Irinotecan Plus 5-FU and Leucovorin in Advanced Colorectal Cancer: North American Trials

ABSTRACT: Both fluorouracil (5-FU) and irinotecan (CPT-11 [Camptosar]) have shown activity in metastatic colorectal cancer and are approved for its treatment in the United States. Preclinical experiments in cell cultures and human tumor xenografts have indicated potential synergy when irinotecan is combined with 5-FU and leucovorin. The synergy appears to be sequence-dependent and is optimal when irinotecan exposure precedes 5-FU exposure by at least 24 hours. Four North American trials have been reported in which the three drugs were used together in either simultaneous, sequential, or alternating schedules. All three schedules showed activity in patients with metastatic colorectal cancer. The concern that diarrhea, which can be a dose-limiting toxicity with both irinotecan and 5-FU, would prevent the two drugs from being combined in reasonable doses has not proven to be a clinical issue. Phase III trials comparing the combination of the three drugs in a variety of schedules against 5-FU plus leucovorin alone are currently under way or in the planning stages.[ONCOLOGY 12(Suppl 6):59-63, 1998]

Introduction

An estimated 131,000 new cases of colorectal cancer were diagnosed in the United States in 1997.[1] The current standard of practice among most North American oncologists is to administer a potentially curative course of chemotherapy in most patients with stage III colorectal cancer and a palliative course of chemotherapy in many patients with stage IV disease. The two drugs that are currently approved by the FDA for the treatment of colon cancer are fluorouracil (5-FU) and irinotecan (CPT-11 [Camptosar]).

Fluorouracil is known to be active in the setting of advanced colorectal cancer. Pooled results from 10 trials in which colorectal cancer patients with advanced disease were treated with intermittent-bolus 5-FU combined with leucovorin led to an overall response rate of 23% and a median survival of 12 months.[2] Fluorouracil plus leucovorin is also an effective adjuvant therapy. In patients with resected stage III colon cancer, 6 months of treatment with 5-FU/leucovorin for 5 consecutive days each month decreased the odds of recurrence by approximately 33%.[3,4]

Irinotecan also is active in advanced colorectal cancer. A recently completed North Central Cancer Treatment Group (NCCTG) trial reported a 26% response rate (95% confidence interval [CI], 11.9% to 44.6%) to irinotecan in patients who had not previously received chemotherapy for advanced colorectal cancer.[5] In the cohort of patients with advanced colorectal cancer that was refractory to 5-FU and leucovorin, this NCCTG trial reported a response rate of 13% with irinotecan (95% CI, 7.1% to 22.1%).[5] Both of these outcomes are typical of published reports cited elsewhere in this monograph.

Based on the NCCTG data and results from other reported studies, in 1996 the FDA approved irinotecan for the treatment of patients with advanced colorectal cancer that is refractory to 5-FU. To date, the activity of irinotecan, either as a single agent or in combination with other chemotherapeutic agents, has not been tested as adjuvant therapy for stage II or III patients; therefore, its potential value in such patients remains unknown. Future trials integrating irinotecan into postsurgical treatment regimens for stage III patients are in the planning stages under the aegis of the National Cancer Institute’s cooperative research group clinical trial program.

The NCCTG and other trials have clearly shown that some tumors that grow during treatment with 5-FU and leucovorin will shrink when exposed to irinotecan. This observation implies a lack of tumor cross-resistance to the two agents, at least in a proportion of metastatic human colon cancers, and led to a number of experiments in which irinotecan and 5-FU were combined in vitro and in human tumor xenograft models.

Promising findings of these experiments led to treatment trials employing a number of different dose and schedule strategies in patients with advanced colorectal cancer. These trials were designed to determine the maximum tolerated doses (MTDs) and response rates of the combination of irinotecan and 5-FU in patients with advanced colorectal cancer, as well as to provide survival data.

Because diarrhea is a major dose-limiting side effect of both irinotecan and 5-FU/leucovorin, the possibility of life- threatening, additive, or even synergistic toxicity exists. Investigators using the combination of the three drugs have taken this into account in their phase I trial designs.

