James F. Nelson

Rapamycin fed late in life extends lifespan in genetically heterogeneous mice

Inhibition of the TOR signalling pathway by genetic or pharmacological intervention extends lifespan in invertebrates, including yeast, nematodes and fruitflies; however, whether inhibition of mTOR signalling can extend lifespan in a mammalian species was unknown. Here we report that rapamycin, an inhibitor of the mTOR pathway, extends median and maximal lifespan of both male and female mice when fed beginning at 600 days of age. On the basis of age at 90% mortality, rapamycin led to an increase of 14% for females and 9% for males. The effect was seen at three independent test sites in genetically heterogeneous mice, chosen to avoid genotype-specific effects on disease susceptibility. Disease patterns of rapamycin-treated mice did not differ from those of control mice. In a separate study, rapamycin fed to mice beginning at 270 days of age also increased survival in both males and females, based on an interim analysis conducted near the median survival point. Rapamycin may extend lifespan by postponing death from cancer, by retarding mechanisms of ageing, or both. To our knowledge, these are the first results to demonstrate a role for mTOR signalling in the regulation of mammalian lifespan, as well as pharmacological extension of lifespan in both genders. These findings have implications for further development of interventions targeting mTOR for the treatment and prevention of age-related diseases.

Rapamycin, But Not Resveratrol or Simvastatin, Extends Life Span of Genetically Heterogeneous Mice

Rapamycin was administered in food to genetically heterogeneous mice from the age of 9 months and produced significant increases in life span, including maximum life span, at each of three test sites. Median survival was extended by an average of 10% in males and 18% in females. Rapamycin attenuated age-associated decline in spontaneous activity in males but not in females. Causes of death were similar in control and rapamycin-treated mice. Resveratrol (at 300 and 1200 ppm food) and simvastatin (12 and 120 ppm) did not have significant effects on survival in male or female mice. Further evaluation of rapamycins effects on mice is likely to help delineate the role of the mammalian target of rapamycin complexes in the regulation of aging rate and age-dependent diseases and may help to guide a search for drugs that retard some or all of the diseases of aging.

Phase II trial of irinotecan in patients with progressive or rapidly recurrent colorectal cancer

PURPOSE To evaluate irinotecan (CPT-11; Yakult Honsha, Tokyo, Japan) in patients with metastatic colorectal carcinoma that had recurred or progressed following fluorouracil (5-FU)-based therapy. PATIENTS AND METHODS Patients were treated with irinotecan 125 to 150 mg/m2 intravenously (IV) every week for 4 weeks, followed by a 2-week rest. Forty-eight patients were entered onto the study and all were assessable for toxicity. Forty-three patients completed one full course of therapy and were assessable for response. RESULTS One complete and nine partial responses were observed (response rate, 23%; 95% confidence interval [CI], 10% to 36%). The median response duration was 6 months (range, 2 to 13). The median survival time was 10.4 months and the 1-year survival rate was 46% (95% CI, 39% to 53%). Grade 4 diarrhea occurred in four of the first nine patients (44%) treated on this study at the 150-mg/m2 dose level. The study was amended to reduce the starting dose of irinotecan to 125 mg/m2. At this dose, nine of 39 patients (23%) developed grade 4 diarrhea. Aggressive administration of loperamide also reduced the incidence of grade 4 diarrhea. Grade 4 neutropenia occurred in eight of 48 patients (17%), but was associated with bacteremia and sepsis in only case. CONCLUSION Irinotecan has significant single-agent activity against colorectal cancer that has progressed during or shortly after treatment with 5-FU-based chemotherapy. The incidence of severe diarrhea is reduced by using a starting dose of irinotecan 125 mg/m2 and by initiating loperamide at the earliest signs of diarrhea. These results warrant further clinical evaluation to define the role of irinotecan in the treatment of individuals with colorectal cancer.

