Minocycline 200 mg or 400 mg versus placebo for mild Alzheimer's disease: the MADE Phase II, three-arm RCT

Notes

Permissions

Chapter 1 Introduction

Alzheimer’s disease (AD) is a progressive neurodegenerative disorder that currently affects 50 million people worldwide,1 with projected numbers of affected people reaching 135.5 million by 2050 and associated costs, just for the USA, at US$1.2T. 2 At the Dementia Summit in 2013, the G8 health ministers committed to identifying a cure or a disease-modifying therapy for dementia by 2025,2 but no treatments have so far been shown to delay the progression of cognitive and functional disability that characterises AD. Failure of treatment approaches directed at preventing the associated build-up of β-amyloid (Aβ) or tau protein deposits has stimulated investigation of alternative treatment approaches, including targeting inflammation in the brain.

The risk of developing AD is associated with immune-related and inflammatory genes, including genes coding for myeloid-specific sialic acid-binding receptor (CD33), triggering receptor expressed on myeloid cell 2 (TREM2), complement receptor 1 (CR1) and bridging integrator 1 (BIN1). 3 Microglial activation is increased in AD. 4 Aβ is a pro-inflammatory agent in AD5 and several microglial surface receptors are also Aβ receptors. 6 In the early stages of AD, microglia clear Aβ by phagocytosis and produce Aβ-degrading enzymes. 7 However, as AD pathology progresses, accumulation of Aβ stimulates the microglial production of pro-inflammatory agents, which drives further neurodegeneration. 7

Two independent systematic reviews of mechanisms, tolerability, brain penetration, epidemiology and early phase trial efficacy data on several classes of repositioned drugs have identified minocycline among the most promising of these agents to progress to clinical trials in patients with AD. 8,9 Minocycline is an anti-inflammatory tetracycline that crosses the blood–brain barrier and inhibits the pro-inflammatory functions of microglia, and there is a substantial body of evidence to indicate that minocycline may be neuroprotective in neurodegenerative diseases. Although the primary neuroprotective target of minocycline in the central nervous system is not known, the principal effects of minocycline include inhibition of microglial activation, attenuation of apoptosis and suppression of the production of reactive oxygen species. 10 In vitro, minocycline protects against Aβ-induced cell death and prevents fibrillisation of Aβ. 11 In transgenic mice, minocycline prevents Aβ deposition and neuronal death,12 reduces tau phosphorylation and insoluble tau aggregates,13 downregulates inducible nitric oxide synthetase, cyclo-oxygenase-2 and Aβ precursor protein-cleaving enzyme 1 (BACE1)14 and protects hippocampal neurogenesis in the presence of Aβ. 15 Minocycline reduces interleukin and tumour necrosis factor alpha (TNFα) levels in mice,16 and neuronal death and learning deficits in rats, following Aβ administration. 17 In stroke patients, open-label treatment with 200 mg per day of minocycline for 5 days after infarct has been reported to improve functional outcome. 18 In animal models of Parkinson’s disease, studies have reported both reduced microglial activation and neuronal death19,20 and reduced microglial activation and worsened neuronal death. 21,22 Pilot clinical trials in Parkinson’s disease at a dose of 200 mg per day of minocycline over 18 months have shown no effect on symptoms and no significant increase in adverse events. 23

Minocycline treatment in the superoxide dismutase 1 transgenic mouse model for amyotrophic lateral sclerosis (ALS) delayed the onset of neurodegeneration and muscle strength decline. 24 A completed Phase III trial in ALS, however, reported worse outcomes with minocycline in terms of faster decline in forced vital capacity and manual muscle strength. 25

Suggested explanations for this faster decline is that the dose of up to 400 mg per day of minocycline may have contributed to fatigue in a highly susceptible population, and that increased levels of glutamate receptor 1 phosphorylation may have promoted glutamate toxicity to motor neurons. 26 In AD, both in vitro and in vivo studies have shown reduced microglial activation, attenuated neuronal death, astrogliosis and improved behavioural performance. 11–13,17,27–31 However, to date, there have been no published clinical trials in AD patients and none is currently registered as recruiting.

The minimum daily dose of minocycline that offers neuroprotection in humans has not been established. A dose of 200 mg per day of minocycline is generally very well tolerated in the long-term treatment of acne32 and has been shown to be neuroprotective in acute stroke,18 spinal cord injury33 and multiple sclerosis. 34 However, 200 mg per day of minocycline, although well tolerated, did not improve outcomes in trials in Parkinson’s disease23 or Huntington’s disease. 35 Some authors have argued that one reason for the failure of some trials may be that such doses of minocycline are too low to be neuroprotective, pointing out that the typical effective dose in animal studies would be equivalent to 3–7 g per day in humans. 10 It would not be feasible or ethical to subject AD participants to such very high doses of minocycline, but the Minocycline in Alzheimer’s Disease Efficacy (MADE) trial includes a comparison of 400 mg per day with 200 mg per day to investigate the tolerability of 400 mg per day and whether or not the higher dose confers increased efficacy.