This review addresses three main topics. First, data on interactions between 5-FU and irinotecan observed in a number of cell culture and human tumor xenograft experiments will be summarized. Second, recent or ongoing North American trials in which irinotecan and 5-FU plus leucovorin have been administered concurrently, sequentially, and in an alternating fashion will be reviewed. Finally, relevant toxicity data, as well as current knowledge about the mechanisms and treatments of irinotecan- and 5-FU-induced gastrointestinal toxicity, will be presented and discussed.

Preclinical Data

Fluorouracil

Fluorouracil inhibits the enzyme thymidylate synthase (TS), thus interfering with the formation of new strands of DNA during replication. Fluorouracil is also incorporated into RNA and DNA, where it inhibits DNA synthesis.[6] Leucovorin is a reduced folate, which, when combined with 5-FU, augments 5-FU cytotoxicity by increasing the inhibition of TS by the 5-FU metabolite 5-fluoro-2¢-deoxyuridine monophosphate (FdUMP).[7]

The role of 5-FU in the treatment of colorectal cancer has been explored for over 40 years. Modulation of 5-FU by leucovorin has been demonstrated to improve response rate, but not median survival, over bolus 5-FU alone.[2] Nevertheless, 5-FU, with or without leucovorin, has become the standard therapy for patients with advanced colon cancer in North America.

Irinotecan

Irinotecan binds and stabilizes topoisomerase I as it works to uncoil DNA during cell division. This stabilized complex of inhibitor-enzyme-DNA halts advancing DNA replication forks, resulting in double-strand DNA breaks and consequent apoptosis.[8] Irinotecan is converted by carboxylesterases to its more active metabolite, 7-ethyl-10 hydroxy-camptothecin, or SN-38.[9,10]

Irinotecan Plus 5-FU and Leucovorin

The clinical activity of both 5-FU/leucovorin and irinotecan in patients with metastatic colorectal cancer makes the combination of these therapies clinically attractive. There is reason to believe, however, that this therapeutic combination may result in antagonism. Fluorouracil is converted to FdUMP, leading to inhibition of TS, depletion of deoxythymidine triphosphate (dTTP) pools, and inhibition of DNA synthesis, resulting in G1/S cell-cycle arrest. The active metabolite of irinotecan, SN-38, stabilizes the covalent complex between topoisomerase I and nuclear DNA, leading to DNA double-strand breaks and accumulation of cells in the G₂ phase of the cell cycle.

Hence, 5-FU may inhibit the DNA synthesis required for the cytotoxicity of SN-38. Conversely, SN-38 may cause cells to accumulate in the G₂ rather than in the S phase, when they would be most sensitive to 5-FU plus leucovorin.

Mullany et al addressed these issues in studies of SN-38 plus 5-FU and leucovorin in HCT-8 cells in vitro. They observed a sequence-dependent interaction of SN-38 and 5-FU plus leucovorin. The combination was less than additive when cells were treated with simultaneous administration of SN-38 and 5-FU plus leucovorin or when 5-FU plus leucovorin was followed by SN-38, but was synergistic when SN-38 was followed by 5-FU plus leucovorin.[11] Other groups have found similar results.[12-18]

Mullany et al further demonstrated that SN-38 resulted in accumulation of cells in the S phase when therapeutically achievable nanomolar concentrations of SN-38 were used for time periods corresponding to three serum half-lives in vivo. Moreover, this S-phase-slowing was accompanied by inhibition of TS activity in the face of constant TS protein levels. Further studies revealed that SN-38 increased dTTP pools.

Taken together, these results are consistent with a model in which elevated dTTP pools account for the observed synergy of SN-38 followed by 5-FU plus leucovorin. The increased dTTP pools after SN-38 therapy inhibit TS by depletion of the normal substrate, deoxyuridine monophosphate (dUMP), and enhance 5-FU-associated inhibition of TS. In addition, the dTTP-associated inhibition of thymidine kinase decreases salvage thymidylate synthesis. Thus, the preclinical studies define a rational schedule for clinical evaluation of irinotecan plus 5-FU and leucovorin and explain, at least in part, the mechanism of this interaction.

