Residual Disease In Patients Research Articles

Molecular predictors of residual disease in patients with ovarian cancer

Objectives: Gross residual disease (RD) following primary cytoreduction is the best predictor of overall survival in patients with high-grade serous ovarian cancer (HGSOC). Accurate identification of patients who will have RD has been elusive, prompting many to undergo unnecessary surgical exploration. Our goal was to identify and validate molecular markers associated with high rates of RD. Methods: We interrogated two publicly available genomic datasets of primary HGSOC for genes consistently differentially expressed in RD and no residual disease (R0) cohorts and significant at a false-discovery rate (FDR) of 10%. Genomic expression was further validated in an independent cohort (chemonaive ovarian tumor tissues) using quantitative reverse-transcriptase polymerase chain reaction followed by a blinded prediction of RD. A one-sided Fisher’s exact test was used to compare RD rates in predicted highand low-risk groups. Results: In the TCGA and Tothill datasets, 491 and 189 patients, respectively, met the criteria for inclusion. As expected, survival was significantly better for patients with R0 resection compared to those with any RD. We identified 47 probesets, representing 38 different genes, significant in both datasets at 10% FDR. These included probesets for FABP4 and ADH1B, which tracked tightly and showed dynamic ranges N16-fold. For these genes, there is a level of expression above which nearly all patients have RD; the distribution of expression values is roughly bimodal. In the validation cohort, using the top quartile of FABP4 polymerase chain reaction values as a prespecified cutoff, we found 30/35 RD cases in the high-expression group and 54/104 in the low-expression group (P= 0.0002). Our predictive method based on FABP4, therefore, correctly identified a cohort of patients with significantly increased rates of RD in an independent test set. Further examination of the ADH1B results showed that predictions using either ADH1B alone or in combination with FABP4 would have produced similar results. Examining the reasons for RD in the high-risk cohort suggests a preponderance of cases with either innumerable sites of disease or unresectable disease involving vital regions. Conclusions: High FABP4 and ADH1B expression are associated with significantly higher risk of RD in HGSOC patients. Patients with high tumoral FABP4 levels may be candidates for neoadjuvant chemotherapy.

The impact and interaction of preoperative disease burden, complex surgery, and residual disease in patients with advanced stage ovarian cancer and the effect on patient survival: A GOG 182 analysis

The impact and interaction of preoperative disease burden, complex surgery, and residual disease in patients with advanced stage ovarian cancer and the effect on patient survival: A GOG 182 analysis

Plasma-derived exosomes mediate immune suppression and serve as a biomarker of response to chemotherapy in Acute Myeloid Leukaemia (AML) (TUM7P.945)

Abstract Exosomes are virus-sized (30-100nm) vesicles formed within endocytic compartment and released into extracellular space. While all cells are capable of releasing exosomes under physiologic conditions, tumor cells are avid exosome producers. We have previously shown that plasma of AML patients at diagnosis contain high levels of exosomes relative to normal controls’ plasma (NC). AML exosomes carry various immunosuppressive molecules, including TGF-β1 related to NK-cell dysfunction in AML. Here we show that molecular content of exosomes isolated from patients’ plasma before, during and after chemotherapy (CT) reflects disease status. Thus, exosomes present in patients’ circulation carry biomarkers that can be useful in evaluating responses to therapy. AML exosomes carry different forms of TGF-β1 (pro-peptide, latency-associated peptide or active form) in different proportions. Only TGF-β1 pro-peptide was seen in exosomes obtained from NC or patients in remission. While TGF-β1 levels tracked protein levels in exosomes, elevated TGF-β1 levels discriminated better between patients receiving CT whose leukemia was likely to recur. In addition, co-incubation of normal NK cells with exosomes carrying active TGF-β1 resulted in down-regulation of NKG2D expression. In conclusion, elevations in exosomal protein and/or TGF-β1 levels during CT may reflect an incomplete response to CT and suggest the presence of residual disease in patients considered to have achieved complete remission.