Alzheimer’s disease is a major public health issue and the imperative to discover and develop treatments that can stop or at least delay disease progression is clear. Symptomatic AD treatments in the form of cholinesterase inhibitors and memantine have been the mainstay of current treatment for > 10 years but do not slow progression of the disease. With a more detailed understanding of the basic biology of the AD process, a wide range of cellular and animal model systems have been developed within which several candidate disease-modifying treatments appear promising, though no such agent has performed successfully in Phase III trials. 36,37 Unfortunately, the development of treatments for AD is a complex and difficult process. The slowness of the neurodegenerative process and the substantial difficulties involved in demonstrating that the process has been changed by treatment are major contributors to this problem. Minocycline was arguably the most promising off-patent candidate for AD modification that was not yet in trials, and it was cheap and well tolerated. The time was known to be right for an adequately powered clinical trial, conducted for a sufficiently long period to demonstrate efficacy on simple cognitive and functional outcomes. The results, even if clearly negative, could be expected to move the field on a significant degree.

Based on this wealth of preclinical research suggesting neuroprotection, this trial investigated whether or not minocycline slows decline in cognitive and functional ability in people with mild AD over a 2-year treatment period. The safety and tolerability of minocycline was also compared at doses of 200 mg and 400 mg per day.

Chapter 2 Methods

Chapter 3 Results

Between 23 May 2014 and 14 April 2016, 554 participants were entered from 32 NHS memory services in England and Scotland. Ten patients did not start trial medication and, as prespecified in the protocol, were excluded from all analyses (Figure 1); one participant had been allocated to 400 mg of minocycline, four to 200 mg of minocycline and five to placebo. Baseline characteristics of the 544 eligible participants were well balanced across the three treatment trial arms (Table 2).

FIGURE 1.

Flow chart of participants through the trial.

The mean age of participants was 74.3 years, 57% (303/544) of whom were male and 84% (455/544) of whom were living with a spouse, partner or relative. The average duration of symptoms was 24 months and the average sMMSE score at baseline was 26.4 points.

The sMMSE assessments were obtained for 542 (99.6%) of the 544 participants at baseline, 498 (92%) of the 544 participants at 6 months, 453 (84%) of the 537 participants at 12 months, 420 (80%) of the 528 participants at 18 months, and 403 (78%) of the 517 participants at 24 months (see Appendix 4, Table 8). There were fewer BADLS than sMMSE assessments, because BADLS assessments are not valid for participants in residential care.

Minocycline at a daily dose of 400 mg was poorly tolerated by participants, with just 29% (53/184) of those allocated 400 mg of minocycline completing 2 years of treatment, significantly fewer participants than in the 200 mg of minocycline (62%; 112/181) or placebo arm (64%; 114/179) (p < 0.0001; see Figures 1 and 7). By contrast, 200 mg of minocycline was well tolerated by participants, with similar discontinuation rates with 200 mg of minocycline and placebo (p = 0.56). When reasons for stopping trial treatment were compared (Table 3), more participants allocated to minocycline than to placebo stopped because of gastrointestinal symptoms (p = 0.0008), dermatological side effects (p = 0.02) and dizziness (p = 0.01).

As a consequence of the higher treatment withdrawal rate, fewer assessments were obtained for the 400 mg of minocycline treatment arm than for the 200 mg of minocycline and placebo arms (see Appendix 4, Table 8). For sMMSE assessments at 24 months, 68% of assessments (119 of the 174 expected) were received for the 400 mg of minocycline arm, 82% (144/176) for the 200 mg of minocycline arm and 84% (140/167) for the placebo arm. Return rates for BADLS assessments were similarly lower for the 400 mg of minocycline arm than for the 200 mg of minocycline and placebo arms (see Appendix 4, Table 8).

The change from baseline in sMMSE scores over time, with standard error bars, is shown in Figure 2. There was an average 4.1-point reduction in the combined minocycline arms compared with a 4.3-point reduction in the placebo arm over the 24-month study period (p = 0.90). The reduction in sMMSE score in the 400 mg of minocycline arm over 24 months was less than that observed in the 200 mg of minocycline arm (i.e. 3.3 vs. 4.7 points), but this difference was not significant (p = 0.08).

FIGURE 2.

Change in sMMSE scores: baseline to 24 months’ follow-up. The graph shows change in mean sMMSE score with standard errors; baseline scores are set to zero; p-values are from tests of the time-by-treatment interaction from repeated measures analyses.

Likewise, the worsening of BADLS scores over 24 months was similar in all treatment arms: 5.7, 6.6 and 6.2 points in the 400 mg of minocycline, 200 mg of minocycline and placebo trial arms, respectively. There were no significant differences in BADLS scores between participants allocated minocycline and those allocated to the placebo arm (p = 0.57) or between those participants allocated 400 mg of minocycline and those allocated 200 mg of minocycline (p = 0.77; Figure 3).

FIGURE 3.

Change in BADLS scores: baseline to 24 months’ follow-up. The graph shows change in mean BADLS scores with standard errors; baseline scores are set to zero; p-values are from tests of the time-by-treatment interaction from repeated measures analyses.

To assess how the higher number of missing outcome assessments in the 400 mg of minocycline treatment arm than in the 200 mg of minocycline or placebo arms (see Appendix 4, Table 8) might have affected outcome comparisons, various sensitivity analyses were run to investigate potential bias from non-random dropout. In particular, there were 41 participants who had a baseline sMMSE score but no further assessments, so did not contribute any information to the primary analysis (see Appendix 4, Figure 8). Those participants who discontinue treatment in AD trials are often atypical, usually having worse cognitive and functional ability than those who continue. 20 This is evident from the scores of the 41 participants with a 6-month sMMSE score but no later assessments.