Phase I and pharmacokinetic trial of weekly CPT-11

PURPOSE We conducted a phase I and pharmacokinetic trial of CPT-11 (irinotecan) to characterize the maximum-tolerated dose (MTD), toxicities, pharmacokinetic profile, and antitumor effects in patients with refractory solid malignancies. PATIENTS AND METHODS We treated 32 patients with CPT-11 administered as a 90-minute intravenous infusion every week for 4 consecutive weeks followed by a 2-week rest period. Dose levels ranged from 50 to 180 mg/m2/wk. We determined concentrations of the lactone (active) and total (lactone plus carboxylate) forms of CPT-11 and its metabolite, SN-38, in the plasma and urine of selected patients during and after drug infusion. RESULTS Grade 4 diarrhea was the dose-limiting toxicity (DLT) at the 180-mg/m2/wk dose level. Other toxicities attributed to CPT-11 included dehydration, nausea, vomiting, and asthenia. Hematologic toxicity was mild in most patients. The terminal plasma half-life for CPT-11 (total) was 7.9 +/- 2.8 hours, for CPT-11 (lactone) 6.3 +/- 2.2 hours, for SN-38 (total) 13.0 +/- 5.8 hours, and for SN-38 (lactone) 11.5 +/- 3.8 hours. We observed significant correlations between drug dose and peak plasma concentration (Cpmax) and between drug dose and area under the concentration curve (AUC) for CPT-11, but not for SN-38. CONCLUSION The MTD for CPT-11 in this patient population was 150 mg/m2/wk when administered on a weekly-times-four schedule repeated every 6 weeks. At dose levels greater than 150 mg/m2/wk, diarrhea is dose-limiting.

Rapamycin-mediated lifespan increase in mice is dose and sex dependent and metabolically distinct from dietary restriction.

Rapamycin, an inhibitor of mTOR kinase, increased median lifespan of genetically heterogeneous mice by 23% (males) to 26% (females) when tested at a dose threefold higher than that used in our previous studies; maximal longevity was also increased in both sexes. Rapamycin increased lifespan more in females than in males at each dose evaluated, perhaps reflecting sexual dimorphism in blood levels of this drug. Some of the endocrine and metabolic changes seen in diet‐restricted mice are not seen in mice exposed to rapamycin, and the pattern of expression of hepatic genes involved in xenobiotic metabolism is also quite distinct in rapamycin‐treated and diet‐restricted mice, suggesting that these two interventions for extending mouse lifespan differ in many respects.

Nordihydroguaiaretic acid and aspirin increase lifespan of genetically heterogeneous male mice

The National Institute on Agings Interventions Testing Program was established to evaluate agents that are purported to increase lifespan and delay the appearance of age‐related disease in genetically heterogeneous mice. Up to five compounds are added to the study each year and each compound is tested at three test sites (The Jackson Laboratory, University of Michigan, and University of Texas Health Science Center at San Antonio). Mice in the first cohort were exposed to one of four agents: aspirin, nitroflurbiprofen, 4‐OH‐α‐phenyl‐N‐tert‐butyl nitrone, or nordihydroguaiaretic acid (NDGA). Sample size was sufficient to detect a 10% difference in lifespan in either sex, with 80% power, using data from two of the three sites. Pooling data from all three sites, a log‐rank test showed that both NDGA (p = 0.0006) and aspirin (p = 0.01) led to increased lifespan of male mice. Comparison of the proportion of live mice at the age of 90% mortality was used as a surrogate for measurement of maximum lifespan; neither NDGA (p = 0.12) nor aspirin (p = 0.16) had a significant effect in this test. Measures of blood levels of NDGA or aspirin and its salicylic acid metabolite suggest that the observed lack of effects of NDGA or aspirin on lifespan in females could be related to gender differences in drug disposition or metabolism. Further studies are warranted to find whether NDGA or aspirin, over a range of doses, might prove to postpone death and various age‐related outcomes reproducibly in mice.

Neuroendocrine involvement in aging: Evidence from studies of reproductive aging and caloric restriction

Neuroendocrine changes contribute to female reproductive aging, but changes in other tissues also play a role. In C57BL/6J mice, neuroendocrine changes contribute to estrous cycle lengthening and reduced plasma estradiol levels, but the midlife loss of cyclicity is mainly due to ovarian failure. Hypothalamic estrogen receptor dynamics and estrogenic modulation of gene expression are altered in middle-aged cycling mice. Although insufficient to arrest cyclicity, these neuroendocrine changes may contribute to other reproductive aging phenomena, such as altered gonadotropin secretion and lengthened estrous cycles. In women, the loss of ovarian oocytes, the cause of menopause, accelerates in the decade before menopause. Accelerated oocyte loss may in turn be caused by a selective elevation of plasma follicle stimulating hormone, and neuroendocrine involvement may thus be implicated in menopausal oocyte loss. Chronic calorie restriction retards both neural and ovarian reproductive aging processes, as well as age-related change in many other physiological systems. The diverse effects of food restriction raises the possibility of an underlying coordinated regulatory response of the organism to reduced caloric intake, possibly effected through alterations of neural and/or endocrine signalling. We are therefore attempting to identify neuroendocrine changes that may coordinate the life prolonging response of animals to food restriction. Our initial focus is on the glucocorticoid system. Food restricted rats exhibit daily periods of hyperadrenocorticism, manifest as elevated free corticosterone during the diurnal peak. We hypothesize that this hyperadrenocortical state potentiates cellular and organismic homeostasis throughout life in a manner similar to that achieved during acute stress, thereby retarding aging processes and extending life span.