Plasma exosomes as markers of therapeutic response in patients with acute myeloid leukemia.

Purpose: Exosomes isolated from the plasma of newly diagnosed acute myeloid leukemia (AML) patients have elevated protein and transforming growth factor-beta 1 (TGF-β1) contents and inhibit natural killer (NK) cell cytotoxicity (Haematologica 96, p. 1302, 2011). A potential role of exosomes in predicting responses to chemotherapy (CT) was evaluated in AML patients undergoing treatment.Experimental Design: Plasma was obtained from AML patients at diagnosis (n = 16); post-induction CT (n = 9); during consolidation CT (n = 10); in long-term remission (Lt-CR, n = 5); and from healthy volunteers (n = 7). Exosomes were isolated by size-exclusion chromatography and ultracentrifugation. The exosomal protein, soluble TGFβ-1 levels (ELISA), and the TGF-β1 profiles (western blots) were compared among patients’ cohorts. The results were correlated with the patients’ cytogenetic profile, percentage of leukemic blast, and outcome.Results: At diagnosis, protein and TGF-β1 levels were higher (p < 0.009 and p < 0.004) in AML than control exosomes. These values decreased after induction CT (p < 0.05 and p < 0.004), increased during consolidation CT (p < 0.02 and p < 0.005), and normalized in Lt-CR. While TGF-β1 and protein levels tracked one another, TGF-β1 pro-peptide, latency-associated peptide (LAP), or mature TGF-β1 differentially decorated exosomes isolated before, during, and after CT. Only TGF-β1 pro-peptide was seen in exosomes of controls or Lt-CR patients. During consolidation CT, exosomes carried TGF-β1 pro-peptide, LAP, and low levels of mature TGF-β1. NK cell co-incubation with AML exosomes carrying all three TGF-β1 forms induced down-regulation of NKG2D expression.Conclusion: Changes in exosomal protein and/or TGF-β1 content may reflect responses to CT. The exosomal profile may suggest the presence of residual disease in patients considered to have achieved complete remission.

The prognostic impact of minimal residual disease in patients with chronic lymphocytic leukemia requiring first-line therapy.

A proportion of patients with chronic lymphocytic leukemia achieve a minimal residual disease negative status after therapy. We retrospectively evaluated the impact of minimal residual disease on the outcome of 255 consecutive patients receiving any front-line therapy in the context of a detailed prognostic evaluation, including assessment of IGHV, TP53, NOTCH1 and SF3B1 mutations. The median follow-up was 73 months (range, 2-202) from disease evaluation. The median treatment-free survival durations for patients achieving a complete response without or with minimal residual disease, a partial response and no response were 76, 40, 11 and 11 months, respectively (P<0.001). Multivariate analysis revealed that three variables had a significant impact on treatment-free survival: minimal residual disease (P<0.001), IGHV status (P<0.001) and β2-microglobulin levels (P=0.012). With regards to overall survival, factors predictive of an unfavorable outcome were minimal residual disease positivity (P=0.014), together with advanced age (P<0.001), unmutated IGHV status (P=0.001), TP53 mutations (P<0.001) and elevated levels of β2-microglobulin (P=0.003). In conclusion, for patients requiring front-line therapy, achievement of minimal residual disease negativity is associated with significantly prolonged treatment-free and overall survival irrespective of other prognostic markers or treatment administered.

Treatment of Microscopic Disease with Hyperthermic Intraoperative Intraperitoneal Chemotherapy After Complete Cytoreduction Improves Disease-Free Survival in Patients with Stage IIIC/IV Ovarian Cancer

