155

Cancer Chronotherapy: Principles, Applications, and Perspectives

Marie-Christine Mormont, Francis Levi, M.D., Ph.D.
Ph.D.

INSERM Chronothe rapeutique des cancers and Service de Cancerologie, Ho pital Paul Brousse (I.C.I.G), Villejuif, Cedex, France.

BACKGROUND. Cell physiology is regulated along the 24-hour timescale by a circadian clock, which is comprised of interconnected molecular loops involving at least nine genes. The cellular clocks are coordinated by the suprachiasmatic nucleus, a hypothalamic pacemaker that also helps the organism adjust to environmental cycles. The rest-activity rhythm is a reliable marker of the circadian system function in both rodents and humans. This circadian organization is responsible for predictable changes in the tolerability and efcacy of anticancer agents, and possibly also may be involved in tumor promotion or growth. METHODS. Expected least toxic times of chemotherapy were extrapolated from experimental models to human subjects with reference to the rest-activity cycle. The clinical relevance of the chronotherapy principle (i.e., treatment administration as a function of rhythms) has been investigated previously in randomized multicenter trials. RESULTS. In the current study, chronotherapeutic schedules were used to safely document activity of the combination of oxaliplatin, 5-uorouracil, and leucovorin against metastatic colorectal carcinoma and to establish new medicosurgical management for this disease, and were reported to result in unprecedented long-term survival. CONCLUSIONS. Chronotherapy concepts appear to offer further potential to improve current cancer treatment options as well as to optimize the development of new anticancer or supportive agents. Cancer 2003;97:155 69. 2003 American Cancer Society. DOI 10.1002/cncr.11040 KEYWORDS: circadian rhythms, chronopharmacology, chronotherapy, colorectal carcinoma, quality of life, survival.

Genetic Basis
The biologic functions of most living organisms are organized along an approximate 24-hour time cycle or circadian rhythm. The endogenicity of the circadian rhythms has been demonstrated in microorganisms such as Neurospora crassa or cyanobacterias, in plants, and in all types of animal species such as ies, mice, rats, and humans. These endogenous rhythms govern daily events such as sleep, activity, hormonal secretion, cellular proliferation, and metabolism. Circadian rhythms are xed genetically. Thus, mutations of the circadian genes (e.g., per in Drosophila or clock in mouse) result in severe disturbances of the rest-activity circadian cycle such as shortening or lengthening of the period or even rhythm suppression.1,2 Studies performed in homozygous or heterozygous human twins further support the role of a genetic basis for circadian rhythms in humans.35 The recent identication of per and clock homologues in

Address for reprints: Francis Le vi, M.D., Ph.D., EPI 0118 INSERM Chronothe rapeutique des cancers, Ho pital Paul Brousse (I.C.I.G), 94800 Villejuif, Cedex, France; Fax: 011-33-1-45-59-36-02; E-mail: levi-m@vjf.inserm.fr Received May 28, 2002; revision received August 2, 2002; accepted August 9, 2002. 2003 American Cancer Society

156

CANCER January 1, 2003 / Volume 97 / Number 1

circadian rhythms with predictable times of peak and trough. These rhythms may inuence the pharmacology and the tolerability of anticancer drugs and/or their antitumor efcacy. Conversely, a lack of synchronization or an alteration of circadian clock function makes rhythm peaks and troughs unpredictable, and may require specic therapeutic measures to restore normal circadian function.

The Rest-Activity Rhythm: A Relevant Marker of the Circadian Clock

Locomotor activity reliably reects circadian clock function in several animal species. Its endogenicity was demonstrated by its persistence in constant environmental conditions in ies, rodents, and humans. This rhythm is controlled by the per gene in Drosophila and by the clock gene in the mouse. Direct pharmacologic actions targeted at the SCN in rodents translate into a phase shift of the rest-activity rhythm of the animals. In rodents, the physical destruction of the SCN results in a complete suppression of the restactivity rhythm, whereas the transplantation of SCN restores circadian rhythmicity.8,9 These and other experimental facts (e.g., mutations of the circadian genes) clearly demonstrate the dependency of this rhythm on SCN function. In humans, the rest-activity rhythm is considered to be and used as a marker of the endogenous circadian clock function in isolation studies, in phase-shift studies, and in psychiatry.10 The rest-activity rhythm can be measured easily using a small-size instrument worn on the wrist called an actigraph. Because this method of monitoring activity is completely noninvasive, to our knowledge there is no restriction to its use in cancer patients, even in an ambulatory setting. The easy recording of rest and/or activity has supported further its use as a reference rhythm for the circadian timing of medications and for the evaluation of circadian clock function.

FIGURE 1. Schematic view of the circadian system. The suprachiasmatic nucleus (SCN) is a biologic clock located at the oor of the hypothalamus. It is able to maintain an approximate 24-hour cycle in its electrical activity in vitro. Its period (cycle duration) is calibrated by the alternation of light (L) directly and darkness (D) through melatonin secretion by the pineal gland. The SCN controls or coordinates the circadian rhythms in the body. The main circadian rhythm is the rest-activity cycle. Cellular metabolism and proliferation also display rhythms in normal tissues, which may be affected by the rest-activity cycle. CNS: central nervous system; PVN: paraventricular nucleus; RHT: retinohypotalamic tract; IGL: intergeniculate leaet; NPY: neuropeptide Y; 5-HT: 5-hydroxytryptamine (or serotonin).
human and rodent cells suggest the ubiquity and similarity of the genetic control of the circadian organization in various species.6

General Organization of the Circadian System

The light perceived by the visual pathways and the secretion of melatonin, a hormone released by the pineal gland during darkness, help to reset the internal clock that regulates the timing of different body functions. A hypothalamic structure, the suprachiasmatic nucleus (SCN), plays a key role in the coordination of circadian rhythms (Fig. 1).7 This temporal organization makes it possible to predict the rhythmic aspects of cellular metabolism and proliferation. Synchronized individuals display

Circadian Rhythms and Cancer Animal data Tumor tissues

Rhythms with periods of approximately 24 hours (for example, ultradian rhythms with a 12-hour or an 8-hour period) have been documented in over 12 murine tumor models. These studies have indicated that the circadian periodicity in cellular proliferation indices or metabolic activity usually is retained in slowgrowing or well differentiated tumors, yet with reduced amplitude and an occasional shift in phase. Conversely, the circadian organization tends to be lost

Cancer Chronotherapy/Mormont and Le vi

157

and possibly replaced with an ultradian periodicity in rapidly growing or advanced stage tumors.11

Clinical Data Tumor tissues

As early as 1953, daily variations in the mitotic index of human mammary carcinoma and squamous or basal cell carcinoma were described, with interindividual variations reported.18,19 Reanalysis of these data validated a circadian rhythm in the group of 6 women with breast carcinoma, with a maximum near 3 p.m.. Ultradian rhythms were found in the group of 31 patients with squamous or basal cell carcinoma.20 Progressive dampening of skin mitotic activity also was suggested in patients with actinic keratoses or skin cancer.21 The cell cycle-related parameters of tumor cells and normal mesothelial cells were studied around the clock within the peritoneal lavage uid from 30 patients with ovarian carcinoma.22 A circadian maximum in the DNA synthesis of both diploid and aneuploid tumor cells was found between noon and 4 p.m.. This time was nearly 12 hours out of phase with the peak of DNA synthesis in mesothelial cells. Ultradian rhythms, with 8-hour and 12-hour periods, also were documented in the aneuploid tumor cell population.23 Twenty-four-hour changes were described for DNA synthesis in malignant lymph nodes from 24 patients with non-Hodgkin lymphoma. The maximum occurred near midnight, whereas the peak of the Sphase in the bone marrow of healthy subjects usually was found between noon and 4 p.m..24 It is interesting to note that when patients were classied according to tumor stage according to the Ann Arbor Classication System, a circadian rhythm in DNA synthesis was validated in the group of patients with early-stage tumor, but not in the group of patients with Stage IV lymphoma. This observation suggested a link between disease stage and circadian rhythm alteration. Noteworthy observations regarding breast cancer in skin surface temperature indicated that circadian rhythms persisted on the surface of breast tumors that were slow-growing and well differentiated. However, rhythms were dampened with the maximum occurring 6 hours earlier than in the noncancerous breast. Conversely, fast-growing, poorly differentiated tumors displayed a shortening in the period of tumor temperature rhythm.25 Another study in 14 patients also reported a phase advance of approximately 6 hours in the rhythm of skin temperature of the cancerous breast compared with the contralateral one.26 Table 1 summarizes the main results published to date regarding human tumor circadian and ultradian rhythms.

