Humanized Mouse Model of Cooley's Anemia.

A novel humanized mouse model of Cooley's Anemia (CA) was generated by targeted gene replacement in embryonic stem (ES) cells. Because the mouse does not have a true fetal hemoglobin, a delayed switching human gamma to beta(0) globin gene cassette (gammabeta(0)) was inserted directly into the murine beta globin locus replacing both adult mouse beta globin genes. The inserted human beta(0) globin allele has a mutation in the splice donor site that produces the same aberrant transcripts in mice as described in human cells. No functional human beta globin polypeptide chains are produced. Heterozygous gammabeta(0) mice suffer from microcytic anemia. Unlike previously described animal models of beta thalassemia major, homozygous gammabeta(0) mice switch from mouse embryonic globin chains to human fetal gamma globin during fetal life. When bred with human alpha globin knockin mice, homozygous CA mice survive solely upon human fetal hemoglobin at birth. This preclinical animal model of CA can be utilized to study the regulation of globin gene expression, synthesis, and switching; the reactivation of human fetal globin gene expression; and the testing of genetic and cell-based therapies for the correction of thalassemia.

Clinical features and mismatch repair gene mutation screening in Chinese patients with hereditary nonpolyposis colorectal carcinoma.

AIM: Hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal dominantly-inherited cancer-susceptibility syndrome that confers an increased risk for colorectal cancer and a variety of other tumors at a young age. It has been associated with germline mutations in five mismatch repair (MMR) genes (hMSH2, hMLH1, hPMS1, hPMS2, and hMSH6/GTBP). The great majority of germline mutations were found in hMSH2 and hMLH1. The purpose of this study was to analyze the clinical features of Chinese HNPCC patients and to screen hMSH2 and hMLH1 gene mutations. METHODS: Twenty-eight independent Chinese families were collected, of which 15 met Amsterdam criteria I and 13 met the Japanese clinical diagnosis criteria. The data were recorded including sex, site of colorectal cancer (CRC), age of diagnosis, history of synchronous and/or metachronous CRC, instance of extracolonic cancers, and histopathology of tumors. Peripheral blood samples were collected from all pedigrees after formal written consents were signed. PCR and denaturing high-performance liquid chromatography (DHPLC) were used to screen the coding regions of hMSH2 and hMLH1 genes. The samples showing abnormal DHPLC profiles were sequenced by a 377 DNA sequencer. RESULTS: One hundred and seventy malignant neoplasms were found in one hundred and twenty-six patients (multiple cancer in twenty-three), including one hundred and twenty-seven CRCs, fifteen gastric, seven endometrial, and five esophageal cancers. Seventy-seven point eight percent of the patients had CRCs, sharing the features of early occurrence (average age of onset, 45.9 years) and of the right-sided predominance reported in the literature. In Chinese HNPCC patients, gastric cancer occurred more frequently, accounting for 11.9% of all cancers patients and ranking second in the spectrum of HNPCC predisposing cancers. Synchronous CRCs occurred less frequently, only accounting for 3.1% of the total CRCs. Twenty percent of the colorectal patients had metachronous CRCs within 10 years after operation. Eight hMSH2 or hMLH1 gene sequence variations were found in twelve families, including the first Mongolian kindred with a hMSH2 gene mutation. CONCLUSION: HNPCC is characterized by an early-age onset, proximal predominance of CRC, multiple metachronous CRCs, and an excess of extra-colonic cancers. Frequent gastric cancer occurrence and less synchronous CRCs are the remarkable features in Chinese HNPCC patients. DHPLC is a powerful tool in hMSH2 and hMLH1 gene mutation screening. hMLH1 gene mutations, especially of the first nine exons, have been found more common than hMSH2 gene mutations in Chinese patients. Three of seven mutations have been found to be novel, and the germline G204X nonsense mutation in the third exon of hMSH2 has become the first MMR gene mutation found in Chinese Mongolian people.

[Clinical features and hMSH2/hMLH1 germline mutation screening of Chinese hereditary nonpolyposis colorectal cancer patients].

OBJECTIVE: To analyze the clinical features of hereditary nonpolyposis colorectal cancer (HNPCC) among Chinese and report the results of screening of hMSH2 and hMLH1 gene mutations. METHODS: The data concerning sex, site of colorectal cancer (CRC), age of diagnosis, history of synchronous and/or metachronous colorectal cancer, instance of extracolonic cancers, and histopathology of tumors of 126 patients from 28 independent families of HNPCC in China were collected, of which 15 met the Amsterdam criteriaI and 13 met the Japanese clinical diagnosis criteria. The genomic DNA was extracted from the peripheral lymphocytes. Polymerase chain reaction (PCR) and denaturing high-performance liquid chromatography (DHPLC) were used to screen the coding region of hMSH2 and hMLH1 genes. Samples showing abnormal DHPLC profiles were sequenced by a 377 DNA sequencer. RESULTS: One hundred and seventy malignant neoplasms were found in the 126 patients, in which 23 of multiple cancers were found. Ninety-eight of the 126 patients (77.8%) had colorectal cancers, with an average age of onset of 45.9 years and a right-sided predominance. Eight hMSH2 or hMLH1 gene sequence variations were found in 12 families, and a germline G204X nonsense mutation in the third exon of hMSH2 was found for the first time, the first mismatch repair gene (MMR) mutation ever found in Chinese Mongolian people. CONCLUSIONS: HNPCC is a typical auto-dominant hereditary disease, characterized by early onset, proximal predominance of colorectal cancer, multiple synchronous and metachronous colorectal cancers, and an excess of extra-colonic cancers. Frequent gastric cancer occurrence and less synchronous colorectal cancers are notable features in Chinese HNPCC patients. DHPLC is a powerful tool in hMSH2 and hMLH1 gene mutation screening. Three novel mutations have been found. hMLH1 gene mutations, especially those of the first nine exons, are more common than hMSH2 gene mutations in Chinese patients.