North American Clinical Trials

There are four reports of North American studies in which irinotecan, 5-FU, and leucovorin have been combined either simultaneously, sequentially, or in an alternating fashion. In all of the American regimens reported to date, 5-FU was given by intermittent bolus injection or short (< 2-hour) infusion. European trials of the three-drug combination are described elsewhere in this monograph . A number of European trials administered the 5-FU as a short infusion over 22 to 48 hours in a manner similar to the schedules commonly used when 5-FU and leucovorin are given without concomitant irinotecan.

The simultaneous schedule was tested in one phase I North American trial, which recruited patients with advanced solid tumors. Additional patients were accrued at the MTD to provide an early evaluation of the regimen’s activity and toxicity. This simultaneous schedule is now being tested as one of the experimental arms of a phase III randomized trial in advanced colorectal cancer. The sequential approach was the subject of one phase I trial, which closed after accruing nine patients, and an ongoing phase I trial in patients with advanced solid tumors, which is currently enrolling subjects. The alternating schedule was tested in a phase II trial in patients with previously untreated advanced colorectal cancer.

Simultaneous Administration Schedule

Trial of Saltz et al--Saltz et al at Memorial Sloan-Kettering Cancer Center developed a program to find the MTD of a combined regimen of irinotecan given with 5-FU and leucovorin. This regimen was based on the administration schedule of irinotecan that is currently FDA approved and that is most often employed in US patients with advanced colorectal cancer.[19] This empirically derived regimen prescribed escalating doses of irinotecan as a 90-minute infusion given weekly for 4 consecutive weeks followed by a 2-week rest. Low-dose leucovorin (fixed dose of 20 mg/m²/d) was given as an intravenous bolus as soon as the irinotecan infusion was completed and immediately preceding bolus 5-FU. Doses of 5-FU were also escalated in typical phase I trial fashion as new patients were recruited and the trial proceeded. Low-dose leucovorin was chosen in an attempt to minimize diarrhea related to 5-FU and leucovorin in the face of simultaneous use of another diarrhea-producing agent, irinotecan.

A secondary goal of this study was to further investigate the finding by Sasaki et al that 5-FU may inhibit conversion of irinotecan to SN-38.[20] To test for this potential drug interaction, patients were given only irinotecan on the first day of the first treatment week. Blood samples for irinotecan and SN-38 pharmacokinetics were collected over 24 hours. Then, on day 2, 5-FU and leucovorin were administered. Thereafter, except on the first day of the second cycle of therapy, all three drugs were given one after the other in sequence, irinotecan, leucovorin, and 5-FU.

At the second week of the study, blood samples for irinotecan and SN-38 pharmacokinetics were collected, and pharmacokinetic parameters were compared to levels seen after administration of irinotecan alone. On the first day of the second cycle of therapy (week 7) leucovorin and 5-FU were given just before irinotecan was administered, and irinotecan and SN-38 pharmacokinetic blood samples were again collected.

Twenty-seven patients were treated with escalating doses and an additional 15 patients were treated at the MTDs, for a total of 42 patients enrolled in this study. The vast majority of registrants had colorectal cancer (38/42). Of the patients with colorectal cancer, two thirds had received one or more prior chemotherapy regimens.

The MTDs were determined to be 125 mg/m²/d of irinotecan, 20 mg/m²/d of leucovorin, and 500 mg/m²/d of 5-FU. Neutropenia and diarrhea were dose limiting. Among the 17 evaluable patients who were treated at the MTDs, 29% experienced grade 4 neutropenia and 18% had grade 4 diarrhea.