In total, there were 252 reported serious adverse events (SAEs), with the most common categories being neuropsychiatric and cardiocirculatory. The number of SAEs was somewhat higher in the placebo arm than in the 400- and 200-mg minocycline treatment arms (Table 4). Given that gastrointestinal symptoms were the main reason for stopping trial treatment, it is reassuring that the numbers of gastrointestinal SAEs in the minocycline arms were low, and no higher than in the placebo arm. Similarly, though more skin-related toxicities, particularly pigmentation, were reported with minocycline than with placebo [36% (130/365) vs. 21% (38/179); p = 0.0007], few participants stopped trial treatment because of such toxicities (see Table 3) and only six skin toxicities were considered severe (three in participants allocated to any dose of minocycline and three in participants to placebo; see Appendix 4, Table 9). There were no differences in the numbers of participants stopping treatment because of impaired renal function, which had been a prior concern, nor in numbers of renal SAEs. Twenty-eight patients died during the study: 10 allocated to the 400-mg minocycline treatment arm, six to the 200-mg minocycline treatment arm and 12 to the placebo arm (see Appendix 4, Table 10 and Figure 11a). Fifteen of these 28 patients had stopped trial treatment prior to dying. One additional patient died without starting trial treatment. Rates of care home admission were low in this mild AD population, with no difference in numbers between trial arms (see Appendix 4, Figures 11b and 11c).

The average decline in sMMSE score from baseline to 6 months in this subset of patients (i.e. those patients without any post-baseline assessments) was 3.9 points, a rate of decline three times higher than the 1.3-point average decline among the 498 patients who had a 6-month sMMSE assessment and went on to complete later assessments. It seems likely, therefore, that those patients without any post-baseline assessments, who do not contribute to the estimate of the rate of decline, also had a worse than average decline in cognitive and functional ability.

To estimate what impact the missing outcome data from the 41 participants with no post-baseline assessments might have had on the trial results, the study’s sensitivity analyses made two different assumptions:

1. It was assumed that for the first 6 months the scores declined at a rate of 3.9 points (as did those scores for participants who had a 6-month sMMSE but no further assessments) and then declined at the average rate of 1.1 points every 6 months for the rest of the trial.

2. It was assumed that for patients who had no postbaseline assessments the scores declined at the average rate of those participants with assessments, that is, 1.3 sMMSE points for the first 6 months and then 1.1 points every 6 months subsequently.

The results from imputation methods 1 and 2 are shown in Appendix 4, Figures 9a and 9c. The results are not qualitatively different from those of the primary analyses. The only borderline-significant (p = 0.06) differences seen in these sensitivity analyses were between 400 and 200 mg of minocycline. However, with 400 mg of minocycline a little better and 200 mg of minocycline a little worse than placebo, and no difference between any dose of minocycline and placebo, this is probably a chance finding.

Because return rates for BADLS assessments were also lower for the 400 mg of minocycline arm than the 200 mg of minocycline and placebo arms, similar sensitivity analyses were run. There were 39 participants with no BADLS assessment post baseline who did not contribute to the primary analysis. Imputation method 1 assumed that their BADLS score worsened (i.e. increased) by 3.7 points over the first 6 months and then by 1.9 points every 6 months for the rest of the trial. Method 2 assumed that their BADLS score worsened by 1.5 points over the first 6 months and then by 1.9 points subsequently. Because BADLS is valid only for community-resident patients, BADLS scores for those who went into residential care were imputed only up until the last time point before moving into care. Results for imputation methods 1 and 2 are shown in Appendix 4, Figures 9b and 9d. Again, results were not qualitatively different from those of the primary analyses of BADLS assessment scores.

To investigate whether or not the efficacy of minocycline varied by baseline characteristics, subgroup analyses of change in sMMSE over 24 months for minocycline (any dose) versus placebo by duration of symptoms, baseline sMMSE, age and gender were conducted (see Appendix 4, Figure 10). There were no indications of any benefit from minocycline treatment in those participants with shorter or longer durations of symptoms, lower or higher baseline sMMSE scores, or in men or women. There was a borderline-significant (i.e. p = 0.04) trend towards greater efficacy in younger than older patients, but this unanticipated finding could be a chance occurrence given the number of subgroup investigations.

Chapter 4 Discussion

The MADE trial has shown that, in patients with mild AD, 24 months of minocycline treatment at the doses tested does not delay the progress of cognitive or functional impairment, as measured by the well-validated and widely used sMMSE and BADLS clinical rating scales. The trial has also established that minocycline at a dose of 400 mg is poorly tolerated in this population, with fewer than one-third of participants completing 24 months of treatment. By contrast, 200 mg per day of minocycline is well tolerated, with participants allocated this treatment being no more likely to withdraw from trial medication than those taking placebo.