We analyze the efficacy of hyperthermic intraoperative intraperitoneal chemotherapy (HIPEC) for microscopic residual disease in patients with stage IIIC/IV ovarian cancer after a complete cytoreduction of their disease. We analyzed the data of 87 consecutive patients diagnosed with stage IIIC/IV ovarian cancer operated between December 1998 and July 2011. In every patient it was possible to achieve a complete cytoreduction of their disease. (Since January 2008, our center has incorporated the use of HIPEC in patients with peritoneal surface malignancies, including patients with peritoneal dissemination of primary ovarian cancer.) Of 87 patients, 52 were treated with HIPEC (paclitaxel 60 mg/m(2), 60 min, 42 °C). After a univariate analysis, factors associated with lower disease-free interval were: performing a gastrointestinal anastomosis, operative time greater than 270 min, poorly differentiated histology, and not being treated with HIPEC. After multivariate analysis, independent prognostic factors included not being treated with HIPEC [hazard ratio (HR) 8.77, 95 % CI 2.76-14.42, p < 0.01] and the presence of poorly differentiated tumors (HR 1.98, 95 % CI 1.45-8.56, p < 0.05). Disease-free survival at 1 and 3 years was 66 and 18 %, respectively, in patients without HIPEC and 81 and 63 %, respectively, in patients treated with HIPEC (p < 0.01). HIPEC administration did not alter the results obtained for disease-free survival in patients with undifferentiated tumors. The treatment of the microscopic disease following complete cytoreduction with HIPEC in patients with advanced ovarian cancer is effective and can prolong disease-free survival. This survival benefit was not seen in undifferentiated tumors.

Prognostic relevance of minimal residual disease in colorectal cancer.

Presence of occult minimal residual disease in patients with colorectal cancer (CRC) has a strong prognostic impact on survival. Minimal residual disease plays a major role in disease relapse and formation of metastases in CRC. Analysis of circulating tumor cells (CTC) in the blood is increasingly used in clinical practice for disease monitoring of CRC patients. In this review article the role of CTC, disseminated tumor cells (DTC) in the bone marrow and micrometastases and isolated tumor cells (ITC) in the lymph nodes will be discussed, including literature published until September 2013. Occult disease is a strong prognostic marker for patient survival in CRC and defined by the presence of CTC in the blood, DTC in the bone marrow and/or micrometastases and ITC in the lymph nodes. Minimal residual disease could be used in the future to identify patient groups at risk, who might benefit from individualized treatment options.

When is a completion axillary lymph node dissection necessary in the presence of a positive sentinel lymph node?

BackgroundThe management of the axilla in the presence of positive sentinel lymph node (SLN) remains controversial. Many centres forgo completion axillary lymph node dissection (cALND) in the presence of micrometastatic disease. The American College of Surgeons Oncology Group (ACOSOG) Z0011 trialists argue for extending this to macrometastasis. The aim of this study was to correlate tumour burden in SLNs with that in the residual lymph node basin to determine the likelihood of residual disease in patients with micro- and macrometastasis in the SLN. MethodsPatients who underwent cALND following a positive SLN were analysed for histopathological features of the primary tumour and burden of axillary disease. ResultsOf 155 patients, 115 (74%) had macrometastases and 40 (26%) micrometastases in the SLNs. Residual axillary disease was detected in 55/155 (35%) patients with macrometastases and 4/40 (10%) with micrometastases. Generally, with increasing size of metastasis in the SLN there was an increasing risk of further disease in residual lymph nodes. Logistic regression analysis showed increased odds ratios for further disease for all groups when compared with the <2mm (micrometastasis) SLN group. ConclusionPatients may be advised to forgo cALND where the SLN contains isolated tumour cells or micrometastasis. Recommendations for proceeding to cALND can be based on the size of metastasis in the SLN, which relates to the risk of further disease in the residual axillary lymph nodes and subsequent regional recurrence.

