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Impact of UGT1A1 genetic polymorphism on toxicity in unresectable pancreatic cancer patients undergoing FOLFIRINOX

Studies have indicated an association between UDP‐glucuronosyltransferase‐1A1 (UGT1A1) genetic polymorphisms and irinotecan‐induced toxicity. We undertook this study to investigate the association between UGT1A1 genetic polymorphisms and toxicity in patients treated with the FOLFIRINOX (comprising oxaliplatin, irinotecan, fluorouracil, and leucovorin) chemotherapy regimen in the JASPAC 06 study. Patients screened for UGT1A1*6 and UGT1A1*28, and treated with either the original FOLFIRINOX (oxaliplatin 85 mg/m2, irinotecan 180 mg/m2, leucovorin 200 mg/m2, bolus 5‐fluorouracil [5‐FU] 400 mg/m2, and continuous 5‐FU 2400 mg/m2) or a modified FOLFIRINOX (oxaliplatin 85 mg/m2, irinotecan 150 mg/m2, leucovorin 200 mg/m2, and continuous 5‐FU 2400 mg/m2) as first‐line chemotherapy were included. Of 199 patients eligible for this analysis, 79 patients were treated with the original FOLFIRINOX regimen and 120 patients were treated with the modified FOLFIRINOX regimen. In the original FOLFIRINOX group, 54 were UGT1A1 WT, and 25 were UGT1A1 heterozygous type (−/*6, 12 patients; −/*28, 13 patients). In the modified FOLFIRINOX group, 64 were UGT1A1 WT and 56 were UGT1A1 heterozygous type (−/*6, 33 patients; −/*28, 23 patients). In the original FOLFIRINOX group, the incidence of diarrhea was significantly higher among patients with UGT1A1 heterozygous type than among those with UGT1A1 WT and the incidence of leukopenia and diarrhea was significantly higher among patients with UGT1A1 −/*6 than among those with UGT1A1 −/*28. Patients with UGT1A1 heterozygous type, especially those with UGT1A1 −/*6, tended to show a higher incidence rate of severe adverse events, but this was not statistically significant. However, for patients who received the modified FOLFIRINOX, there was no difference in the frequency of adverse events due to UGT1A1 status. In conclusion, patients with heterozygous UGT1A1 polymorphisms treated with the original FOLFIRINOX regimen experienced severe toxicity more frequently than patients with WT UGT1A1.


