Circadian Disruption and Cancer: Making the Connection

Web Sites

HHMI: Biological Clocks
Animations demonstrating the mechanisms of biological clocks in humans, all mammals, and the fruit fly Drosophila.

International Agency for Research on Cancer
Organization that coordinates and conducts research on the causes and mechanisms of human cancer and develops scientific strategies for cancer prevention and control.

International Dark Sky Association
Organization whose mission is to preserve and protect the nighttime environment through environmentally responsible outdoor lighting.

Journal of Biological Rhythms
Journal that publishes research into all aspects of biological rhythms, with emphasis on circadian and seasonal rhythms.

Journal of Circadian Rhythms
Journal that publishes research on all aspects of circadian and nycthemeral rhythms in living organisms.

Life Over Cancer
Website devoted to Keith Block's book, Life Over Cancer: The Block Center Program for Integrative Cancer Treatment.

Journal Articles

Additional Background

Stevens RG, Blask DE, Brainard GC, et al. 2007. Meeting report: the role of environmental lighting and circadian disruption in cancer and other diseases. Environ. Health Perspect. 115: 1357-1362.

Richard Stevens

Kloog I, Haim A, Stevens RG, et al. 2008. Light at night co-distributes with incident breast but not lung cancer in the female population of Israel. Chronobiol. Int. 25: 65-81.

Stevens RG. 2009. Electric light causes cancer? Surely you're joking, Mr. Stevens. Mutat Res. Jan 16. [Epub ahead of print]

Stevens RG. 2009. Light-at-night, circadian disruption and breast cancer: assessment of existing evidence. Int. J. Epidemiol. Apr 23. [Epub ahead of print]

Straif K, Baan R, Grosse Y, et al. 2007. Carcinogenicity of shift-work, painting, and fire-fighting. Lancet Oncol. 8: 1065-1066.

Eva S. Schernhammer

Schernhammer ES, Hankinson SE. 2009. Urinary melatonin levels and postmenopausal breast cancer risk in the Nurses' Health Study cohort. Cancer Epidemiol. Biomarkers Prev. 18: 74-79.

Viswanathan AN, Schernhammer ES. 2009. Circulating melatonin and the risk of breast and endometrial cancer in women. Cancer Lett. 281: 1-7.

Viswanathan AN, Hankinson SE, Schernhammer ES. 2007. Night shift work and the risk of endometrial cancer. Cancer Res. 67: 10618-10622. Full Text

George C. Brainard

Brainard GC, Sliney D, Hanifin JP, et al. 2008. Sensitivity of the human circadian system to short-wavelength (420-nm) light. J. Biol. Rhythms 23: 379-386.

Hanifin JP, Brainard GC. 2007. Photoreception for circadian, neuroendocrine, and neurobehavioral regulation. J. Physiol. Anthropol. 26: 87-94. (PDF, 90.4 KB) Full Text

Jasser SA, Hanifin JP, Rollag MD, Brainard GC. 2006. Dim light adaptation attenuates acute melatonin suppression in humans. J. Biol. Rhythms 21: 394-404.

Zaidi FH, Hull JT, Peirson SN, et al. 2007. Short-wavelength light sensitivity of circadian, pupillary, and visual awareness in humans lacking an outer retina. Curr. Biol. 17: 2122-2128.

Steven M. Hill

Lai L, Yuan L, Cheng Q, et al. 2008. Alteration of the MT1 melatonin receptor gene and its expression in primary human breast tumors and breast cancer cell lines. Breast Cancer Res Treat. Nov 4.

Lai L, Yuan L, Chen Q, et al. 2008. The Galphai and Galphaq proteins mediate the effects of melatonin on steroid/thyroid hormone receptor transcriptional activity and breast cancer cell proliferation. J. Pineal Res. 45: 476-488.

Xiang S, Coffelt SB, Mao L, et al. 2008. Period-2: a tumor suppressor gene in breast cancer. J. Circadian Rhythms 6: 4. Full Text

David E. Blask

Blask DE. 2009. Melatonin, sleep disturbance and cancer risk. Sleep Med. Rev. 13: 257-264.

Dauchy RT, Blask DE, Dauchy EM, et al. 2009. Antineoplastic effects of melatonin on a rare malignancy of mesenchymal origin: melatonin receptor-mediated inhibition of signal transduction, linoleic acid metabolism and growth in tissue-isolated human leiomyosarcoma xenografts. J. Pineal Res. May 22.

Dauchy RT, Dauchy EM, Davidson LK, et al. 2007. Inhibition of fatty acid transport and proliferative activity in tissue-isolated human squamous cell cancer xenografts perfused in situ with melatonin or eicosapentaenoic or conjugated linoleic acids. Comp. Med. 57: 377-382.

Patricia A. Wood and Xiaoming Yang

Wood PA, Yang X, Taber A, et al. 2008. Period 2 mutation accelerates ApcMin/+ tumorigenesis. Mol. Cancer Res. 6: 1786-1793.

Yang X, Wood PA, Ansell CM, et al. 2009. Beta-catenin induces beta-TrCP-mediated PER2 degradation altering circadian clock gene expression in intestinal mucosa of ApcMin/+ mice. J. Biochem. 145: 289-297.