Healthy tissues
Twenty-four-hour changes in cellular proliferation and metabolism also were investigated in healthy tissues of tumor-bearing animals. All studies suggest that the alterations of circadian rhythms appear progressively and vary as a function of the experimental model; however, to our knowledge, its chronology remains to be dened.11 For instance, rhythms of DNA synthesis were studied in various organs of mice bearing a transplanted Lewis lung carcinoma.12 In all organs, the changes observed were more pronounced in mice with 10-day-old and 14-day-old tumors than in mice with 6-day-old tumors. Nevertheless, in bone marrow, a second peak of DNA synthesis already appeared for mice with a 6-day-old tumor compared with controls. This peak was found to be sharper and occurred earlier for the animals with a 10-day-old tumor. Finally, ultradian rhythms characterized DNA synthesis of animals bearing 14-day-old tumors. In a diethylnitrosamine (DEN)-induced rat liver carcinoma model after a two-thirds partial hepatectomy, the circadian rhythms of both proliferation and cholesterol 7--hydroxylase activity were slightly dampened after a 2-week treatment, and were abolished thoroughly after a 6-week exposure to DEN.13 Nearly normal corticosterone rhythms were observed in Wistar rats 1 week after the cessation of a 6-week oral administration of DEN; however, mean levels of plasma corticosterone were diminished drastically, and circadian variations were found to be lacking 3 months later when neoplastic nodules were growing.14 Changes in body temperature rhythms also were reported. Both the 24-hour mean and amplitude of the rectal temperature were found to be decreased in Fisher rats transplanted with methylcholantrene-induced sarcomas, as well as in Holtzman rats bearing Walker 256 carcinomas compared with controls.15 Conversely, a 24-hour rhythm in intraperitoneal temperature, as assessed by telemetry in immunocytomabearing rats, persisted for 7 weeks, with only a slight advance in phase noted. Circadian rhythmicity was suppressed only during the week preceding death.16 More recently, circadian rhythms in the tumor blood ow of sarcoma-bearing rats were depicted, with a maximal ow detected in the rest span, at a time when proliferative activity was maximum, which suggests that both variables might be related.17

Healthy tissues
The mean proportion of bone marrow cells in the S-phase was signicantly greater at noon when com-

158

CANCER January 1, 2003 / Volume 97 / Number 1

TABLE 1 Circadian and Ultradian Rhythms in Human Tumors

Circadian rhythm Yes Yes Yes Yes Yes (variable according to ploidy) Yes (variable according to stage) Peak different from normal tissue Yes Yes Not specied Yes Yes

Malignancy Breast

Variable Surface temperature 32 P uptake Mitotic index Surface temperature Cell cycle distribution

Cervix Ovary

Non-Hodgkin lymphoma

Cell cycle distribution

Yes

pared with midnight in both 19 healthy subjects and 11 patients with advanced cancer.2728 A concurrent disruption of the normal rhythms in cortisol secretion and in bone marrow S-phase cells was found in four cancer patients.28 This nding supports the occurrence of altered circadian coordination in some cancer patients. The circadian rhythm in plasma melatonin was found to be dampened in patients with an estrogen receptor-positive breast carcinoma, but not in those with an estrogen receptor-negative tumor.29 Rhythms in plasma melatonin, cortisol, prolactin, thyroid-stimulating hormone, growth hormone, luteinizing hormone, and follicle-stimulating hormone were found to be altered in patients with hormone-dependent breast tumors, but not in the case of hormone-insensitive breast tumors.30 Similarly, rhythm alterations were found to be slight in patients with an early-stage prostate carcinoma whereas major disturbances were found in patients with advanced-stage tumors.30 Studies regarding cortisol and other blood circadian rhythms were performed in 51 patients with advanced or metastatic carcinoma of the ovary, breast, the colon or rectum, with a minimum of 10 blood samples obtained to estimate individual rhythmicity as well.3134 Once more, a circadian rhythm was validated statistically for each group of patients (Fig. 2). Nevertheless, the 24-hour rhythms in plasma cortisol and other variables were found to be prominent in some patients and apparently suppressed in others, despite the lack of any glucocorticoid medication (Fig. 3). These rhythm alterations were found for the most part in patients with a poor performance status (PS) (graded as 2 4 according to the World Health Organization scale) and/or a large tumor burden. A recent study in 104 breast carcinoma patients found that abnormalities in the diurnal cortisol rhythm were associated with reduced survival.35 However, to our

FIGURE 2. Total serum cortisol (mean the standard error of the mean) as a function of circadian sampling time in a group of 19 healthy subjects (E), 18 patients with metastatic colorectal carcinoma (), and 20 patients with advanced ovarian carcinoma (). Rhythms were validated statistically with both analysis of variance and cosinor. Reprinted with permission from Mormont MC, Hecquet B, Bogdan A, et al. Selection and validation of a non-invasive test of cortisol circadian rhythm in cancer patients and control subjects. Int J Cancer. 1998;78:421 424.
knowledge the mechanism by which altered circadian rhythms may impact on PS and survival in patients with breast carcinoma is unclear. In a population of 200 patients with metastatic colorectal carcinoma who were eligible for clinical trials (i.e., those with a PS 2), a study was undertaken to estimate the frequency of circadian system alterations, as assessed from rest-activity and cortisol rhythms. This circadian system assessment was as least invasive as possible, and did not require hospitalization. Motor activity was monitored continuously for 3 days using an actigraph worn on the patients wrist. The strength of the circadian component was assessed with two robust parameters: an autocorrelation coefcient at 24 hours (r24)36 and a dichotomy index comparing amounts of activity when the patient was in bed and out of bed (I O).37 For cortisol assessment, a blood sample was obtained at 8 a.m.

Cancer Chronotherapy/Mormont and Le vi

159

quality of life outcomes. The circadian rest-activity rhythm appeared to be a strong predictor of both tumor response and survival in patients with metastatic colorectal carcinoma. Each of the rest-activityrelated variables provided additional prognostic information concerning maximum response to treatment and survival, to that of the other well known clinical factors, reecting tumor burden and general condition. The patients with poor circadian rhythmicity (i.e., those with a r24 [Fig. 5A] or I O [Fig. 5B] in the lowest quartile) were found to have a 5-fold higher risk of dying within 2 years compared with the patients with better circadian rhythm. Furthermore, the prognostic value of I O remained statistically signicant in the subgroups of patients with a PS of 0 or 1, both on univariate and multivariate analysis. This result demonstrated that low rest-activity rhythm parameters did not merely reect poor PS.38 The circadian distribution of activity also was found to be correlated with several quality of life parameters from the European Organization for Research and Treatment of Cancer (EORTC) QLQ-30 questionnaire.38,39These results suggest that circadian system function itself may play an important role in the outcome of patients with cancer, an issue that deserves further investigation.