The failure of minocycline to slow the progression of cognitive and functional decline in mild AD is disappointing given the evidence suggesting that neuroinflammation is instrumental in AD progression,7 minocycline’s established anti-inflammatory and neuroprotective effects and the positive data from several experimental animal models of AD. 11–17 NSAIDs have similarly failed to slow disease progression in clinical trials,54 despite long-term use being associated with a lower risk of developing AD in observational studies55 and promising data from transgenic animal models. 56 The study’s findings also parallel those of clinical trials of minocycline in other neurodegenerative disorders in which, despite preclinical research suggesting neuroprotection, minocycline worsened outcomes in ALS,25 had no effect in Huntington’s disease57 and multiple system atrophy,58 had negative symptoms in schizophrenia59 and had only short-term benefits in multiple sclerosis. 60

Therefore, there could be three broad potential explanations for the negative results of this trial. First, although there is good evidence for neuroinflammation in AD,7 this may be more a reaction to pathology than an important driver of progressive neurodegeneration, particularly in patients who are still at the mild stage of dementia. Second, even if neurodegeneration is accelerated by neuroinflammation, minocycline at the doses administered in the MADE study may not have had sufficient activity against these processes to show efficacy. Animal studies, from which much of the evidence for minocycline’s activity as an anti-inflammatory and anti-AD agent has come, have generally used much higher doses of minocycline than used in the MADE trial (i.e. typically equivalent to 3–7 g per day in humans)10 and, so, it could be that trial participants were not exposed to a sufficiently high dose for efficacy. However, if doses of 200–400 mg per day are insufficient for neuroprotection, the difficulties with tolerability experienced by participants allocated 400 mg of minocycline indicate that use of higher doses in this patient population is not feasible.

Minocycline is potentially neuroprotective through several anti-inflammatory processes (suppression of microglial proliferation and activation, reduced release of interleukins 1β and 6 and of tumour necrosis factor alpha, decreased chemokine expression and decreased activity of metalloproteases) as well as anti-apoptotic and anti-oxidant effects. 11–17 A study of 15 patients with traumatic brain injury found reduced microglial activation, as visualised with ¹¹C-PBR28 positron emission tomography (PET),61 following 12 weeks of treatment with 200 mg of minocycline per day, indicating that the dose ranges used in the MADE trial can have a measurable effect on anti-inflammatory targets. The relationship between minocycline-sensitive microglial activation and neurodegeneration may, however, be complicated. Minocycline treatment in the traumatic brain injury study61 was also associated with increased plasma levels of neurofilament light, considered a marker of neurodegeneration. The faster progression seen with minocycline in ALS25 also suggests that some activated microglia might have a reparative function so that their inhibition could accelerate neurodegeneration. This study’s results do not suggest that reduced microglial activation with minocycline worsens neurodegeneration in AD.

A third plausible explanation for the negative results of the MADE study could be that minocycline did have some efficacy against progression of AD but treatment effects were too small to be detectable in the trial. It is difficult to discount this possibility. The MADE trial was, however, powered to detect minimal clinically important differences between minocycline and placebo, so smaller differences might not be considered of clinical relevance.

This pragmatic trial had a number of strengths. It was based in a broad network of academic and NHS memory services and the wide eligibility criteria facilitated the recruitment of participants who were representative of patients with very mild AD diagnosed within the NHS. Outcome measures were limited in number, practical and easy to administer reliably by trial staff and chosen because any differences between minocycline and placebo treatment would have unambiguous clinical relevance. The trial recruited to target and was sufficiently large to detect a clinically meaningful treatment effect, and the trial arms were well matched on potentially important variables at baseline. This streamlined trial design could usefully be applied to test other putative disease-modifying therapies.

Potential limitations of the study include that biomarkers were not used to confirm AD diagnosis, because these are not routinely available within the NHS and, therefore, there was no access to these data. Nonetheless, no diagnoses were revised during the study and rates of decline were as expected in a mild AD population. Compliance was also problematic, with few patients in the 400-mg arm completing 2 years of treatment and only moderate compliance in the 200-mg and placebo arms. A recommendation to take trial medication once rather than twice daily in the event of perceived side effects helped improve compliance but was introduced only late in the trial when the problem with 400-mg compliance was identified.

Although the trial protocol specified that outcome assessments should be obtained irrespective of treatment compliance, this could not always be achieved despite the vigorous efforts of the trial team. Consequently, differential follow-up rates could have biased the study’s results. However, despite the large number of treatment withdrawals in the 400-mg arm, and consequent loss to follow-up of some participants, results were essentially unchanged in sensitivity analyses investigating potential bias from missing data. Thus, the study’s conclusion that 2 years of minocycline treatment for patients with mild AD does not result in any clinically meaningful difference in the rate of decline of cognitive and functional ability is disappointing but robust.

Acknowledgements

The study team would like to thank the following people and organisations for their contributions:

• Rebecca Gathercole for helping to manage the trial

• Keith Anderson for his input in designing and programming the study database

• Nicholas Woodthorpe, Ajay Macharouthu, Anilkumar Pillai, Vandana Mate, Demi Onalaja, Simon Thacker, Richard Brown, Anna Green, Santanu Chakrabarti, Latha Velayudhan, Abdul Patel, Ban Al-Kaissy, Iracema Leroi, Judy Rubinsztein, Vandana Menon, Paul Koranteng, Robert Barber, Rob Jones, Sujata Das, Rohan Van Der Putt, Ejaz Nazir, Jeremy Isaacs, Paul Loughlin, Divya Tiwari, Vanessa Raymont, Tarun Kuruvilla, Rosalind Ward, Marisa Wray, Wendy Neil, Robert Lawrence, Farhad Huwez, Bryan Corridan, Tarik Qassem, Vijayendra Waykar and Aparna Prasanna for recruiting participants.