Yang X, Wood PA, Oh EY, et al. 2008. Down regulation of circadian clock gene Period 2 accelerates breast cancer growth by altering its daily growth rhythm. Breast Cancer Res. Treat. Jul 24. [Epub ahead of print]

William J.M. Hrushesky

Du-Quiton J, Wood PA, Burch JB, et al. 2009. Actigraphic assessment of daily sleep-activity pattern abnormalities reflects self-assessed depression and anxiety in outpatients with advanced non-small cell lung cancer. Psychooncology Feb 6. [Epub ahead of print]

Levin RD, Daehler MA, Grutsch JF, et al. 2005. Circadian function in patients with advanced non-small-cell lung cancer. Br. J. Cancer 93: 1202-1208. (PDF, 116 KB) Full Text

Oh EY, Wood PA, Du-Quiton J, Hrushesky WJ. 2008. Seasonal modulation of post-resection breast cancer metastasis. Breast Cancer Res. Treat. 111: 219-228.

H. Phillip Koeffler

Gery S, Koeffler HP. 2007. The role of circadian regulation in cancer. Cold Spring Harb. Symp Quant. Biol. 72: 459-464.

Gery S, Virk RK, Chumakov K, et al. 2007. The clock gene Per2 links the circadian system to the estrogen receptor. Oncogene 26: 7916-7920.

Gery S, Gombart AF, Yi WS, et al. 2005. Transcription profiling of C/EBP targets identifies Per2 as a gene implicated in myeloid leukemia. Blood 106: 2827-2836. Full Text

Georg Bjarnason

Georg A. Bjarnason GA, Mackenzie RG, Nabid A, et al; National Cancer Institute of Canada Clinical Trials Group (HN3). 2009. Comparison of toxicity associated with early morning versus late afternoon radiotherapy in patients with head-and-neck cancer: a prospective randomized trial of the National Cancer Institute of Canada Clinical Trials Group (HN3). Int. J. Radiat. Oncol. Biol. Phys. 73: 166-172.

Innominato PF, Focan C, Gorlia T, et al; Chronotherapy Group of the European Organization for Research and Treament of Cancer. 2009. Circadian rhythm in rest and activity: a biological correlate of quality of life and a predictor of survival in patients with metastatic colorectal cancer. Cancer Res. 69: 4700-4707.

Tyvin A. Rich

Iurisci I, Rich T, Lévi F, et al. 2007. Relief of symptoms after gefitinib is associated with improvement of rest/activity rhythm in advanced lung cancer. J. Clin. Oncol. 25: e17-19.

Rich TA. 2007. Symptom clusters in cancer patients and their relation to EGFR ligand modulation of the circadian axis. J. Support Oncol. 5: 167-174; discussion 176-177.

Rich T, Innominato PF, Boerner J, et al. 2005. Elevated serum cytokines correlated with altered behavior, serum cortisol rhythm, and dampened 24-hour rest-activity patterns in patients with metastatic colorectal cancer. Clin. Cancer Res. 11: 1757-1764. Full Text

Steven W. Lockley

Flynn-Evans EE, Stevens RG, Tabandeh H, et al. 2009. Total visual blindness is protective against breast cancer. Cancer Causes Control. [Epub ahead of print]

Lockley SW, Dijk DJ, Kosti O, et al. 2008. Alertness, mood and performance rhythm disturbances associated with circadian sleep disorders in the blind. J. Sleep Res. 17: 207-216.

Lockley SW, Arendt J, Skene DJ. 2007. Visual impairment and circadian rhythm disorders. Dialogues Clin. Neurosci. 9: 301-314.

Zaidi FH, Hull JT, Peirson SN, et al. 2007. Short-wavelength light sensitivity of circadian, pupillary, and visual awareness in humans lacking an outer retina. Curr. Biol. Dec 17: 2122-2128.

Elizabeth Filipski

Filipski E, Innominato PF, Wu M, et al. 2005. Effects of light and food schedules on liver and tumor molecular clocks in mice. J. Natl. Cancer Inst. 97: 507-517.

Iurisci I, Filipski E, Reinhardt J, et al. 2006. Improved tumor control through circadian clock induction by Seliciclib, a cyclin-dependent kinase inhibitor. Cancer Res. 66: 10720-10728.

Lévi F, Filipski E, Iurisci I, et al. 2007. Cross-talks between circadian timing system and cell division cycle determine cancer biology and therapeutics. Cold Spring Harb. Symp. Quant. Biol. 72: 465-475.

Keith I. Block

Block KI, Gyllenhaal C, Tripathy D, et al. 2009. Survival impact of integrative cancer care in advanced metastatic breast cancer. Breast J. May 12. [Epub ahead of print]

Block KI, Jonas WB. 2006. "Top of the hierarchy" evidence for integrative medicine: what are the best strategies? Integr. Cancer Ther. 5: 277-281.

Block KI, Block P, Gyllenhaal C. 2004. The role of optimal healing environments in patients undergoing cancer treatment: clinical research protocol guidelines. J. Altern. Complement Med. 10 Suppl 1: S157-S170.

Russel J. Reiter

Erren TC, Reiter RJ. 2008. A generalized theory of carcinogenesis due to chronodisruption. Neuro. Endocrinol. Lett. 29: 815-821.

Korkmaz A, Reiter RJ. 2008. Epigenetic regulation: a new research area for melatonin? J. Pineal Res. 44: 41-44.

Reiter RJ, Tan DX, Korkmaz A, et al. 2007. Light at night, chronodisruption, melatonin suppression, and cancer risk: a review. Crit. Rev. Oncog. 13: 303-328.

Sponsorship

This conference has been made possible through the generous support of

The Mushett Family Foundation

and Ambulatory Monitoring, Inc.

