, although its safety and efficacy for this purpose are poorly understood.

Purpose: To evaluate evidence about the effects of growth hormone on athletic performance in physically fit, young individuals.

Data sources: MEDLINE, EMBASE, SPORTDiscus, and Cochrane Collaboration databases were searched for English-language studies published between January 1966 and October 2007.

Study selection: Randomized, controlled trials that compared growth hormone treatment with no growth hormone treatment in community-dwelling healthy participants between 13 and 45 years of age.

Data extraction: 2 authors independently reviewed articles and abstracted data.

Data synthesis: 44 articles describing 27 study samples met inclusion criteria; 303 participants received growth hormone, representing 13.3 person-years of treatment. Participants were young (mean age, 27 years [SD, 3]), lean (mean body mass index, 24 kg/m2 [SD, 2]), and physically fit (mean maximum oxygen uptake, 51 mL/kg of body weight per minute [SD, 8]). Growth hormone dosage (mean, 36 microg/kg per day [SD, 21]) and treatment duration (mean, 20 days [SD, 18] for studies giving growth hormone for >1 day) varied. Lean body mass increased in growth hormone recipients compared with participants who did not receive growth hormone (increase, 2.1 kg [95% CI, 1.3 to 2.9 kg]), but strength and exercise capacity did not seem to improve. Lactate levels during exercise were statistically significantly higher in 2 of 3 studies that evaluated this outcome. Growth hormone-treated participants more frequently experienced soft tissue edema and fatigue than did those not treated with growth hormone.

Limitations: Few studies evaluated athletic performance. Growth hormone protocols in the studies may not reflect real-world doses and regimens.

Conclusion: Claims that growth hormone enhances physical performance are not supported by the scientific literature. Although the limited available evidence suggests that growth hormone increases lean body mass, it may not improve strength; in addition, it may worsen exercise capacity and increase adverse events. More research is needed to conclusively determine the effects of growth hormone on athletic performance.

On 13 January 2014 the Spanish Agency for the Protection of Health in Sport (Agencia Española de Protección de la Salud en el Deporte, AEPSAD) decided to impose a 2 year period of ineligibility and a € 3001,- fine on the respondent E10 after his A and B samples tested positive for the prohibited substance 6-hydroxybromantan (bromantan).

Hereafter in February 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The respondent claimed that the substance 6-hydroxybromantan was not mentioned on the list of the prohibited substances and he disputed the reliability of the laboratory and the test method.

The Tribunal concludes that in the certified and accredited laboratory the metabolite 6-hydroxybromantan was detected in the respondent’s sample consistent with the use of the prohibited substance bromantan.
Therefore on 9 April 2015 the Tribunal decides to dismiss the respondent’s appeal and confirmes the AEPSAD decision of 13 January 2014.

On 19 November 2013 the Spanish Agency for the Protection of Health in Sport (Agencia Española de Protección de la Salud en el Deporte, AEPSAD) decided to impose a 4 year period of ineligibility on the respondent E06 after his A and B samples tested positive for the prohibited substance benzoylecgonine (metabolite of cocaine).

Hereafter in January 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The Tribunal concludes that the respondent’s argument is weak to request for a reduction of the imposed sanction considering the respondent’s doping history and repeated infringements.
Therefore on 12 March 2014 the Tribunal decides to dismiss the respondent’s appeal and confirmes the AEPSAD decision of 19 November 2013.

On 13 January 2014 the Spanish Agency for the Protection of Health in Sport (Agencia Española de Protección de la Salud en el Deporte, AEPSAD) decided to impose a 2 year period of ineligibility on the respondent after his sample tested positive for the prohibited substance methylhexaneamine (dimethylpentylamine). The Athlete stated that he had used a prepared drink with Hemo Rage from a team member.

Hereafter in February 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The respondent argued that the sanction was disproportional; he acted without fault or negligence; had not intention to enhance his sport performance; and he didn’t know he ingested a prohibited substance.
He stated that he ingested the drink in the morning on Saturday while the competition took place on Sunday after 17:00 hours. Therefore he claimed that 3 hours after ingestion the substance does not enhance sport performance anymore. Also the respondent referred to a similar case with a Portugese duathlete who was sanctioned only for six months.

The Tribunal finds that the respondent had admitted the use of the prohibited substance an acted negligence without ensuring that the drink does not contain a prohibited substance. He also showed insufficient evidence that he acted without intention to enhance sport performance.
Therefore the Tribunal dismiss the respondent’s appeal and decides to confirm the AEPSAD decision of 13 January 2014.

On 3 Febuary 2014 the Spanish Agency for the Protection of Health in Sport (Agencia Española de Protección de la Salud en el Deporte, AEPSAD) decided to impose a 1 year period of ineligibility on the respondent E16 after her sample tested positive for the prohibited substances chlorothiazide and hydrochlorothiazide without a TUE.