The trial was supported by a grant from the Efficacy Mechanisms and Evaluation Board funded by NIHR and the Medical Research Council following external peer and board review. MODEPHARMA Limited manufactured the IMP and placebo that was distributed to participating cites by Polar Speed.

References

1. Alzheimer’s Disease International . World Alzheimer Report 2015: The Global Impact of Dementia 2015.
2. Vradenburg G. A pivotal moment in Alzheimer’s disease and dementia: how global unity of purpose and action can beat the disease by 2025. Expert Rev Neurother 2015;15:73-82. https://doi.org/10.1586/14737175.2015.995638.
3. Karch CM, Goate AM. Alzheimer’s disease risk genes and mechanisms of disease pathogenesis. Biol Psychiatry 2015;77:43-51. https://doi.org/10.1016/j.biopsych.2014.05.006.
4. Edison P, Donat CK, Sastre M. In vivo imaging of glial activation in Alzheimer’s disease. Front Neurol 2018;9. https://doi.org/10.3389/fneur.2018.00625.
5. VanItallie TB. Alzheimer’s disease: innate immunity gone awry?. Metab Clin Exp 2017;69S:S41-9. https://doi.org/10.1016/j.metabol.2017.01.014.
6. Yu Y, Ye RD. Microglial Aβ receptors in Alzheimer’s disease. Cell Mol Neurobiol 2015;35:71-83. https://doi.org/10.1007/s10571-014-0101-6.
7. Heneka MT, Carson MJ, El Khoury J, Landreth GE, Brosseron F, Feinstein DL, et al. Neuroinflammation in Alzheimer’s disease. Lancet Neurol 2015;14:388-405. https://doi.org/10.1016/S1474-4422(15)70016-5.
8. Corbett A, Pickett J, Burns A, Corcoran J, Dunnett SB, Edison P, et al. Drug repositioning for Alzheimer’s disease. Nature Revi Drug Discovery 2012;11:833-46. https://doi.org/10.1038/nrd3869.
9. Appleby BS, Cummings JL. Discovering new treatments for Alzheimer’s disease by repurposing approved medications. Curr Top Med Chem 2013;13:2306-27. https://doi.org/10.2174/15680266113136660162.
10. Plane JM, Shen Y, Pleasure DE, Deng W. Prospects for minocycline neuroprotection. Arch Neurol 2010;67:1442-8. https://doi.org/10.1001/archneurol.2010.191.
11. Familian A, Boshuizen RS, Eikelenboom P, Veerhuis R. Inhibitory effect of minocycline on amyloid beta fibril formation and human microglial activation. Glia 2006;53:233-40. https://doi.org/10.1002/glia.20268.
12. Seabrook TJ, Jiang L, Maier M, Lemere CA. Minocycline affects microglia activation, Abeta deposition, and behavior in APP-tg mice. Glia 2006;53:776-82. https://doi.org/10.1002/glia.20338.
13. Noble W, Garwood C, Stephenson J, Kinsey AM, Hanger DP, Anderton BH. Minocycline reduces the development of abnormal tau species in models of Alzheimer’s disease. FASEB J 2009;23:739-50. https://doi.org/10.1096/fj.08-113795.
14. Ferretti MT, Allard S, Partridge V, Ducatenzeiler A, Cuello AC. Minocycline corrects early, pre-plaque neuroinflammation and inhibits BACE-1 in a transgenic model of Alzheimer’s disease-like amyloid pathology. J Neuroinflammation 2012;9. https://doi.org/10.1186/1742-2094-9-62.
15. Biscaro B, Lindvall O, Tesco G, Ekdahl CT, Nitsch RM. Inhibition of microglial activation protects hippocampal neurogenesis and improves cognitive deficits in a transgenic mouse model for Alzheimer’s disease. Neurodegener Dis 2012;9:187-98. https://doi.org/10.1159/000330363.
16. Garcez ML, Mina F, Bellettini-Santos T, Carneiro FG, Luz AP, Schiavo GL, et al. Minocycline reduces inflammatory parameters in the brain structures and serum and reverses memory impairment caused by the administration of amyloid β (1-42) in mice. Prog Neuropsychopharmacol Biol Psychiatry 2017;77:23-31. https://doi.org/10.1016/j.pnpbp.2017.03.010.
17. Choi Y, Kim HS, Shin KY, Kim EM, Kim M, Kim HS, et al. Minocycline attenuates neuronal cell death and improves cognitive impairment in Alzheimer’s disease models. Neuropsychopharmacology 2007;32:2393-404. https://doi.org/10.1038/sj.npp.1301377.
18. Lampl Y, Boaz M, Gilad R, Lorberboym M, Dabby R, Rapoport A, et al. Minocycline treatment in acute stroke: an open-label, evaluator-blinded study. Neurology 2007;69:1404-10. https://doi.org/10.1212/01.wnl.0000277487.04281.db.
19. Du Y, Ma Z, Lin S, Dodel RC, Gao F, Bales KR, et al. Minocycline prevents nigrostriatal dopaminergic neurodegeneration in the MPTP model of Parkinson’s disease. Proc Natl Acad Sci USA 2001;98:14669-74. https://doi.org/10.1073/pnas.251341998.
20. Radad K, Moldzio R, Rausch WD. Minocycline protects dopaminergic neurons against long-term rotenone toxicity. Can J Neurol Sci 2010;37:81-5. https://doi.org/10.1017/s0317167100009690.