FIGURE 3. Individual 24-hour variation in plasma cortisol and total proteins and in circulating leukocytes in two patients with metastatic breast carcinoma. Reprinted with permission from Touitou Y, Le vi F, Bogdan A, et al. Circadian desynchronization of blood variables in patients with metastatic breast cancer. Role of prognostic factors. J Cancer Res Clin Oncol. 1995;121:181188.
and at 4 p.m. on 2 consecutive days from each patient because the relative difference between cortisol levels at 8 a.m. and at 4 p.m. has previously been shown to be a good estimator of the circadian amplitude of this rhythm, with 40% being the lower limit of normal.31 Approximately 30% of the 200 patients had an abnormal cortisol rhythm using this criterion.38 The restactivity pattern ranged from marked to completely disrupted 24-hour rhythmicity. Approximately 30% of the patients were found to have a profoundly disturbed cycle, with a r24 0.30, as illustrated in Figure 4.38 Nevertheless, only a weak correlation was found between cortisol rhythm and rest-activity cycle alterations. This suggests that the two rhythms are controlled by different circadian oscillators and/or circadian clock pathways. The latter study prospectively investigated the relevance of circadian system function for survival and

Chronopharmacology of Anticancer Drugs Laboratory rodents Toxicity

Circadian dosing time appears to inuence the extent of toxicity of approximately 30 anticancer drugs, including cytostatics and cytokines, in mice or rats (Fig. 6). For all these drugs, the survival rates are reported to vary by 50% according to a circadian dosing time of a potentially lethal dose. Such a large difference is observed irrespective of injection route (intravenous or intraperitoneal) or the number of injections (single or repeated).40,41 The methodology that was used to demonstrate such phenomena has consisted mainly of the administration of the same drug dose to different groups of animals, with each one corresponding to a different circadian stage, also called circadian time. Six circadian stages, usually located 4 hours apart, commonly are tested. Time usually is expressed in hours after light onset (HALO). Nocturnally active mice or rats previously are synchronized with an alternation of 12 hours of light and 12 hours of darkness. A 3-week span appears necessary for the synchronization of biologic rhythms to a new light/dark regimen. Neither the knowledge of therapeutic class of a drug nor of its main target organs for toxicity is sufcient to predict the least toxic administration time of

160

CANCER January 1, 2003 / Volume 97 / Number 1

FIGURE 4. Example of individual actigraphy records of ve patients with metastatic colorectal carcinoma. The three subjects on the left side of the gure had a high mean activity level. The patient described in the upper left displayed a high activity level during the day, which decreased during the night (rest) period; he thus had high rhythm parameters (dichotomy index [activity in and out of bed (I O)*] and autocorrelation coefcient at 24 hours [r24]). The second patient had a r24 above the median (i.e., his activity pattern was highly reproducible) but a low I O (i.e., poor sleep), and the third patient had a low r24 (i.e., his activity pattern was not reproducible from one day to the next) but a high I O (he slept well when he was in bed). The two subjects on the right side of the gure had a low mean activity level. The rst maintained a marked circadian rhythm (high I O and r24) whereas the second patient had low circadian parameters, with apparently altered periods of activity and rest. (*The dichotomy index I O is the percent of the activity counts measured when the patient is in bed that are inferior to the median of the activity counts measured when the patient is out of bed). PS: performance status.

an anticancer drug. Platinum complex analogs (cisplatin, carboplatin, and oxaliplatin [l-OHP]) all appear to be best tolerated near the middle of the nocturnal activity span of mice or rats, although the respective target tissues for toxicity of these compounds differ; cisplatin is toxic chiey to both kidney and bone marrow, carboplatin to bone marrow and colonic mucosa, and l-OHP to jejunal mucosa and bone marrow.42 44 Conversely, pirarubicin, an anthracycline compound, for the most part exerts myelosuppressive effects, which reportedly are least after dosing in the second half of the diurnal rest span (approximately 7 HALO),45 whereas mitoxantrone, an anthracycline-related compound, displays the least hematologic toxicity 8 hours later (15 HALO).46 The same holds true for vinca alkaloids, with times of least toxicity found at 14 HALO, 18 HALO, and 20 HALO, respectively, for vincristine, vinblastine, and vinorelbine.47,48

Antitumor efcacy
Quite strikingly, the administration of a drug at a circadian time when it is best tolerated usually achieves best antitumor activity. This was found to be true for antimetabolites such as arabinofuranosylcytosine, 5-uorouracil (5-FU), or oxuridine (FUDR); for intercalating agents such as doxorubicin; and for alkylating drugs such as melphalan or cisplatin.49 Similarly, docetaxel tolerability and antitumor efcacy against PO3 pancreatic carcinoma were enhanced by drug dosing in the second half of the rest phase (711 HALO).50 Among newer anticancer drugs, vinorelbine was simultaneously less toxic and more efcient against P388 leukemia between 19 23 HALO. With regard to irinotecan, best results with regard to both tolerance and efcacy against Glasgow osteosarcoma were observed when the agent was administered between 711 HALO.51 Such an improvement in efcacy

Cancer Chronotherapy/Mormont and Le vi

161

FIGURE 6. Circadian rhythms in anticancer drug tolerability in laboratory mice or rats. The least toxic dosing time is indicated for each cytostatic or immunologic agent as a function of the rest-activity cycle. iv: intravenous; TNF: tumor necrosis factor.
usually has been achieved because the drug doses could be safely and selectively increased by 30 50% at the circadian time of best tolerability. Nevertheless, chronoefcacy has also been demonstrated with non toxic dose levels. The reproducible coincidence between times of highest efcacy and least toxicity for the majority of anticancer agents suggest that common mechanisms may be involved.

Mechanisms involved
The 24-hour rhythms in drug tolerability may relate to plasma pharmacokinetics; low maximal plasma concentration (CMAX) and an increased area under the curve (AUC) could favor both improved tolerability and efcacy. Despite their technical limitations in small rodents, reproducible circadian variations in plasma or urinary pharmacokinetics were demonstrated for several anticancer drugs. Times of high toxicity were found to correspond to longest elimination half-lives for methotrexate, cisplatin, carboplatin, and mitoxantrone.40 These rhythms also may result from circadian changes in the susceptibility of target tissues. Rhythms of cell proliferation in tumors, as well as cancer-associated circadian rhythm changes in healthy organs, also may play a role in the chronoefcacy of anticancer drugs. Recent studies have demonstrated the prominent role of the enzymes involved in metabolism or cell cycle regulation as major determinants of anticancer drug chronopharmacology (Table 2).50 54

FIGURE 5. KaplanMeier estimates of survival as a function of the 24-hour

rhythm parameters (A) autocorrelation coefcient at 24 hours (r24) and (B), dichotmoy coefcient (I O) assigned to 1 of 4 categories according to quartiles. Group 1: very high ( 75% quartile); Group 2: high ( 50% and 75% quartile); Group 3: low ( 25% and 50% quartile); and Group 4: very low ( 25% quartile). Comparison of the survival curves with the log-rank test was statistically signicant for both circadian rhythm parameters (r24: P 10-4; and I O: P 10-4). Reprinted with permission from Mormont MC, Bleuzen P, Waterhouse J, et al. Marked 24-h rest/activity rhythms are associated with better quality of life, better response and longer survival in patients with metastatic colorectal cancer and good performance status. Clin Cancer Res. 2000;6:3038 3045.