The pharmacokinetic goals of the study were to determine whether the maximal concentration and area under the concentration-time curve (AUC) of SN-38 and irinotecan varied based on the sequence of drug administration. No differences were discerned when irinotecan was given alone, before, or after 5-FU plus leucovorin. There also was no correlation between the SN-38 AUC and the magnitude of granulocytopenia observed. A small but significant decrease in SN-38 AUC was observed by Saltz, but this was not clinically significant.

Although response was not the primary end point of this study, tumor responses were observed. In the 35 evaluable patients (out of 38 entered) who had metastatic colorectal cancer and measurable disease, a response rate of 17% was observed. This included patients treated at less than the MTD and may have underestimated the true response rate. In addition, only 12 previously untreated patients were included in the series, two of whom responded to treatment.

Currently, Saltz and colleagues, in conjunction with investigators at Pharmacia and Upjohn, are conducting a trial that randomly assigns patients with previously untreated measurable colon cancer to one of three treatment arms; (1) irinotecan alone, (2) irinotecan, 5-FU, and leucovorin as described above, or (3) 5-FU and leucovorin. As of January 1998, over 500 of the 660 patients needed to meet the study’s goals have been enrolled. It is likely that the results will be available in 1999 or shortly thereafter.

Doses Used in Simultaneous Schedule--It is notable that the dose of irinotecan (125 mg/m²/d) that can be given concomitantly with 5-FU is identical to the dose that can be administered safely when the drug is given as a single agent, while the dose of 5-FU that can be given concomitantly with irinotecan is five sixths of the dose used for monotherapy (500 mg/m²/d instead of 600 mg/m²/d). The leucovorin dose, which is used to modulate 5-FU in the weekly regimen of 600 mg/m²/d, is 500 mg/m²/d rather than the 20 mg/m²/d employed in the Saltz regimen. It is difficult to determine whether the difference in the leucovorin dose is of clinical importance. The potential for overlapping toxicity has not proven to be as problematic in this schedule as was anticipated.

Since the drugs are administered simultaneously in the Saltz regimen, it does not take advantage of the potential synergistic effects of sequential administration of irinotecan 24 hours before 5-FU noted in vitro and discussed above. (These findings were reported after the initiation of the Saltz trial.) However, because there is more experience with the Saltz regimen than with other regimens that combine irinotecan, 5-FU, and leucovorin, plans are under way to evaluate the Saltz regimen in the adjuvant setting. A randomized trial in which the combination will be compared to a 5-FU and leucovorin-containing regimen in patients with resected stage III colon cancer is being designed.

Sequential Administration

Trial of Parnes et al--Parnes and colleagues attempted a phase I trial in which leucovorin (500 mg/m²/d) and 5-FU (500 mg/m²/d) were administered weekly, with doses of irinotecan escalated for 4 consecutive weeks in 6-week cycles.[21] Irinotecan was administered over 90 minutes 48 hours prior to 5-FU on the first and fourth weeks of treatment beginning at a dose of 25 mg/m²/d. Grade 4 diarrhea proved to be dose limiting at a dose of 50 mg/m²/d, and only nine patients were entered on the study. The authors concluded that this dosing strategy was unproductive and recommended the investigation of other schedules. The higher dose of leucovorin used in this trial may have led to the increased severity of diarrhea seen here as compared to that seen by Saltz.

Mayo Clinic Trial--A phase I trial that combines irinotecan with 5-FU and leucovorin is in progress at the Mayo Clinic.[22] The drugs are given sequentially, based on the work of Mullany et al. In the Mayo Clinic program, irinotecan is given as a 90-minute intravenous infusion on day 1. This is followed, after a 24-hour interval, by leucovorin, given as a standard 20-mg/m²/d bolus, which is followed immediately by 5-FU as a 90-minute intravenous infusion on days 2 through 5. Doses of both irinotecan and 5-FU are escalated according to typical phase I trial design, wherein three patients are accrued and followed for toxicity at each dose level before increasing the doses of either agent.

