Background

X-linked agammaglobulinemia (XLA) is an X-linked inherited disease caused by genetic mutations in the Bruton tyrosine kinase (BTK) gene [1, 2], which suppress the development of mature B lymphocytes. The human BTK gene encompasses 37.5 kb containing 19 exons. BTKbase is an up-to-date database compiling 1796 entities showing 917 unique BTK mutations from 1749 individuals, two thirds of which are from unrelated families, while one third are believed to be sporadic cases [3]. These mutations are found throughout the entire BTK gene sequence. The incidence of XLA varies between 1/200,000 and 1/20,000,000 in Western countries, whereas it has not been calculated in China yet [4]. Based on estimation, there should be more than 1000 cumulative XLA cases below 14 years of age [5].

XLA patients are characterized by insufficient number of normal circulating B lymphocytes and immunoglobulins [6]. The concurrent hallmark symptoms and complications of XLA include lower respiratory tract infection (bronchitis/pneumonia), otitis media, persistent diarrhea and skin infections [7, 8]. XLA patients are also susceptible to certain types of cancer including colorectal cancer (CRC) [9,10,11,12,13,14,15]. According to the National Comprehensive Cancer Network (NCCN) guidelines for colon cancer and rectal cancer, 5-fluorouracil (5-FU)-based drugs are recommended and commonly used for first-line chemotherapy [16]. But patients receiving 5-FU-based chemotherapy, alone or in a combination regimen, may experience drug-related toxicities (DRT) involving hand-foot syndrome, leukopenia, neutropenia, thrombocytopenia, diarrhea, nausea and vomiting [17]. Severe DRT not only leads to an early termination of chemotherapy but also causes safety issues. With previous evidence of lymphopenia being an independent factor associated with first-line chemotherapy induced hematologic toxicities in CRC patients [18, 19], we assume that XLA patients, who are characterized by low levels of B lymphocytes and immunoglobulins, are more likely to develop DRTs when they are diagnosed with CRC and receive chemotherapy [11]. Therefore, gene sequencing, therapeutic drug monitoring and any possible measurement to predict DRT should be considered before making chemotherapy regimens for XLA patients with CRC.

Case presentation

Presenting concerns

A 21-year-old man with XLA was hospitalized for fecal occult blood, epigastric pain and bronchitis in 2016. The patient was not married. The timeline of hospitalization is shown in Fig. 1a.

Fig. 1
figure 1

The timeline, diagnosis and progression focus. a The timeline of hospitalization: This patient was diagnosed with XLA at the age of 4. He suffered upper abdominal pain on Oct 2016, and was diagnosed with advanced colorectal cancer with liver metastasis by liver biopsy ((b), magnification, × 100) and PET-CT scan (c). Abdominal CT scans on Oct 21th 2016 (d) and Jan 9th 2017 (e) showed that the tumor was stabilized after the first two treatment cycles. However, abdominal CT scans on Mar 24th 2017 (f) and May 22th 2017 (g) showed metastatic tumor progressed. Blood calcium/potassium levels (h) and tumor load (i) were recorded throughout all treatment cycles. The dotted lines indicate the starting time of each chemotherapy treatment cycle. The dark dotted line indicates the sixth treatment cycle when oxaliplatin was replaced by irinotecan

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Clinical findings

This patient was diagnosed with XLA when he was 4 years old. He had no family history of XLA. Regular intravenous immunoglobulin (IVIG) replacement therapy was applied since the diagnosis. The admission physical examination found no positive symptoms of XLA.

Diagnostic focus and assessment

Abdominal ultrasonography showed multiple hepatic parenchymal lesions, gallbladder stones, splenomegaly and a hypoechoic mass in the right lower abdomen. Colonoscopy showed a cauliflower-like mass in the ascending colon. Needle biopsy of focal liver lesions (Fig. 1b) and PET-CT (Fig. 1c) suggested metastatic adenocarcinoma. The Eastern Cooperative Oncology Group (ECOG) Performance Status was one [20].