From Mice to Cancer Patients Chronopharmacokinetics

Short intravenous infusions of cisplatin, carboplatin, doxorubicin, 5-FU, or methotrexate or oral intake of

162

CANCER January 1, 2003 / Volume 97 / Number 1

TABLE 2 Role of Cellular Determinants of Circadian Rhythms in Tolerance for Cancer Chemotherapy
Biologic function Reduced glutathione Nonprotein sulfydryl groups Enzymatic activities Dehydropyrimidine dehydrogenase Deoxythymidine kinase Dihydrofolate reductase Topoisomerase I O6-alkylguanine methyltransferase Cellular proliferation DNA synthesis (S-phase) Drug Cisplatin, oxaliplatin

5-Fluorouracil, oxuridine Methotrexate Irinotecan Cystemustine 5-Fluorouracil Theprubicin Irinotecan Docetaxel Docetaxel

BCL-2 expression

healthy human subjects. For all these tissues, lower mean values occur between midnight and 4 a.m., and higher mean values are reported to occur between 8 a.m. and 8 p.m.27,40,65 These mechanisms of anticancer drug chronopharmacology display a similar phase relation with the rest-activity cycle in mice and in humans, despite the fact that the former are active at night and the latter during the day. Similarly, DPD activity peaks during early light in mice or rats and at early night in humans. For instance, the proportion of S-phase bone marrow cells peaks in the second half of darkness in mice and near 4 p.m. in humans. In addition, a constant-rate infusion of 5-FU results in a circadian rhythm in the plasma level both in mice and in cancer patients. Peak concentrations in 5-FU occur in the early rest span in both species if the drug is infused continuously over 1 week.

busulfan or 6-mercaptopurine (but not methotrexate) were associated with modications of plasma and/or urinary pharmacokinetics according to dosing time. Interpatient variability in circadian time-dependent pharmacokinetics also were observed.40 The most striking results stemmed from continuous intravenous infusion of 5-FU. In 5 clinical studies, when 5-FU was infused at a at rate for 24 hours to 5 days, the mean plasma 5-FU was increased from 50% to 100%, from approximately midday to approximately 1 a.m. or 4 a.m.55 60 Despite a at infusion rate, circadian changes in plasma drug levels also were observed at an equilibrium state for doxorubicin (2-day, 4-day, or 4 6-week infusion) or vindesine (4-day infusion).61 63

Chronomodulation of Chemotherapy Prerequisites

The apparent coupling between the circadian restactivity cycle and several chronopharmacology mechanisms across species has been the basis for the chronotherapy schedules that have been given to cancer patients. As a working hypothesis, expected times of least toxicity in human patients were extrapolated from those experimentally demonstrated in mice or rats, by referring them to the respective rest-activity cycle of each species (e.g., with an approximately 12hour time lag). For example, least toxicity of 5-FU occurred near 5 HALO in mice and was predicted to correspond to 4 a.m. in human subjects, resting from 11 p.m. to 7 a.m. Multichannel, programmable in-time pumps have allowed testing of the clinical relevance of the chronotherapy principle in fully ambulatory patients. For this purpose, the same chronomodulated schedule is applied to all cancer patients registered in each protocol. Today, the sinusoidal delivery of up to four anticancer drugs can be performed routinely in the patients home or during their usual activities.

Coupling of cellular rhythms to the rest-activity cycle

To our knowledge, the mechanisms that may account for circadian variations in the tolerance and efcacy of anticancer drugs have only been addressed in a few human studies published to date. The activity of dihydropyrimidine dehydrogenase (DPD), the initial enzyme for the catabolism of 5-FU, was studied around the clock in the peripheral blood mononuclear cells of seven patients diagnosed with a gastrointestinal tumor; a highly signicant group circadian rhythm was demonstrated, with an apparent inverse relation with plasma 5-FU levels during continuous infusion.64 Cell proliferation also is likely to be one mechanism involved because cells that are engaged into DNA synthesis usually display an increased susceptibility to antimetabolites or intercalating agents. The proportion of bone marrow, gut, skin, and oral mucosa cells engaged in the S-phase of the cell division cycle vary by 50% along the 24-hour time scale in

Early Clinical Trials

Early clinical trials had suggested signicant clinical benets from specic circadian timing of chemotherapy (combined vinblastine, cyclophosphamide, and methotrexate or 5-FU) or radiotherapy.66,67 The survival rate of children with acute lymphoblastic leukemia (ALL) was found to differ markedly depending on the time of maintenance chemotherapy.68 Thus, 80% of the patients dosed with 6-mercaptopurine and methotrexate in the evening were alive and disease-free 5 years after disease onset compared

Cancer Chronotherapy/Mormont and Le vi

163

with 40% of the children receiving the same drugs in the morning (P 0.001). Although this study was not randomized, the magnitude of the time-related difference is impressive. These ndings suggested that residual malignant lymphoblasts might be more susceptible to antimetabolites in the evening than in the morning. Two clinical trials compared the toxicity of two dosing times of anthracyclines and cisplatin in 30 patients with advanced ovarian carcinoma. Both randomized studies demonstrated that doxorubicin or theprubicin given 6 a.m. and cisplatin given between 4 p.m. and 8 p.m. produced signicantly fewer severe events of hematologic suppression and renal toxicity than treatment given 12 hours apart.69,70 Encouraging results also have been obtained using chronotherapy for renal carcinoma with FUDR or -interferon71,72; breast carcinoma with mitoxantrone, 5-FU, and leucovorin (LV)73; and lung carcinoma with 5-FU, LV, and cisplatin or carboplatin.74

FIGURE 7. Three drug chronotherapy against colorectal cancer. Prole of the

chronomodulated infusion of 5-uorouracil (5-FU), leucovorin (LV), and oxaliplatin (l-OHP) over 24 hours. This cycle usually is repeated automatically for 4 or 5 consecutive days.

Metastatic Colorectal Carcinoma

The clinical relevance of the chronotherapy principle was tested in a large population of patients with metastatic colorectal carcinoma using the standard methodology of clinical trials. Metastatic colorectal carcinoma is the second most common cause of cancer death in both genders and its conventional treatment methods did not appear to offer many therapeutic possibilities other than the reference combination chemotherapy of 5-FU and LV used until the mid1990s. The chronomodulated protocols involved the time-qualied infusion of 5-FU and LV, and eventually were associated with l-OHP, an active drug recognized only more recently. The maximum delivery rate of 5-FU and LV was scheduled at 4 a.m., and that of l-OHP was scheduled at 4 p.m., based on an extrapolation from experimental data (Fig. 7). Courses lasted 4 or 5 days and were repeated every 2 or 3 weeks. The tolerability, maximum dose intensities, and antitumor activity of these chronotherapy schedules were evaluated in Phase I, II, and III clinical trials, involving 2000 patients with metastatic colorectal carcinoma (Table 3).75 83 In a Phase II monoinstitutional trial, 93 patients (46 of whom had received previous chemotherapy) were treated with the chronomodulated combination of 5-FU, LV, and l-OHP over 5 days every 3 weeks; this treatment resulted in a 58% response rate (95% condence interval, 48 68%).79 A rst randomized multicenter study in 92 previously untreated patients compared at rate infusion with the chronomodulated administration of 5-FU, LV, and l-OHP. The chronomodulated regimen achieved an objective response rate of 53% compared with a rate of 32% in patients receiving at infusion (P 0.038).80 These gures were conrmed in a subsequent multicenter trial involving 186 patients; chronotherapy was found to reduce the incidence of severe mucositis vefold, halved that of functional impairment from peripheral sensory neuropathy, and reduced threefold the incidence of World Health Organization (WHO) Grade 4 toxicity requiring hospitalization compared with the at infusion regimen. This improvement in tolerability was accompanied by a signicant increase in the objective response rate from 29% to 51% (Table 4).81 The good tolerability of chronotherapy further allowed its dose intensication through the administration of a 4-day cycle every 2 weeks, and by increasing the dose of 5-FU (Fig. 8). This was achieved rst in a Phase II study involving 50 patients,82 and conrmed in a multicenter study, with apparent improvements noted in both the objective response rate and survival, which were increased to 66% and 18.5 months, respectively.83 To our knowledge, both endpoints rank among the highest that have been reported to date for the treatment of metastatic colorectal carcinoma in a multicenter setting. Taken together, the results from chronotherapy multicenter trials were suggestive of a survival improvement (Fig. 9). This issue currently is under investigation within the EORTC. Furthermore, patients with initially unresectable liver metastases could be reconsidered for liver surgery after responding to chronomodulated chemotherapy with 5-FU, LV, and l-OHP. In a group of 151 patients, retrospective analysis demonstrated that 77