Like the Saltz regimen, which is based on a commonly used schedule for administration of single-agent irinotecan, the Mayo Clinic trial repeats treatments every 3 weeks in an attempt to replicate the every 3 week schedule of irinotecan that is often given in Europe and that was used in a previous Mayo Clinic phase I study. In that phase I trial of single-agent irinotecan administered every 3 weeks, the MTD was 320 mg/m²/d.[23]

The MTDs of the three drugs in combination in the current Mayo Clinic phase I trial have not been reached. As of January 1998, 28 patients have been enrolled in the current trial. The primary tumor sites are colorectal cancer in 18 patients, other gastrointestinal cancers in 5 patients, breast cancers in 2 patients, head and neck cancer in 1 patient, and lung cancers in 2 patients. The median number of cycles to date is 4, with a range of 1 to 19 cycles. The current cohort is being treated at doses of 250 mg/m²/d of irinotecan, 20 mg/m²/d of leucovorin, and 350 mg/m²/d of 5-FU.

Toxicity to date has included one patient with grade 3 and one patient with grade 4 diarrhea. One case each of grade 3 and grade 4 neutropenia has been noted. Other toxicities have included one case of grade 3 fatigue and one case of grade 3 dyspnea.

Response is not the primary trial end point of this study. Although responses have been observed, the response data will not be reported in detail until the trial has concluded and a phase II dose can be recommended.

Alternating Schedule

Trial of Rothenberg et al--Rothenberg and colleagues developed a regimen in which irinotecan and 5-FU plus leucovorin were administered in alternating cycles. The rationale for this schedule was to ensure that full doses of all three agents could be administered by avoiding the potential for overlapping side effects that may occur with concomitant drug administration. [reference 24 and Mace Rothenberg, MD, personal communication, December, 1997]

In this trial, irinotecan was administered at 100 mg/m²/d each week for 4 weeks, followed by a 2-week rest period. It is noteworthy that the irinotecan dose used by Rothenberg et al is lower than that recommended for single-agent therapy in the package insert (125 mg/m²/d). It is also lower than the dose employed in the previously discussed Saltz regimen (125 mg/m²/d), which combines irinotecan with 5-FU plus leucovorin. The regimen of 5-FU plus leucovorin used by Rothenberg et al is that popularized by Poon et al, which calls for leucovorin 20 mg/m²/d and 5-FU 425 mg/m²/d on days 1 to 5 every 4 weeks.[25]

Of the 71 patients enrolled in the trial, 23 had partial or complete remissions (32%; 95% CI, 21.5% to 43.3%). The median time to tumor progression was 6.9 months. The median survival is 17.6 months.

The principal toxicities were diarrhea, nausea and vomiting, neutropenia, and stomatitis. During the irinotecan segment of therapy, 20% of patients developed grade 3 or 4 diarrhea, as compared with 5% who experienced grade 3 to 4 diarrhea after 5-FU plus leucovorin. The grade 3 and 4 toxicity statistics for irinotecan and 5-FU/leucovorin were 3% and 0%, respectively, for vomiting; 16% and 12%, respectively, for neutropenia; and 0% and 3%, respectively, for stomatitis.

The authors concluded that both the activity and toxicity patterns of the alternating schedule were similar to those seen when each regimen is given alone. The median survival data are intriguing. Confirmation of these findings in a randomized trial seems to be warranted to discern whether the high median survival was due to treatment effect or to inadvertent selection of patients with better prognoses or more indolent disease.

Diarrhea in Patients Treated With Irinotecan Plus 5-FU/Leucovorin

Diarrhea is one of the dose-limiting toxicities for patients treated with either irinotecan or 5-FU/leucovorin programs. Therefore, when combining the two regimens, the potential exists for severe gastrointestinal toxicity. If the agents produce synergistic diarrhea, combining the drugs safely in clinically meaningful doses may be impossible.

The trial by Parnes et al described above is an example of a study in which this toxicity proved to be enough of a problem that the investigators chose to abandon their administration schedule. The actual outcomes of therapy in the three other trials described above indicated that irinotecan and 5-FU plus leucovorin could be combined at doses that are the same or nearly the same as those used when the drugs are administered alone without producing undue gastrointestinal toxicity.

