Matt Coward, MD FACSMale Reproductive Medicine and Surgery, University of North Carolina
- University of North Carolina
- United States of America
About Matt Coward, MD FACS
R. Matthew Coward, MD FACS is an Associate Professor of Urology and Clinical Associate Professor of Obstetrics and Gynecology at the UNC School of Medicine, Director of Male Reproductive Medicine and Surgery at UNC Fertility, and Program Director for the UNC Andrology Fellowship. He earned his MD with Distinction followed by urology residency at UNC, and he completed fellowship in Male Reproductive Medicine and Surgery at the Baylor College of Medicine.
He is an Interactive Associate for Fertility and Sterility, and he serves on the Board of Directors for the ASRM's Society for Male Reproduction and Urology as well as the Society for the Study of Male Reproduction. He has published more than 75 peer-reviewed scientific articles and chapters.
Dr. Coward’s clinical practice offers comprehensive reproductive urology services on-site at UNC Fertility that includes the full array of surgical sperm extraction procedures including microdissection testicular sperm extraction. Dr. Coward’s research interests include clinical trials in reproductive medicine, male infertility, vasectomy and vasectomy reversal, surgical education, and urologic imaging. He has received grants from the NIH, the SMSNA, the Doris Duke Foundation, and UNC Healthcare, and he has served as a co-investigator with the Reproductive Medicine Network of the NICHD.
Yet another flawed antioxidant trial; a plea for rigorous evaluation of antioxidant therapy for male infertility involving oxidative stress
The role of oxidative stress in the aetiology of male infertility has been appreciated since the 1920’s when it was demonstrated that depriving rats of natural vitamin E, the major antioxidant vitamin, generated testicular degeneration and infertility (1,2). Subsequently, a great deal of research has been conducted on the role of oxidative stress in male infertility and there is now a general consensus that reactive oxygen species play a significant role in the creation of defective sperm function and the induction of sperm DNA damage (3-6). Recent papers clearly reveal roles for oxidative stress in a variety of clinical situations including the aetiology of non-obstructive azoospermia (7), defective sperm quality in teratozoospermia (8), the origins of sperm DNA damage (9), the toxic impact of parabens (10) and the pathophysiology of varicocele (11). If oxidative stress is such an important cause of male infertility, then surely antioxidant therapy should be part of the cure (12).
In animal models, this is certainly the case. For example, the GPx5 (glutathione peroxidase 5) knockout mouse suffers from localized oxidative stress in the epididymis (13). As a consequence of this stress, the spermatozoa exhibit clear signs of oxidative damage including impaired DNA compaction and high levels of oxidative DNA damage. When Gpx5-deficient males were mated to normal females, the presence of oxidatively damaged spermatozoa was associated with higher incidences of miscarriage and developmental defects in the offspring. However, when such GPx5-/- males were treated with a carefully engineered antioxidant formulation, oxidative DNA damage in the spermatozoa was found to return to control levels (14). Similarly, treatment with the same antioxidant formulation almost completely restored the fertility of mice rendered infertile by a transient testicular hyperthermia (14). Indeed there is a wealth of animal literature demonstrating that antioxidants can restore fertility and testicular function in animals treated with a wide range of factors capable of causing oxidative stress in the male reproductive tract including cadmium (15), phthalate esters (16), lead, organophosphate pesticides (17), induced diabetes (18), acrylamide exposure (19), mental stress (20) and so on.
Against such a background of supportive data in animal models, it is perhaps surprising that extension of these studies into the clinical domain has not provided evidence of a clear therapeutic benefit of antioxidant administration in man. Several meta-analyses addressing this question have been performed. Recent systematic Cochrane analyses, for example, (21,22) concluded that there was low quality evidence for a positive effect on clinical pregnancy and live birth rates but emphasized the flaws in existing studies based on ‘on serious risk of bias due to poor reporting of methods of randomisation, failure to report on the clinical outcomes, live birth rate and clinical pregnancy, often unclear or even high attrition, and also imprecision due to often low event rates and small overall sample sizes’. Both Cochrane analyses called for large, well-designed, randomised placebo-controlled trials to clarify these results. It was, therefore, with interest that we encountered the paper in Fertility and Sterility by Steiner et al. (23) reporting the results of a large randomized placebo-controlled trial of antioxidant therapy in male infertility. While this study has many excellent features, disappointingly, it turned out to be as fundamentally flawed as its predecessors.
