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L265P mutation of the MYD88 gene is frequent in Waldenström's macroglobulinemia and its absence in myeloma.

Mori N, Ohwashi M, Yoshinaga K, Mitsuhashi K, Tanaka N, Teramura M, Okada M, Shiseki M, Tanaka J, Motoji T - PLoS ONE (2013)

Bottom Line: We next tested for the mutation with BSiE1 digestion and allele-specific polymerase chain reaction in the 28 patients and 38 patients with myeloma.No siginificant difference was observed in the incidence of the L265P mutation between BSiE1 digestion and allele-specific polymerase chain reaction (p=0.32).BSiE1 digestion and allele-specific polymerase chain reaction may detect a small fraction of mutated cells in some cases.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Tokyo Women's Medical University, Tokyo, Japan.

ABSTRACT
L265P mutation in the MYD88 gene has recently been reported in Waldenström's macroglobulinemia; however the incidence has been different according to the methods used. To determine the relevance and compare the incidence by different methods, we analyzed the L265P mutation in bone marrow mononuclear cells from lymphoid neoplasms. We first performed cloning and sequencing in 10 patients: 8 Waldenström's macroglobulinemia; 1 non-IgM-secreting lymphoplasmacytic lymphoma; and 1 low grade B-cell lymphoma with monoclonal IgG protein. The L265P mutation was detected in only 1/8 Waldenström's macroglobulinemia patients (2 of 9 clones). To confirm these results, direct sequencing was performed in the 10 patients and an additional 17 Waldenström's macroglobulinemia patients and 1 lymphoplasmacytic lymphoma patient. Nine of 28 patients (7/25 Waldenström's macroglobulinemia, 1/2 lymphoplasmacytic lymphoma, and B-cell lymphoma) harbored the mutation. We next tested for the mutation with BSiE1 digestion and allele-specific polymerase chain reaction in the 28 patients and 38 patients with myeloma. Aberrant bands corresponding to the mutation were detected by BSiE1 digestion in 19/25 patients with Waldenström's macroglobulinemia (76%), 1/2 lymphoplasmacytic lymphoma and B-cell lymphoma, but not in the 38 myeloma patients. The L265P mutation was more frequent in patients with Waldenström's macroglobulinemia than in those with myeloma (p=1.3x10(-10)). The mutation was detected by allele-specific polymerase chain reaction in 18/25 Waldenström's macroglobulinemia patients (72%). In the 25 Waldenström's macroglobulinemia patients, the L265P was more frequently detected by BSiE1 digestion than by direct sequencing (p=5.3x10(-4)), and in males (15/16, 94%) than in females (4/9, 44%) (p=1.2x10(-2)). No siginificant difference was observed in the incidence of the L265P mutation between BSiE1 digestion and allele-specific polymerase chain reaction (p=0.32). These results suggest that the L265P mutation is involved in the majority of Waldenström's macroglobulinemia. BSiE1 digestion and allele-specific polymerase chain reaction may detect a small fraction of mutated cells in some cases.

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Sequence analysis of the MYD88 gene in Waldenström’s macroglobulinemia.(A) Sequencing revealed a T to C transition resulting in a leucine to proline substitution at amino acid position 265 (WM5).(B) Wild-type sequences (T) are shown as a control (WM2). (C) Direct sequencing showed both wild-type and mutant alleles in WM5 and WM9, and the wild-type allele only in WM2. (D) Sensitivity of direct sequencing. L265P-positive DNA (WM5) was diluted into wild-type DNA (WM3) before amplification. Aberrant bands were detected in samples containing 10% or more of the L265P mutation.
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pone-0080088-g001: Sequence analysis of the MYD88 gene in Waldenström’s macroglobulinemia.(A) Sequencing revealed a T to C transition resulting in a leucine to proline substitution at amino acid position 265 (WM5).(B) Wild-type sequences (T) are shown as a control (WM2). (C) Direct sequencing showed both wild-type and mutant alleles in WM5 and WM9, and the wild-type allele only in WM2. (D) Sensitivity of direct sequencing. L265P-positive DNA (WM5) was diluted into wild-type DNA (WM3) before amplification. Aberrant bands were detected in samples containing 10% or more of the L265P mutation.

Mentions: We first selected 10 patients including 8 WM (WM1-WM8), 1 non-IgM-secreting LPL (NHL1), and 1 low grade B-cell lymphoma with IgG M-protein (NHL3) for cloning (Table 2). Sequencing was performed using at least four clones in each patient. The nucleotide change, the T to C transition resulting in the L265P mutation, was detected in 2 of 9 clones from 1 patient with WM (WM5), while it was absent in any of the clones from the other 9 patients (Figure 1 A-B). Because of the low frequency of the mutation, we next performed direct sequencing in the 10 patients and an additional 17 patients with WM and 1 LPL patient. The T to C transition was detected in 9 of 28 patients (32%) (Table 2, Figure 1 C). It was found in 7 of the 25 WM patients (28%), one of the 2 LPL, and 1 low grade B-cell lymphoma with IgG M-protein. All of the 9 patients with the transition also had wild-type sequences. To determine sensitivity, DNA from the L265P-positive clone (WM5) was serially diluted into DNA from a wild-type clone (WM3) to the following percentages: 0%, 0.1%, 0.5%, 1%, 5%, 10%, 20%, and 30%. Sensitivity to the L265P mutation by direct sequencing was 10% (Figure 1 D).


L265P mutation of the MYD88 gene is frequent in Waldenström's macroglobulinemia and its absence in myeloma.