21. Diguet E, Fernagut PO, Wei X, Du Y, Rouland R, Gross C, et al. Deleterious effects of minocycline in animal models of Parkinson’s disease and Huntington’s disease. Eur J Neurosci 2004;19:3266-76. https://doi.org/10.1111/j.0953-816X.2004.03372.x.
22. Yang L, Sugama S, Chirichigno JW, Gregorio J, Lorenzl S, Shin DH, et al. Minocycline enhances MPTP toxicity to dopaminergic neurons. J Neurosci Res 2003;74:278-85. https://doi.org/10.1002/jnr.10709.
23. NINDS NET-PD Investigators . A pilot clinical trial of creatine and minocycline in early Parkinson disease: 18-month results. Clin Neuropharmacol 2008;31:141-50. https://doi.org/10.1097/WNF.0b013e3181342f32.
24. Zhu S, Stavrovskaya IG, Drozda M, Kim BY, Ona V, Li M, et al. Minocycline inhibits cytochrome c release and delays progression of amyotrophic lateral sclerosis in mice. Nature 2002;417:74-8. https://doi.org/10.1038/417074a.
25. Gordon PH, Moore DH, Miller RG, Florence JM, Verheijde JL, Doorish C, et al. Efficacy of minocycline in patients with amyotrophic lateral sclerosis: a phase III randomised trial. Lancet Neurol 2007;6:1045-53. https://doi.org/10.1016/S1474-4422(07)70270-3.
26. Huntington Study Group . Minocycline safety and tolerability in Huntington disease. Neurology 2004;63:547-9. https://doi.org/10.1212/01.WNL.0000133403.30559.FF.
27. Hunter CL, Quintero EM, Gilstrap L, Bhat NR, Granholm AC. Minocycline protects basal forebrain cholinergic neurons from mu p75-saporin immunotoxic lesioning. Eur J Neurosci 2004;19:3305-16. https://doi.org/10.1111/j.0953-816X.2004.03439.x.
28. Familian A, Eikelenboom P, Veerhuis R. Minocycline does not affect amyloid beta phagocytosis by human microglial cells. Neurosci Lett 2007;416:87-91. https://doi.org/10.1016/j.neulet.2007.01.052.
29. Cuello AC, Ferretti MT, Leon WC, Iulita MF, Melis T, Ducatenzeiler A, et al. Early-stage inflammation and experimental therapy in transgenic models of the Alzheimer-like amyloid pathology. Neurodegener Dis 2010;7:96-8. https://doi.org/10.1159/000285514.
30. Ryu JK, McLarnon JG. Minocycline or iNOS inhibition block 3-nitrotyrosine increases and blood-brain barrier leakiness in amyloid beta-peptide-injected rat hippocampus. Exp Neurol 2006;198:552-7. https://doi.org/10.1016/j.expneurol.2005.12.016.
31. Parachikova A, Vasilevko V, Cribbs DH, LaFerla FM, Green KN. Reductions in amyloid-beta-derived neuroinflammation, with minocycline, restore cognition but do not significantly affect tau hyperphosphorylation. J Alzheimers Dis 2010;21:527-42. https://doi.org/10.3233/JAD-2010-100204.
32. Goulden V, Glass D, Cunliffe WJ. Safety of long-term high-dose minocycline in the treatment of acne. Br J Dermatol 1996;134:693-5. https://doi.org/10.1111/j.1365-2133.1996.tb06972.x.
33. Casha S, Zygun D, McGowan D, Yong VW, Hurlbert RJ. Neuroprotection with minocycline after spinal cord injury: results of a double blind, randomized, controlled pilot study. Neurosurgery 2009;65:410-11. https://doi.org/10.1227/01.neu.0000358700.03703.a5.
34. Metz LM, Li D, Traboulsee A, Myles ML, Duquette P, Godin J, et al. Glatiramer acetate in combination with minocycline in patients with relapsing–remitting multiple sclerosis: results of a Canadian, multicenter, double-blind, placebo-controlled trial. Mult Scler 2009;15:1183-94. https://doi.org/10.1177/1352458509106779.
35. Bonelli RM, Hodl AK, Hofman P, Kapfhammer HP. Neuroprotection in Huntington’s disease: a 2-year study on minocycline. Int Clin Psychopharmacol 2004;19:337-42. https://doi.org/10.1097/00004850-200411000-00004.
36. Green RC, Schneider LS, Amato DA, Beelen AP, Wilcock G, Swabb EA, et al. Effect of Tarenflurbil on cognitive decline and activities of daily living in patients with mild Alzheimer’s disease. A randomised controlled trial. JAMA 2009;302:2557-64. https://doi.org/10.1001/jama.2009.1866.
37. Aisen PS, Gauthier S, Ferris SH, Saumier D, Haine D, Garceau D, et al. Tramiprosate in mild-to-moderate Alzheimer’s disease – a randomized, double-blind, placebo-controlled, multi-centre study (the Alphase Study). Arch Med Sci 2011;7:102-11. https://doi.org/10.5114/aoms.2011.20612.
38. World Medical Association . Declaration of Helsinki: ethical principles for medical research involving human subjects. JAMA 2013;310:2191-4. https://doi.org/10.1001/jama.2013.281053.
39. Molloy DW, Alemayehu E, Roberts R. Reliability of a Standardized Mini Mental State Examination compared with the traditional Mini Mental State Examination. Am J Psychiatry 1991;148:102-5. https://doi.org/10.1176/ajp.148.1.102.
40. Molloy DW, Standish TI. A guide to the standardized Mini Mental State Examination. Int Psychogeriatr 1997;9:87-94. https://doi.org/10.1017/s1041610297004754.
