Interventions to improve the cognitive health of older adults are of critical importance. In the current protocol, ithe investigators conducted a short-term (two months) randomized clinical trial to examine the influence of the nutraceutical NT-020 on the cognitive performance of older adults. Participants were tested with a battery of cognitive tasks at baseline and two months after being randomized to the interventional drug or placebo.

SUMMARY OF STUDY PLAN

The proposed clinical study is a Phase II, randomized, double-blinded, placebo-controlled trial in adults 65-85 years of age with no evidence of cognitive impairment. A total of 120 persons will be randomized to the study, with the goal of completing 100 evaluable subjects.

At the screening assessment, participants will complete the informed consent; questionnaires on basic demographic information including personal medical history and allergies to drug compounds; a mental ability test called the Mini-Mental Status Examination (MMSE); the SF-12 Health survey; the Centers for Epidemiologic Studies Depression Scale (CES-D) and distribute instructions and forms for diet survey and gastrointestinal questionnaires.

At the baseline/randomization assessment, study staff will (a) Confirm eligibility based on screening tests; (b) collect completed diet and gastrointestinal questionnaire survey forms; (c) collect blood for CBC/CMP and antioxidant biomarkers; (d) battery of cognitive tests including Rey Auditory Verbal Learning Test, Educational Testing Service - Identical Pictures Test, Educational Testing Service - Number Comparison Test, Educational Testing Service - Vocabulary Test, Trailmaking A & B, Wechsler Adult Intelligence Scale - Digit Symbol Test, Wechsler Intelligence Scale - Digit Span, Letter Fluency, Category Fluency. Subjects will be equally randomized (n=60 per arm) to blinded treatment with either NT-020 (Natura Therapeutics Nutrastem) or matching placebo, and an initial supply of study drug will be dispensed. Participants will be supplied with one-month supply of blinded treatment (n=60/arm) and be given the study agent intake and symptom log form. The planned intervention period is 2 months.

At specified intervals (weeks 1, 3, 5, 7), participants will be contacted to inquire about the correct administration of the supplement or placebo as well as to assess for any adverse events.

At one month (midpoint), participants will return to campus to (a) collect blood for CBC/CMP and antioxidant biomarkers, (b) collect unused nutritional supplement or placebo and completed study agent intake and symptom log form; (c) collect completed diet survey; (d) supply participant with new one-month supply of nutritional supplement or placebo and new study agent intake and symptom log form.

At the end of the intervention (2 months), we will (a) collect blood for CBC/CMP and antioxidant biomarkers; (b) collect unused nutritional supplement or placebo and completed study agent intake and symptom log form; (c) collect completed diet survey; (d) battery of cognitive tests including Rey Auditory Verbal Learning Test, Educational Testing Service - Identical Pictures Test, Educational Testing Service - Number Comparison Test, Educational Testing Service - Vocabulary Test, Trailmaking A & B, Wechsler Adult Intelligence Scale - Digit Symbol Test, Wechsler Intelligence Scale - Digit Span, Letter Fluency, Category Fluency; ; (e) complete eyeblink conditioning paradigm; (f) complete timed interval tapping paradigm; and (g) Memory Abilities Questionnaire (MAQ-SR);

The primary endpoint of the study is a comparison of the delay eyeblink acquisition performance between persons in the placebo and intervention arms. Other endpoints include: assessing the safety of NT-020 under the proposed dosing regimen; investigating the effect of NT-020 on additional measures of cognitive performance. An additional exploratory endpoints is to examine results from serum and urine diagnostic markers (C-reactive protein, IL-1B, TNFa) after 2 months of intervention with nutritional supplement vs. placebo.

STATISTICAL CONSIDERATIONS

Study Design/Endpoints

The central hypothesis for this Phase II clinical trial is that adults who receive NT-020 will exhibited faster acquisition of the delay eyeblink conditioning, as compared to adults on the placebo.

Sample Size/Accrual Rate We anticipate screening 120 persons, randomizing 100, and having 88 persons complete the two-month study period. We powered the analysis for the repeated measures ANOVA for the eyeblink conditioning paradigm. Power was estimated to be .80 (α = .05), assuming six blocks of trials during the acquisition of the CR, an effect size of f = .16 for the group X trials interaction, and a sample size of 44 per group, assuming an attrition rate of 12% over the two month period.

Randomization and Stratification During the baseline/randomization visit, subjects will be randomized using the stratified randomization method prior to intervention by utilizing a web-based randomization system.

Primary Endpoints The primary efficacy endpoint is to compare the acquisition of delay eyeblink conditioning between persons treated with NT-020 vs. placebo. The planned intervention period is two months. Eyeblink conditioning outcomes will be examined using a 2 (group; NT-020, placebo) X 6 (trial) repeated measures ANOVA to examine differences in the acquisition of the conditioned response from the eyeblink conditioning.

Secondary Endpoint(s) Is the facilitation of the acquisition of the delay and trace human eyeblink response in the nutritional supplementation group mediated by changes in anti-oxidant markers? In order to examine the mediation of the intervention group effects on eyeblink acquisition, we will adopt Baron and Kenny's approach to regression-based statistical mediation. In this model, there are several requirements that must be satisfied in order to claim statistical mediation. We will examine whether intervention group is associated with eyeblink acquisition, whether intervention group is associated with changes in biomarkers, as well as the critical step of whether the influence of intervention group on eyeblink acquisition is reduced or made not statistically significant by the inclusion of changes to biomarker values in the regression analysis.

Evaluate the effect of NT-020 treatment on changes in paper and pencil measures of cognitive performance: A 2 (time; baseline, two-months) X 2 (group; NT-020, placebo) ANOVA will be computed on each of the cognitive test scores. The presence of a group X time interaction will suggest that the two groups exhibit differential change across the two time points.

Inclusion Criteria:

• Adults 65-85 years of age (inclusive).
• Able to understand and sign the informed consent.
• Native English-speaking.
• No evidence of dementia (MMSE >=23).
• Each subject must be willing to give consent after being informed of the experimental nature of therapy, alternatives, potential benefits, side-effects, risks and discomforts.
• All subjects must be willing to be monitored for adequacy of nutritional intake during the intervention, as is the current standard of clinical practice.

Exclusion Criteria:

• Use of high dose antioxidant supplements other than what is provided in the trial.
• History of known allergy to components of the study supplements.
• Mental illness judged by the PI to preclude a successful completion of the trial.
• Persons who are not native English speakers are excluded from the current proposal because many of the cognitive tests that are administered are language-based and are heavily influenced by first language.
• Other acute or chronic medical or psychiatric condition or laboratory abnormality that may increase risk associated with study participation or study drug administration, or may interfere with interpretation of study results, and in the judgment of the investigator would make the subject inappropriate for entry into this study.

Description is limited to persons who completed the entire study. The eight subjects who did not complete the study are excluded from this table.