Dihydropyrimidine Dehydrogenase (DPD) Pharmacogenetics

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This entry is adapted from the peer-reviewed paper 10.3390/jpm10030113

The dihydropyrimidine dehydrogenase (DPD), encoded by the DPYD gene, is the enzyme mainly involved in the catabolism of fluoropyrimidines (FP). DPYD polymorphisms increase the risk of severe FP-related toxicity and DPYD-pharmacogenetics (DPYD-PGx) is recommended before starting the FP-based chemotherapy.

Other factors influence FP safety, therefore phenotyping methods, such as measurement of plasmatic 5-fluorouracil (5-FU) clearance and DPD activity, could complement the DPYD-PGx.

Here, authors reported eleven clinical cases in whom a combined genotyping/phenotyping approach, together with careful clinical monitoring was used to optimise the FP-based treatment. In addition, authors performed a systematic review of the literature concerning the use of DPYD-PGx, together with phenotyping methods to personalise such a chemotherapy.

DPYD Pharmacogenetics 5-fluorouracil Therapeutic drug monitoring

1. Introduction

Fluoropyrimidines (FP), including 5-fluorouracil (5-FU) and its oral prodrug capecitabine, are cytotoxic antineoplastic agents belonging to the class of antimetabolites. They are commonly used to treat solid cancer types such as gastrointestinal, head-neck and breast cancers associated or not to other chemotherapeutics and both cytotoxic and biologic drugs ^[1]^[2]. The administration of the FP may cause severe, even life-threatening, adverse drug reactions (ADR), including myelosuppression, mucositis/stomatitis, diarrhoea and hand–foot syndrome (HFS). Indeed, it has been estimated that an increased risk of severe ADR (grade > 2) involves 10–30% of treated patients, although these data greatly depend on the therapeutic regimen used ^[1]^[3]^[4].

The rate-limiting step of FP catabolism is the conversion of fluorouracil to dihydrofluorouracil, which is catalysed by an enzyme called dihydropyrimidine dehydrogenase (DPD), encoded by a highly polymorphic gene (i.e., DPYD). Several single-nucleotide polymorphisms (SNPs) have been associated to an alteration of the DPYD sequence, and some of them may determine a partial or complete DPD deficiency, leading to FP severe toxicity ^[1].

The Clinical Pharmacogenetics Implementation Consortium (CPIC) and the Dutch Pharmacogenetics Working Group (DPWG) ^[5]^[6] have published guidelines for FP dosing based on the pharmacogenetic testing of four DPYD polymorphisms that are DPYD*2A (rs3918290), DPYD*13 (rs55886062), DPYD c.2846A>T (rs67376798) and c.1129-5923C>G (rs75017182 and HapB3). The latter is the most common variant, with ~4% allelic frequency; the DPYD*2A, c.2846A>T and DPYD*13 are present in ~2.0%, 1.4% and 0.1%, respectively, of Caucasian patients ^[5]. Recently, a new polymorphism, DPYD*6 (rs1801160), has been associated with both gastrointestinal and haematological FP-ADR ^[7].

Notably, other DPYD genetic variants may lead to dangerous clinical consequences, although their frequency is very low ^[5]^[6]^[8].

With the main aim of reducing the risk of severe FP-induced toxicity, the CPIC and DPWG have implemented a gene activity score (DPYD-AS), which ranges from 0 (complete DPD deficiency) to 2 (normal DPD activity). In patients who are homozygous for one or more of the aforementioned SNPs, the recommendation is to avoid the use of FP. However, if alternative drugs are not considered a suitable option, the FP dosage should be markedly reduced while establishing a therapeutic drug-monitoring (TDM) approach. Patients who are heterozygous should receive a 50% dose reduction at the first cycle of chemotherapy, followed by a titration dose, while monitoring the patient’s clinical conditions and possibly performing TDM ^[5]^[6].

However, it has been estimated that 30–50% of the patients experience severe ADR, despite not having a DPD deficit associated with such DPYD polymorphisms. In fact, there are several factors, including comorbidities, polytherapy, other variants in the DPYD and other genes, that can play an important role ^[9]. Besides DPYD polymorphisms, two SNPs in the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene ^[10] and a tandem repeat in the thymidylate synthase enhancer region (TYMS-TSER) could concur in predicting FP-related toxicity ^[11]. Moreover, a SNP in glutathione S-transferase-p1 (GSTP1) has been suggested as a genetic factor able to influence the response to oxaliplatin, a drug frequently administered with FP ^[12].

Several strategies complementing the DPYD pharmacogenetics (DPYD-PGx) have been proposed to prevent FP-related severe ADR associated with DPD deficit. Among others, the measurement of the plasmatic dihydrouracil/uracil ratio (UH₂/U) and the monitoring of 5-FU clearance are considered valid approaches ^[13]^[14].

2. Cases Presentation

Case 1 was a Caucasian 55-year-old male former smoker with a history of hypertension. The patient had stage IV colorectal adenocarcinoma with metastases in the lymph nodes, lungs, liver and kidneys. The tumour mutational profile identified no mutations in the KRAS, NRAS or BRAF genes. The patient was treated with the combination of 5-FU, leucovorin and oxaliplatin (FOLFOX6) regimen, plus cetuximab. After three cycles of chemotherapy, the patient reported grade 1 thrombocytopenia and paraesthesia; grade 2 stomatitis, rash and leukopenia and grade 3 neutropenia and mucositis.

A post-therapeutic DPYD-PGx was performed, revealing that the patient was heterozygous for DPYD*2A. Moreover, the patient was homozygous (TT) for MTHFR-C677T, homozygous TYMS-TSER-2R/2R and homozygous (AA) for GSTP1-A313G. The plasmatic UH2/U ratio was 4.52. Based on these results and the reported toxicity, both the 5-FU and cetuximab doses were reduced. Specifically, the total dosage of 5-FU was reduced to 50%, according to the CPIC and DPWG guidelines.