The respondent uses prescribed medication for her chronic illness and was sanctioned without a TUE on 28 November 2011 with a 3 month period of ineligibility.
After she tested positive in june 2012 her case was withdrawn due to a temporary TUE was alreay applied and approved.
The respondent applied for a new TUE on 10 June 2013 when tested out-of-competion on 4 July 2013. The TUE application for the respondent’s was approved for the period 12 July 2013 up to and including 7 June 2017 after she was tested.

Hereafter in March 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The respondent argued that she was without fault or negligence and without intention to enhance sport performance; the sanction was disproportional. With medical reports she showed the necessity to use the prescribed medication for her chronic illness including an approved TUE.

The Tribunal considers the respondent’s arguments and agrees that there was no intention to enhance sport perfomacne and there is an approved TUE with retrospective effect for the use of the prescribed medication as treatment for her illness.
However the Tribunal concludes that the respondent failed to apply for a new TUE in 2012 after her first TUE had expired in June 2012. The respondent can’t claim ignorance and that she was tested out-of-competition. She acted negligently because she waited almost a year until she applied for a new TUE on 10 June 2013. The respondent was tested before and therefore fully aware that the prescribed medication would result in a positive test and the necessity for a TUE.
Therefore the Tribunal dismiss the respondent’s appeal and decides on 8 May 2014 to confirm the AEPSAD decision of 3 February 2014.

On 3 Febuary 2014 the Spanish Agency for the Protection of Health in Sport (Agencia Española de Protección de la Salud en el Deporte, AEPSAD) decided to impose a 2 year period of ineligibility on the respondent after his sample tested positive for the prohibited substances terbutaline.
The respondent stated that he used the product Terbasmin as self-medication because of respiratory difficulties, purchased in a pharmacy without prescription.

Hereafter in April 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The respondent filed several arguments in his defence regarding the disproportionality of the imposed sanction; the violations of the rights for defence and against the AEPSAD allegations.

Considering the principle of strict liability the Tribunal concludes that there where no grounds to reduce the imposed sanction; that the respondent failed to apply for a TUE for his medication without a prescription; and that his rights in this case were not violated.
Therefore the Tribunal dismiss the respondent’s appeal and decides on 25 April 2014 to confirm the AEPSAD decision of 3 February 2014.

Related cases:

• AEPSAD 2015 AEPSAD vs respondent E02
  April 10, 2015
• AEPSAD 2015 AEPSAD vs respondent E03
  March 30, 2015
• AEPSAD 2015 AEPSAD vs respondent E04
  March 30,
• AEPSAD 2015 AEPSAD vs respondent E05
  March 30, 2015
• AEPSAD 2015 AEPSAD vs respondent E06
  May 20, 2015
• TAD 2014_074 Respondent E11 vs AEPSAD
  June 6, 2015
• TAD 2015_077 Respondent E02 vs AEPSAD
  June 26, 2015
• TAD 2015_083 Respondent E03 vs AEPSAD
  July 13, 2015
• TAD 2015_086 Respondent E06 vs AEPSAD
  July 30, 2015
• AEPSAD 2015 AEPSAD vs respondent E41
  January 21, 2016

In March 2014 the Spanish police arrested 13 people in the police action operation Jimbo. Several Athletes were arrested whom operated in Lucena, Cantabria, Silla (Valencia), Marbella (Malaga), Almonte (Huelva) and Sevilla. After house searches the police confiscated blood bags, syringes, growth hormone, EPO and other doping substances.

After news reports in the national media about opertion Jimbo the Agencia Española de Protección de la Salud en el Deporte, AEPSAD) reported an anti-doping rule violation against the respondent E08 for the possession and trafficking of S2 class prohibited substances (peptide hormones, growth factors and related substances). Therefore a provisional suspension was ordered on 14 March 2014.

Hereafter in March 2014 in April 2014 the respondent appealed the AEPSAD decision with the Tribunal Administrativo del Deporte (TAD), the Spanish Disciplinary Committee for Sports.
The respondent objected to the provisional suspension, denied the allegations and argued that the house search conducted by the police at his place was negative for prohibited subsances or other relevant evidence.

The respondent was surprised that EAPSAD ordered a provisional suspension based on news reports in the national media and requested lifting of the imposed provisional sanction.

The Tribunal notes that AEPSAD suspended the proceedings against the respondent already in March 2014 pending information from the police and the provisional suspension was imposed to prevent the participation in any competition. In April the respondent and AEPSAD received files from the national police which respondent and AEPSAD could use for the disciplinary proceedings.

Considering the serious allegations the Tribunal finds that ordering a provisional suspension was justified.
Therefore the Tribunal dismiss the respondent’s appeal and decides on 6 June 2014 to confirm the AEPSAD decision of 14 March 2014.