This eBriefing was sponsored by the Cancer and Signaling Discussion Group

Media Partners

Nature

International Society of Chronobiology

American Association of Medical Chronobiology and Chronotherapeutics (AAMCC)

Lancet

Integrative Cancer Therapies

Society for Research on Biological Rhythms

Cancer Decisions

Nurses Association

European Sleep Research Society

The Oncology Times

American Society of Clinical Oncology

Advance for Sleep

Can the increased cancer risks of shift workers be reduced using therapies such as melatonin supplementation to promote more robust circadian rhythms?

Could the reduction of light pollution lead to lower cancer rates in industrialized societies?

Will the proteins that constitute the circadian clock represent good targets for intervention in cancer?

What is the role of tumor-specific circadian rhythms in the development and progression of cancer?

Why are circadian gene expression patterns so different between men and women?

Will paying greater attention to the role of circadian rhythms in quality of life parameters for cancer patients lead to better treatment outcomes?

Until quite recently in evolutionary history, night was a time of darkness, varied only by the presence of a full moon or when clouds obscured the stars. Daytime was, by contrast, a period of much greater brightness. Humans, animals, and even unicellular organisms developed circadian rhythms that let them follow this day/night, light/dark pattern very closely, prompting them to be active during the day and sleep at night, or vice versa, depending on their affinity for a nocturnal, diurnal, or other type of lifestyle.

With Edison's invention of the light bulb, however, the sharp delineation between day and night ended in industrialized societies. Humans are now often exposed to quite high levels of light at night, whether through light pollution, because they are awake longer in the evening, or because they must stay awake to work the night shift. At the same time, daytime exposure to light has decreased as more workers stay indoors in often poorly lit environments. Coinciding with the rise of light at night, industrialized countries have seen a steady increase in the incidence of cancer, prompting epidemiologists and other scientists to ask if there is a connection between exposure to light at night and the development of cancer.

On June 19, 2009, a group of researchers studying circadian rhythms and the physiological, biochemical, and molecular mechanisms behind them gathered at the New York Academy of Sciences to ponder this question and its implications for human health. The symposium was organized by William Hrushevsky of the University of South Carolina and David Blask of Tulane University School of Medicine.

A bright new world

The first session examined evidence for the possible link between light at night and cancer risk and the role of the hormone melatonin, which is the primary link between light perceived by the eyes and the internal circadian clock. Epidemiologists Richard Stevens of the University of Connecticut Health Center and Eva Schernhammer of Harvard Medical School described a large body of research linking light exposure at night, particularly among shift workers, to increased cancer risk, including often hormonally driven cancers such as breast and prostate. George Brainard of Jefferson Medical College described his work on the physiological and biophysical aspects of light transmission through the eye and the effects of light at night on melatonin production in humans. Steve Hill of Tulane Cancer Center presented work on the melatonin receptor, MT-1, and its role in modulating cell signaling pathways that are also known to be involved in cancer. David Blask described metabolic rhythms that are tied to melatonin levels, and that implicate consumption of the fatty acid linoleic acid, commonly found in so-called junk foods like potato chips and pastries, in the mechanisms of cancer development.

In the second session, researchers presented work on the molecular components of the circadian clock, and how they might interact with the pathways involved in cancer to promote or prevent it. Much of this work focuses on the role of the Period genes (abbreviated Per) which produce the key clock proteins PER1 and PER2. Patricia Wood of the University of South Carolina described her work on the interactions between Per gene mutations and mutations of the Apc gene of mice, which work together to promote the development of cancerous polyps in the small intestine and colon. Xiaoming Yang, also of the University of South Carolina, discussed his work comparing the rhythmic patterns of PER1 and PER2 expression in normal tissues with those in tumors, investigating the molecular basis for the strong circadian rhythms observed in tumor growth rates. H. Phillip Koeffler of the University of California, Los Angeles, described work linking Per mutations to certain types of hematologic malignancies, including acute myelogenous leukemia and diffuse large B-cell lymphoma.

The third session examined the link between circadian rhythms and cancer from the completely opposite perspective, as researchers explored how the development of cancer leads to the disruption of circadian rhythms in humans, and whether mitigation of this disruption could lead to better cancer treatment and outcomes. William Hrushesky described his ongoing studies of the effects of circadian rhythm disruption on advanced lung cancer patients. Georg Bjarnason, of the Toronto-Sunnybrook Regional Cancer Center, presented work in colorectal patients which has led to enhanced awareness of gender differences in the genes and proteins whose expression is controlled by the circadian clock. Tyvin Rich of the University of Virginia discussed his work on the connection between the greatly increased expression of cytokines in cancer patients and circadian disruptions. Elisabeth Filipski of the French Institute of Health and Medical Research described her work using mouse models of disrupted circadian rhythm to explore effects on tumor growth in different cancer types. Keith Block provided information on his Center for Integrative Cancer Treatment, which combines interventions intended to restore circadian rhythms with a wide array of other pharmacological and non-pharmacological interventions to improve cancer treatment.

The keynote address, given by Russ Reiter of the University of Texas Health Science Center, San Antonio, pulled much of this research together from the perspective of a pioneer in the field of melatonin research. Reiter spoke for many in saying that the weight of accumulated evidence strongly suggests that the disruption of circadian rhythms is a major factor in cancer and perhaps other diseases as well. Reiter said that "repeated perturbations of these systems has biological consequences," or, in other words, "there is a price to pay" for disrupting the work of these ancient regulatory systems in our bodies.