A randomized controlled trial for the use of Thymus Honey in decreasing Salivary Gland Damage following Radioiodine Therapy for Thyroid cancer: research protocol

To test the effectiveness of thymus honey as a complementary intervention for decreasing the salivary gland damage due to Radioiodine ((131) I) therapy. Radioiodine is the treatment of choice in people diagnosed with thyroid cancer following total thyroidectomy. Although its value has been acknowledged in eradicating remnant thyroid tissue and treating residual disease in patients with visible, inoperable, iodine-avid metastases, it has been associated with various salivary gland side effects. This is a randomized controlled trial with a 2 × 3 mixed between-within subjects design. In total, 120 participants of postsurgical differentiated thyroid cancer, who will be referred to this centre for (131) I therapy to ablate the remnant thyroid tissue or to treat metastatic tumour, will be prospectively studied under varying regimens of lemon candy (standard treatment) and thymus honey mouthwashes (experimental intervention). Patients will be randomized in four equally numbered groups based on the assumptions and hypothesis of the study. The recruiting process will be informed by predefined inclusion and exclusion criteria. Mixed statistical modelling will be adopted taking into consideration between and within subjects' effects and repeated measures. The recommended intervention protocol is expected to improve the comprehensive management of salivary gland-related side effects induced by the radioiodine treatment in people diagnosed with thyroid cancer. Through the methodological approach chosen, the ideal intervention protocol in terms of the time to initiate the intervention and the frequency of the intervention to acquire optimal results in minimizing salivary glands damage will be tested.

Combined ROR1 and CD160 Detection For Improved Minimal Residual Disease In Patients With Chronic Lymphocytic Leukemia (CLL)

IntroductionOver the past decade, treatments for patients with chronic lymphocytic leukemia (CLL) have produced complete remissions (CR) without evidence for minimal residual disease (MRD), particularly for younger and/or fitter patients. In this setting, achieving an MRD-negative CR has prognostic implications, yielding longer progression free survival (PFS) and overall survival (OS) than for patients who achieve a CR with persistent MRD. Complicating efforts to incorporate testing for MRD in clinical practice has been the lack of a defined consensus on the methods of MRD detection. In this study, we report on a novel combination of mAbs for MRD detection by flow cytometry based on two antigens: the NK-cell receptor and tumor specific antigen, CD160; and the tumor associated antigen, receptor tyrosine kinase-like orphan receptor 1 (ROR-1). ObjectiveTo compare a novel single-tube, tumor-specific (CD160+ROR1) targeted approach to MRD detection against the previously published CD160 flow cytometric assay (CD160FCA) (Farren et al, 2011) and the new, single-tube 8-color ERIC assay. MethodsBetween October 2012 and July 2013, prospective assessment of MRD was performed on peripheral blood in 56 patients (86 samples). We developed a flow cytometric assay using mAbs specific for CD160 or ROR1 (Fukuda et al, 2008). For this we used the following mAb from BD Biosciences: CD2 FITC, CD5 Pe-Cy7, CD19 PerCP5.5, CD45V500, CD160PE, ROR-1 AF647 (“ROR-160FCA”) and a sequential gating strategy. This was compared with CD160FCA and the 8-color ERIC consortium protocol (unpublished). Light chain analysis (LCR) was performed in all cases and reported where detectable. A proof of concept spiking experiment simulating MRD was prepared by mixing CLL and normal peripheral blood leukocytes in a serial dilution to a level of 10-5(n=3). Statistical analysis was performed using Spearman Rank correlation coefficients, Mann-Whitney t-test, and Bland-Altman method comparison. Significance was set at <0.05%. ResultsTo establish the proof of principle, MRD levels ranging from 0.001% to 100% (Neat CLL) were prepared by serial dilutions, in which MRD levels could be established by the ROR-160FCA to 10-5 (n=3). Assessment of the observed incidence against expected incidence of CLL MRD demonstrated a highly significant correlation (R2=0.96, p=0.01). In the study, the range of detectable disease went from <0.01% to 38.59%, of which 37% of samples had levels below <0.01%. Analyzing all flow cytometric methods, a highly significant correlation was observed between all three: CD160FCA vs ERIC: Spearman R=0.96 (95%CI: 0.93 - 0.97, p<0.001); CD160FCA vs ROR-160FCA: Spearman R=0.97 (95%CI: 0.96 – 0.99, p<0.001); ROR-160FCA vsERIC: Spearman R=0.97 (95%CI: 0.94 – 0.98, p<0.001).54 samples had levels of disease <1%. Bland-Altman assay comparison in these patients again demonstrated significant associations between the assays (CD160FCA vs ERIC: mean 0.08 ±0.15; CD160FCA vs ROR-160FCA: mean 0.04 ±0.19; ROR-160FCA vs ERIC: mean 0.03 ±0.25). Light chain restriction was detectable in 24 patients (size of the restricted population ranged from 0.2% to 47% of all cellular events). This sub-group of patients also demonstrated excellent correlation between level of LCR and detectable disease by CD160FCA (Spearman R=0.96, 95%CI: 0.92-0.98, p<0.001), ERIC (R=0.95, 95%CI: 0.92-0.98, p<0.01) and ROR-160FCA (R=0.97, 95%CI 0.93-0.98, p<0.001). ConclusionMonitoring minimal residual disease in CLL is a key focus for clinical trials, as MRD is an important prognostic marker in CLL in terms of PFS and OS. Here we provide a single tube assay, ROR-160FCA, which is unique by targeting two antigens restricted to malignant B-cells, CD160 and ROR1. ROR-160FCA is equivalent in MRD detection compared to both CD160FCA and the current ERIC assay under development. The two tumor-specific antigens give ROR-160FCA the potential for improved sensitivity, particularly where limited sample is available. Furthermore, it only requires a simple sequential gating strategy, is rapid, and appears more cost effective than other Methods.ReferencesFarren TW, Giustiniani J, Liu FT et al. Blood. 2011;118 (8):2174-2183.Fukuda T, Chen L, Endo T et al. Proc Natl Acad Sci U S A. 2008;105 (8):3047-3052. Disclosures:Farren:BD Biosciences: Research Funding. Warner:BD Biosciences: Employment, Research Funding. Agrawal:BD Biosciences: Research Funding.

