Minimal Residual Disease Marker Research Articles

Application of droplet digital PCR in minimal residual disease monitoring of rare fusion transcripts and mutations in haematological malignancies

Leukaemia of various subtypes are driven by distinct chromosomal rearrangement or genetic abnormalities. The leukaemogenic fusion transcripts or genetic mutations serve as molecular markers for minimal residual disease (MRD) monitoring. The current study evaluated the applicability of several droplet digital PCR assays for the detection of these targets at RNA and DNA levels (atypical BCR::ABL1 e19a2, e23a2ins52, e13a2ins74, rare types of CBFB::MYH11 (G and I), PCM1::JAK2, KMT2A::ELL2, PICALM::MLLT10 fusion transcripts and CEBPA frame-shift and insertion/duplication mutations) with high sensitivity. The analytical performances were assessed by the limit of blanks, limit of detection, limit of quantification and linear regression. Our data demonstrated serial MRD monitoring for patients at molecular level could become “digitalized”, which was deemed important to guide clinicians in treatment decision for better patient care.

Multiomic Single-Cell Sequencing May Distinguish BCR-ABL-Mutated Acute Myeloid Leukemia (AML) from Blast Crisis Chronic Myelogenous Leukemia (BC-CML)

Introduction The existence of BCR-ABL+ AML as an entity distinct from BC-CML is controversial. The WHO recently recognized BCR-ABL+ AML, but with the caveat that it is poorly characterized (Khoury, 2022). Clinically, BCR-ABL+ AML is associated with high risk of relapse (Neuendorff, 2018). Further, acquired BCR-ABL mutations have been identified as a cause of relapse on FLT3 inhibitors (McMahon, 2019). We hypothesize that BCR-ABL+ AML can be distinguished as a distinct biologic entity from BC-CML by the observation of BCR-ABL as a sub-clonal rather than a founder mutation in leukemogenesis. Methods To test the utility of multi-omic single cell (SC) sequencing in determining tumor phylogeny and identification of BCR-ABL+ AML vs BC-CML, we analyzed bone marrow samples from 4 patients (pts): 1 relapsed AML with clinical development of a new BCR-ABL fusion at the time of relapse, and 3 newly diagnosed BC-CML (2 myeloid, 1 lymphoid). We performed high-throughput SC DNA sequencing with simultaneous cell-surface immunophenotyping and targeted RNA sequencing for BCR-ABL fusion proteins (Mission Bio Tapestri). All SC DNA sequencing included pt-specific primers for BCR-ABL DNA breakpoints, which were previously identified using nanopore-based technology for sequencing long strands of DNA. Results We analyzed 19,197 single cells (median 1995 cells/pt). Multi-omic SC technology differentiated BCR-ABL+ AML from BC-CML by confirming the fusion arose as a branch mutation from a founding leukemic clone and was not present in all leukemic populations. In Patient 1, who was sampled at time of AML relapse, we identified pathogenic FLT3 ITD and NRAS Q61K mutations. FLT3 co-mutated with either the BCR-ABL DNA breakpoint or NRAS in the same single cell and clone, while the BCR-ABL DNA breakpoint and NRAS were not co-mutated. Phylogenetic reconstruction determined FLT3 to be a founding mutation, which later branched into FLT3/BCR-ABL and FLT3/ NRAS co-mutant subclones. Further, the FLT3-ITD/BCR-ABL and FLT3-ITD/ NRAS subclones had distinct immunophenotypes. Blasts harboring the BCR-ABL DNA breakpoint were predominantly identified in the CD117+/CD33+ population (60.2% of cells) compared to CD34+ (16.4% of cells, p <0.001) and blasts with monocytic differentiation (19.1% of cells, p = 0.02). Notably, BCR-ABL+ cells were not identified in T cells. NRAS mutations were predominantly identified in both CD34+ and monocytic blasts. Together, these data support BCR-ABL arising late within the evolutionary history of this pt's AML. By contrast, in Patients 2 and 3 with myeloid BC-CML, the BCR-ABL DNA breakpoint and corresponding BCR-ABL fusion were detectable in all cell populations, including blasts as well as immunophenotypically normal T cells, B cells, and macrophages. In Patient 2, we identified an IDH2 R132C mutation co-mutated with BCR-ABL. Phylogenetic reconstruction determined the IDH2 mutation arose as a branch from a founding BCR-ABL+clone. Unlike the BCR-ABL breakpoint, which was present in all cell populations, the IDH2 mutation was only identified in immunophenotypically-defined CD33+/CD34+/CD117+ myeloid blasts and was absent from bystander T cells and macrophages. Finally, Patient 4had a clinical diagnosis of lymphoid BC-CML. Like Patients 2 and 3, the BCR-ABL DNA breakpoint and corresponding RNA fusion were present in all immunophenotypically-defined blast populations. Unlike Patients 2 and 3, however, BCR-ABL was absent from immunophenotypically normal CD4+ and CD8+ T cells, macrophages, and monocytes. Additional pts are needed to determine whether this pattern is consistent with all lymphoid BC-CML or whether this pt's biology may have been closer to BCR-ABL+ acute lymphoblastic leukemia (ALL). Conclusions Multi-omic SC technology can distinguish BCR-ABL+ AML from myeloid BC-CML and provide direct confirmation that lineage-restriction of BCR-ABL potentially clarifies the diagnosis of discrete Ph+ leukemias. Future studies are needed to understand the role of RT-PCR based BCR-ABL1 testing as a marker of minimal residual disease in pts with BCR-ABL+ subclones, the impact of BCR-ABL as a potential driver of AML resistance, and the utility of BCR-ABL inhibitors in leukemias with late-arising BCR-ABL+ clones. Support: U10CA180821, U10CA180882, U24CA196171; https://acknowledgments.alliancefound.org. Clinicaltrials.gov Identifiers: NCT00048958, NCT00899223, NCT00900224