41. South London and Maudsley NHS Foundation Trust . Minocycline in Alzheimer’s Disease Efficacy Trial: The MADE Trial 2013. https://gtr.ukri.org/projects?ref=MC_PC_13091.
42. Folstein MF, Folstein SE, McHugh PR. ‘Mini-mental state’. A practical method for grading the cognitive state of patients for the clinician. J Psychiatr Res 1975;12:189-98. https://doi.org/10.1016/0022-3956(75)90026-6.
43. Tombaugh TN, McIntyre NJ. The Mini Mental State Examination: a comprehensive review. J Am Geriatr Soc 1992;40:922-35. https://doi.org/10.1111/j.1532-5415.1992.tb01992.x.
44. Howard R, Phillips P, Johnson T, O’Brien J, Sheehan B, Lindesay J, et al. Determining the minimum clinically important differences for outcomes in the DOMINO trial. Int J Geriatr Psychiatry 2011;26:812-17. https://doi.org/10.1002/gps.2607.
45. Courtney C, Farrell D, Gray R, Hills R, Lynch L, Sellwood E, et al. Long-term donepezil treatment in 565 patients with Alzheimer’s disease (AD2000): randomised double-blind trial. Lancet 2004;363:2105-15. https://doi.org/10.1016/S0140-6736(04)16499-4.
46. Feldman H, Gauthier S, Hecker J, Vellas B, Subbiah P, Whalen E. Donepezil MSAD Study Investigators Group . A 24-week, randomized, double-blind study of donepezil in moderate to severe Alzheimer’s disease. Neurology 2001;57:613-20. https://doi.org/10.1212/wnl.57.4.613.
47. Howard RJ, Juszczak E, Ballard CG, Bentham P, Brown RG, Bullock R, et al. Donepezil for the treatment of agitation in Alzheimer’s disease. N Engl J Med 2007;357:1382-92. https://doi.org/10.1056/NEJMoa066583.
48. Bucks RS, Ashworth DL, Wilcock GK, Siegfried K. Assessment of activities of daily living in dementia: development of the Bristol Activities of Daily Living Scale. Age Ageing 1996;25:113-20. https://doi.org/10.1093/ageing/25.2.113.
49. Lavori PW. Clinical trials in psychiatry: should protocol deviation censor patient data?. Neuropsychopharmacology 1992;6:39-48.
50. McKhann GM, Knopman DS, Chertkow H, Hyman BT, Jack CR, Kawas CH, et al. The diagnosis of dementia due to Alzheimer’s disease: recommendations from the National Institute on Aging–Alzheimer’s Association workgroups on diagnostic guidelines for Alzheimer’s disease. Alzheimers Dement 2011;7:263-9. https://doi.org/10.1016/j.jalz.2011.03.005.
51. European Commission . EU Guidelines to Good Manufacturing Practice: Medicinal Products for Human and Veterinary Use – Annex 13: Investigational Medicinal Products 2010.
52. Great Britain . Data Protection Act 1998 1998.
53. Great Britain . Medicines for Human Use (Clinical Trials) Regulations 2004 2004.
54. Miguel-Álvarez M, Santos-Lozano A, Sanchis-Gomar F, Fiuza-Luces C, Pareja-Galeano H, Garatachea N, et al. Non-steroidal anti-inflammatory drugs as a treatment for Alzheimer’s disease: a systematic review and meta-analysis of treatment effect. Drugs Aging 2015;32:139-47. https://doi.org/10.1007/s40266-015-0239-z.
55. Breitner JC, Gau BA, Welsh KA, Plassman BL, McDonald WM, Helms MJ, et al. Inverse association of anti-inflammatory treatments and Alzheimer’s disease: initial results of a co-twin control study. Neurology 1994;44:227-32. https://doi.org/10.1212/wnl.44.2.227.
56. McGeer PL, McGeer EG. NSAIDs and Alzheimer disease: epidemiological, animal model and clinical studies. Neurobiol Aging 2007;28:639-47. https://doi.org/10.1016/j.neurobiolaging.2006.03.013.
57. Huntington Study Group DOMINO Investigators . A futility study of minocycline in Huntington’s disease. Mov Disord 2010;25:2219-24. https://doi.org/10.1002/mds.23236.
58. Dodel R, Spottke A, Gerhard A, Reuss A, Reinecker S, Schimke N, et al. Minocycline 1-year therapy in multiple-system-atrophy: effect on clinical symptoms and [(11)C] (R)-PK11195 PET (MEMSA-trial). Mov Disord 2010;25:97-107. https://doi.org/10.1002/mds.22732.
59. Deakin B, Suckling J, Barnes TRE, Byrne K, Chaudhry IB, Dazzan P, et al. The benefit of minocycline on negative symptoms of schizophrenia in patients with recent-onset psychosis (BeneMin): a randomised, double-blind, placebo-controlled trial. Lancet Psychiatry 2018;5:885-94. https://doi.org/10.1016/S2215-0366(18)30345-6.
60. Metz LM, Li DKB, Traboulsee AL, Duquette P, Eliasziw M, Cerchiaro G, et al. Trial of minocycline in a clinically isolated syndrome of multiple sclerosis. N Engl J Med 2017;376:2122-33. https://doi.org/10.1056/NEJMoa1608889.
61. Scott G, Zetterberg H, Jolly A, Cole JH, De Simoni S, Jenkins PO, et al. Minocycline reduces chronic microglial activation after brain trauma but increases neurodegeneration. Brain 2018;141:459-71. https://doi.org/10.1093/brain/awx339.