At the fourth cycle of therapy, the pharmacokinetic analysis revealed a trough 5-FU plasma concentration of 950 ng/mL. The CT scan demonstrated an overall stable disease, according to the RECIST criteria v1.1. The patient was still treated with the same doses of 5-FU. At the sixth cycle of therapy, the 5-FU plasma concentration was 400 ng/mL. The following cycles (fifth to eighth) of chemotherapy were administered at the same drug doses. A new CT scan demonstrated no evidence of disease progression. The ADR were grade 1 leukopenia, neutropenia, thrombocytopenia and mucositis and grade 2 HFS. Following a further two cycles of therapy, the reported ADR were grade 1 paraesthesia, erythematous maculopapular rash and grade 2 cutaneous and mucous fissures. Lastly, following a further two treatment cycles, a new CT scan showed disease progression. The treatment was stopped, and the administration of a new chemotherapeutic regimen was planned. Sadly, the patient died before starting a second line of treatment.

Case 2 was a Caucasian 48-year-old male with no comorbidity. He had stage IV colorectal adenocarcinoma with metastases in the lymph nodes and liver. The tumour mutational profile highlighted the presence of a KRAS mutation; thus, a treatment with the FOLFOX6 regimen plus bevacizumab was planned. A pretherapeutic DPYD-PGx was requested, and the patient was identified as DPYD*2A heterozygous. In addition, he was wild type for MTHFR-C677T and MTHFR-A1298C, heterozygous TYMS TSER-2R/3R and homozygous (GG) for GSTP1-A313G. The plasmatic UH2/U ratio was 3.22. Based on these results, a 50% dose reduction of 5-FU was planned for the first cycle of FOLFOX administration, according to the CPIC and DPWG guidelines. After the first cycle of treatment, the plasmatic 5-FU clearance was 474 ng/mL. Following three cycles of therapy, a stable disease was found, according to the RECIST criteria v1.1, and no adverse events were reported. The patient was still treated with the same doses of chemotherapeutic agents for an additional seven cycles of therapy. Grade 1 paraesthesia and mucositis and grade 2 HFS but no severe ADR were reported, and the CT scan demonstrated a stable disease. Afterward, the patient was treated up to the twelfth cycle with a FOLFOX regimen plus bevacizumab, still obtaining, at revaluation, a stable disease. Then, he was a candidate for a maintenance therapy with capecitabine plus bevacizumab. Following 16 cycles of this therapy, the patient reported grade 1 paraesthesia and mucositis, and no severe ADR were recorded.

Case 3 was a Caucasian 60-year-old male former smoker with no comorbidities. He had stage IV rectal adenocarcinoma with liver metastases. The tumour mutational profile did not identify mutations in either KRAS, NRAS or BRAF. Based on the tumour profile and stage, the patient was a candidate for a FOLFOX regimen plus cetuximab. A pretherapeutic DPYD-PGx was performed, and the patient was found heterozygous for DPYD c2846A>T SNP. Therefore, according to the CPIC and DPWG guidelines, he started chemotherapy with a 50% dose reduction of 5-FU. Moreover, he was homozygous (TT) for MTHFR-C677T, heterozygous TYMS TSER-2R/3R and heterozygous for GSTP1-A313G. The plasmatic UH2/U ratio was 1.77.

A grade 2 diffuse maculopapular rash was reported, and, based on such an ADR, the dose of cetuximab was also reduced to 50% for the second cycle of therapy. The plasmatic 5-FU clearance was 811 ng/mL—still high, notwithstanding the 5-FU dose reduction. The patient reported no improvement of the skin rash and grade 2 diarrhoea. At the third cycle of therapy with the same drugs doses, the 5-FU plasma level was 1093 ng/mL. Grade 1 nausea and grade 3 diarrhoea were reported. Based on these results, the 5-FU dose was further reduced by an additional 10% at the fourth cycle of therapy. However, the 5-FU plasma concentration was still high (1048 ng/mL), and grade 4 diarrhoea was reported. Hence, it was decided not to administer 5-FU in a continuous infusion, leaving the administration of 5-FU in bolus. Nevertheless, the 5-FU plasma concentration was still high (i.e., 934 ng/mL), and grade 3 diarrhoea was reported.

A CT scan showed a partial response according to the RECIST criteria with a reduction of hepatic lesions. It was decided to carry out a further cycle with oxaliplatin plus cetuximab.

After this cycle, the hepatic lesions were resected. After one month from surgery, a CT scan demonstrated the development of a new hepatic lesion. The patient was a candidate to start a new treatment with 5-FU plus irinotecan as a modified 5-FU, leucovorin and irinotecan (FOLFIRI) regimen, since 5-FU was administered with a 50% dose reduction and without continuous infusion. The patient performed six cycles of FOLFIRI plus bevacizumab. Grade 3 diarrhoea was reported. As a consequence, the 5-FU administration was stopped, and only irinotecan and bevacizumab were further administered. After four cycles of this treatment, the CT scan demonstrated a progression of the disease. The patient died after 11.2 months from starting treatment with irinotecan plus bevacizumab.

Table 1 and Table 2 report the main characteristics and the occurrence of grade ≥ 3 ADR of 3/11 and 8/11 clinical cases, respectively. Table 1 describes three clinical cases for whom either pretherapeutic DPYD-PGx or post-therapeutic DPYD-PGx were performed. As phenotypic characteristics, the UH2/U ratio values and plasmatic 5-FU clearance were reported.

Table 1. Reports of the main characteristics of three patients with the occurrence of grade ≥ 3 ADR.