| INTRODUC TI ON
Pancreatic cancer is one of the most lethal types of common cancer and the eighth and ninth leading cause of cancer-related death among men and women worldwide, respectively. 1 A randomized phase II/III study found that improvement in overall survival, progression-free survival, and response rate among patients with metastatic pancreatic cancer were significantly greater among patients given a combination chemotherapy regimen comprising oxaliplatin, irinotecan, fluorouracil, and leucovorin (FOLFIRINOX) compared to those treated with gemcitabine. However, a higher incidence of adverse events was associated with the FOLFIRINOX regimen. 2 To improve tolerability of the FOLFIRINOX regimen, a modified version has been used in practice. Several reports have suggested improved safety and maintained efficacy under the modified FOLFIRINOX regimen. 3,4 Irinotecan is a prodrug, and its active metabolite SN-38 shows antitumor activity and toxicity. 5,6 SN-38 is inactivated into SN-38 glucuronide mainly by UDP-glucuronosyltransferase 1A1 (UGT1A1). 7 Genetic polymorphisms in UGT1A1, such as UGT1A1*28 and UGT1A1*6, contribute to interpatient variability in the pharmacokinetics and toxicity of irinotecan, particularly severe neutropenia. Decreased UGT1A1 activity is commonly attributed to the insertion of TA in the TATA box of the UGT1A1 gene promoter, and this polymorphism is named UGT1A1 *28. 8 Several meta-analyses have examined the impact of the *28 allele on irinotecan-based therapy toxicity. [9][10][11] A further polymorphism, UGT1A1 *6, characterized by a single nucleotide replacement in exon 1 of the UGT1A1 gene, is the most frequent and important polymorphism among the Asian population and is rarely found among Caucasians. 12 Many studies have reported a significant association between the UGT1A1 *6/*6 type and severe neutropenia, although no association was found with UGT1A1 WT. 13-17 UGT1A1 *28 occurs with a frequency of 26%-31% among Caucasians, 42%-56% among African Americans, and only 9%-16% among Asians. 18,19 UGT1A1 *6 has allele frequencies of 0%, 13%, 23%, and 23% among German, Japanese, Korean, and Chinese populations, respectively. 20 In Japan, three genotypes were identified based on the following types of UGT1A1 *6 and *28 genetic polymorphisms: WT (−/−), heterozygous type (−/*6 and −/*28), and homozygous type (*6/*6, *28/*28). Compound heterozygous type (*6/*28) was classified as homozygous based on the results of previous studies. 14,21,22 Studies have shown an association between UGT1A1 genetic polymorphisms and irinotecan-induced toxicity, although the only cancer type specifically investigated was colorectal cancer. The association between UGT1A1 genetic polymorphisms and FOLFIRINOX-induced toxicity remains unclear. Furthermore, to our knowledge, there are few studies on the association between toxicity and heterozygous type or WT, 10,13,16,17,23 nevertheless the frequency of heterozygous UGT1A1 polymorphisms is approximately 40%. 24 We undertook this modified FOLFIRINOX (oxaliplatin 85 mg/m 2 , irinotecan 150 mg/m 2 , leucovorin 200 mg/m 2 , and continuous 5-FU 2400 mg/m 2 ) as first-line chemotherapy were included. Of 199 patients eligible for this analysis, 79 patients were treated with the original FOLFIRINOX regimen and 120 patients were treated with the modified FOLFIRINOX regimen. In the original FOLFIRINOX group, 54 were UGT1A1 WT, and 25 were UGT1A1 heterozygous type (−/*6, 12 patients; −/*28, 13 patients). In the modified FOLFIRINOX group, 64 were UGT1A1 WT and 56 were UGT1A1 heterozygous type (−/*6, 33 patients; −/*28, 23 patients). In the original FOLFIRINOX group, the incidence of diarrhea was significantly higher among patients with UGT1A1 heterozygous type than among those with UGT1A1 WT and the incidence of leukopenia and diarrhea was significantly higher among patients with UGT1A1 −/*6 than among those with UGT1A1 −/*28. Patients with UGT1A1 heterozygous type, especially those with UGT1A1 −/*6, tended to show a higher incidence rate of severe adverse events, but this was not statistically significant. However, for patients who received the modified FOLFIRINOX, there was no difference in the frequency of adverse events due to UGT1A1 status. In conclusion, patients with heterozygous UGT1A1 polymorphisms treated with the original FOLFIRINOX regimen experienced severe toxicity more frequently than patients with WT UGT1A1.

K E Y W O R D S
chemotherapy, FOLFIRINOX, pancreatic cancer, toxicity, UGT1A1 study to investigate the association between UGT1A1 genetic polymorphisms and toxicity in patients treated with FOLFIRINOX in the JASPAC 06 study 25 .

| Patients
An exploratory analysis was carried out using pooled data from the JASPAC 06 study, a nationwide multicenter observational study of Patients with homozygous (*6/*6, *28/*28) or compound heterozygous (*6/*28) UGT1A1 polymorphisms were excluded. This study protocol was approved by the Shizuoka Cancer Center Institutional Review Board (Shizuoka, Japan) and carried out in accordance with the Ethical Guidelines for Epidemiological Research. This trial was registered at the UMIN Clinical Trials Registry as UMIN000014658.

| Evaluation
Toxicity was graded by the Common Toxicity Criteria for Adverse Events version 4.0.

| Statistical analysis
Comparisons between categorical variables were carried out using Fisher's exact test. Median values of variables were compared using the Mann-Whitney U test. A P-value < .05 was considered statistically significant. Statistical analysis was undertaken using the software EZR version 1.32 (Saitama Medical Center, Jichi Medical University, Saitama, Japan). 26

| Patient selection and characteristics
Of the 399 eligible patients in the JASPAC 06 study, 200 patients were excluded for the following reasons: previous chemotherapy (n = 146), use of a different chemotherapy regimen that is reduced more than the modified FOLFIRINOX regimen (n = 50), lack of genetic testing (n = 11), and UGT1A1 homozygous type (n = 9) or compound heterozygous type (n = 6) polymorphisms; there were some overlaps between these criteria ( Figure 1). Finally, 199 patients were analyzed in the present study. Of these, 79 were treated with the original FOLFIRINOX and 120 patients were treated with the modified FOLFIRINOX therapy as initial treatment. Patient characteristics are shown in Table 1. Of the 199 patients, 118 (59%) were WT for UGT1A1 polymorphisms, 45 (23%) were heterozygous for UGT1A1*6, and 36 (18%) were heterozygous for UGT1A1*28. In the original FOLFIRINOX group, 54 (68%) patients had UGT1A1 WT, and 25 (32%) had UGT1A1 heterozygous type (−/*6 in 12 and −/*28 in 13 patients). In the modified FOLFIRINOX group, 64 (53%) patients had UGT1A1 WT, and 56 (47%) had UGT1A1 heterozygous type (−/*6 in 33 and −/*28 in 23 patients). In each treatment group, most patient characteristics were comparable between the heterozygous type and WT.