Mechanisms of Drug-Induced Diarrhea

The finding that gastrointestinal toxicity does not preclude administration of reasonable doses of both irinotecan and 5-FU is both encouraging and surprising. An explanation for this may be found in some data in the literature, which suggest that the diarrhea produced by the two agents results from different mechanisms of damage to the gastrointestinal tract.

Fluorouracil Plus Leucovorin--In a study of 16 patients with advanced colorectal cancer who were assessed by the cellobiose/mannitol test before and after 5-FU treatment, 5-FU plus leucovorin was shown to increase small intestine permeability.[26] After chemotherapy, small intestine permeability increased from 0.016 ± 0.011 to 0.029 ± 0.025 (P < .05). A correlation was noted between the number of days of diarrhea and the increased intestinal permeability score (P = .05).

The authors concluded that diarrhea due to 5-FU plus leucovorin appears to be related to small intestinal damage. In this circumstance, the use of concomitant medications to both diminish small intestinal permeability and slow intestinal transit time in order to maximize fluid and electrolyte reabsorption could reduce diarrhea.

A number of investigators have reported that octreotide (Sandostatin) has beneficial effects on 5-FU-related diarrhea.[27,28] The principal mode of action by which octreotide is thought to reduce the diarrhea noted in patients with VIPomas is suppression of the associated small bowel hypersecretory state that correlates with elevated levels of vasoactive intestinal peptide (VIP). The finding that octreotide suppresses 5-FU-induced diarrhea appears to substantiate the role of increased small bowel permeability as the mechanism of 5-FU-induced diarrhea. In the trial by Cascinu et al, octreotide was more effective than loperamide in suppression of diarrhea.[27]

Irinotecan--Ikuno et al have administered irinotecan to mice and examined intestinal cell proliferation, differentiation of colonocytes, and apoptosis, as well as levels of intestinally active secretogogues, such as serotonin and VIP.[29] Histologic evaluation revealed vacuolation of absorptive epithelial cells associated with apoptosis in the mucosal cells of the ileum and goblet-cell hyperplasia in the cecum. The former two processes cause water and electrolyte malabsorption in the ileum, while the latter process causes mucin hypersecretion from the cecum. There was neither an increase in the number of intestinal enterochromaffin cells nor an increase in VIP levels, indicating that irinotecan-induced diarrhea likely results from a different mechanism from that thought to account for 5-FU-induced diarrhea.

Araki et al found evidence of hemorrhagic enterocolitis in small and large bowel samples obtained after intraperitoneal injection of irinotecan in athymic mice.[30] The maximal change was noted in the cecum at day 10.

Loperamide is effective in the treatment of irinotecan-induced diarrhea and allows dose escalation to occur that is not possible without this antidiarrheal therapy.[31] The trials reported by Saltz et al, Parnes et al, Fonseca et al, and Rothenberg et al have prescribed the intensive loperamide treatment program described by Abigerges et al.[31]

The ability to administer relatively high doses of both irinotecan and 5-FU plus leucovorin has been somewhat surprising. It does suggest, however, that it may be possible to take advantage of the augmented activity observed in human tumor xenografts and in vitro in the clinical setting.

Conclusions

A number of recent and ongoing clinical trials in North America and elsewhere have investigated combinations of irinotecan plus 5-FU and leucovorin in patients with advanced colorectal cancer. The drugs have been administered in a simultaneous, sequential, or alternating fashion. All of these regimens have been associated with tumor responses. Since none of the regimens has been compared in a phase III setting, the optimal dose and schedule from among those tested to date is not yet known.

An intergroup phase III trial in which several irinotecan and 5-FU plus leucovorin regimens are to be compared to 5-FU plus leucovorin alone in patients with metastatic, previously untreated colorectal cancer is in the planning stages. The three-drug combination program is also slated to be tested in the adjuvant setting in patients with stage III colon cancer in a North American intergroup trial.

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