Pt	Sex	Age (years)	Tumor Type and Stage	Chemotherapy Regimen	Pre-Therapeutic DPYD-PGx	Post-Therapeutic DPYD-PGx	DPYD Genotype	UH2/U Ratio	5-FU Dosage	5-FU Clearance	ADR ≥ 3	Total Toxicity
1	M	55	CRC (metastatic)	Folfox plus cetuximab	/	Yes (C4)	Heterozygous for DPYD*2A	4.52	C1-C3: 100% C4-C6: 50%	950 ng/mL (C4) 400 ng/mL (C6)	G3 mucositis (C3) G3 neutropenia (C3) (neuthrophils: 830.58 /mm³)	7
2	M	48	CRC (metastatic)	Folfox plus bevacizumab	Yes	/	Heterozygous for DPYD*2A	3.22	C1-C8: 50%	474 ng/mL (C1)	/	6
3	M	60	Rectal cancer (metastatic)	Folfox plus cetuximab)	Yes	/	Heterozygous for c.2846A>T	1.77	C1-C3: 50% C4: 40% C5-C6: 40% (only bolus)	811 ng/mL (C2) 1093 ng/mL (C3) 1048 ng/mL (C4) 934 ng/mL (C5)	G3 diarrhoea (C3) G4 diarrhoea (C4) G3 diarrhoea (C5)	3

Pt, patient; M, male; CRC, colorectal cancer; FOLFOX, 5-fluorouracil plus leucovorin plus oxaliplatin; DPYD-PGx, DPYD pharmacogenetics; 5-FU, 5-fluorouracil; C, cycle; plasmatic UH2/U ratio, dihydrouracil/uracil ratio; ADR, adverse drug reaction; G, grade and total toxicity, number of ADR regardless of the grade of severity.

Table 2. Reports of 8 clinical cases for whom either pretherapeutic DPYD-PGx or post-therapeutic DPYD-PGx were performed. As phenotypic characteristics, the UH2/U ratio values were reported.

Pt	Sex	Age (Years)	Tumor Type and Stage	Chemotherapy Regimen	Pre-Therapeutic DPYD-PGx	Post-Therapeutic DPYD-PGx	DPYD Genotype	UH2/U Ratio	5-FU Dosage	ADR ≥3
1	F	63	Stomach cancer (locally advanced)	Folfox	/	Yes (C2)	Heterozygous for DPYD*2A	7.09	C1-C2: 100% C3: 5-FU withdrawal	G3 vomit (C2)
2	M	43	CRC (metastatic)	Folfiri with bevacizumab	/	Yes (C8)	Heterozygous for c.2846A>T	3.88	C1-C8: 100% C9: 50%	G3 vomit (C8)
3	M	63	Kidney cancer (metastatic)	Xeloda	yes	/	Heterozygous for c.2846A>T	6.57	C1-C6: 50%	/
4	M	68	CRC (metastatic)	Xelox	yes	/	Heterozygous for c.2846A>T	4.4	C1-C7: 50%	/
5	M	78	CRC (local)	Xelox	yes	/	Heterozygous for c.2846A>T	3.37	C1-C2: 50%	/
6	M	72	CRC (locally advanced)	Xelox	yes	/	Heterozygous for DPYD*2A	5.15	C1-C8: 50%	/
7	F	52	Vulva carcinoma (local)	Xeloda with cisplatin	yes	/	Heterozygous for DPYD*2A	7.38	C1-C5: 50%	/
8	M	76	Rectosigmoid cancer (locally advanced)	Folfox	yes	/	Heterozygous for DPYD*2A	2.44	C1-C5: 50%	/

Pt, patient; M, male; F, female; CRC, ColoRectal Cancer; FOLFOX, 5-Fluorouracil plus leucovorin plus oxaliplatin; FOLFIRI, 5-Fluorouracil plus leucovorin plus irinotecan; XELOX, capecitabine plus oxaliplatin; XELODA, Capecitabine; DPYD-PGx, DPYD pharmacogenetics; UH2/U RATIO, dihydrouracil/uracil ratio; ADR, Adverse Drug Reaction; C, cycle; G, Grade.

A pretherapeutic DPYD-PGx was performed in two out of three cases, while one patient (case 1) had already started chemotherapy before requesting DPYD-PGx. Importantly, the patients were monitored during all treatment cycles.

Besides these three clinical cases, the history of other eight patients is briefly reported below, and their main characteristics are listed in Table 2.

All subjects were monitored for at least four treatment cycles. In two out of eight subjects, the DPYD-PGx was required after the occurrence of severe toxicity (post-therapeutic DPYD-PGx), while in six out of eight, pharmacogenetic testing was performed before the treatment started (pretherapeutic DPYD-PGx). All patients were identified as carriers of DPYD variants—precisely, four out of eight were DPYD*2A heterozygous, and four out of eight were DPYD c.2846 heterozygous.

The two patients for whom the DPYD-PGx was performed after 5-FU administration experienced grade 3 ADR with a different timing, and both were then revealed as DPYD-variant carriers. More in detail, one patient with stage III gastric cancer, treated with FOLFOX, suffered from grade 3 vomit after the second cycle; he was then identified as DPYD*2A heterozygous and continued to be treated only with oxaliplatin. Moreover, the patient was homozygous (TT) for MTHFR-C677T, homozygous TYMS TSER-3R/3R and homozygous (AA) for GSTP1-A313G.

The other one with stage IV colon cancer, treated with FOLFIRI plus bevacizumab, showed grade 3 vomit after the eighth cycle of chemotherapy. The patient was identified as DPYD c.2846 heterozygous, and the 5-FU dosage was halved. With regards to the other SNPs, the patient was homozygous (TT) for MTHFR-C677T, heterozygous TYMS TSER-2R/3R and wild type for UGT1A1*28 SNP. The latter polymorphism is routinely analysed in patients treated with irinotecan.

Conversely, in the other patients, a pretherapeutic DPYD-PGx was performed; thus they were treated with a starting halved dose of 5-FU, and no severe ADR were reported.

3. Systematic Review

A systematic review was performed to analyse the studies investigating the variability of responses to FP-based chemotherapy by DPYD genotyping combined with phenotyping methods and/or clinical monitoring.