Whole exome sequencing (WES) was conducted using blood and liver tumor tissue. It revealed 10 somatic (Table 1) and 200 germline SNVs, including one on the BTK gene (c.340_347del, p.F114delX115) (Additional file 2: Figure S1). Somatic mutation was absent in KRAS, NRAS, BRAF, or PIK3CA. There is also no mutation in genes related to efficacy or safety of 5-FU-based drugs (Additional file 3: Table S1).

Table 1 Somatic variants may initiate tumor development
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Therapeutic focus and assessment

Standard first-line chemotherapy was started on Nov. 18th, 2016. The regimen is as follows: 130 mg/m2 oxaliplatin iv on day 1, 1000 mg/m2 capecitabine (Cap) twice po daily for 14 days, which was repeated every 3 weeks, and 500 mg/m2 cetuximab iv over 2 h on day 1 every 2 weeks. The tumor was stabilized after the first two treatment cycles (Fig. 1d-e), and the abdominal pain ceased, the abdominal lump was reduced, and CA-199 (from 1410 to 157 U/ml) and CEA (from 113.6 to 20.1 μg/L) levels decreased. During the second treatment cycle, TDM on Cap and its metabolites was conducted. Compared to 10 other random CRC patients without XLA, the XLA patient showed delayed Cmax of Cap, lower doxifluridine (5′-DFUR) level and higher 5-FU level (Fig. 2). These results suggested that the XLA patient had a slower ability to absorb Cap and increased thymidine phosphorylase (TYMP) activity. Since both low 5′-DFUR levels and induced TYMP activity are negatively associated with Cap-related toxicity [21, 22] and this patient did not show any sign of DRT, the original chemotherapy plan was not adjusted.

Fig. 2
figure 2

Therapeutic drug monitoring on Cap and its metabolites. a A simplified metabolic pathway of Cap. Metabolites and enzymes are indicated in black and red, respectively. Plasma concentrations of Cap (b) and its metabolites 5′-dFCR (c), 5′-dFUR (d), 5-FU (e), and FUH2 (f) were measured before and 1–4 h after the administration of Cap. The concentrations are presented as the mean ± SEM. Abbreviations: Cap, capecitabine; 5′-dFCR, 5′-deoxy-5-fluorocytidine; 5′-dFUR, doxifluridine; 5-FU, 5-fluorouracil; FUH2, 5-fluoro-5,6-dihydrouracil; CES, carboxylesterase; CDA, cytidine deaminase; TYMP, thymidine phosphorylase; and DPYD, dihydropyrimidine dehydrogenase

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Follow-up and outcomes

After the fifth treatment, the metastatic tumor progressed (PFS 4.2 m) (Fig. 1f-g). The patient also experienced upper abdominal pain, an upper respiratory tract infection and a severe rash. From the sixth treatment, oxaliplatin was changed to irinotecan (180 mg/m2 iv over 30–90 min on day 1). After the sixth treatment cycle, hypocalcemia (mean, 5.8 mg/dL; range, 4.3–8.2 mg/dL; diagnostic lower limit, 14.0 mg/dL) and a tendency of hypokalemia (mean, 142.4 mEq/L; range, 118.6–161.1 mEq/L; diagnostic lower limit, 136.5 mEq/L) were observed (Fig. 1h). They were not relieved by intravenous calcium gluconate supplementation (1 g, twice daily).

Prior to the eighth treatment, a CT examination revealed an increase in total liver mass (Fig. 1i), which indicated PD (PFS 1.3 m). Unfortunately, the condition of the patient deteriorated with severe ascites and infection. Finally, best supportive care lasted for 1 month before he died (OS was 7.5 m).

Discussion and conclusions

Primary immunodeficiency caused by the BTK mutation is the main reason for XLA-associated diseases, which include recurrent bacterial infections, arthritis and types of cancer [23, 24]. In this study, the diagnosis of XLA was confirmed by WES, where a hemizygous pathogenic mutation (c.340_347del, p.F114delX115) of the BTK gene was found (Additional file 1). The 8 bp deletion of TTCTCCCC resulted in a frameshift mutation that affected general membrane targeting and the regulatory function of BTK [25]. Immunohistochemistry showed similar low expression of BTK in the tumor and tumor-adjacent tissue (Additional file 4: Figure S2).