164

CANCER January 1, 2003 / Volume 97 / Number 1 TABLE 3 Clinical Trials of Chronotherapy in 1275 Patients with Metastatic Colorectal Carcinoma
Phase of study I Drug(s) 5-FU 5-FU and LV 1-OHP 5-FU, LV, and 1-OHP Schedule 5 days every 3 wks 5 days every 3 wks 5 days every 3 wks 4 days every 2 wks No. of patients 35 34 25 114 Reference Le vi et al., 199575 Garu et al., 199776 Caussanel et al., 199077 Unpublished data

II m m m m 5-FU and LV id. id. 1-OHP 5-FU, LV, and 1-OHP id. id. id. id. 5 days every 3 wks 4 days every 2 wks 14 days every 4 wks 5 days every 3 wks 5 days every 3 wks 4 days every 2 wks 4 days every 2 wks 4 days every 2 wks 4 days every 2 wks 43 100 67 30 93 54 50 62 90 Cure et al., 200088 Cure et al., 200289 Bjarnason et al., 199890 Proc. ASCO Le vi et al., 199378 Le vi et al., 199279 Brienza et al., 199391 Proc. ASCO Bertheault-Cvitkovic et al., 199682 Le vi et al., unpublished data Le vi et al., 199983

m m m III m m m

5-FU, LV, and 1-OHP at vs. chrono id. Chrono 5-FU and LV 1-OHP

5 days every 3 wks 5 days every 3 wks

92 186 200

Le vi et al., 199480 Le vi et al., 199781 Giacchetti et al., 200092 and Giacchetti et al., 200293

5-FU: 5-uorouracil; id: identical; LV: leucovorin; 1-OHP: oxaliplatin; m: multicentric; chrono: chronomodulated.

TABLE 4 Main Results of a Randomized Multicenter Trial Comparing Flat Versus Chronomodulated Chemotherapy in 186 Patients With Metastatic Colorectal Carcinoma
Effect Hospitalization for toxicities Severe mucositis Functional impairment (periph sensory neuro) Tumor response 50% Flat 31a 76 31 29 Chrono 10 14 16 51 P value 0.001 0.0001 0.01 0.003

DISCUSSION AND PERSPECTIVES

Mechanisms of the Chronopharmacology of Anticancer Drugs
Malignant tumors and cancer-bearing hosts may exhibit nearly normal or markedly altered circadian rhythms. Rhythm alterations appear to depend on tumor type, growth rate, and level of differentiation, both in animal and human tumors, and they usually worsen along the course of disease progression. Some available experimental and clinical articles have provided interesting hypotheses for understanding the relevance of biologic rhythms for cancer treatment. Comparative human studies22-24 evidenced a phase opposition between DNA synthesis rhythms in healthy target organs and in tumor; this nding suggests the involvement of cell cycle regulation mechanisms in the chronopharmacology of anticancer drugs. The same holds true for the rhythms in the activity of catabolic enzymes for cytostatics, as demonstrated by the study of DPD.64 The majority of tumors also produce cytokines and hematologic growth factors, which can modify the central circadian system, or peripheral circadian rhythms, as shown in healthy subjects.85 The interactions between the circadian system and a growing tumor will require accurate study

Flat: at infusion; Chrono: chronomodulated infusion; periph sensory neuro: peripheral sensory neuropathy. a Percentage of patients. Data adapted from: Le vi F, Zidani R, Misset JL, for the International Organization for Cancer Chronotherapy. Randomized multicentre trial of chronotherapy with oxaliplatin, uorouracil, and folinic acid in metastatic colorectal cancer. Lancet. 1997;350:681 686.

patients could benet from surgery after responding to chronotherapy, and that among these patients, 58 patients could undergo a complete resection of their metastases; the survival rate of the patients who underwent surgery was 50% after 5 years.84 These results clearly show that chronotherapy, either alone or associated with surgery, improves the therapeutic index of chemotherapy for metastatic colorectal carcinoma.

Cancer Chronotherapy/Mormont and Le vi

165

FIGURE 9. The median survival expressed as a function of the response rate FIGURE 8. Relation between infusion modality, dose intensity of 5-uorouracil (5-FU), and response outcome in six clinical trials involving patients with metastatic colorectal carcinoma who received either 5-FU and leucovorin (LV), or 5-FU, LV, and oxaliplatin (l-OHP) as rst-line chemotherapy for metastases. Treatment modalities were comprised of at infusion chemotherapy over 5 days with a 16-day interval between cycles, chronomodulated chemotherapy over 5 days with a 16-day interval between cycles, chronomodulated chemotherapy over 4 days with a 10-day interval between cycles, and dose intensication. Each dot represents a treatment group from one clinical trial. The addition of l-OHP, chronomodulation, and dose intensication was shown to clearly increase the response rate. in ve clinical trials involving patients with metastatic colorectal carcinoma who received 5-uorouracil (5-FU), leucovorin (LV), and oxaliplatin (l-OHP) as rstline chemotherapy for metastases. The modalities were the same as those shown in Figure 8. Higher response rates appeared to be associated with longer median survival times.

through experimental models, which allow manipulation of the circadian system to mimic pathologic situations. Currently available models include mutant mice with longer circadian period than controls,2 light-induced functional rhythm alterations in rats,86 lesions of the SCN in mice,87 and computer simulation. Finally, the search for periodic genes in healthy tissues and tumors is one of the next challenging steps in developing a greater understanding of the mechanisms of anticancer drug chronopharmacology, both in experimental models and in cancer patients. In humans, the functioning of the circadian system is reected by behavioral and physiologic circadian rhythms, such as locomotor activity, body temperature, and several neuroendocrine secretions. However, the majority of these rhythms cannot be evaluated in large cohort studies. This is the case for measures of proliferation rhythms in tumor or healthy tissue, which imply repeated biopsies, or for the assessment of the rhythm in central temperature, which requires the patient to wear a rectal probe for at least 72 hours. Occasionally, a two-time sampling procedure can provide an estimate of a given rhythm, if one

of the timepoints corresponds to the group maximum. This approach was validated in an analysis of cortisol circadian rhythm both in controls and in cancer patients, in which blood sampling at 8:00 a.m. and 4 p.m. provided an accurate estimate of individual circadian amplitude.31 Similar conclusions were suggested for DNA synthesis in human bone marrow.27,28 Nevertheless, studies with few timepoints will not allow the detection of short-period rhythms, which often have been suspected to replace circadian rhythms in late cancer stages. The collection of dense longitudinal time series is a necessary step toward understanding cancer-associated circadian system alterations. To our knowledge to date, only the assessment of the restactivity rhythm by wrist actigraphy meets such methodologic constraints in cancer patients.

Synchronization of Patients with Altered Circadian Rhythms

The rest-activity rhythm was found to be a positive prognostic factor of both tumor response and survival in 200 patients with metastatic colorectal carcinoma.38 The rest-activity rhythm provided additional prognostic information regarding patients maximum response to treatment and survival potential to that of well-known clinical factors, which reect tumor burden and patient general condition. The same study also documented the existence of a link between the rest-activity rhythm and the welfare of cancer patients. Marked rest-activity rhythms were associated

166

CANCER January 1, 2003 / Volume 97 / Number 1

with high functional scores and low symptom scores.39 This was not surprising because several variables evaluated in quality of life questionnaires, such as locomotor activity, sleep, and psychomotor performance, are organized along the 24-hour time scale. The statistical analysis indicated that the rest-activity rhythm did not simply reect confounding factors such as fatigue or pain. To our knowledge the current study is the rst to establish that an output rhythm of the circadian clock is a signicant predictor of survival in a prospective clinical trial. Although the issue of a causal relation was not addressed, these results provide novel perspectives toward a better understanding of the impact of cancer-induced circadian system alterations on host physical and psychologic balance. Furthermore, the above-mentioned investigation indicates that circadian function in each individual patient may provide a pertinent explanation for interindividual differences in the outcome of patients with metastatic colorectal carcinoma. The scope of application of this concept now needs to be assessed with regard to other human tumors and other chemotherapy schedules. These results also call for devising specic therapies to restore the circadian rest-activity rhythm; such therapies could include chronobiotics such as melatonin and its analogs, light therapy, sleep management, and psychosocial support. Such specic treatments for circadian dysfunctions may help to improve the status and/or the outcome of cancer patients, and contribute to enhancing the therapeutic efcacy of chemotherapy. This issue will have to be addressed in prospective clinical trials.

modulated versus conventional therapy are being planned for the adjuvant treatment of colorectal carcinoma, as well as for carcinomas of the head and neck and biliary duct. More Phase III trials will be needed to rmly establish chronotherapy in medical oncology.94,95 Therefore, a chronotherapy study group was created in 1996 within the EORTC. This group has established links with the Radiation Therapy Oncology Group (RTOG) in the U.S. and the National Cancer Institute in Canada.