Of the potential 112 articles assessed for eligibility, after considering the inclusion and exclusion criteria, 22 studies were included in the analysis ^[15]^[16]^[17]^[18]^[19]^[20]^[21]^[22]^[23]^[24]^[25]^[26]^[27]^[28]^[29]^[30]^[31]^[32]^[33]^[34]^[35]^[36]. Table 3 shows such studies subdivided with respect to the analysed DPYD polymorphisms (DPYD-PGx), the used phenotyping methods and the presence of clinical monitoring. A DPYD-PGx/clinical monitoring combination was present in 11, and DPYD-PGx/phenotyping in three, surveys. A DPYD-PGx/phenotyping/clinical monitoring combined approach was made in eight studies (Table 3).

Table 3. The table reports the studies included in the systematic review and subdivided into three groups: DPYD-PGx/clinical monitoring combination, DPYD-PGx/phenotyping and DPYD-PGx/phenotyping/clinical monitoring. Abbreviations: PBMC, peripheral blood mononuclear cells; HPLC-UV, high-performance liquid chromatography-UV detector; LC-MS/MS, liquid chromatography-tandem mass spectrometry; 5-FUDR, 5-FU degradation rate; UHPLC-MS/MS, ultra-high-performance liquid chromatography-tandem mass spectrometry; PK, pharmacokinetics; FP, fluoropyrimidines; DPD, dihydropyrimidine dehydrogenase; UH2/U ratio, dihydrouracil/uracil ratio and AAS, atomic absorption spectrometry.

First Author‘s Name (Published Year)	Enrolled Patients (n)	Outcomes	DPYD-PGx/Clinical Monitoring	DPYD-PGx/Phenotyping	DPYD-PGx/Phenotyping/Clinical Monitoring
Kuilenburg et al. (2000) ^[15]	37	DPD activity and overall toxicity; DPYD genotyping in patients with reduced DPD activity.			DPYD2A, c.2846A>T, DPYD6, DPYD*9A, c.496A>G/ UH2/U ratio in PBMC/ADR until two treatment months.
Schwab et al. (2008) ^[16]	683	Overall toxicity; DPYD, TYMS, MTHFR genotyping; sequencing of DPYD exome; influence of sex and promoter methylation on DPD expression in human liver.	DPYD2A, c.2846A>T, c.623G>T, DPYD4, DPYD*6, and c.2858G>C/ ADR reported until the second cycle of treatment.
Kristensen et al.(2010) ^[17]	68	Relationship between UH₂/U plasma ratio and 5-FU-related early toxicity; relationship between 5-FU concentration and toxicity; IVS14+1G>A mutation screening.			DPYD*2A/ UH₂/U ratio in plasma 5-FU clearance by HPLC-UV/ ADR reported until the second cycle of treatment.
Deenen et al. (2011) ^[18]	568	Relationships between SNPs and toxicity, SNPs and dose modification of capecitabine, DPYD haplotypes and toxicity, DPYD SNPs and haplotypes and survival.	DPYD*2A, c.2846A>T and c.1236G>A [HapB3]/ ADR reported until the second cycle of treatment.
Deenen et al. (2016) ^[19]	2038	Feasibility, safety and cost of DPYD*2A genotype-guided dosing.	DPYD*2A/ADR reported until the sixth cycle of treatment.
Sistonen et al. (2014) ^[20]	28	Relationship between UH₂/U plasma ratio and DPYD genetic variation; plasma concentration of 5-FU and corresponding AUC; toxicity.			c.234-123G>C, c.496A>G, c.775A>G, c.1129-5923C>G [Hap B3], DPYD13, DPYD2A and c.2846A>T/ UH₂/U ratio in plasma- 5-FU clearance by LC-MS/MS/ ADR reported until the second cycle of treatment.
Lee et al.(2014) ^[21]	2886	Relationship between DPYD variants and toxicity.	DPYD2A, DPYD13, c.2846A>T/ ADR until the twelfth cycle of treatment.
Gentile et al. (2015) ^[22]	156	Correlation between degradation rate of 5-FU with detected SNPs.		DPYD2A, DPYD13, c.2846A>T/5-FUDR assay in PBMC by HPLC-MS/MS
Joerger et al. (2015) ^[23]	140	Quantitative effect of 44 gene polymorphism in 16 drug pathway associated genes on progression free survival (PFS), on chemotherapy toxicity, on objective response rate (ORR), on overall survival (OS).			DPYD13, DPYD2A, c.2846A>T, DPYD*9A, c.1896T>C/5-FU clearance by AAS and HPLC/ADR until disease progression.
Lunenburg et al. (2016) ^[24]	275	Evaluation of requests of prospective DPYD screening and results with a dose recommendation; estimation of the follow up of the dose recommendations.	DPYD2A, DPYD13, c.2846A>T, c.1236G>A [HapB3]/ ADR reported until the second cycle of treatment.
Galarza et al. (2016) ^[25]	60	Estimation of the use of plasma and saliva; Uracil to UH₂ metabolic ratio and DPYD genotyping.			DPYD 2A, 13, c.557A>G, DPYD *7/ UH₂/U ratio in plasma/ ADR reported until the third cycle of treatment.
Milano et al. (2016) ^[26]	243	Sequencing of DPYD exome and frequence of G3, G4 toxicity over cycle 1-2.	DPYD2A, DPYD13, c.2846A>T, c.1774C>T, c.1475C>T, D342G/ ADR reported until the second cycle of treatment.
Boisdron-Celle et al.(2017) ^[27]	85	UGT1A1 and DPYD genotyping; UH₂/U ratio; follow up of efficacy and tolerance.			DPYD2A, DPYD13, c.2846A>T, DPYD*7/ UH₂/U ratio in plasma/ADR every two weeks until three months.
Etienne-Grimaldi et al.(2017) ^[28]	243	DPYD sequencing; relationship between toxicity and DPYD variants; DPD phenotyping.			DPYD2A, DPYD13, c.2846A>T/ UH₂/U ratio in plasma/ ADR reported until the second cycle of treatment.
Liu et al.(2017) ^[29]	661	Relationship between UGT1A1 and DPYD polymorphism and incidence of severe neutropenia and diarrhea; relationship between UGT1A1 and DPYD variants and objective response rate, disease control rate, overall and progression free survival.	DPYD5, c.1896 T > C, and DPYD2A/ ADR reported every two-three cycles or whenever patient’s condition changed.
Henricks et al.(2018) ^[30]	1181	Frequency of severe overall FP-related toxicity; pharmacokinetics of fluoropyrimidines in DPYD variant allele carriers; DPD enzyme activity; cost analysis on individualised dosing by upfront DPYD genotyping.			DPYD2A, c.2846A>T, DPYD13 and c.1236G>A [Hap B3]/ UH₂/U ratio in PBMC/PK data by UHPLC-MS/MS/ADR until toxicity resolution.
Cremolini et al. (2018) ^[31]	443	Relationship between DPYD and UGT1A1 genotyping and toxicity.	DPYD2A, 13, c.2846A>T/ADR reported until the fourth cycle of treatment.
Jacobs et al.(2019) ^[32]	237	Pharmacokinetics of capecitabine and 5-FU in DPYD variant allele carriers.		DPYD*2A, c.2846A>T, c.1236G>A [HapB3]/5-FU clearance by LC MS/MS.
Iachetta et al.(2019) ^[33]	1827	Relationship between DPYD and toxicity.	DPYD13, DPYD2A, c.2846A>T, DPYD*6 /ADR reported until the eleventh cycle of treatment.
Kleinjan et al. (2019) ^[34]	185	DPYD genotyping and toxicity.	DPYD2A, c.2846A>T, DPYD13 and c.1236G>A [HapB3] /ADR reported until the second cycle of treatment.
Negarandeh et al.(2020) ^[35]	88	Relationship between DPYD genotyping and toxicity.	DPYD2A, c.2846A>T, DPYD6/ADR reported following 227 cycles for 88 patients.
Nicolas Pallet et al.(2020) ^[36]	5886	Relationship between DPYD genotyping and [U] and UH₂/U ratio in plasma.		DPYD2A, DPYD13, c.2846A>T, c.1236G>A [HapB3]/ [U] and UH₂/U ratio in plasma.