Speakers:
Richard Stevens, University of Connecticut Health Center
Eva Schernhammer, Harvard Medical School
George Brainard, Jefferson Medical College at Thomas Jefferson University
Steve Hill, Tulane Cancer Center
David Blask, Tulane University School of Medicine

Highlights

• Different clocks control circadian rhythms that operate on an organismal versus the cellular level.
• There is growing evidence that disrupted circadian rhythms increases cancer risk.
• Higher melatonin levels were associated with a 41% reduction in the risk of breast cancer in the second Nurses' Health Study.
• Very low levels of light are needed to affect human melatonin production.
• Uptake of linoleic acid, a fatty acid whose metabolite increases cell proliferation, is regulated by melatonin.

The rhythms of life

In humans, the central biologic clock is a region in the anterior hypothalamus of the brain known as the suprachiasmatic nucleus (SCN). This region controls multiple circadian rhythms at the organismal level, including behavioral rhythms such as sleep, hunger, and thirst, and physiological rhythms such as blood pressure, body temperature, and hormone production. But this is not the only clock that is present in humans. The clock mechanism itself, which consists of a complex cycle of gene–gene and gene–protein interactions, is found in many cells in the body, where it governs rhythms that are specific to the functions of different cells and tissue types. Tissues such as the lungs, liver, pancreas, spleen, thymus, and skin have all been shown to produce cyclical patterns of gene expression that follow a circadian rhythm.

Light entrains the biologic clock.

The human circadian clock is entrained to the day/night cycle by the amount and type of light that reaches special receptor cells in the retina of the eye. These cells form part of a sensory system for light that is completely separate from the one that is responsible for vision. In the absence of light, these cells send a message to the pineal gland, also located in the brain, to produce the hormone melatonin. Melatonin is the biologic indicator of darkness: its production peaks during the night and is low during the day. Humans who are exposed to light at night produce less melatonin, and thus may have disrupted circadian rhythms.

The melatonin factor

Richard Stevens of the University of Connecticut Health Center recounted much of the epidemiologic evidence that links the disruption of circadian rhythms to the development of cancer. He noted that the incidence of breast cancer continues to rise around the world, and is especially high in the U.S. and Western Europe. Decades of intensive research have failed to confirm the once popular hypothesis that a high fat diet is responsible for this increase. Melatonin reduces the production of reproductive hormones such as estrogen and progesterone, which led to the hypothesis that at least some of the increased cancer risk in industrialized societies, particularly for hormonally driven cancers like breast and prostate, is due to light at night and the consequent reduction in circulating melatonin levels.

This hypothesis makes a number of predictions, including that shift workers would be at higher risk of breast cancer, and that profoundly blind women would be at lower risk. Multiple epidemiologic studies have shown that these predictions are correct, and have uncovered a link between light at night and prostate cancer as well. The evidence is compelling enough that the International Agency for Research on Cancer (IARC) concluded in a 2007 review article that "shift work that involves circadian disruption is probably carcinogenic in humans." Stevens also reviewed recent genetic studies of the association between circadian clock gene polymorphisms and the risk of developing cancer. He said that multiple such associations have been found but thus far they have been modest in magnitude, and it is not yet known if they are replicable between studies.

Eva Schernhammer of Harvard Medical School is a researcher involved in the Nurses' Health Study, which consists of two long-term prospective studies of women's health that have produced key evidence for the link between circadian rhythm disruptions and cancer. Schernhammer described how melatonin levels vary among individuals, based on factors such as age, diet, body mass index (BMI), lack of sleep, or the need to be awake at night due to shiftwork. Studies have shown that people who work the night shift have lower levels of melatonin. Melatonin has a number of possible roles in cancer prevention: it may act as an antioxidant, as an immunomodulator, or it may reduce the levels of reproductive hormones. Reduced melatonin levels may also affect the expression of multiple other chemical messengers, in turn leading to increased risk for many different types of cancers.

In the Nurses' Health Study, shift workers had a 36%–79% increased risk of breast cancer.

The Nurses' Health Study included two prospective cohort studies, one beginning in 1976 and the other in 1989, for a total of around 250,000 women. When the data were adjusted for BMI, alcohol intake, and exercise levels, these studies showed that shift workers had a 36%–79% increased risk of breast cancer. Shift workers also had a 35% increase in the risk of colorectal cancer and a 43% increase in the risk of endometrial cancer. The 1989 study also showed that higher melatonin levels were associated with a 41% reduction in the risk of breast cancer. Many of these findings have been confirmed and extended by other studies.

Schernhammer said that it is estimated that 15 million Americans work night shifts. More African Americans work night shifts than whites, and most are men, except in certain professions such as nursing. Protecting the health of these workers is an important public health problem. At this time, it is unknown whether interventions such as melatonin supplementation or providing special light conditions are safe and would reduce cancer risks. It is also unknown whether there is a specific time of life when humans are more vulnerable to cancers caused by light at night, or whether changes such as limiting the number of years on the night shift might mitigate risks. These are topics of future research in this area.

George Brainard of Jefferson Medical College at Thomas Jefferson University presented his work on the characteristics of light transmission by the eye and how they are related to melatonin production in humans. He and his group are examining multiple aspects of ocular physiology, including behavioral characteristics such as gaze, physical characteristics of the lens and pupil, and the location and sensitivity of photoreceptors responsible for melatonin regulation.

Brainard described highly detailed studies intended to uncover how much light and which part of the light spectrum is most important for inhibiting melatonin production. Human subjects were exposed to monochromatic light during the night under very controlled conditions and their melatonin levels measured. The researchers found that melatonin production is reduced most by light in the blue region of the spectrum, compared to the visual system for which responses peak in the yellow to green region. They also found that only very low levels of light are needed to affect human melatonin production. Their findings could have important implications for understanding how light exposure in the real world affects melatonin production, how light at night might affect human health, and how these effects might best be mitigated.