REFERENCES
1. Plautz JD, Kaneko M, Hall JC, et al. Independent photoreceptive circadian clocks throughout Drosophila. Science. 1997;278:16321635. Vitaterna MH, King DP, Chang AM, et al. Mutagenesis and mapping of a mouse gene, Clock, essential for circadian behavior. Science. 1994;264:719 725. Reinberg A, Touitou Y, Restoin A, Migraine C, Le vi F, Montagner H. The genetic background of circadian and ultradian rhythm patterns of 17-hydroxycorticosteroids: a cross-twin study. J Endocrinol. 1985;105:247253. Linkowski P, Van Onderbergen A, Kerkhofs M, Bosson D, Mendlewicz J, Van Cauter E. Twin study of the 24-h cortisol prole: evidence for genetic control of the human circadian clock. Am J Physiol. 1993;264:E173E181. Mendlewicz J, Linkowski P, Kerkhofs M, Leproult R, Copinschi G, Van Cauter E. Genetic control of 24-hour growth hormone secretion in man: a twin study. J Clin Endocrinol Metab. 1999;84:856 862. Tei H, Okamura H, Shigeyoshi Y, et al. Circadian oscillation of a mammalian homologue of the Drosophila period gene. Nature. 1997;389:512516. Moore RY, Eichler VB. Central neural mechanism in diurnal rhythm regulation and neuroendocrine responses to light. Psychoneuroendocrinology. 1972;1:265279. Stephan FK, Zucker I. Circadian rhythms in drinking behaviour and locomotor activity of rats are eliminated by hypothalamic lesions. Proc Natl Acad Sci U S A. 1972;69:1583 1586. Ralph MR, Foster RG, Davis FC, Menaker M. Transplanted suprachiasmatic nucleus determines circadian period. Science. 1990;247:975978. Rosenwasser AM, Wirz-Justice A. Circadian rhythms and depression: clinical and experimental models. In: Redfern P, Lemmer B, editors. Physiology and pharmacology of biological rhythms. Berlin: Springer-Verlag, 1997:457 485. Mormont MC, Le vi F. Circadian-system alterations during cancer processes: a review. Int J Cancer. 1997;70:241247. Burns RE, Scheving LE, Tsai TH. Circadian rhythms in DNA synthesis and mitosis in normal mice and in mice bearing the lewis lung carcinoma. Eur J Cancer. 1979;15:233242. Barbason H, Betz EH. Liver cell control after discontinuation of DENA feeding in hepatocarcinogenesis. Eur J Cancer. 1981;17:149 156. Bouzahzah B. Inuence de diffe rents promoteurs sur la prolife ration cellulaire du foie de rat normal et pre cance reux : ro le des cortico des et des rythmes circadiens [Ph.D. thesis]. Lia ` ge, Belgium: Universite de Lie ` ge, 1990.

2.

3.

4.

5.

6.

7.

Development of New Clinical Trials

The concept of cancer chronotherapy, which has comprised of the extrapolation of the least toxic times of chemotherapy from mice to cancer patients, has been validated in Phase III clinical trials for the treatment of patients with metastatic colorectal carcinoma. According to previous experience, there are two ways by which the chronomodulation of chemotherapy can be benecial to the patients. The dosing timerelated reduction in chemotherapy toxicity may result in an improvement in quality of life, although high doses of chemotherapy are being delivered. Conversely, the administration of a higher maximum tolerated dose at the least toxic circadian time, compared with other dosing times, may result in an improvement in efcacy outcomes (i.e., tumor response and/or survival). The clinical relevance of chronotherapy currently is being investigated along those lines in the outcome of patients with metastatic breast or pancreatic carcinomas. Multicenter clinical trials comparing chrono8.

9.

10.

11. 12.

13.

14.

Cancer Chronotherapy/Mormont and Le vi

15. Kampfschmidt RF, Upchurch HF. Daily variation of body temperature, liver catalase activity, and plasma iron concentration in normal and tumor-bearing rats. Proc Soc Exp Biol Med. 1970;134:527529. 16. Sothern RB, Hrushesky WJM, Halberg F, Haus E, Kennedy BJ. Circadian temperature rhythm in LOU rats with and without immunocytoma during polychronochemotherapy with adriamycin and cis-diamminedichloroplatinum. Chronobiologia. 1978;5:216. 17. Hori K, Zhang QH, Saito S. Variation of growth rate of rat tumor during a light-dark cycle: correlation with circadian uctuation in tumor blood ow. Br J Cancer. 1995;71:1163 1168. 18. Voutilainen A. Uber die 24-stunden-rhythmik der mitozfrequenz in malignen tumoren. Acta Pathol Microbiol Scand. 1953;99(Suppl):1104. 19. Tahti E. Studies of the effect of X-irradiation on 24 hour variations in the mitotic activity in human malignant tumors. Acta Pathol Microbiol Scand. 1956;117:1 61. 20. Garcia-Sainz M, Halberg F. Mitotic rhythms in human cancer reevaluated by electronic computer programs. Evidence for chronopathology. J Natl Cancer Inst.. 1966;37:279 292. 21. Zagula-Mally ZW, Cardoso SS, Williams D, et al. Time point differences in skin mitotic activity of actinic keratoses and skin cancers. In: Reinberg A, Halberg F, editors. Chronopharmacology. New York: Pergamon Press, 1979:399 402. 22. Klevecz R, Shymko R, Braly P. Circadian gating of S phase in human ovarian cancer. Cancer Res. 1987;47:6267 6271. 23. Klevecz R, Braly P. Circadian and ultradian cytokinetic rhythms of spontaneous human cancer. Ann NY Acad Sci. 1991;618:257276. 24. Smaaland R, Lote K, Sothern RB, et al. DNA synthesis and ploidy in non-Hodgkins lymphomas demonstrate variation depending on circadian stage of cell sampling. Cancer Res. 1993;53:3129 3138. 25. Gautherie M, Gros C. Circadian rhythm alteration of skin temperature in breast cancer. Chronobiologia. 1974;4:117. 26. Manseld CM, Carabasi RA, Wells W, et al. Circadian rhythm in the skin temperature of normal and cancerous breasts. Int J. Chronobiol. 1973;1:235243. 27. Smaaland R, Laerum OD, Lote K, et al. DNA synthesis in human bone marrow is circadian stage dependent. Blood. 1991;77:26032611. 28. Smaaland R, Abrahamsen JF, Svardal AM, et al. DNA cell cycle distribution and glutathione (GSH) content according to circadian stage in bone marrow of cancer patients. Br J Cancer, 1992;66:39 45. 29. Tamarkin L, Danforth D, Lichter A, et al. Decreased nocturnal plasma melatonin peak in patients with estrogen receptor positive breast cancer. Science. 1982;216:10031005. 30. Bartsch C, Bartsch H, Flu chter SH, Mecke D, Lippert TH. Diminished pineal function coincides with disturbed circadian endocrine rhythmicity in untreated primary cancer patients. Ann N Y Acad Sci. 1994;719:502525. 31. Mormont MC, Hecquet B, Bogdan A, et al. Selection and validation of a non-invasive test of cortisol circadian rhythm in cancer patients and control subjects. Int J Cancer. 1998; 78:421 424. 32. Touitou Y, Le vi F, Bogdan A, et al. Circadian desynchronization of blood variables in patients with metastatic breast cancer. Role of prognostic factors. J Cancer Res Clin Oncol. 1995;121:181188. 33. Touitou Y, Bogdan A, Le vi F, et al. Disruption of the circadian patterns of serum cortisol in breast and ovarian cancer

167

34.

