Ig heavy chain PCR of serum and plasma fails as a reliable source of minimal residual disease (MRD) in patients with B cell intermediate grade lymphoma
The accurate identification of patients destined to fail a treatment during or soon after therapy allows for custom tailored changes in treatment. The presence of residual tumor in a defined reservoir at specific time points may be prognostic. PCR of tumor derived sequences is one method of detecting MRD. PCR for Ig heavy chain gene (IgH) rearrangements can detect a malignant population at the ICr5 level. Frickhofen et al.(Blood, 1997;90:4953-60) have reported that IgH PCR of plasma and serum DNA reliably detects tumor-derived DNA in 47% of patients with B cell malignancies. A limited number of patient were also studied at multiple time points with the suggestion that persistent or recurrent PCR positivity might be a predictor for relapse. A wide variety of diseases were evaluated without specific attention to subtypes. Because of the ease of IgH PCR and the accessibility of plasma and serum, we sought to confirm these findings in a uniformly treated population of patients with relapsed and refractory diffuse large cell lymphoma. RESULTS: Patients undergoing 3 cycles of second line chemotherapy with Rituximab (day -4 and -2 on cycle 1 and day -2 on cycle 2 and 3) followed by ICE (ifosfamide, carboplatinum, etoposide) on days 1 to 3 had cells, serum and plasma evaluated by IgH PCR before each cycle and at leukopheresis which followed the last cycle. Semi-nested PCR for both Framework 2 and 3 rearrangements were performed. Amplifiable DNA was present in both serum and plasma for nearly all patients at all time points. Although 12/13 patients had clearly detectable circulating clonal cells pre-treatment, the mononuclear fraction was PCR negative after the first treatment in all but 1 patient who became PCR negative after the second treatment. In contrast, in only 1 patient was an identically sized band amplified by IgH PCR pre-treatment in both the mononuclear cells and plasma. However, sequencing revealed that these were not identical. In 6 patients IgH PCR revealed randomly sized bands at multiple time points. In 6 patients only control sequences were amplified. For comparison, we identified identical IgH rearrangements in cells and plasma or serum in 1 patient with mantle cell lymphoma (ALC 63K) and 2 patients with CLL (ALC 25 and 12. CONCLUSIONS : We conclude that IgH rearrangements are not readily detectable in plasma or serum of patients with B cell large lymphoma and that this is not a useful method of tracking MRD disease in this patient population were there is not a high circulating tumor burden. Whether this approach is feasible in diseases such as ALL, CLL or leukemic phase NHL requires additional testing.