How is it possible to design a study determining the efficacy of antioxidant therapy when no attempt is made to determine levels of oxidative stress in the patients before or after the administration of treatment? This makes no sense. It is like giving insulin to everyone who comes into hospital in a coma. Some will improve, some will continue to deteriorate and, overall, any therapeutic benefit will be lost in the noise. The males selected in this study exhibited at least one abnormal semen parameter in the previous 6 months [sperm concentration <15 million/mL (oligospermia), total motility <40% (asthenospermia), normal morphology <4% (teratospermia), or DNA fragmentation >25%]. Naturally, there are many different reasons for males to exhibit such defects and only in a proportion will this be due to oxidative stress. The authors clearly recognise this fundamental defect in their study but justify the omission of oxidative stress measurements on the basis that the random allocation of antioxidant treatment to infertile patients, irrespective of need, represents the current standard of clinical practice. While this statement is clearly correct, surely we should be encouraging higher standards of clinical implementation, rather than just reinforcing the status quo.
Similar arguments could be directed towards the DNA damage component of the study which focused on the vulnerability of the chromatin to low pH (the sperm chromatin structure assay) rather than the formation of oxidative base adducts (8-Oxo-2'-deoxyguanosine) which has been recently shown to be rather a common trait of infertile male patients (24) and to concern specific paternal chromosomal regions that could be linked to developmental defects in the offspring (25).
The other major factor that needs to be considered is the choice of formulation used in the trial. There are numerous male fertility supplements available in the market worldwide, greatly varying by both the nature of antioxidants incorporated into the formulation and the doses used. So why this particular composition? The authors justify the ingredients and doses used because similar ones had been previously evaluated individually in clinical trials and demonstrated some benefit. However, other studies have demonstrated serious concerns with either the isoformic nature of the ingredients or high doses of the individual ingredients used (14). As an example, high doses of antioxidants such as vitamin C or Zinc, especially in the situation where oxidative stress is absent or minimal, can lead to reductive stress (12) causing sperm DNA decondensation (26-28). Reductive stress arising from over-supplementation can therefore be just as damaging to spermatozoa as oxidative stress and should be avoided. This is another reason why understanding a patient’s oxidative stress status is important to the success of a clinical trial, so patients with low or no oxidative stress can be excluded from the trial.
The field desperately needs to support large scale placebo-controlled studies of the type conducted by Steiner et al (23). However, we shall learn nothing if we do not select the patients on the basis of criteria reflecting levels of oxidative stress in the male reproductive tract, treat accordingly with an evidence-based formulation and measure the performance of the treatment against those oxidative stress markers. Whether such treatment will also increase pregnancy rates will depend on a variety of other factors that will not necessarily be impacted by antioxidant therapy. However, if antioxidant therapy can, as the animal models demonstrate, reverse levels of oxidative damage in both the male and female germ lines (29) then at least one component of the overall pregnancy equation will have been successfully addressed. Furthermore, reducing levels of oxidative DNA damage in the male germ line prior to conception is not just about establishing pregnancies per se, but making sure that the mutational load carried by the resultant offspring is kept as low as possible.
In conclusion, the paper by Steiner et al (23) clearly does indicate that the prevailing clinical practice of randomly prescribing antioxidant therapy to the subpopulation of males attending infertility clinics is unlikely to generate any overall benefit. However, this does not mean that carefully formulated antioxidant preparations are without significant therapeutic merit. They just have to be given to patients where there is evidence of oxidative stress as a causative factor in the patients’ infertility profile. This simple requirement has been met in previous small-scale studies and positive outcomes observed (30). The time has now arrived not for us to abandon this approach to therapy but to replicate such studies at scale, using a randomized, placebo-controlled, double-blinded study design, ensuring that the appropriate patients are selected for treatment and the appropriate assessment criteria are in place to measure the impact of antioxidant intervention.