Molecular mechanisms

Steve Hill of the Tulane Cancer Center is investigating the molecular mechanisms that might link melatonin to the prevention or treatment of cancer. In vitro, melatonin has been shown to have direct effects on cancer cells—for example, suppressing the growth and proliferation of breast cancer cells in an effect that is mediated by the melatonin receptor, MT-1. Hill and his group have found that a portion of the MT-1 receptor population is localized to lipid rafts, cell membrane platforms that form gathering places for important signaling molecules, including many of the receptors and kinases involved in cancer. They are working to identify and trace specific signaling pathways that might link MT-1 to cancer, including well known pathways that are aberrant in cancer such as Raf, ERK/MEK, and AKT. The estrogen receptor, whose activity promotes the growth of many breast cancers, is modulated by the activities of many of these pathways.

Hill and his group have found that melatonin reduces the expression of estrogen receptor-α, and also modulates the expression of a variety of other nuclear and/or steroid hormone receptors. These results and others led Hill to test 9-cis-retinoic acid, a ligand of the nuclear retinoid X receptors (RXRs), in combination with melatonin as a treatment for carcinogen-induced tumor formation in rats. RXRs are thought to be involved in the regulation of a large number of metabolic and developmental pathways, many of which are implicated in cancer. The 9-cis-retinoic acid/melatonin combination reduced tumor incidence from 90% to 5%–12%, and also induced complete or partial tumor regression in many cases. The overall response rate to this treatment was 98% if tumor stasis was also taken into account, suggesting a high potential payoff for therapies based on this approach.

The activity of melatonin, the melatonin receptor, and clock genes suggest many potential drug targets against cancer.

Hill and his group are pursuing many other aspects of the melatonin–cancer link. They found that melatonin also appears to play a role in tumor invasion and metastasis, a role that is mediated by the p38 MAP kinase signaling pathway and whose downstream effects may be on the expression of matrix metalloproteases, which are important for tissue invasion. They are also investigating the roles of clock genes such as Per2, Cry2, and Sirt1 in the control of cell proliferation, cell cycle, and apoptosis in breast cancer cells. Their research has identified many points at which melatonin, the melatonin receptor, and/or clock genes could be involved in cancer initiation, promotion, and progression, suggesting multiple new targets for drugs that might prevent or treat primary cancers or metastases.

David Blask and his group at the Tulane University School of Medicine are interested in the interactions between light, circadian timing disruption, and diet in the development of cancer. They have focused on the role of linoleic acid, which is the most common fatty acid in our diets. Linoleic acid consumption has risen at the same time as the use of electric light has increased. This fatty acid provides an excellent source of caloric energy for cancers to use as they increase their biomass, and also plays an important signaling role in cell proliferation.

At the cellular level, uptake of linoleic acid is initiated by activation of the protein kinase A (PKA) signaling pathway. Once it enters the cell, a metabolite of linoleic acid, known as 13-HODE, participates in further signaling pathways that promote cell proliferation. Melatonin downregulates PKA, thus reducing linoleic acid uptake and inhibiting these growth-promoting pathways. This phenomenon has been shown to occur in human breast, head and neck, and urogenital cancers.

Blask and his group are investigating the details of this phenomenon by studying the influence of light on human breast cancer xenografts grown in nude mice, which lack intact immune systems and are unable to reject foreign tissue. If the mice are kept on a regular light/dark schedule, linoleic acid uptake and 13-HODE production are rhythmic in the tumors, peaking late in the day. But if the mice are exposed to light at night, this rhythm is lost and these levels stay high all of the time. Tumor growth rates were also found to be rhythmic. The tumors still grew in the presence of an intact light/dark cycle, but growth rates were dramatically increased when this cycle was disrupted or absent. The investigators are working to confirm these findings in humans, and to extend them to other cancers, including prostate.

Speakers:
William Hrushesky, University of South Carolina
Georg Bjarnason, Sunnybrook Odette Cancer Center, Toronto
Tyvin Rich, University of Virginia
Steve Lockley, Harvard Medical School
Elisabeth Filipski, French Institute of Health and Medical Research
Keith Block, Block Center for Integrative Cancer Treatment
Russel Reiter, The University of Texas Health Science Center

Highlights

• Lung cancer patients with higher levels of circadian disruption showed higher rates of anxiety and depression and scored lower on quality of life indices.
• There are large differences in rhythmic gene expression in men versus women.
• Administration of melatonin can help blind people entrain to the day/night cycle, and thus may also be useful in regulating the circadian rhythms of individuals with cancer.
• Cytokines can affect the SCN and the high levels produced in disease situations may cause the mood and sleep disorders associated with them.
• Meal timing can induce stable circadian rhythms in mice without an SCN, suggesting that behavioral approaches may also promote more stable rhythms in cancer patients.
• Holistic approaches that include consideration of circadian rhythms may improve the effectiveness and tolerability of cancer treatment.
• Melatonin has many overlapping actions in addition to clock entrainment, including roles in the immune system and as an antioxidant.

Improving quality of life

In addition to arising from circadian disruptions, cancer and cancer treatments frequently cause circadian disruptions, leading to a greatly reduced quality of life for many cancer patients. Many patients, particular those with advanced cancer, suffer from poor sleep at night, daytime fatigue, reduced appetite, muscle wasting, reduced mental and physical functioning, anxiety, depression, and other debilitating symptoms. Many researchers believe that treating circadian disruption in these patients will improve their quality of life and may also lead to improved survival.