Appendix 1 Protocol changes

Table 5 is a summary of the changes made to the original MADE trial protocol, which have been approved by the REC and the Medicines and Healthcare products Regulatory Agency (where relevant).

Appendix 2 Literature search

Table 6 presents a summary of the number of articles identified by each search engine.

The following URL provides a link to the PROSPERO record for the search:

 www.crd.york.ac.uk/prospero/display_record.php?RecordID = 90377 9 (accessed April 2018).

The following search terms were used in March 2018 to perform the search:

 minocycline AND (AD OR alzheimer OR MCI OR “mild cognitive impairment” OR dementia).

Appendix 3 Outcome measure response sheets and sample participant responses

FIGURE 4.

Bristol Activities of Daily Living Scale: outcome measure responses sheet.

FIGURE 5.

Sample participant responses: BADLS responses for 12 months’ follow-up.

FIGURE 6.

Sample participant responses: sMMSE responses for 12 months’ follow-up.

Appendix 4 Additional figures and tables

FIGURE 7.

Proportion of participants taking trial treatment over time: Kaplan–Meier plot.

FIGURE 8.

Flow chart showing the completeness over time of participant follow-up. Colour coding to show assessments split by treatment arm: navy font is 400 mg of minocycline, light blue font is 200 mg of minocycline and red font is placebo.

FIGURE 9.

Change in sMMSE and BADLS scores from baseline to month 24 using imputation methods 1 (a and b) and 2 (c and d). A to estimate scores for patients with no follow-up past baseline. Graph shows change in mean sMMSE and BADLS scores with standard errors. Baseline scores are set to zero and p-values are from tests for time-by-treatment interaction from repeated measures analyses. (a) Average change in sMMSE score from baseline to month 24, using imputation (baseline scores are 26.3 points for 400 mg of minocycline, 26.5 points for 200 mg of minocycline and 26.4 points for placebo). (b) Average change in BADLS score from baseline to month 24, using imputation (baseline scores are 5.6 points for 400 mg of minocycline, 4.9 points for 200 mg of minocycline and 5.5 points for placebo). (c) Average change in sMMSE score from baseline to month 24 using imputation. (d) Average change in BADLS score from baseline to month 24, using imputation.

FIGURE 10.

Subgroup analyses of change in sMMSE score over 24 months for minocycline (any dose) vs. placebo by baseline characteristics: duration of symptoms, baseline sMMSE score, age and gender. Results are derived from a repeated measures model, with p-values from tests for interaction between treatment and the selected subgroup. Min., minimum. The p-value is from the test statistic for testing the interaction between the treatment and any subgroup variable.

FIGURE 11.

Probability of (a) overall survival, (b) institutionalisation and (c) time to death or institutionalisation by treatment arm: Kaplan–Meier survival plots.

FIGURE 12.

Average decline of sMMSE score split by baseline sMMSE score (i.e. a score of 24–26 or 27–30^a points). a, Eight patients start with a sMMSE score of 30 points and have a 24-month sMMSE score of 30 points.

List of abbreviations

Aβ

β-amyloid

AD

Alzheimer's disease

ALS

amyotrophic lateral sclerosis

BADLS

Bristol Activities of Daily Living Scale

DeNDRoN

Dementias and Neurodegenerative Diseases Research Network

GP

general practitioner

IMP

investigational medicinal product

MADE

Minocycline in Alzheimer's Disease Efficacy

MMSE

Mini Mental State Examination

MR

modified release

NDPH

Nuffield Department of Population Health

NIA–AA

National Institute on Aging–Alzheimer’s Association

NIHR

National Institute for Health Research

NRES

National Research Ethics Service

NSAID

non-steroidal anti-inflammatory drug

PI

principal investigator

QP

qualified person

REC

Research Ethics Committee

SAE

serious adverse event

SD

standard deviation

SLE

systemic lupus erythematosus

sMMSE

standardised Mini Mental State Examination

TSC

Trial Steering Committee