Among the studies with a DPYD-PGx/clinical monitoring combination, five out of eleven studies confirmed the importance of DPYD variants in predicting FP-related toxicity, although a too-short clinical monitoring was performed (only two treatment cycles) ^[16]^[18]^[24]^[26]^[34]. Two studies ^[19]^[29] analysed only DPYD*2A of the DPYD SNPs currently recommended. The first confirmed a strong association between DPYD*2A and a severe and potentially fatal toxicity ^[19]. The second did not analyse this type of association because of the too-low DPYD*2A allelic frequency found in the study population ^[29].

Lee et al., by testing 2886 patients, reported a significant association between DPYD*2A and DPYD c.2846A>T and grade ≥3 FP ADR ^[21]. Similarly, Cremolini et al., in a large cohort of colon cancer patients who were treated with FOLFIRI or FOLFOXIRI plus bevacizumab, demonstrated that DPYD*2A and DPYD c.2846A>T predicted FP-associated clinically relevant ADR ^[31]. Iachetta et al. analysed 668 out of 1827 patients enrolled in their study. The authors, first, confirmed the clinical relevance of DPYD c.2846A>T and DPYD*13 in predicting FP safety. Second, they found a significant association between DPYD*6 and severe neutropenia. Notably, no patients carrying DPYD*2A (1.7%) had started a FP-based treatment ^[33]. Negarandeh et al. screened the presence of DPYD c.2846A>T, DPYD*6 and DPYD*2A in a population of 73 Iranian patients. DPYD c.2846A>T and DPYD*6 were not found in the patient population analysed. However, a high allelic frequency for DPYD*2A (5.5%) was reported. Surprisingly, Negarandeh et al. did not find any significant association between DPYD*2A and severe toxicity ^[35].

A DPYD-PGx/phenotyping combined approach was performed in three studies that underlined the importance of complementing the DPYD-PGx with a phenotyping analysis. Gentile et al., by measuring the 5-FUDR (degradation rate) in peripheral blood mononuclear cells (PBMCs) utilising HPLC with MS/MS, found a significant correlation between several polymorphisms, including DPYD*2A and DPYD c.2846A>T, and this phenotyping marker ^[22]. Jacobs et al. determined 5-FU clearance in the plasma of patients treated with capecitabine, finding that the presence of DPYD*2A led to a 21.5% reduction in 5-FU elimination. Pallet et al. evaluated an approach based on the combination of the plasmatic UH2/U ratio and uracil concentration with genotyping of the four recommended DPYD SNPs. The main finding of this study was that complementing the DPYD-PGx with the plasmatic UH2/U ratio increased the possibility to identify patients at higher risks of severe FP-related toxicity ^[36]. Among the studies that performed a DPYD-PGx/phenotyping approach with clinical monitoring, four out of eight studies reported information about FP-related ADR until a maximum of three treatment cycles.

Joerger et al. confirmed the importance of DPYD genotyping to identify patients at high risks of severe FP-related toxicity. Unfortunately, because of the low sample size of the study, they did not show conclusive results about the usefulness of plasmatic 5-FU clearance determination in improving the predictive potential of DPYD-PGx ^[23].

Galarza et al. found that salivary and plasmatic UH2 concentrations were inversely correlated with the ADR grade. However, given the low number of patients enrolled in the study, no DPYD variant allele carriers were identified ^[25].

Boisdron et al. conducted a phase II study in 85 patients to test the efficacy of a pharmacogenetic-guiding dosing approach combined with the UH2/U ratio measurement. Despite a very large increase in drug dosages, a low incidence of severe ADR was shown in patients who used a guiding dosing approach. However, also in this case, it was not possible to conclude if this phenotyping analysis enhanced the predictability of DPYD genotyping because of the low sample size of the study ^[27]. Etienne et al. failed to demonstrate a correlation between DPYD variants and the plasmatic UH2/U ratio values. The authors concluded that only an extension of the genetic panel may improve the performance of DPYD-PGx for predicting severe and life-threatening ADR associated with capecitabine ^[28].