William Hrushesky initiated the discussion of this topic with a description of his clinical studies that are intended to improve the last year of life for patients with advanced lung cancer. Previous studies in colorectal cancer showed that many patients had increased levels of nocturnal activity and daytime sleeping, and that higher levels of circadian disruption were associated with reduced survival rates. Hrushesky and his group are in the initial stages of a study to investigate whether the same holds true for patients with lung cancer, and whether interventions that improve circadian rhythm can improve quality of life and survival rates for these patients.

He and his group have designed a research program that combines objective monitoring of patient activity by actigraphy with more subjective tools, such as patient questionnaires, that ask about quality of life, fatigue, anxiety and depression, sleep habits, and other potentially related symptoms. This wide array of research tools was used to assess the degree to which circadian rhythms were disrupted in 84 lung cancer patients, half of whom were hospitalized and half of whom were at home. Completion of these baseline assessments showed that most of these patients slept very badly, particularly when they were in the hospital, and that patients whose sleep patterns showed higher levels of circadian disruption also showed higher rates of anxiety and depression and scored lower on quality of life indices.

Lung cancer patients, especially those in the hospital have much worse scores on the Pittsburg Sleep Quality Index (PSQI), which measures seven components related to sleep.

These assessments have validated circadian disruption as an important therapeutic target in advanced lung cancer patients. Further studies will assess the effects of interventions intended to improve circadian rhythm, which may include exercise, exposure to full spectrum light at certain times of the day, optimization of sleep hygiene, melatonin supplementation, and others.

Georg Bjarnason of the Sunnybrook Odette Cancer Center in Toronto is investigating the role of circadian rhythm in the success of colorectal cancer treatment. He and his collaborators compared the effect of giving colorectal cancer patients a well known treatment regimen, known as FOLFOX, at undefined times, with the same treatment regimen optimized for circadian timing, known as ChronoFLOX. While they found no difference in survival for the ChronoFLOX treated group as a whole, they found that this treatment dramatically improved survival in men but had the opposite effect in women, even though the men and women had no major differences in their sleep/activity patterns.

The expression patterns of most rhythmic genes differ between men and women.

Bjarnason and his group are investigating the biological basis for this phenomenon by examining genomic and proteomic differences between men and women in the expression of rhythmic genes. They have found that out of about 2000 rhythmic genes, only 200 were expressed in patterns that overlapped between men and women. Other investigators have found that about half of rhythmically expressed proteins were associated with RNAs that were not rhythmically expressed. These findings suggest that there is still much to be learned about when and how the circadian clock controls gene and protein expression, and how that control is affected by gender.

Bjarnason and his group are also looking at the differences in rhythmically expressed genes between cancer cells from male and female patients. These findings have important implications for drug development because drug candidates are often tested in cell cultures, which do not have circadian rhythms, or in rats and mice, which are nocturnal and are likely to have very different patterns of gene expression at the time of day when they receive the experimental agents compared with humans, who are diurnal.

The development of cancer is often accompanied by a cytokine storm, that is, greatly increased levels of a wide range of immunological and inflammatory messenger molecules, resulting in the disruption of normal physiological functioning. Tyvin Rich of the University of Virginia is studying the effects of this storm and its relationship to the disruption of circadian rhythm in cancer patients. High levels of cytokines and other immunomodulatory molecules can result in pain, increased drug toxicity and resistance, psychobehavioral changes, and other symptoms, and may promote cancer recurrence and progression as well. Rich suggested that the effects of these cytokines on the SCN, which contains cytokine receptors, may explain the clustering of common cancer symptoms such as fatigue, depression, and loss of appetite.

He and his group are studying the relationship of fatigue and other quality of life indicators to serum levels of cytokines in cancer patients with a wide range of tumor types. Their studies have shown that in general, cancer patients have higher levels of the cytokines IL-6 and IL-8 than normal controls. However, patients in their study who had more robust circadian rhythms had lower levels of IL-6, as well as less fatigue and loss of appetite. Patients whose circadian rhythms were disrupted had the opposite characteristics. The researchers also found that patients with disrupted rhythms had elevated levels of EGFR ligands and VEGF in their blood, a finding that has important implications for the use of targeted cancer therapies that depend on blocking the receptors for these molecules.

The success of interventions to promote more robust circadian rhythms in cancer patients depends on a good understanding of these rhythms and how they are entrained by exposure to melatonin and light. Steve Lockley of Harvard Medical School shared his expertise on melatonin cycles in humans and how to use melatonin supplementation and/or light therapy most effectively.

Lockley noted that melatonin secretion does not require periods of light and darkness to be rhythmic, but that these signals are what entrains the internal clock to the external day/night cycle. Blind people often produce melatonin in a rhythmic pattern, but because their cycles are usually more or less than 24 hours their rhythm drifts away from the actual day/night cycle. This drift can be treated with melatonin, which when given at the appropriate time can either advance or delay the cycle as needed. Giving exogenous melatonin changes the time of the melatonin peak and thus promotes sleep at night when it is more appropriate. Melatonin given in the early evening advances the clock, while melatonin given in the morning delays the clock. In cancer patients who are sighted, light could also be used similarly to advance or delay the clock and promote more robust rhythms.