Hematopoietic Cell Transplantation for Mantle Cell Lymphoma: Predictive Value of Pretransplant Positron Emission Tomography/Computed Tomography and Bone Marrow Evaluations for Outcomes

The prognostic roles of 18F-fludeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) imaging and marrow involvement evaluation on outcomes following autologous and allogeneic hematopoietic cell transplantation (HCT) for mantle cell lymphoma (MCL) are uncertain and require more data. We categorized 66 patients with MCL who received HCT (38 autologous and 28 allogeneic) on the basis of pre-HCT residual disease (RD) status as assessed by marrow MCL morphology and flow/molecular analysis and PET/CT imaging to RD positive (RD(+)) (either or both measures positive) and RD(-) (both negative). We analyzed the predictive value of these RD detection methods on transplant outcomes. The 2-year relapse rate after autograft was significantly higher in pre-HCT RD(+) patients (46% [95% CI 16-77%]) than in patients who were RD(-) (19% [95% CI 0-42%]; P = .02), leading to worse 5-year disease-free survival (DFS) in RD(+) patients (46% [95% CI 14%-73%] vs. 68% [95% CI 33-87%], P = .04). In multivariate analysis, RD(+) status was associated with a reduction in DFS (hazard ratio, 5.6; P = .02). Most allogeneic HCT recipients had advanced disease and most were RD(+) (12 PET/CT(+); 5 marrow-positive). The 5-year DFS and relapse rates after allogeneic HCT were 34% and 25% for all patients and 40% and 33% for RD(+) recipients, suggesting that active disease at the time of allograft does not preclude long-term remissions in advanced MCL. Both autologous and allogeneic HCT lead to promising long-term survival. RD detected prior to autograft was associated with increased relapse and worse 5 year DFS. Allograft recipients had favorable long-term outcomes even in presence of pre-HCT detectable disease.