35.

36.

37.

38.

39.

40.

41.

42.

43.

44.

45.

46.

47.

48.

49.

50.

patients. Relationships with tumor marker antigens. Br J. Cancer. 1996;74:1248 1252. Benavides M. Cancer avance de lovaire: approche chronobiologique comme nouvelle strate gie du traitement et de la surveillance clinique et biologique [Ph.D. thesis]. Paris: Universite Paris XI, 1991. Sephton SE, Sapolsky RM, Kraemer HC, Spiegel D. Diurnal cortisol rhythm as a predictor of breast cancer survival. J Natl Cancer Inst. 2000;92:994 1000. Dowse HB, Ringo JM. The search for hidden periodicities in biological time series revisited. J Theor Biol. 1989;139:487 515. Minors DS, Akerstedt T, Atkinson G, et al. The difference between activity when in bed and out of bed. I. Healthy subjects and selected patients. Chronobiol Int. 1996;13:27 34. Mormont MC, Bleuzen P, Waterhouse J, et al. Marked 24-h rest/activity rhythms are associated with better quality of life, better response and longer survival in patients with metastatic colorectal cancer and good performance status. Clin Cancer Res. 2000;6:3038 3045. Mormont MC, Waterhouse J. Contribution of the rest-activity circadian rhythm to quality of life in cancer patients. Chronobiol Int. 2002;19:313323. Mormont MC, Boughattas N, Le vi F. Mechanisms of circadian rhythms in the toxicity and the efcacy of anticancer drugs: relevance for the development of new analogs. In: Lemmer B, editor. Chronopharmacology: cellular and biochemical interactions. New York: Marcel Dekker, 1989:395 437. Le vi F. Chronopharmacology of anticancer agents. In: Redfern P, Lemmer B, editors. Physiology and pharmacology of biological rhythms. Berlin: Springer-Verlag, 1997:299 331. Le vi F, Hrushesky W, Blomquist CH, et al. Reduction of cis-diamminedichloroplatinum nephrotoxicity in rats by optimal circadian drug timing. Cancer Res. 1982;42:950 955. Boughattas NA, Le vi F, Hecquet B, et al. Circadian time dependence of murine tolerance for carboplatin. Toxicol Appl Pharmacol. 1988;96:233247. Boughattas N, Le vi F, Fournier C, et al. Circadian rhythm in toxicities and tissue uptake of 1,2-diamminocyclohexane (trans-1) oxalatoplatinum (II) in mice. Cancer Res. 1989;49: 33623368. Le vi F, Mechkouri M, Roulon A, et al. Circadian rhythm in tolerance of mice for the new anthracycline analog 4-tetrahydropyranyladriamycin (THP). Eur J Cancer Clin Oncol. 1985;121:12451251. Le vi F, Tampellini M, Metzger G, et al. Circadian changes in mitoxantrone toxicity in mice: relationship with plasma pharmacokinetics. Int J Cancer. 1994;59:543547. Mormont MC, Beretska JS, Mushiya T, et al. Circadian dependency of vinblastine toxicity. Ann Rev Chronopharmacol. 1986;3:187190. Filipski E, Amat S, Lemaigre G, et al. Relationship between circadian rhythm of vinorelbine toxicity and efcacy in P388-bearing mice. J Pharmacol Exp Ther. 1999;289:231 235. Granda TG, Le vi F. Tumor-based rhythms of anticancer efcacy in experimental models. Chronobiol Int. 2002;19:21 41. Tampellini M, Filipski E, Liu X-H, et al. Circadian rhythm in docetaxel tolerability and efcacy in mice. Cancer Res. 1998; 58:3896 3904.

168

CANCER January 1, 2003 / Volume 97 / Number 1

human solid tumors. Cancer Chemother Pharmacol. 1979;3: 197202. Deka AC. Application of chronobiology to radiotherapy to tumours of the oral cavity [M.D. thesis]. Chandigarh, India: Post-Graduate Institute of Medical Education and Research, 1975. Rivard G, Infante-Rivard C, Hoyoux C, et al. Maintenance chemotherapy for childhood acute lymphoblastic leukemia: better in the evening. Lancet 1985;ii:1264 1266. Hrushesky W. Circadian timing of cancer chemotherapy. Science. 1985;228:7375. Le vi F, Benavides M, Chevelle C, et al. Chemotherapy of advanced ovarian cancer with 4-0-tetrahydropyranyl adriamycin (THP) and cisplatin: a phase II trial with an evaluation of circadian timing and dose intensity. J Clin Oncol. 1990;8:705714. Hrushesky W, von Roemeling R, Lanning RM, et al. Circadian-shaped infusions of oxuridine for progressive metastatic renal cell carcinoma. J Clin Oncol. 1990;8:1504 1513. Depre s-Brummer P, Le vi F, Di Palma M, et al. A phase I trial of 21-day continuous venous infusion of alpha-interferon at circadian rhythm modulated rate in cancer patients. J Immunother. 1991;10:440 447. Depre s-Brummer P, Bertheault-Cvitkovic F, Le vi F, et al. Circadian rhythm modulated (CRM) chemotherapy of metastatic breast cancer with mitoxantrone, 5-uorouracil and folinic acid: preliminary results of a Phase I trial. J Infus Chemother. 1995;5:144 147 Focan C, Denis B, Kreutz F, Focan-Henrard D, Le vi F. Ambulatory chronotherapy with 5-uorouracil, folinic acid and carboplatin for advanced non-small cell lung cancer. A phase II feasibility trial. J Infus Chemother. 1995;5:148 152. Le vi F, Soussan A, Adam R, et al. A Phase I-II trial of ve-day continuous intravenous infusion of 5-uorouracil delivered at circadian rhythm modulated rate in patients with metastatic colorectal cancer. J Infus Chemother. 1995;5:153158. Garu C, Le vi F, Aschelter AM, et al. A Phase I trial of ve day chronomodulated infusion of 5-uorouracil and l-folinic acid in patients with metastatic colorectal cancer. Eur J Cancer. 1997;33:1566 1571. Caussanel JP, Le vi F, Brienza S, et al. Phase I trial of 5-day continuous venous infusion of oxaliplatin at circadian rhythm-modulated rate compared with constant rate. J Natl Cancer Inst. 1990;82:1046 1050. Le vi F, Perpoint B, Garu C, et al. Oxaliplatin activity against metastatic colorectal cancer. A Phase II study of 5-day continuous venous infusion at circadian rhythm modulated rate. Eur J Cancer. 1993;29:1280 1284. Le vi F, Misset JL, Brienza S, et al. A chronopharmacologic Phase II clinical trial with 5-uorouracil, folinic acid and oxaliplatin using an ambulatory multichannel programmable pump. High antitumor effectiveness against metastatic colorectal cancer. Cancer. 1992;69:893900. Le vi F, Zidani R, Vannetzel JM, et al. Chronomodulated versus xed infusion rate delivery of ambulatory chemotherapy with oxaliplatin, 5-uorouracil and folinic acid in patients with colorectal cancer metastases. A randomized multiinstitutional trial. J Natl Cancer Inst. 1994;86:1608 1617. Le vi F, Zidani R, Misset JL, for the International Organization for Cancer Chronotherapy. Randomized multicentre trial of chronotherapy with oxaliplatin, uorouracil, and folinic acid in metastatic colorectal cancer. Lancet. 1997;350: 681 686.