| Incidence of grade III-IV adverse events according to UGT1A1 status in each cycle
Incidence of grade III-IV major adverse events according to UGT1A1 status (WT vs heterozygous type vs −/*6 type vs −/*28 type) among patients given either the original FOLFIRINOX or the modified FOLFIRINOX regimen are summarized in Figures 2 and 3, respectively.
Cycle 1 had one of the highest incidences of adverse events. In the modified FOLFIRINOX group, the incidence of grade III-IV adverse events tended to be lower than in the original FOLFIRINOX group.

| D ISCUSS I ON
The present study is the first report investigating the association between UGT1A1 genetic polymorphisms, specifically UGT1A1*6 and UGT1A1*28 heterozygosity, and toxicity among Japanese patients with pancreatic cancer treated with FOLFIRINOX.
TA B L E 2 Treatment delivery and adverse events among 199 patients with unresectable pancreatic cancer treated with oxaliplatin, irinotecan, fluorouracil, and leucovorin (FOLFIRINOX), according to UGT1A1 (WT vs heterozygous)

P value n (%) n (%) n (%) n (%)
Adverse events (grade III-IV) Our study showed that in the original FOLFIRINOX group the incidence of diarrhea was significantly higher among patients with UGT1A1 heterozygous type than among those with UGT1A1 WT,    and the incidence of leukopenia and diarrhea was significantly higher among patients with UGT1A1 −/*6 than among those with UGT1A1 −/*28. In addition, we analyzed the incidence of grade III-IV adverse events according to UGT1A1 status in each treatment cycle. Cycle 1 had one of the highest incidences of adverse events regardless of UGT1A1 status or treatment regimen (original FOLFIRINOX or modified FOLFIRINOX). One of the reasons is that dose reduction of chemotherapeutic drugs was undertaken due to toxicity. As several factors such as dose modification or treatment course influence the incidence of adverse events, we attempted to analyze the incidence of adverse events in cycle 1 to adjust these factors. The incidence of leukopenia and neutropenia in cycle 1 was significantly higher among patients with UGT1A1 −/*6 than among those with UGT1A1 −/*28.
Patients with UGT1A1 heterozygous type, especially those with UGT1A1 −/*6 tended to show a higher incidence rate of se- The FOLFIRINOX regimen, using the same doses and schedule as in the PRODIGE 4/ACCORD 11 trial, was approved in Japan for advanced pancreatic cancer in 2013. However, hematological toxicity in the phase II FOLFIRINOX trial for Japanese patients was reported to be higher than in the PRODIGE 4/ACCORD 11 trial. 2,27 This outcome might have been caused by the fact that patients with the UGT1A1*6 variant were analyzed in addition to those with the UGT1A1*28 variant.
Our study also showed that, in the modified FOLFIRINOX group, the incidence of adverse events was comparable among patients with UGT1A1 heterozygous type and UGT1A1 WT or among patients with UGT1A1 −/*6 and −/*28 regardless of treatment cycle. In addition, severe hematological adverse events occurred less frequently among patients in the modified FOLFIRINOX group than among those in the original FOLFIRINOX group.
These findings might be due to the initial irinotecan dose reduction. Several meta-analyses reported that UGT1A1 *28/*28 was associated with an increased risk of neutropenia at high doses (180-350 mg/m 2 ) of irinotecan, although this association has not been clearly established at low doses (<150 mg/m 2 ). 9,11 However, further meta-analysis evaluating predominantly Asian populations reported that patients with UGT1A1 *6/*6 or UGT1A1 −/*6 had an increased incidence of severe neutropenia and this was associated with any dose of irinotecan. 30 Although there are contradictory reports, our findings suggest that the association between increased risk of adverse events and UGT1A1 status might not be established in patients treated with low-dose irinotecan.
In clinical practice in Japan, the initial irinotecan dose reduc-

ACK N OWLED G EM ENTS
We thank all patients, clinicians, and support staff who participated in this study.