Acarbose, 17-α-estradiol, and nordihydroguaiaretic acid extend mouse lifespan preferentially in males

Four agents — acarbose (ACA), 17‐α‐estradiol (EST), nordihydroguaiaretic acid (NDGA), and methylene blue (MB) — were evaluated for lifespan effects in genetically heterogeneous mice tested at three sites. Acarbose increased male median lifespan by 22% (P < 0.0001), but increased female median lifespan by only 5% (P = 0.01). This sexual dimorphism in ACA lifespan effect could not be explained by differences in effects on weight. Maximum lifespan (90th percentile) increased 11% (P < 0.001) in males and 9% (P = 0.001) in females. EST increased male median lifespan by 12% (P = 0.002), but did not lead to a significant effect on maximum lifespan. The benefits of EST were much stronger at one test site than at the other two and were not explained by effects on body weight. EST did not alter female lifespan. NDGA increased male median lifespan by 8–10% at three different doses, with P‐values ranging from 0.04 to 0.005. Females did not show a lifespan benefit from NDGA, even at a dose that produced blood levels similar to those in males, which did show a strong lifespan benefit. MB did not alter median lifespan of males or females, but did produce a small, statistically significant (6%, P = 0.004) increase in female maximum lifespan. These results provide new pharmacological models for exploring processes that regulate the timing of aging and late‐life diseases, and in particular for testing hypotheses about sexual dimorphism in aging and health.

An aging Interventions Testing Program: study design and interim report

The National Institute on Agings Interventions Testing Program (ITP) has developed a plan to evaluate agents that are considered plausible candidates for delaying rates of aging. Key features include: (i) use of genetically heterogeneous mice (a standardized four‐way cross), (ii) replication at three test sites (the Jackson Laboratory, TJL; University of Michigan, UM; and University of Texas, UT), (iii) sufficient statistical power to detect 10% changes in lifespan, (iv) tests for age‐dependent changes in T cell subsets and physical activity, and (v) an annual solicitation for collaborators who wish to suggest new interventions for evaluation. Mice in the first cohort were exposed to one of four agents: aspirin, nitroflurbiprofen (NFP), 4‐OH‐α‐phenyl‐N‐tert‐butyl nitrone (4‐OH‐PBN), or nordihydroguiaretic acid (NDGA). An interim analysis was conducted using survival data available on the date at which at least 50% of the male control mice had died at each test site. Survival of control males was significantly higher, at the interim time‐point, at UM than at UT or TJL; all three sites had similar survival of control females. Males in the NDGA group had significantly improved survival (P = 0.0004), with significant effects noted at TJL (P < 0.01) and UT (P < 0.04). None of the other agents altered survival, although there was a suggestion (P = 0.07) of a beneficial effect of aspirin in males. More data will be needed to determine if any of these compounds can extend maximal lifespan, but the current data show that NDGA reduces early life mortality risks in genetically heterogeneous mice at multiple test sites.

Evaluation of Resveratrol, Green Tea Extract, Curcumin, Oxaloacetic Acid, and Medium-Chain Triglyceride Oil on Life Span of Genetically Heterogeneous Mice

The National Institute on Aging Interventions Testing Program (ITP) was established to evaluate agents that are hypothesized to increase life span and/or health span in genetically heterogeneous mice. Each compound is tested in parallel at three test sites. It is the goal of the ITP to publish all results, negative or positive. We report here on the results of lifelong treatment of mice, beginning at 4 months of age, with each of five agents, that is, green tea extract (GTE), curcumin, oxaloacetic acid, medium-chain triglyceride oil, and resveratrol, on the life span of genetically heterogeneous mice. Each agent was administered beginning at 4 months of age. None of these five agents had a statistically significant effect on life span of male or female mice, by log-rank test, at the concentrations tested, although a secondary analysis suggested that GTE might diminish the risk of midlife deaths in females only.