51. Granda TG, DAttino RM, Filipski E, et al. Circadian optimization of irinotecan and oxaliplatin efcacy in mice with Glasgow osteosarcoma. Br J Cancer. 2002;86:999 1005. 52. Zhang R, Lu Z, Liu T, et al. Relationship between circadiandependent toxicity of 5-uorodeoxyuridine and circadian rhythms of pyrimidine enzymes: possible relevance to uoropyrimidine chemotherapy. Cancer Res. 1993;53:2816 2822. 53. Li XM, Metzger G, Filipski E, et al. Pharmacologic modulation of reduced glutathione (GSH) circadian rhythms by buthionine sulfoximine (BSO): relationship with cisplatin (CDDP) toxicity in mice. Toxicol Appl Pharmacol. 1997;143:281290. 54. Ohdo S, Makinosumi T, Ishizaki T, et al. Cell cycle-dependent chronotoxicity of irinotecan hydrochloride in mice. J Pharmacol Exp Ther. 1997;283:13831388. 55. Petit E, Milano G, Le vi F, et al. Circadian varying plasma concentration of 5-FU during 5-day continuous venous infusion at constant rate in cancer patients. Cancer Res. 1988; 48:1676 1679. 56. Thiberville L, Compagnon C, Moore N, et al. Plasma 5-uorouracil and alpha-uoro-beta-alanin accumulation in lung cancer patients treated with continuous infusion of cisplatin and 5-uorouracil. Cancer Chemother Pharmacol. 1994;35:64 70. 57. Metzger G, Massari C, Etienne MC, et al. Spontaneous or imposed circadian changes in plasma concentrations of 5-uorouracil coadministered with folinic acid and oxaliplatin: relationship with mucosal toxicity in cancer patients. Clin Pharmacol Ther. 1994;56:190 201. 58. Fleming GF, Schilsky RL, Mick R, et al. Circadian variation of 5-uorouracil (5-FU) and cortisol plasma levels during continuous-infusion 5-FU and leucovorin (LV) in patients with hepatic or renal dysfunction. Proc Am Soc Clin Oncol. 1994; 13:139. 59. Bressole F, Joulia JM, Pinguet F, et al. Circadian rhythm of 5-uorouracil population pharmacokinetics in patients with metastatic colorectal cancer. Cancer Chemother Pharmacol. 1999;44:295302. 60. Takimoto CH, Yee LK, Venzon DJ, et al. High inter- and intrapatient variation in 5-uorouracil plasma concentrations during a prolonged drug infusion. Clin Cancer Res. 1999;5:13471352. 61. Le vi F, Metzger G, Bailleul F, et al. Circadian-varying plasma pharmacokinetics of doxorubicin (DOX) despite continuous infusion at constant rate [abstract]. Proc Am Assoc Cancer Res. 1986;27:693. 62. Focan C, Doalto L, Mazy V, et al. Vindesine en perfusion continue de 48 heures (suivie de cisplatine) dans le cancer pulmonaire avance . Donne es chronopharmacocine tiques et efcacite clinique. Bull Cancer. 1989;76:909 912. 63. Squalli A, Oustrin J, Houin G, et al. Clinical chronopharmacokinetics of doxorubicin (DXR). Ann Rev Chronopharmacol. 1989;5:393396. 64. Harris B, Song R, Soong S, Diasio RB. Relationship between dihydropyrimidine dehydrogenase activity and plasma 5-uorouracil levels: evidence for circadian variation of plasma drug levels in cancer patients receiving 5-uorouracil by protracted continuous infusion. Cancer Res. 1990;50: 197201. 65. Buchi KN, Moore JG, Hrushesky WJM, et al. Circadian rhythm of cellular proliferation in the human rectal mucosa. Gastroenterology. 1991;101:410 415. 66. Focan C. Sequential chemotherapy and circadian rhythm in

67.

68.

69. 70.

71.

72.

73.

74.

75.

76.

77.

78.

79.

80.

81.

Cancer Chronotherapy/Mormont and Le vi

82. Bertheault-Cvitkovic F, Jami A, Ithzaki M, et al. Biweekly intensied ambulatory chronomodulated chemotherapy with oxaliplatin, 5-uorouracil and folinic acid in patients with metastatic colorectal cancer. J Clin Oncol. 1996;14: 2950 2958. 83. Le vi F, Zidani R, Brienza S, et al. A multicenter evaluation of intensied, ambulatory chronomodulated chemotherapy with oxaliplatin, 5-uorouracil and leucovorin as initial treatment of patients with metastatic colorectal carcinoma. Cancer. 1999;85:25322540. 84. Giacchetti S, Itzhaki M, Gruia G, et al. Long term survival of patients with unresectable colorectal cancer liver metastases following infusional chemotherapy with 5-uorouracil, leucovorin, oxaliplatin and surgery. Ann Oncol. 1999;10:663 669. 85. Born J, Lange T, Hansen K, Mo lle M, Fehm HL. Effects of sleep and circadian rhythm on human circulating immune cells. J Immunol.1997;158:4454 4464. 86. Depre s-Brummer P, Le vi F, Metzger G, Touitou Y. Lightinduced suppression of the rat circadian system. Am J Physiol. 1995;268:r1111r1116. 87. Filipski E, King VM, Li XM, et al. Host circadian clock as a control point in tumor progression. J Natl Cancer Inst. 2002; 94:690 697. 88. Cure H, Chevalier V, Adenis A, et al. Phase II trial of chronomodulated infusion of high-dose 5-uorouracil and lfolinic acid in previously untreated patients with metastatic colorectal cancer. Anticancer Res. 2000;20:4649 4654. 89. Cure H, Chevalier V, Adenis A, et al. Phase II trial of chronomodulated infusion of high-dose 5-uorouracil and l-

169

90.

91.

92.

93.

94. 95.

folinic acid in previously untreated patients with metastatic colorectal cancer. J Clin Oncol. 2002;20:11751181. Bjarnason GA, Marsh R, Chu NM, Kerr I, Franssen E. Phase II study of 5-uorouracil (FU) and leucovorin (LV) by a 14 day chronomodulated infusion in patients with metastatic colorectal cancer (abstr. #1058). 34th Annual Meeting Am. Soc. Clin. Oncol., Orlando, USA, May 16 19, 1998. Proc Am Soc Clin Oncol 1998;17:275a. Brienza S, Le vi F, Valori VM, et al. Intensied (every 2 weeks) chronotherapy with 5-uorouracil (5-FU), folinic acid (FA) and oxaliplatin (L-OHP) in previously treated patients with metastatic colorectal cancer. (abstr. #577). 29th Annual Meeting Am. Soc. Clin. Oncol., Orlando, USA, May 16 18, 1993. Proc Am Soc Clin Oncol. 1993;12:197. Giacchetti S, Perpoint B, Zidani R, et al., for the International Organization of Cancer. Phase III multicenter randomized trial of oxaliplatin added to chronomodulated uorouracilleucovorin as rst line treatment of metastatic colorectal cancer. J Clin Oncol. 2000;18:136 147. Giacchetti S, Perpoint B, Zidani R, et al., for the International Organization of Cancer. Phase III multicenter randomized trial of oxaliplatin added to chronomodulated uorouracilleucovorin as rst line treatment of metastatic colorectal cancer. Update. Classic papers and current comments, highlights of gastrointestinal cancer research. A best of JCO. J Clin Oncol. 2002;6:885 897. Le vi F. From circadian rhythms to cancer chemotherapeutics. Chronobiol Int. 2002;19:119. Condert B, Focan C, Donato di Paola E, Le vi F for the EORTC chronotherapy Group (CTG). Its time for chronotherapy! Eur J Cancer. 2002;38(Suppl 4):50 53.