Mori N, Ohwashi M, Yoshinaga K, Mitsuhashi K, Tanaka N, Teramura M, Okada M, Shiseki M, Tanaka J, Motoji T - PLoS ONE (2013)

Sequence analysis of the MYD88 gene in Waldenström’s macroglobulinemia.(A) Sequencing revealed a T to C transition resulting in a leucine to proline substitution at amino acid position 265 (WM5).(B) Wild-type sequences (T) are shown as a control (WM2). (C) Direct sequencing showed both wild-type and mutant alleles in WM5 and WM9, and the wild-type allele only in WM2. (D) Sensitivity of direct sequencing. L265P-positive DNA (WM5) was diluted into wild-type DNA (WM3) before amplification. Aberrant bands were detected in samples containing 10% or more of the L265P mutation.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3818242&req=5

pone-0080088-g001: Sequence analysis of the MYD88 gene in Waldenström’s macroglobulinemia.(A) Sequencing revealed a T to C transition resulting in a leucine to proline substitution at amino acid position 265 (WM5).(B) Wild-type sequences (T) are shown as a control (WM2). (C) Direct sequencing showed both wild-type and mutant alleles in WM5 and WM9, and the wild-type allele only in WM2. (D) Sensitivity of direct sequencing. L265P-positive DNA (WM5) was diluted into wild-type DNA (WM3) before amplification. Aberrant bands were detected in samples containing 10% or more of the L265P mutation.
Mentions: We first selected 10 patients including 8 WM (WM1-WM8), 1 non-IgM-secreting LPL (NHL1), and 1 low grade B-cell lymphoma with IgG M-protein (NHL3) for cloning (Table 2). Sequencing was performed using at least four clones in each patient. The nucleotide change, the T to C transition resulting in the L265P mutation, was detected in 2 of 9 clones from 1 patient with WM (WM5), while it was absent in any of the clones from the other 9 patients (Figure 1 A-B). Because of the low frequency of the mutation, we next performed direct sequencing in the 10 patients and an additional 17 patients with WM and 1 LPL patient. The T to C transition was detected in 9 of 28 patients (32%) (Table 2, Figure 1 C). It was found in 7 of the 25 WM patients (28%), one of the 2 LPL, and 1 low grade B-cell lymphoma with IgG M-protein. All of the 9 patients with the transition also had wild-type sequences. To determine sensitivity, DNA from the L265P-positive clone (WM5) was serially diluted into DNA from a wild-type clone (WM3) to the following percentages: 0%, 0.1%, 0.5%, 1%, 5%, 10%, 20%, and 30%. Sensitivity to the L265P mutation by direct sequencing was 10% (Figure 1 D).

Bottom Line: We next tested for the mutation with BSiE1 digestion and allele-specific polymerase chain reaction in the 28 patients and 38 patients with myeloma.No siginificant difference was observed in the incidence of the L265P mutation between BSiE1 digestion and allele-specific polymerase chain reaction (p=0.32).BSiE1 digestion and allele-specific polymerase chain reaction may detect a small fraction of mutated cells in some cases.

View Article: PubMed Central - PubMed

Affiliation: Department of Hematology, Tokyo Women's Medical University, Tokyo, Japan.

ABSTRACT
L265P mutation in the MYD88 gene has recently been reported in Waldenström's macroglobulinemia; however the incidence has been different according to the methods used. To determine the relevance and compare the incidence by different methods, we analyzed the L265P mutation in bone marrow mononuclear cells from lymphoid neoplasms. We first performed cloning and sequencing in 10 patients: 8 Waldenström's macroglobulinemia; 1 non-IgM-secreting lymphoplasmacytic lymphoma; and 1 low grade B-cell lymphoma with monoclonal IgG protein. The L265P mutation was detected in only 1/8 Waldenström's macroglobulinemia patients (2 of 9 clones). To confirm these results, direct sequencing was performed in the 10 patients and an additional 17 Waldenström's macroglobulinemia patients and 1 lymphoplasmacytic lymphoma patient. Nine of 28 patients (7/25 Waldenström's macroglobulinemia, 1/2 lymphoplasmacytic lymphoma, and B-cell lymphoma) harbored the mutation. We next tested for the mutation with BSiE1 digestion and allele-specific polymerase chain reaction in the 28 patients and 38 patients with myeloma. Aberrant bands corresponding to the mutation were detected by BSiE1 digestion in 19/25 patients with Waldenström's macroglobulinemia (76%), 1/2 lymphoplasmacytic lymphoma and B-cell lymphoma, but not in the 38 myeloma patients. The L265P mutation was more frequent in patients with Waldenström's macroglobulinemia than in those with myeloma (p=1.3x10(-10)). The mutation was detected by allele-specific polymerase chain reaction in 18/25 Waldenström's macroglobulinemia patients (72%). In the 25 Waldenström's macroglobulinemia patients, the L265P was more frequently detected by BSiE1 digestion than by direct sequencing (p=5.3x10(-4)), and in males (15/16, 94%) than in females (4/9, 44%) (p=1.2x10(-2)). No siginificant difference was observed in the incidence of the L265P mutation between BSiE1 digestion and allele-specific polymerase chain reaction (p=0.32). These results suggest that the L265P mutation is involved in the majority of Waldenström's macroglobulinemia. BSiE1 digestion and allele-specific polymerase chain reaction may detect a small fraction of mutated cells in some cases.

Show MeSH
Related in: MedlinePlus