It has been hypothesized that male XLA patients tend to have an early-onset CRC [11, 12]. B-cell deficiency is a risk factor for a narrow range of solid cancers, including CRC [26]. However, the correlation between CRC and XLA has not yet been statistically confirmed with a reasonable sample size. One study found colorectal adenomatous polyps from 2 out of 4 XLA patients in the Netherlands [27]. However, two other studies found that among 44 and 27 XLA patients from the United Kingdom and mainland China, respectively, none of the patients developed CRC [5, 28]. In the case studied in this report, the onset of CRC was more likely due to sporadic colorectal mutations. First, the XLA patient had no family history of immunodeficiency within 3 generations. Second, among the 10 somatic variants, the most convincing candidate CRC-driver mutations in ABCA6, C6, and PAX3 provided by the DriverDBv2 database are not functionally related [29]. C6 encodes for one of the membrane attack proteins, which plays a key role in the innate and adaptive immune responses by forming pores in the plasma membrane of target cells. It has been shown that dextran-sulfate-sodium (DSS) induced colitis was aggravated in C6-deficient mice with a series of enhanced production of pro-inflammatory mediators, including IL-1β, IL-6, CXCL-1, CCL-3, TGF-β1 and IL-17F, compared with wild-type mice [30]. In addition, exogenous C6 could ameliorate DSS-induced colitis in C6-deficient mice [30]. Since both colitis and enhanced inflammatory are risk factors of colorectal cancer, the deficiency of C6 may also participate in CRC development [31]. PAX3 encodes a transcription factor with an N-terminal DNA binding domain consisting of a paired box. It acts as a transcriptional regulator to activate or repress target genes of carcinogenesis [32, 33]. ABCA6 is a member of the ATP-binding cassette transporter family. It is ubiquitously expressed in the liver, heart and brain, contributing to drug resistance and tumor metastasis [34, 35]. In addition to these cancer-promoting mutations, the mutation (c.857G > A) of N-acetyltransferase 2 (NAT2) has been shown to be associated with slow acetylator phenotypes, which is normally correlated with the effectiveness of drugs and xenobiotic toxicity [36].

Because the incidence of CRC in China has increased rapidly over the past two decades [37, 38], and XLA patients are more prone to carcinogenesis. It is suggested to begin surveillance programs on XLA patients for CRC screening, especially for patients over the age of 20 and 30. As most XLA patients with gastric cancer or CRC have been found at these ages [23].

Regarding severe hypocalcemia, it is speculated to be a joint result of a number of germline mutations in genes related to calcium metabolism and the administration of irinotecan. First, based on the enrichment analysis of genes having germline mutations, the largest enriched group was calcium-related proteins (n = 48, p = 1.2E-7) (Additional file 5 and Additional file 6: Table S2). This group contains genes that bind at least one calcium atom or proteins whose function is calcium-dependent [39,40,41]. Among them, 11 genes are directly involved in calcium metabolism and transportation (Table 2). Mutations in these genes may make the patient prone to abnormal calcium metabolism such as hypocalcemia. Second, the onset of hypocalcemia occurred immediately after the administration of irinotecan. Several studies have reported that hypocalcemia may be one of the rare DRTs of irinotecan (Table 3). Most of these cases had concurrent electrolyte abnormalities such as hypokalemia and hypomagnesemia. Consistently, the patient in this study also showed a trend of hypokalemia (Fig. 2j). However, the magnesium level was not measured in this study.

Table 2 Germline variants related to calcium metabolism
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Table 3 Reported cases showing a positive association between irinotecan and hypocalcemia
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In conclusion, this study reported the first written record of a Chinese XLA patient with metastatic CRC and severe hypocalcemia following irinotecan administration. The chemotherapy regimen was carefully determined based on TDM and WES. Based on WES results and bioinformatic analysis, a germline mutation of BTK was confirmed to be the cause of XLA, and somatic mutations of ABCA6, C6, and PAX3 may contribute to the onset and metastasis of CRC. The administration of irinotecan and a number of germline mutations on genes related to calcium metabolism might collectively cause hypocalcemia.