Pharmacogenetics of ABCG2 and adverse reactions to gefitinib

Gefitinib is an inhibitor of the epidermal growth factor receptor (EGFR) tyrosine kinase with activity in non-small-cell lung cancer. Diarrhea and skin toxicity are prominent gefitinib-related adverse events that potentially limit its use. Gefitinib is a substrate for ABCG2 (ABCP, BCRP, MXR), a polymorphic efflux transporter protein that is highly expressed in the intestines and liver. Here we investigated associations between allelic variants of EGFR, ABCG2, and the transporter protein ABCB1 with diarrhea and skin toxicity in gefitinib-treated patients. One variant, a common functional single-nucleotide polymorphism (SNP) in the ABCG2 gene, was associated with diarrhea in 124 patients treated with oral gefitinib 250 mg once daily; seven (44%) of 16 patients heterozygous for ABCG2 421C>A (Q141K) developed diarrhea, versus only 13 (12%) of 108 patients homozygous for the wild-type sequence (P = .0046). However, this SNP was not associated with skin toxicity (P = .99). The finding suggests that patients with reduced ABCG2 activity due to a common genetic variant are at increased risk for substrate drug-induced diarrhea, with implications for optimizing treatment with such agents.     

Active antimetastatic immunotherapy in Lewis lung carcinoma with self EGFR extracellular domain protein in VSSP adjuvant

The epidermal growth factor receptor (EGFR) plays a central role in regulating neoplastic processes. The EGFR overexpression in many human epithelial tumors has been correlated with disease progression and bad prognosis. Passive EGFR-directed immunotherapy, but not active specific approaches, has already been introduced in medical oncology practice. Then we wonder if mice immunization with the extracellular domain of murine EGFR (mEGFR-ECD) in adjuvants can circumvent tolerance to self EGFR, by inducing an immune response with consequent antitumor effect. The present study demonstrated that despite mEGFR expression in thymus, strong DTH response was induced by inoculation of mice with the mEGFR-ECD. This self-immunization, using both CFA and very small sized proteoliposomes from Neisseria meningitidis (VSSP), promoted highly specific IgG titers, predominantly IgG2a and IgG2b. Sera from mice immunized with mEGFR-ECD/VSSP not only recognized EGFR+ tumor cell lines by FACS, but also inhibited their in vitro growth, even in the absence of complement. Noteworthy, vaccination of mice with mEGFR-ECD/VSSP stimulated a potent antimetastatic effect in the EGFR+ Lewis lung carcinoma model, while reproduction-associated side effects were absent. Curiously, mice immunized with the human EGFR-ECD (Her1-ECD) in VSSP though induced highly specific IgG antibodies with strong in vitro cytotoxic effect over EGFR+ human cell lines, showed low cross-reactivity with the mEGFR-ECD. These results further encouraged the development of the Her1-ECD/VSSP vaccine project for patients with EGFR+ tumors.     

Impaired inflammatory response and increased oxidative stress and neurodegeneration after brain injury in interleukin-6-deficient mice

In order to determine the role of the neuropoietic cytokine interleukin-6 (IL-6) during the first 3 weeks after a focal brain injury, we examined the inflammatory response, oxidative stress and neuronal survival in normal and interleukin-6-deficient (knockout, IL-6KO) mice subjected to a cortical freeze lesion. In normal mice, the brain injury was followed by reactive astrogliosis and recruitment of macrophages from 1 day postlesion (dpl), peaking at 3-10 dpl, and by 20 dpl the transient immunoreactions were decreased, and a glial scar was present. In IL-6KO mice, the reactive astrogliosis and recruitment of macrophages were decreased throughout the experimental period. The expression of the antioxidant and anti-apoptotic factors metallothionein I+II (MT-I+II) was increased prominently by the freeze lesion, but this response was significantly reduced in the IL-6 KO mice. By contrast, the expression of the antioxidants Cu/Zn-superoxide dismutase (Cu/Zn-SOD), Mn-SOD, and catalase remained unaffected by the IL-6 deficiency. The lesioned mice showed increased oxidative stress, as judged by malondialdehyde (MDA) and nitrotyrosine (NITT) levels and by formation of inducible nitric oxide synthase (iNOS). IL-6KO mice showed higher levels of MDA, NITT, and iNOS than did normal mice. Concomitantly, in IL-6KO mice the number of apoptotic neurons was significantly increased as judged by TUNEL staining, and regeneration of the tissue was delayed relative to normal mice. The changes in neuronal tissue damage and in brain regeneration observed in IL-6KO mice are likely caused by the IL-6-dependent decrease in MT-I+II expression, indicating IL-6 and MT-I+II as neuroprotective factors during brain injury.     

