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Between 10% and 25% of chronic lymphocytic leukemia (CLL) patients have episodes of autoimmune hemolytic anemia (AIHA) during the course of their disease. The anti-erythrocyte autoantibodies in most cases are polyclonal and express a different heavy chain isotype than the malignant clone, indicating that they are secreted by normal autoreactive B lymphocytes. To further investigate the pathogenesis of the AIHA in CLL, we analyzed the lg heavy (H) chain variable region genes expressed by leukemic cells from CLL patients with and without AIHA. Two VH genes were preferentially expressed by the leukemic cells in the CLL cases with AIHA and were present in 9 of the 12 investigated cases. The 51p1/DP-10 gene was expressed in 5 of these cases and was absent in the control group of 12 consecutive CLL cases without AIHA, whereas the DP-50 gene was present in 4 CLL-AIHA cases and only once in the control CLL group. A strikingly similar H-chain CDR3 region that contained a single reading frame of the DXP4 DH gene segment, and N- encoded proline at the DH/JH boundary, and a tyrosine-rich region encoded by the JH6 gene segment was observed in four CLL-AIHA cases. The preferential expression of two VH gene segments and a particular CDR3 region by the leukemic cells of patients with AIHA suggests that the antibodies produced by the CLL cells are directly involved in the pathogenesis of the hemolytic anemia.

Through the procedure of gene amplification combined with hybridization to synthetic 19 base pair (bp) oligonucleotide probes, it has been possible to identify nine different mutations in the DNA of 47 subjects from Turkey and Northern Cyprus with a beta-thalassemia homozygosity. The IVS-I nucleotide (nt) 110 G----A and the IVS-I nt 6 T----C substitutions and the frameshift at codon 8 were most frequently observed. Direct correlations were made between these data and clinical observations; mild disease was associated with homozygosity for IVS-I nt 6 T----C, for frameshift at codon 8, for the C----G substitution at nt -87, and for IVS-I nt 5 G----T, and for a double heterozygosity for some of these conditions. Moderate disease, observed in some of the patients, could be explained by combinations of specific mutations. All mutations were associated with specific haplotypes, while in some the observed beta zero-thalassemia was of the mild type due to a considerable production of Hb F.

The Atlanta type of hereditary persistence of fetal hemoglobin (HPFH) is characterized by a mild elevation of Hb F (2% to 5% in heterozygotes), almost exclusively of the G gamma type (more than 90%). Gene-mapping analysis has identified this condition as a -G gamma-G gamma- arrangement with the -158 (C-->T) substitution in the promoters of both G gamma genes. We have reevaluated this condition in members of two families. Sequence analysis identified only two changes in the 3' gamma gene as compared with the A gamma gene from a chromosome with haplotype no. 3 (or Senegal), namely the -158 (C-->T) promoter mutation and the C-->G change in codon (CD) 136, which accounts for the -G gamma- G gamma- phenotype. The absence of any other nucleotide (nt) substitution provides genetic evidence that the -158 (C-->T) change is primarily responsible for the elevated Hb F levels associated with this condition. A quantitative reverse transcription/polymerase chain reaction (RT/PCR) procedure, presented in detail in this report, was developed to determine the effect of this mutation on the transcription of individual gamma genes in four individuals with the Atlanta type of HPFH. Both gamma-globin genes, ie, the (5') G gamma and the (3') G gamma-Atlanta genes of the Atlanta type of HPFH chromosome, expressed elevated amounts of transcripts, which were present in nearly equal amounts. This shows that the -158 (C-->T) mutation exerts its effect on the transcriptional rate of the gene with which it is associated.