Lockley described a study of 1367 blind women from the U.S. and Canada who had varying degrees of light perception. They found that profoundly blind women, who had no light perception at all, had about half the risk of breast cancer of blind women with some residual light perception, after adjustment for other factors including reproductive parameters such as exposure to estrogen. Increased estrogen exposure is a known risk factor for breast cancer. Their study also showed that profoundly blind girls tended to reach menarche earlier, and thus are exposed to more estrogen over the course of their lives, yet their risk of breast cancer was still lower than in women with some light perception. Lockley and his group are working to better understand the relationship between light perception, melatonin levels, and estrogen exposure as they relate to breast cancer risk.

Breast cancer risk is associated with suppressed melatonin levels and disrupted melatonin rhythms.

There are many potential benefits to using light or melatonin therapy to stabilize circadian rhythms in blind people, cancer patients, or other populations. Such interventions can standardize sleep-wake timing with the rest of society, improve mood and alertness, and make other changes in quality of life parameters. Lockley also suggested that a stabilized cycle might be valuable in cancer patients by allowing health care providers to match the patient's clock to optimal treatment times.

Alterations in daily rhythms of rest-activity or in blood levels of the hormone cortisol have been shown to predict poor survival in patients with metastatic colorectal, metastatic breast, and early stage lung cancer. Elisabeth Filipski of the French Institute of Health and Medical Research is studying this phenomenon in a number of experimental mouse models. Circadian rhythms can be disrupted in mice using a variety of methods, including destruction of the SCN, introducing clock mutations, or providing unevenly varying patterns of light and darkness that put mice in a state resembling chronic jet lag. Filipski is using these models to investigate the connections between tumor growth and circadian disruption.

Enhancing the circadian clock in peripheral tissues, including the tumor itself, may have beneficial effects.

She and her group have shown that mice whose SCNs have been ablated show faster growth in at least two types of tumors, Glasgow osteosarcoma and pancreatic adenocarcinoma. Mice with chronic jet lag show changes in clock gene expression in their livers, and tumors in their livers grow faster. Filipski has also shown that meal timing can be used to entrain mice whose SCNs are ablated back to a stable rhythm, reducing tumor growth rates. This type of circadian reinforcement also reduced tumor growth rates in mice with chronic jet lag. These results suggest that enhancement of the clock in peripheral tissues, including the tumor itself, may have beneficial effects that are just as important as reinforcing the rhythm of the central clock.

Keith Block of the Block Center for Integrative Cancer Treatment, Chicago, rounded out the session by describing his efforts to include treatment for circadian disruption into an integrative model of cancer treatment that also includes diet and lifestyle interventions; treatment to improve biological factors such as oxidative stress, inflammation, and immune disruptions; and treatment of the pathological aspects of cancer. This approach includes multiple interventions that are intended to produce and maintain circadian integrity, both as a means of improving quality of life and potentially improving the odds for survival.

Block described how quality of life issues such as fatigue, insomnia, constipation, and lack of appetite can be related to biochemical disruptions, such as changes in inflammation, growth factors, and immune status, and clinical complications, such as appetite suppression, muscle wasting, pneumonia, headache or pain. These symptoms may exacerbate treatment side effects and sometimes even prevent appropriate treatment from being administered. Such issues may lead to the recurrence or progression of cancer, or to more serious, life-threatening complications that might otherwise have been avoided. Additionally, research demonstrates that patients who are unable to maintain optimal chemotherapy dosing, suffer from interrupted treatment scheduling, or are simply unable to complete a full course of treatment, face poorer response and shorter survival. Thus health status and other factors impacting quality of life issues can be directly related to patient survival.

Many of these issues can be addressed by minimizing circadian disruption and prescribing therapies to reset a patient's clock. Block and his coworkers have designed an individualized program that assesses quality of life and circadian disruptions and addresses as many issues as possible for each cancer patient. Their program includes not only change in the content of diet but also when particular foods are to be eaten. Exercise is used to promote better rest-activity cycling as well as nutritional supplements, and pharmacological interventions are given according to available research on the best times of day for such treatment. Preliminary research on a cohort of 90 metastatic breast cancer patients (Breast Journal, July 2009) demonstrates favorable outcome. Time will tell if this holistic approach is replicable and produces better outcomes for patients treated at the center.

In a final session, keynote speaker Russ Reiter of the University of Texas Health Science Center, considered the father of pineal gland and melatonin research, provided an evolutionary perspective on melatonin and tied together many of the topics of the day. He said that melatonin is 2.5 to 3 billion years old and is found in bacteria, birds, reptiles, and insects in addition to mammals. Melatonin is produced in other tissues besides the pineal gland in mammals, including the female ovary and the retina, but only the pineal gland releases it into the rest of the body to provide central clock regulation. He noted that melatonin has many overlapping actions in addition to clock entrainment, including roles in the immune system and as an antioxidant.

Humans living in industrialized society are melatonin-deficient, he said, because they are exposed to less darkness. Given melatonin's many roles, it is not hard to imagine that the disruption of circadian rhythms may be source of many of the disorders of modern life, including not only cancer but also diabetes and obesity, depression, and other disorders as well.

Speakers:
Patricia Wood, University of South Carolina
Xiaoming Yang, University of South Carolina
H. Phillip Koeffler, University of California, Los Angeles
William Hrushesky, University of South Carolina

Highlights

• Mice with mutations in the circadian clock gene Per2 develop tumors that are resistant to radiation-induced apoptosis.
• The PER2 protein may act as a tumor suppressor by regulating the beta-catenin signaling pathway.
• Tumors appear to have their own internal clocks.
• Per2 gene expression is regulated by the C/EBP family of transcription factors.