John Aitken (RJA), Parviz Gharagozloo (PG), Joel Drevet (JD), Jorge Hallak (JH), Alfonso Gutierrez-Adan (AGA), Juan Alvarez, Rafael Ambar, Gihan Bareh, Sheryl Homa, Sergey I. Moskovtsev, Mohammad Hossein Nasr-Esfahani, Cristian O'Flaherty, Suresh Sikka, Steven Somkuti, Paul Turek, Armand Zini.
DECLARATION OF INTEREST
PG is CEO of a company, CellOxess biotechnology, involved in the design and manufacture of antioxidant formulations for male and female infertility. RJA, JD, JH and AGA are honorary advisors to this company.
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- Mason KE Minimal requirements of male and female rats for vitamin E. Am J Physiol 1940;131:268-280.
- Aitken RJ, De Iuliis GN. On the possible origins of DNA damage in human spermatozoa. Mol Hum Reprod 2010;16:3-13.
- Aitken RJ, Drevet JR. The importance of oxidative stress in determining the functionality of mammalian spermatozoa: a two-edged sword. Antioxidants (Basel) 2020;9: pii: E111.
- Bisht S, Faiq M, Tolahunase M, Dada R. Oxidative stress and male infertility. Nat Rev Urol. 2017;14: 470-485.
- Drevet JR, Aitken RJ. Oxidation of sperm nucleus in mammals a physiological necessity to some extends with adverse impacts on oocyte and offspring. Antioxidants (Basel) 2020;9: pii: E95.
- Cito G, Becatti M, Natali A, Fucci R, Picone R, Cocci A, et al. Redox status assessment in infertile patients with non-obstructive azoospermia undergoing testicular sperm extraction: A prospective study. Andrology 2020;8:364-371.
- Ammar O, Mehdi M, Muratori M. Teratozoospermia: its association with sperm DNA defects, apoptotic alterations and oxidative stress. Andrology 2020 ( in press).
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- Samarasinghe SVAC, Krishnan K, Naidu R, Megharaj M, Miller K, Fraser B et al. Parabens generate reactive oxygen species in human spermatozoa. Andrology 2018;6:532-541.
- Altintas R, Ediz C, Celik H, Camtosun A, Tasdemir C, Tanbek K, Tekin S, Colak C, Alan C. The effect of varicocoelectomy on the relationship of oxidative stress in peripheral and internal spermatic vein with semen parameters. Andrology 2016;4:442-446.
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- Chabory E, Damon C, Lenoir A, Kauselmann G, Kern H, Zevnik B, et al. Epididymis seleno-independent glutathione peroxidase 5 maintains sperm DNA integrity in mice. J Clin Invest 2009;119:2074-2085.
- Gharagozloo P, Gutiérrez-Adán A, Champroux A, Noblanc A, Kocer A, Calle A, et al. A novel antioxidant formulation designed to treat male infertility associated with oxidative stress: promising preclinical evidence from animal models. Hum Reprod 2016;31:252-62.
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- Naderi N, Souri M, Nasr Esfahani MH Hajian M, Tanhaei Vash N. Ferulago angulata extract ameliorates epididymal sperm toxicity in mice induced by lead and diazinon. Andrology 2019 (in press).
- Bahmanzadeh M, Goodarzi MT, Rezaei Farimani A, Fathi N, Alizadeh Z. Resveratrol supplementation improves DNA integrity and sperm parameters in streptozotocin-nicotinamide-induced type 2 diabetic rats. Andrologia 2019;51:e13313.
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- Fahim AT, Abd El-Fattah AA, Sadik NAH, Ali BM. Resveratrol and dimethyl fumarate ameliorate testicular dysfunction caused by chronic unpredictable mild stress-induced depression in rats. Arch Biochem Biophys 2019;665:152-165.
- Showell MG, Mackenzie-Proctor R, Brown J, Yazdani A, Stankiewicz MT, Hart RJ. Antioxidants for male subfertility. Cochrane Database Syst Rev 2014:12:CD007411.