Fluorescence in situ hybridization panel for monitoring of minimal residual disease in patients with double minute chromosomes

A double minute chromosome (dmin) is a small fragment of extrachromosomal DNA bearing amplified genes observed in malignancies. We investigated the incidence and characteristics of dmins in hematologic malignancies, and the quantitative changes during the treatment follow-up. Once a dmin was observed in conventional G-banding, it was characterized using fluorescence in situ hybridization (FISH) with the panel of MYC, NMYC, and MLL probes. Quantitative changes of malignant cells were measured using G-banding and FISH during the follow up. Dmins were observed in 1.23% of patients (6/489) at the initial diagnosis including 4 with MYC amplification, 1 with MLL and 1 with NMYC. All 6 had complex karyotypes and showed short overall survival (7.7 months). In follow-up specimens, FISH detected dmins in 11 cases out of which G-banding detected dmins in 9 cases. The number of dmins detected by FISH and G-banding did not correlate well. Amplification of NMYC in dmins is reported for the first time. A FISH panel composed of frequently amplified oncogenes (MYC, NMYC, and MLL) in dmins is useful for characterization of dmins. FISH is a sensitive method in detecting dmins and will be useful in monitoring of the minimal residual disease.

Detecting minimal residual disease in patients with chronic lymphocytic leukemia using 8‐color flow cytometry protocol in routine hematological practice

Minimal residual disease (MRD) detection has become increasingly important for the assessment of therapy response in chronic lymphocytic leukemia (CLL). However, current MRD analysis methods, both molecular genetic and flow cytometric, are time-consuming and require experienced laboratory staff. To reduce the demands of flow cytometric MRD detection in CLL, we have introduced a novel flow cytometric 8-color protocol. The MRD analysis results using this protocol were then compared with the commonly employed 4-color protocol and the molecular genetic (real-time quantitative allele-specific oligonucleotide IGH polymerase chain reaction; RQ-ASO IGH PCR) approach. Forty-two CLL patient samples were repeatedly analyzed after allogeneic stem cell transplantation (n=20) or after fludarabine-based therapy (n=22), and 100% concordance was found using both flow cytometric protocols. Furthermore, there was a strong correlation (r=0.94) between flow cytometric and RQ-ASO IGH PCR results in MRD detection. Flow cytometry is less time-consuming, less financially demanding, and moreover, MRD assessment using our novel 8-color protocol is less complicated than the 4-color approach and molecular methods.

Gemtuzumab ozogamicin can reduce minimal residual disease in patients with childhood acute myeloid leukemia

Gemtuzumab ozogamicin (GO) is an active agent for the treatment of CD33-postive acute myeloid leukemia (AML) and may improve the outcome of specific patient subgroups when combined with conventional chemotherapy. However, to the best of the authors' knowledge, the effects of GO on levels of minimal residual disease (MRD) are unknown. Pediatric patients with AML who received GO, either alone or in combination with chemotherapy on the AML02 multicenter trial, were analyzed to determine the effects of GO on MRD and outcome. Among 17 patients who received GO alone because of persistent leukemia, 14 had a reduction in their MRD level and 13 became MRD negative. Of the 29 who received chemotherapy in combination with GO after responding poorly to chemotherapy, 28 demonstrated reduced MRD and 13 became MRD negative. Treatment with GO effectively reduced MRD before hematopoietic stem cell transplantation and was not found to be associated with increased treatment-related mortality after transplantation. GO is effective in reducing MRD levels in pediatric patients with AML and may improve the outcome of those patients at high risk of disease recurrence.

Detection of minimal residual disease in patients with multiple myeloma using clonotype-specific PCR primers designed from DNA extracted from archival bone marrow slides