Altered central nervous system cytokine-growth factor expression profiles and angiogenesis in metallothionein-I+II deficient mice.

To study the importance of metallothionein-I and -II (MT-I+II) for brain inflammation and regeneration, the authors examined normal and MT-I+II knock-out (MT-KO) mice subjected to a cortical freeze injury. Normal mice showed profound neurodegeneration, inflammation, and gliosis around the injury, which was repaired by 20 days postlesion (dpl). However, in MT-KO mice the lesion-associated inflammation was still present as late as 90 dpl. Scanning electron microscopy demonstrated that the number of capillaries was lower, and ultrastructural preservation of the lesioned parenchyma was poorer in MT-KO mice, suggesting an altered angiogenesis. To gain insight into the mechanisms involved, a number of cytokines and growth factors were evaluated. The number of cells expressing the proinflammatory cytokines IL-1beta, IL-6, and TNF-alpha was higher in MT-KO mice than in normal mice, which was confirmed by RNase protection analysis, whereas the number of cells expressing the growth factors bFGF, TGFbeta1, VEGF, and NT-3 was lower. Increased expression of proinflammatory cytokines could be involved in the sustained recruitment of CD-14+ and CD-34+ inflammatory cells and their altered functions observed in MT-KO mice. Decreases in trophic factors bFGF, TGFbeta1, and VEGF could mediate the decreased angiogenesis and regeneration observed in MT-KO mice after the freeze lesion. A role for MT-I+II in angiogenesis was also observed in transgenic mice expressing IL-6 under the control of the promoter of glial fibrillary acidic protein gene (GFAP-IL6 mice) because MT-I+II deficiency dramatically decreased the IL-6-induced angiogenesis of the GFAP-IL6 mice. In situ hybridization analysis indicated that the MT-III expression was not altered by MT-I+II deficiency. These results suggest that the MT-I+II isoforms have major regulatory functions in the brain inflammatory response to injury, especially in the angiogenesis process.     

Hepatitis G virus infection in hemodialysis patients and its relationship with hepatitis C virus infection

Our aim was to study the characteristics of hepatitis G virus (HGV) infection in hemodialysis (HD) patients. We evaluated 108 patients from two different units (A: 67 patients; B: 41 patients). HGV RNA and HCV RNA were detected by PCR. Nineteen patients (17.6%) were HGV RNA positive (20.9% in unit A and 12.2% in unit B (NS)). HCV RNA was positive in 19 patients (17.6%) (28.4% in unit A and 0 in unit B (p < 0.01)). Eight patients were HGV RNA and HCV RNA positive (group I), 11 HGV RNA positive (group II), 11 HCV RNA positive (group III), and 78 negative for both viruses (group IV). Time on HD was 51.3 +/- 37.0 months for group I, 36.0 +/- 27.9 months for group II, 63.5 +/- 40.2 months for group III, and 26.4 +/- 27.1 months for group IV (p < 0.01 for I and III). Seven patients (87.5%) from group I, 9 (81.8%) from group II, 10 (90.9%) from group III, and 44 (56.4%) from group IV had a history of transfusion (p < 0.03 for I, II and III). Two patients (25%) from group I, none from group II, 5 (45.4%) from group III, and 6 (7.7%) from group IV had chronic ALAT elevation (p < 0.01 for I and III). We conclude that HGV infection was frequent in our HD patients, related to transfusions and independent of HCV prevalence, and that HGV infection itself was not a cause of ALAT elevation suggesting chronic hepatitis.