Elimination of neoplastic cells from peripheral blood progenitor cells (PBPCs) is an important issue in transplantation-based high-dose chemotherapy in non Hodgkin’s lymphoma (NHL). The capacity to reliably assess the presence of residual lymphoma cells in PBPCs is mandatory in designing this type of protocols. Polymerase chain reaction (PCR) amplification of molecular rearrangements is widely used to detect minimal residual disease (MRD) in NHL patients. Although concordant data can be obtained in most of the cases from peripheral blood (PB) and bone marrow (BM) at diagnosis, the relationship between these two compartments and the role of their analysis in predicting the molecular status of PBPCs is still an open issue. Here we report data about MRD analysis in BM, PB and PBPCs in a series of mantle cell and indolent NHL patients who underwent high-dose chemotherapy: discordant results were obtained comparing PB, BM and PBPC molecular data. In addition, differences were noted among these results if molecular analysis was performed using well-known rearrangements (i.e., bcl-1/IgH and bcl-2/IgH) or patient specific oligonucleotides. We conclude that neither BM nor PB are reliable in predicting the molecular status of PBPCs and that caution must be adopted in interpreting molecular data obtained using patient specific oligonucleotides.

It is not known if immune response to T cell–defined human histocompatibility leukocyte antigen (HLA) class I–restricted melanoma antigens leads to an expanded peripheral pool of T cells in all patients, affects cytotoxic T lymphocyte (CTL) generation, and correlates with anti-tumor response in metastatic lesions. To this end, a limiting dilution analysis technique was developed that allowed us to evaluate the same frequency of peptide-specific T cells as by staining T cells with HLA–peptide tetrameric complexes. In four out of nine patients, Melan-A/Mart-127–35–specific CTL precursors (CTLp) were ≥1/2,000 peripheral blood lymphocytes and found mostly or only in the CD45RO+ memory T cell subset. In the remaining five patients, a low (<1/40,000) peptide-specific CTLp frequency was measured, and the precursors were only in the CD45RA+ naive T cell subset. Evaluation of CTL effector frequency after bulk culture indicated that peptide-specific CTLs could be activated in all patients by using professional antigen-presenting cells as dendritic cells, but CTLp frequency determined the kinetics of generation of specificity and the final number of effectors as evaluated by both limiting dilution analysis and staining with HLA-A*0201–Melan-A/Mart-1 tetrameric complexes. Immunohistochemical analysis of 26 neoplastic lesions from the nine patients indicated absence of tumor regression in most instances, even in patients with an expanded peripheral T cell pool to Melan-A/Mart-1 and whose neoplastic lesions contained a high frequency of tetramer-positive Melan-A/Mart-1–specific T cells. Furthermore, frequent lack of a “brisk” or “nonbrisk” CD3+CD8+ T cell infiltrate or reduced/absent Melan-A/Mart-1 expression in several lesions and lack of HLA class I antigens were found in some instances. Thus, expansion of peripheral immune repertoire to Melan-A/Mart-1 takes place in some metastatic patients and leads to enhanced CTL induction after antigen-presenting cell–mediated selection, but, in most metastatic lesions, it does not overcome tumor escape from immune surveillance.