Polymerase chain reaction (PCR)-negative molecular complete remission (mCR) can be induced by stem cell transplantation in some patients with multiple myeloma (MM) and is associated with long-term progression-free survival (PFS). The detection of molecular minimal residual disease (MRD), however, requires fresh or frozen materials for designing clone-specific primers, which are not always readily available. In this study, we used DNA extracted from archival bone marrow (BM) slides for PCR to detect MRD in 50 patients with MM who received various induction therapies and autologous peripheral blood stem cell transplantation (ASCT). Clonotype-specific immunoglobulin (Ig) H PCR primers were prepared for 32 of 50 cases (64%) using BM slides, and for 9 of 14 cases (64%) using fresh BM cells. DNA in peripheral blood stem cell autografts of the 22 patients who achieved at least a partial response after ASCT was subjected to PCR to amplify clonotype-specific rearranged IgH gene sequences. The median PFS of the eight patients with MRD-positive autografts was 18 months, whereas that of 14 patients with MRD-negative autografts was not reached at a median follow-up of 27 months (p = 0.012). Post-ASCT PFS of the four patients who achieved mCR was 100% at a median follow-up of 47 months. These results indicate that archival BM slides can serve as a source of DNA for preparing clonotype-specific primers for MRD monitoring in patients with MM whose cryopreserved myeloma cells are not available for DNA preparation. Our results also suggest that patients with MM who received MRD-negative autografts and achieved mCR have a long PFS.

Clinical significance of minimal residual disease in patients with acute leukaemia undergoing haematopoietic stem cell transplantation

In patients with acute leukaemia, the relative risk of relapse influences the choice between chemotherapy and haematopoietic stem cell transplantation (HSCT). The demonstration that minimal residual disease (MRD) is the strongest overall prognostic indicator and can identify patients who are unlikely to be cured by standard chemotherapy has added a powerful new factor to consider when making this decision. There is substantial data indicating that the likelihood of relapse after transplant is directly correlated with levels of MRD before transplant. This knowledge can be used to adjust the timing of HSCT, and guide the selection of donor, conditioning regimen, and post-HSCT strategies to maximize the graft-versus-leukaemia effect. Because MRD emerging post-transplant carries a dire prognosis, its detection can trigger withdrawal of immunosuppression, additional cellular and molecular therapies, or preparations for a second HSCT. Although it is not yet clear whether any of these actions will significantly improve outcome, it is likely that they will be most effective for patients with a relatively low tumour burden, who can be identified only through MRD testing. In this article, we review the clinical significance of MRD in the context of autologous and allogeneic HSCT.

Susceptibility of multiple myeloma stem cells and their progeny to NK‐cell mediated cytotoxicity

BackgroundThe purpose of our study was to examine sensitivity of MM cells and their SC progenitors (which are characterized by negativity for CD138 and active efflux of Hoechst 33342 dye) to natural killer (NK)‐cell mediated cytotoxicity.MethodsNK cells were generated from peripheral blood mononuclear cells of healthy volunteers and co‐cultured with MM (RPMI 8226) cells for 4 hours. NK cell cytotoxicity was assessed by flow cytometry assay. Three different NK/target cell ratios were used. MMSC represented by side population (SP) (negative for Hoechst and CD138) were quantified as percentage of total viable MM cells.ResultsNK cells exhibited significant cytotoxicity against RPMI 8226 cells in all three ratios. Surprisingly, the strongest cytotoxicity was demonstrated when minimal NK/MM ratio (2.5:1) was used. We hypothesize that this phenomenon could be due to NK‐NK cell fratricide. Remarkably, proportion of the overall viable target cell population, represented by MMSC, decreased.ConclusionsMM cells are sensitive to NK‐mediated cytotoxicity. The finding of major importance was potential susceptibility of MMSC to NK‐cell mediated cytotoxicity which could make NK cells additive armamentarium in treating minimal residual disease in patients with MM.

Influence of imatinib interruption and rechallenge on the residual disease in patients with advanced GIST: results of the BFR14 prospective French Sarcoma Group randomised, phase III trial