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Reply to “Yet another flawed antioxidant trial; a plea for rigorous evaluation of antioxidant therapy for male infertility involving oxidative stress” by John Aitken et al.
No doubt there would be immense praise for the MOXI Trial by the fertility supplement industry if the trial had shown a benefit to antioxidants. However, it is not surprising to receive criticism when the results don’t align with the interests of owners and associates of companies with financial interest in the effectiveness of antioxidant formulations for the treatment of male infertility. The CEO, chief scientific officer, advisory board members, and associates of Celloxcess LLC, along with the co-founder and owner of Alphasperm and a part-owner of Yad-Tech, are among the recent critics of MOXI. Celloxcess produces and sells Fertilix, which shares 8 of its 10 vitamin ingredients with all 8 vitamins in the supplement utilized in the MOXI trial. Tremendous planning went into the selection of the commercial antioxidant formulation for MOXI. We performed a complete market analysis of widely available supplements with attention toward identifying a formulation that was both representative of marketed supplements, as well as which contained pure, non-proprietary antioxidants, which had previously been shown to improve semen parameters.
The NICHD Reproductive Medicine Network tested the hypothesis that antioxidants would improve male fertility in a randomized, double blinded fashion. The trial was designed to study a prevailing clinical practice based on dogma, observations from animal studies (dating from 1926!), and underpowered clinical trials. We agree with Aitken, Gharagozloo, and associates that the MOXI trial found no benefit to support the practice of prescribing antioxidant therapy to infertile males, exactly in the manner in which their supplements (Fertilix, Alphasperm, and Fertil Pro) and many others are marketed.
Fertilix is broadly marketed as a vitamin supplement “for every man; those preparing to conceive naturally, or those requiring added support” in order “to support a normal reproductive redox balance, essential for sperm development and DNA integrity” (1). We agree that a trial evaluating the use of antioxidants in a highly select group of patients with high levels of oxidative stress in the male reproductive tract would be interesting and informative. Unfortunately, this approach is neither feasible nor representative of current practice patterns or direct-to-consumer marketing by the supplement industry. Although direct measurement of oxidative stress in semen is possible, it is not widely available nor routinely performed on fresh semen samples. Therefore, the pragmatic MOXI trial protocol implemented the more reliable and widely accepted surrogate measure of oxidative stress in sperm, DNA fragmentation — which is one of the specifically marketed benefits promulgated by Fertilix’s advertising. Antioxidant therapy did not improve the DNA fragmentation index for the entire treatment group or the subgroup of men with high DNA fragmentation at baseline, in a multicentered, double-blind, randomized, placebo-controlled clinical trial.
Given the revenue generated by the antioxidant supplement industry, coupled with the strong opinions and expertise of its owners, scientific officers, and advisory board members, perhaps they can design and fund an appropriately designed, adequately powered, randomized placebo-controlled trial with primary outcomes of pregnancy and live birth in the subgroup they espouse as needed. Until then, the negative MOXI Trial, along with the negative FAZST Trial (2), will be the definitive clinical trials on this subject. In the meantime, will Celloxcess and others only market their supplements to those with documented high levels of reactive oxygen species in the semen and not make claims about their effects on sperm parameters, DNA integrity, and fertility outcomes? Continuing to prescribe these products to all subfertile males, including preconception patients, no longer appears to be appropriate clinical practice.
R. Matthew Coward, MD FACS
Anne Z. Steiner, MD, MPH
Karl Hansen, MD, PhD
Kurt Barnhart, MD
Marcelle Cedars, MD
Richard Legro, MD
Michael Diamond, MD
Nanette Santoro, MD
Esther Eisenberg, MD, MPH
Heping Zhang, PhD
For the Reproductive Medicine Network
- “Men’s Preconceptuals | Fertilix.” Fertilix.com. Accessed March 27, 2020. https://www.fertilix.com/men.
- Schisterman EF, Sjaarda LA, Clemons T, et al. Effect of Folic Acid and Zinc Supplementation in Men on Semen Quality and Live Birth Among Couples Undergoing Infertility Treatment: A Randomized Clinical Trial. JAMA. 2020;323(1):35–48.