Kuilenburg et al. measured the UH2/U ratio in PBMCs as an indirect assessment of DPYD activity. They demonstrated that patients with a low DPD activity experienced a more rapid onset of toxicity as compared to those with a normal enzymatic activity. Moreover, grade 4 neutropenia occurred in a substantial percentage (55%) of the patients with a decreased DPD activity as compared to that (13%) of subjects with a normal DPD activity. Notably, eleven out of fourteen patients suffering from severe ADR with a decreased enzymatic activity were identified as carriers of DPYD polymorphisms. In particular, six, four and one out of eleven patients carried DPYD*2A, DPYD*9A and DPYD*6 in homozygosis, respectively ^[15]. Kristensen et al. also showed a significant correlation between the plasmatic UH2/U ratio and the presence of DPYD*2A ^[17].

Finally, in a prospective study, Henricks et al. analysed all the four recommended DPYD SNPs and performed two phenotyping tests by measuring the UH2/U ratio in PBMCs and plasmatic 5-FU pharmacokinetics (PK) by UHPLC-MS/MS. The patients carrying DPYD c.1236G>A and DPYD c.2846A>T were more likely to manifest FP-related severe toxicity as compared to wild-type subjects. In addition, the mean DPD enzyme activity was significantly lower in patients bearing these two genetic variants, as well as DPYD*2A, as compared to other patients. Only one patient carrying DPYD*13 showed a 60% DPD activity reduction. This patient was treated with a reduced 5-FU dosage for three treatment cycles, and no severe ADR occurred.

4. Conclusions

Nowadays, the regulatory agencies recommend carrying out the DPYD-PGx, including four DPYD polymorphisms (i.e., rs3918290, rs55886062, rs67376798 and rs75017182, HapB3) in patients who need to be treated with FP.

Despite that these DPYD variants are strongly associated with treatment toxicity, other genetic and nongenetic factors concur to determine the variable response to FP-based chemotherapy.

A pretherapeutic DPYD-PGx offers the possibility to avoid early ADR. Nonetheless, severe and even fatal FP-related toxicity may happen anytime during the therapy also in subjects having no DPD deficit attributable to the four recommended DPYD SNPs.

On the other hand, measuring the plasmatic 5-FU clearance—currently, the best method to perform TDM—does not permit to diagnose a possible DPD deficit prior to starting the treatment.

Therefore, because both genetic and phenotypic tests show advantages and disadvantages, a combined genotyping/phenotyping approach, together with careful and continuous clinical monitoring, is the best diagnostic method to optimise the therapy with FP.