BackgroundWe previously demonstrated that interruption of imatinib mesylate (IM) in responding patients (pts) with advanced gastrointestinal stromal tumours (GISTs) results in rapid reprogression. The impact of interruption on residual tumour, quality of response and secondary resistance has not been fully investigated. Patients and methodsWithin the BRF14 study, 71 non-progressing patients were randomly assigned in the interruption arms after 1, 3 or 5 years. IM was resumed in the case of progressive disease (PD). Tumour status at randomisation, relapse and after IM rechallenge, progression-free survival (PFS) and time to secondary resistance were analysed. ResultsAt data cut-off, 51 of 71 patients had restarted IM following documented PD. Eighteen patients (35%) progressed on known lesions only, while 33 patients (65%) had new lesions, with concomitant progression of known lesions in 17 patients. Only 8 (42%) of complete remission (CR) patients and 12 (52%) of partial response (PR) patients at randomisation achieved a new CR and PR. Patients progressing rapidly after interruption had a poorer prognosis. Tumour status at randomisation influenced time to progression after rechallenge. ConclusionIn advanced GIST patients interrupting IM, quality of response upon reintroduction did not reach the tumour status observed at randomisation. Rapid progression after imatinib interruption is associated with poor PFS after reintroduction.

Serial Measurement of WT1 Expression and Decrement Ratio Until Hematopoietic Cell Transplantation as a Marker of Residual Disease in Patients with Cytogenetically Normal Acute Myelogenous Leukemia

Using real-time quantitative PCR, we monitored Wilms tumor gene 1 (WT1) expression from diagnosis to hematopoietic stem cell transplantation (HSCT) in adult patients with cytogenetically normal acute myelogenous leukemia (CN-AML) and FLT3-ITD and NPM1 mutations. The values at diagnosis were evaluated in 104 patients. Data collected after induction chemotherapy were available for all patients, but only 68 patients were treated with HSCT. Significant WT1 expression cut-offs were determined by receiver operation characteristic curve analysis, and rates of overall survival (OS) and disease-free survival (DFS) were estimated. WT1 decrement ratios (DR) at postinduction chemotherapy and at pre- and post-HSCT compared with the diagnostic level were calculated. Higher WT1 expression at diagnosis, postinduction chemotherapy, and pre-HSCT showed inferior OS (P = .015, <.001, and .002) and DFS (P = .006, <.001, and .003). The cut-offs were determined at the median for diagnostic WT1 expression and at the 25% level from the top for other time points excluding post-HSCT. The WT1 DR ≥ 1-log after induction chemotherapy showed superior OS and DFS (P = .009 and .002) and WT1 DR ≥ 1-log preceding HSCT also showed superior OS and DFS (P = .009 and .003). Results of WT1 DR were consistently applicable in each subgroup with higher (≥1.0) and lower (<1.0) WT1 expression at diagnosis and also in NPM1-wild-type/FLT3-ITD–negative CN-AML. The WT1 DR therefore predicted survival outcomes after HSCT more accurately than did the diagnostic WT1 expression. WT1 expression may serve as a reliable marker for residual disease and WT1 DR as a prognostic indicator, particularly in NPM1-wild-type/FLT3-ITD–negative CN-AML. These measures may be applied throughout the course of treatment and even after HSCT.

Prospective evaluation of gene mutations and minimal residual disease in patients with core binding factor acute myeloid leukemia

AbstractNot all patients with core binding factor acute myeloid leukemia (CBF-AML) display a good outcome. Modern risk factors include KIT and/or FLT3 gene mutations and minimal residual disease (MRD) levels, but their respective values have never been prospectively assessed. A total of 198 CBF-AML patients were randomized between a reinforced and a standard induction course, followed by 3 high-dose cytarabine consolidation courses. MRD levels were monitored prospectively. Gene mutations were screened at diagnosis. Despite a more rapid MRD decrease after reinforced induction, induction arm did not influence relapse-free survival (RFS) (64% in both arms; P = .91). Higher WBC, KIT, and/or FLT3-ITD/TKD gene mutations, and a less than 3-log MRD reduction after first consolidation, were associated with a higher specific hazard of relapse, but MRD remained the sole prognostic factor in multivariate analysis. At 36 months, cumulative incidence of relapse and RFS were 22% vs 54% (P < .001) and 73% vs 44% (P < .001) in patients who achieved 3-log MRD reduction vs the others. These results suggest that MRD, rather than gene mutations, should be used for future treatment stratifications in CBF-AML patients. This trial was registered at EudraCT as #2006-005163-26 and at www.clinicaltrials.gov as #NCT 00428558.