Immunoglobulin V_H gene sequence analysis of spontaneous murine immunoglobulin‐secreting B‐cell tumours with clinical features of human disease

Abstract

The 5T series of multiple myelomas (MM) and Waldenstrsöm's macroglobulinaemia-like lymphomas (WM), which developed spontaneously in ageing mice of the C57BL/KaLwRij strain, shows clinical and biological features that closely resemble their corresponding human diseases. In order to compare the patterns of somatic mutation in VH genes of mouse tumours with those of human counterparts, we have determined and analysed sequences of immunoglobulin VH genes in five cases of murine MM, two of WM and one of biclonal benign monoclonal gammopathy (BMG). Four of five MM and 2/2 WM cases used VH genes of the large J558 family; one MM used a gene of the VGAM3.8 family, and both clones of the BMG used genes of the 36-60 family. N-region insertions were observed in all cases, but D-segment genes were only identified in 6/9 cases, which were all from the D-SP family and translated in reading frame 3. Compared with human MM, in which the VH genes have been found to be consistently hypermutated (mean% +/- SD = 8.8 +/- 3.2), the degree of somatic mutation in the murine tumours was significantly lower (mean% +/- SD = 2.9 +/- 2.3). There was no significant evidence of clustering of replacement mutations in complementarity determining regions (CDR), a feature considered to be characteristic of antigen-selected sequences. However, one clone of the biclonal BMG case showed intraclonal variation, a feature described in some cases of human BMG. These results indicate that murine VH genes in mature tumours differ from human counterparts in the level and distribution of somatic mutations, but support the concept that BMG may be distinct from MM.

Myeloma isotype-switch variants in the murine 5T myeloma model: evidence that myeloma IgM and IgA expressing subclones can originate from the IgG expressing tumour

“…The 5T33MM-VDJ sequence differed slightly (only three nucleotides) from the published sequence 19 ( Figure 1). …”

“…The Ig heavy chain sequence expressed by the 5T2MM and 5T33MM tumour cells has already been published. 19 Tumour-specific primers and probes were designed in the CDR1 and the CDR3 region: 5T2MM-CDR1 sense primer: 5Ј CAC-AAC-TGC-TTG-GAA-TGC-AGT-G 3Ј; 5T2MM-CDR3 probe: 5Ј CCG-CCT-GGT-TTG-CTT-ACT-GG 3Ј; 5T33MM-CDR1 sense primer: 5Ј CAC-TAA-TTA-CTT-GAT-AGA-GTG-G 3Ј and 5T33MM-CDR3 probe: 5Ј GCA-GTT-ACC-ATA-AGC-CTC-TC 3Ј. 5T2MM and 5T33MM myeloma-related sequences were amplified using the specific CDR1 primers together with different Ig constant region-specific primers: C : 5Ј GCT-CTC-GCA-GGA-GAC-GAG-GGG-GA 3Ј; C␥: 5Ј GCG-AAT-TCC-CTT-GAC-CAG-GCA-TCC 3Ј and C␣: 5Ј CTC-GGA-TCC-TCA-CAT-TCA-TCG-TGC-C 3Ј.…”

Selective initial in vivo homing pattern of 5T2 multiple myeloma cells in the C57BL/KalwRij mouse

“…For the detection of the 5T2 V H sequences (Zhu et al, 1998) a PCR was performed making use of an antisense FR4 primer (5′-GGGGGATCCTGCAGAGACAGTGACCAGAGT) in combination with a sense FR1 primer (5′-GGGGGATCCACAGATCCAG-TTGGTGCAGT) in a 25 µl reaction containing: 2 mM magnesium chloride, 0.2 mM dNTP, 0.6 µM of each primer and 1.25 U Taq polymerase (Gibco, Life Technologies). Forty PCR cycles were applied after a 5 min incubation at 94°C with the following steps: 30 s at 94°C, 30 s at 55°C and 30 s at 72°C.…”

Dual effects of macrophage inflammatory protein-1α on osteolysis and tumor burden in the murine 5TGM1 model of myeloma bone disease

“…23,24 Cytogenetic and molecular genetic analyses of these murine myelomas have shown that they closely resemble human myelomas. [25][26][27] We reported previously that when 5T33 myelomaderived 5TGM1 cells are inoculated intravenously in syngeneic or bg-nu-xid mice, they home almost exclusively to the skeleton and infrequently to the spleen. 28 Expansion within these sites results in radiographically detectable lytic bone lesions, markedly elevated serum titers of the monoclonal paraprotein, and splenomegaly.…”