References

Longley, D.B.; Harkin, D.P.; Johnston, P.G. 5-Fluorouracil: Mechanisms of Action and Clinical Strategies. Nat. Rev. Cancer 2003, 3, 330–338.
Venook, A.P.; Niedzwiecki, D.; Lenz, H.-J.; Innocenti, F.; Fruth, B.; Meyerhardt, J.A.; Schrag, D.; Greene, C.; O’Neil, B.H.; Atkins, J.N.; et al. Effect of First-Line Chemotherapy Combined with Cetuximab or Bevacizumab on Overall Survival in Patients With KRAS Wild-Type Advanced or Metastatic Colorectal Cancer: A Randomized Clinical Trial. JAMA 2017, 317, 2392–2401.
Hoff, P.M.; Ansari, R.; Batist, G.; Cox, J.; Kocha, W.; Kuperminc, M.; Maroun, J.; Walde, D.; Weaver, C.; Harrison, E.; et al. Comparison of Oral Capecitabine versus Intravenous Fluorouracil plus Leucovorin as First-Line Treatment in 605 Patients with Metastatic Colorectal Cancer: Results of a Randomized Phase III Study. J. Clin. Oncol. Off. J. Am. Soc. Clin. Oncol. 2001, 19, 2282–2292.
Van Kuilenburg, A.B.P.; Dobritzsch, D.; Meijer, J.; Meinsma, R.; Benoist, J.-F.; Assmann, B.; Schubert, S.; Hoffmann, G.F.; Duran, M.; de Vries, M.C.; et al. Dihydropyrimidinase Deficiency: Phenotype, Genotype and Structural Consequences in 17 Patients. Biochim. Biophys. Acta 2010, 1802, 639–648.
Amstutz, U.; Henricks, L.M.; Offer, S.M.; Barbarino, J.; Schellens, J.H.M.; Swen, J.J.; Klein, T.E.; McLeod, H.L.; Caudle, K.E.; Diasio, R.B.; et al. Clinical Pharmacogenetics Implementation Consortium (CPIC) Guideline for Dihydropyrimidine Dehydrogenase Genotype and Fluoropyrimidine Dosing: 2017 Update. Clin. Pharmacol. Ther. 2018, 103, 210–216.
Lunenburg, C.A.T.C.; van der Wouden, C.H.; Nijenhuis, M.; Crommentuijn-van Rhenen, M.H.; de Boer-Veger, N.J.; Buunk, A.M.; Houwink, E.J.F.; Mulder, H.; Rongen, G.A.; van Schaik, R.H.N.; et al. Dutch Pharmacogenetics Working Group (DPWG) Guideline for the Gene–Drug Interaction of DPYD and Fluoropyrimidines. Eur. J. Hum. Genet. 2020, 28, 508–517.
Del Re, M.; Cinieri, S.; Michelucci, A.; Salvadori, S.; Loupakis, F.; Schirripa, M.; Cremolini, C.; Crucitta, S.; Barbara, C.; Di Leo, A.; et al. DPYD*2A and c.2846A>T: A Comprehensive Analysis in 1254 Patients. Pharm. J. 2019, 19, 556–563.
Van Kuilenburg, A.B.P.; Meijer, J.; Mauer, D.; Dobritzsch, D.; Meinsma, R.; Los, M.; Knegt, L.C.; Zoetekouw, L.; Jansen, R.L.H.; Dezentjé, V.; et al. Severe Fluoropyrimidine Toxicity Due to Novel and Rare DPYD Missense Mutations, Deletion and Genomic Amplification Affecting DPD Activity and MRNA Splicing. Biochim. Biophys. Acta Mol. Basis Dis. 2017, 1863, 721–730.
Coenen, M.J.H.; Paulussen, A.D.C.; Breuer, M.; Lindhout, M.; Tserpelis, D.C.J.; Steyls, A.; Bierau, J.; van den Bosch, B.J.C. Evolution of Dihydropyrimidine Dehydrogenase Diagnostic Testing in a Single Center during an 8-Year Period of Time. Curr. Ther. Res. Clin. Exp. 2019, 90, 1–7.
Ulvik, A.; Ueland, P.M.; Fredriksen, A.; Meyer, K.; Vollset, S.E.; Hoff, G.; Schneede, J. Functional Inference of the Methylenetetrahydrofolate Reductase 677C > T and 1298A > C Polymorphisms from a Large-Scale Epidemiological Study. Hum. Genet. 2007, 121, 57–64.
Lecomte, T.; Ferraz, J.-M.; Zinzindohoué, F.; Loriot, M.-A.; Tregouet, D.-A.; Landi, B.; Berger, A.; Cugnenc, P.-H.; Jian, R.; Beaune, P.; et al. Thymidylate Synthase Gene Polymorphism Predicts Toxicity in Colorectal Cancer Patients Receiving 5-Fluorouracil-Based Chemotherapy. Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res. 2004, 10, 5880–5888.
McLeod, H.L.; Sargent, D.J.; Marsh, S.; Green, E.M.; King, C.R.; Fuchs, C.S.; Ramanathan, R.K.; Williamson, S.K.; Findlay, B.P.; Thibodeau, S.N.; et al. Pharmacogenetic Predictors of Adverse Events and Response to Chemotherapy in Metastatic Colorectal Cancer: Results from North American Gastrointestinal Intergroup Trial N9741. J. Clin. Oncol. Off. J. Am. Soc. Clin. Oncol. 2010, 28, 3227–3233.
Boisdron-Celle, M.; Remaud, G.; Traore, S.; Poirier, A.L.; Gamelin, L.; Morel, A.; Gamelin, E. 5-Fluorouracil-Related Severe Toxicity: A Comparison of Different Methods for the Pretherapeutic Detection of Dihydropyrimidine Dehydrogenase Deficiency. Cancer Lett. 2007, 249, 271–282.
Van Staveren, M.C.; Jan Guchelaar, H.; Van Kuilenburg, A.B.P.; Gelderblom, H.; Maring, J.G. Evaluation of Predictive Tests for Screening for Dihydropyrimidine Dehydrogenase Deficiency. Pharm. J. 2013, 13, 389–395.
Van Kuilenburg, A.B.; Haasjes, J.; Richel, D.J.; Zoetekouw, L.; Van Lenthe, H.; De Abreu, R.A.; Maring, J.G.; Vreken, P.; van Gennip, A.H. Clinical Implications of Dihydropyrimidine Dehydrogenase (DPD) Deficiency in Patients with Severe 5-Fluorouracil-Associated Toxicity: Identification of New Mutations in the DPD Gene. Clin. Cancer Res. Off. J. Am. Assoc. Cancer Res. 2000, 6, 4705–4712.
Schwab, M.; Zanger, U.M.; Marx, C.; Schaeffeler, E.; Klein, K.; Dippon, J.; Kerb, R.; Blievernicht, J.; Fischer, J.; Hofmann, U.; et al. Role of Genetic and Nongenetic Factors for Fluorouracil Treatment-Related Severe Toxicity: A Prospective Clinical Trial by the German 5-FU Toxicity Study Group. J. Clin. Oncol. 2008, 26, 2131–2138.
Kristensen, M.H.; Pedersen, P.; Mejer, J. The Value of Dihydrouracil/Uracil Plasma Ratios in Predicting 5-Fluorouracilrelated Toxicity in Colorectal Cancer Patients. J. Int. Med. Res. 2010, 38, 1313–1323.