Periodicity

The operation of the circadian clock itself and the mechanisms by which changes in clock genes and proteins could be involved in causing cancer or promoting its growth are topics of intense investigation. Patricia Wood of the University of South Carolina described her research on the role of the Period genes Per1, Per2, and Per3. These genes, together with the Cry, Clock, and Bmal1 genes, form the central mechanism of the circadian clock by participating in a complex series of negative and positive feedback loops that control their expression levels in a daily rhythm. Genetic studies have linked changes in Period genes, and the Period (PER) proteins that they encode, to the developmental pathways of cancer. Mice with mutations in the Per2 gene develop tumors that are resistant to radiation-induced apoptosis. Wood hypothesized that Per2 mutations might act synergistically to accelerate tumor formation in mice that also had a mutation in the Apc gene, which creates a genetic predisposition for the development of intestinal tumors. They found that mice carrying mutations in both genes did in fact develop almost twice as many tumors as mice carrying the Apc mutation alone.

They investigated the mechanism of this synergistic effect, focusing on two reciprocal hypotheses. One hypothesis is that Per2 affects beta-catenin signaling, which is known to be increased in mice with Apc mutations and is part of the mechanism of tumor formation. The other hypothesis is that Apc mutations might cause abnormalities in circadian clock function, thus affecting Per2 function. Further experiments have provided support for both hypotheses. When unmutated, Per2 was found to suppress the formation of intestinal tumors in mice through a mechanism that involves the regulation of beta-catenin and its downstream targets. On the reciprocal side, they found that both the daily levels and the rhythm of PER2 protein expression were depressed in mice with Apc mutations. These results contribute to a large, developing body of research that suggests that PER proteins act as tumor suppressors, and that reinforces their value as potential targets for anticancer therapies.

Xiaoming Yang, who works with Wood at the University of South Carolina, presented his work, which is intended to further elucidate the role of the Period genes and their encoded proteins in tumors and in normal tissues. Yang described evidence that PER proteins are tumor suppressors, including the observation that overexpression of PER1 or PER2 can inhibit breast cancer growth, as well as the fact that Per mutations, low PER protein expression, and alterations in the epigenetic regulation of Per genes are found in many types of cancers.

PER protein expression is lower in the tumors, with a peak that is 5-fold lower than expression in the liver. The time of peak expression is shifted compared to that in the liver.

Yang found that PER1 and PER2 were expressed with a circadian rhythm in normal mouse liver. Expression of these proteins was high at night and low in the daytime. He found that tumor growth in mice followed a circadian rhythm as well, with peaks in growth rate occurring once at night and once during the day. PER protein expression was lower in the tumors, and the time of peak expression was shifted compared to normal tissues. Yang found alterations in the expression other clock-controlled genes in the tumor as well. Reducing the expression of Per genes in the tumors both increased their growth and altered the circadian growth rhythm. Yang's findings suggest that tumors have their own internal clocks, but that they can respond to signals from the organism's clock as well. The details of these complex interactions remain to be determined. Yang's results also suggest that it is important to take circadian rhythm into account when measuring tumor growth rates, since they change at different times in the daily cycle.

Hematologic malignancies are among the cancers that have been shown to have reduced PER1 and PER2 expression. H. Phillip Koeffler and his group at the University of California, Los Angeles, are studying the role of abnormalities in Per2 gene expression in such malignancies. They are particularly interested in the C/EBP family of transcription factors, which play important roles in energy metabolism, adipogenesis, and myelopoiesis, and have been implicated in the control of Per gene expression as well.

Using cultured cells, Koeffler and his group have shown that two members of this family, C/EBP-α and C/EBP-ε, directly regulate the Per2 promoter. Increased expression of these transcription factors upregulates Per2 expression in human leukemia cell lines. They are investigating the expression of these transcription factors and Per2 in tissue samples from patients with a variety of leukemias and lymphomas. Lymph node and bone marrow samples from acute myeloid leukemia patients show that many have reduced Per2 expression. Patients with diffuse large B-cell lymphomas show reduced Per2 expression as well as dysregulation of C/EBP-α levels. However, these types of changes were not seen in patients with follicular non-Hodgkins lymphoma or mantle cell leukemia. These findings suggest a role for circadian clock genes in the initiation and/or progression of some, but not all, types of leukemia and lymphoma.

William Hrushesky of the University of South Carolina assessed what is known and what needs to be discovered in order to manipulate the circadian clock to control or prevent cancer. He said that in humans it is clear that circadian disruptions alter the host/cancer balance and promote cancer growth. This conclusion can be drawn from many lines of evidence, including epidemiologic research on people with circadian disruptions, genetic research on the associations between cancer risk and clock gene polymorphisms, and from the fact that many human tumors contain clock gene mutations. Much information has also been gathered about the potential role of melatonin in cancer suppression, and preceding speakers had provided significant detail on the experimental systems that have been developed to investigate the mechanisms behind these observations.

Unanswered questions about the role of circadian disruption in cancer.

Yet quite a few unanswered questions remain. What are the relative contributions of the host's versus the tumor's clock in cancer growth? Can clock gene mutations be used as prognostic indicators in human cancers? Melatonin and many clock proteins also have non-clock functions; what is the importance of these functions in cancer initiation and progression? What is the role of host and tumor melatonin receptors in these processes?

Hrushesky noted that cancer might be caused by general circadian disruption, or by the presence of specific clock gene changes that promote abnormal cell growth and proliferation. In the first case, diminishing the disruption and restoring a normal rhythm might be sufficient to prevent or control cancer. This could be done using behavioral, endocrinologic, or pharmacologic interventions. In the second case, however, cancer therapies will need to be targeted at specific genes or gene products to successfully prevent or treat cancer via circadian mechanisms.