Deenen, M.J.; Tol, J.; Burylo, A.M.; Doodeman, V.D.; De Boer, A.; Vincent, A.; Guchelaar, H.-J.; Smits, P.H.M.; Beijnen, J.H.; Punt, C.J.A.; et al. Relationship between Single Nucleotide Polymorphisms and Haplotypes in DPYD and Toxicity and Efficacy of Capecitabine in Advanced Colorectal Cancer. Clin. Cancer Res. 2011, 17, 3455–3468.
Deenen, M.J.; Meulendijks, D.; Cats, A.; Sechterberger, M.K.; Severens, J.L.; Boot, H.; Smits, P.H.; Rosing, H.; Mandigers, C.M.P.W.; Soesan, M.; et al. Upfront Genotyping of DPYD*2A to Individualize Fluoropyrimidine Therapy: A Safety and Cost Analysis. J. Clin. Oncol. 2016, 34, 227–234.
Sistonen, J.; Büchel, B.; Froehlich, T.K.; Kummer, D.; Fontana, S.; Joerger, M.; Van Kuilenburg, A.B.P.; Largiadèr, C.R. Predicting 5-Fluorouracil Toxicity: DPD Genotype and 5,6-Dihydrouracil:Uracil Ratio. Pharmacogenomics 2014, 15, 1653–1666.
Lee, A.M.; Shi, Q.; Pavey, E.; Alberts, S.R.; Sargent, D.J.; Sinicrope, F.A.; Berenberg, J.L.; Goldberg, R.M.; Diasio, R.B. DPYD variants as predictors of 5-fluorouracil toxicity in adjuvant colon cancer treatment (NCCTG N0147). J. Natl. Cancer Inst. 2014, 106.
Gentile, G.; Botticelli, A.; Lionetto, L.; Mazzuca, F.; Simmaco, M.; Marchetti, P.; Borro, M. Genotype-Phenotype Correlations in 5-Fluorouracil Metabolism: A Candidate DPYD Haplotype to Improve Toxicity Prediction. Pharm. J. 2016, 16, 320–325.
Joerger, M.; Huitema, A.D.R.; Boot, H.; Cats, A.; Doodeman, V.D.; Smits, P.H.M.; Vainchtein, L.; Rosing, H.; Meijerman, I.; Zueger, M.; et al. Germline TYMS Genotype Is Highly Predictive in Patients with Metastatic Gastrointestinal Malignancies Receiving Capecitabine-Based Chemotherapy. Cancer Chemother. Pharmacol. 2015, 75, 763–772.
Lunenburg, C.A.T.C.; Van Staveren, M.C.; Gelderblom, H.; Guchelaar, H.-J.; Swen, J.J. Evaluation of Clinical Implementation of Prospective DPYD Genotyping in 5-Fluorouracil- or Capecitabine-Treated Patients. Pharmacogenomics 2016, 17, 721–729.
Galarza, A.F.A.; Linden, R.; Antunes, M.V.; Hahn, R.Z.; Raymundo, S.; da Silva, A.C.C.; Staggemeier, R.; Spilki, F.R.; Schwartsmann, G. Endogenous Plasma and Salivary Uracil to Dihydrouracil Ratios and DPYD Genotyping as Predictors of Severe Fluoropyrimidine Toxicity in Patients with Gastrointestinal Malignancies. Clin. Biochem. 2016, 49, 1221–1226.
Milano, G. Highlight on DPYD Gene Polymorphisms and Treatment by Capecitabine*. Scand. J. Clin. Lab. Invest. 2016, 76, S30–S33.
Boisdron-Celle, M.; Metges, J.P.; Capitain, O.; Adenis, A.; Raoul, J.L.; Lecomte, T.; Lam, Y.H.; Faroux, R.; Masliah, C.; Poirier, A.L.; et al. A Multicenter Phase II Study of Personalized FOLFIRI-Cetuximab for Safe Dose Intensification. Semin. Oncol. 2017, 44, 24–33.
Etienne-Grimaldi, M.-C.; Boyer, J.-C.; Beroud, C.; Mbatchi, L.; Van Kuilenburg, A.; Bobin-Dubigeon, C.; Thomas, F.; Chatelut, E.; Merlin, J.-L.; Pinguet, F.; et al. New Advances in DPYD Genotype and Risk of Severe Toxicity under Capecitabine. PLoS ONE 2017, 12.
Liu, D.; Li, J.; Gao, J.; Li, Y.; Yang, R.; Shen, L. Examination of Multiple UGT1A and DPYD Polymorphisms Has Limited Ability to Predict the Toxicity and Efficacy of Metastatic Colorectal Cancer Treated with Irinotecan-Based Chemotherapy: A Retrospective Analysis. BMC Cancer 2017, 17.
Henricks, L.M.; Lunenburg, C.A.T.C.; de Man, F.M.; Meulendijks, D.; Frederix, G.W.J.; Kienhuis, E.; Creemers, G.-J.; Baars, A.; Dezentjé, V.O.; Imholz, A.L.T.; et al. DPYD Genotype-Guided Dose Individualisation of Fluoropyrimidine Therapy in Patients with Cancer: A Prospective Safety Analysis. Lancet Oncol. 2018, 19, 1459–1467.
Cremolini, C.; Del Re, M.; Antoniotti, C.; Lonardi, S.; Bergamo, F.; Loupakis, F.; Borelli, B.; Marmorino, F.; Citi, V.; Cortesi, E.; et al. DPYD and UGT1A1 Genotyping to Predict Adverse Events during First-Line FOLFIRI or FOLFOXIRI plus Bevacizumab in Metastatic Colorectal Cancer. Oncotarget 2018, 9, 7859–7866.
Jacobs, B.A.W.; Deenen, M.J.; Joerger, M.; Rosing, H.; de Vries, N.; Meulendijks, D.; Cats, A.; Beijnen, J.H.; Schellens, J.H.M.; Huitema, A.D.R. Pharmacokinetics of Capecitabine and Four Metabolites in a Heterogeneous Population of Cancer Patients: A Comprehensive Analysis. CPT Pharmacomet. Syst. Pharmacol. 2019, 8, 940–950.
Iachetta, F.; Bonelli, C.; Romagnani, A.; Zamponi, R.; Tofani, L.; Farnetti, E.; Nicoli, D.; Damato, A.; Banzi, M.; Casali, B.; et al. The Clinical Relevance of Multiple DPYD Polymorphisms on Patients Candidate for Fluoropyrimidine Based-Chemotherapy. An Italian Case-Control Study. Br. J. Cancer 2019, 120, 834–839.
Kleinjan, J.P.; Brinkman, I.; Bakema, R.; Van Zanden, J.J.; Van Rooijen, J.M. Tolerance-Based Capecitabine Dose Escalation after DPYD Genotype-Guided Dosing in Heterozygote DPYD Variant Carriers: A Single-Center Observational Study. Anticancer Drugs 2019, 30, 410–415.
Negarandeh, R.; Salehifar, E.; Saghafi, F.; Jalali, H.; Janbabaei, G.; Abdhaghighi, M.J.; Nosrati, A. Evaluation of Adverse Effects of Chemotherapy Regimens of 5-Fluoropyrimidines Derivatives and Their Association with DPYD Polymorphisms in Colorectal Cancer Patients. BMC Cancer 2020, 20, 560.
Pallet, N.; Hamdane, S.; Garinet, S.; Blons, H.; Zaanan, A.; Paillaud, E.; Taieb, J.; Laprevote, O.; Loriot, M.-A.; Narjoz, C. A Comprehensive Population-Based Study Comparing the Phenotype and Genotype in a Pretherapeutic Screen of Dihydropyrimidine Dehydrogenase Deficiency. Br. J. Cancer 2020.

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