MRD Monitoring By Euroclonality IGH Based NGS Approach Predicts Outcome in Follicular Lymphoma Patients Lacking a Conventional BCL2::IGH Marker: A Substudy from the Fondazione Italiana Linfomi (FIL) FOLL12 Trial

Background. Despite durable clinical responses achieved by chemo-immunotherapy in follicular lymphoma (FL) patients, most of them eventually relapse. Actually, minimal residual disease (MRD) analysis based on the detection of BCL2::IGH rearrangement by PCR is the most widely used and standardized approach, able to early identify patients at high risk of disease reappearance. Nevertheless, this tool fails the molecular marker definition in up to 45% of cases, hampering the disease monitoring in a large group of patients. In the last decade, the continuous improvement of next-generation sequencing (NGS) techniques allowed to introduce these novel tools for MRD monitoring in some hematologic diseases, as acute lymphoblastic leukemia and multiple myeloma, thus reducing PCR limitations and increasing the number of patients eligible for MRD. Nonetheless, applications of NGS tools for MRD detection in FL are still scant. Aims. Therefore, we tested the feasibility of the EuroClonality (EC) IGH-based NGS approach [Brüggeman, Leukemia 2019] to identify novel molecular markers suitable for MRD monitoring in the context of the large prospective trial “FOLL12” (EudraCT: 2012-003170-60) by the Fondazione Italiana Linfomi (FIL), where 57% of patients were already studied for MRD by classic PCR methods targeting the BCL2::IGH rearrangement.[Luminari, JCO2022] Methods. Advanced FL patients received upfront treatment with either R-CHOP or BR, followed by a combined PET/MRD response-based maintenance with rituximab. Bone marrow (BM) samples were centralized to the FIL MRD Network for standardized MRD analysis. Baseline gDNA of “no marker” patients was screened by IGH-based NGS using EC-IGH two-steps PCR approach in the EuroMRD certified lab of Torino university; then MRD was monitored in those patients harboring a IGH clonal marker at baseline by one step EC-IGH method. The identification of baseline clonotypes and MRD clones was performed by ARRest/Interrogate tool. Results. 343/786 enrolled patients (43%) lacked a conventional BCL2::IGH marker and 161/343 (47%) showed morphologic BM lymphoma infiltration: of these, 124 (77%) had an available BM baseline sample for IGH screening and were thus included in the analysis. Overall, IGH clonotypes suitable as novel molecular markers were identified in 75/124 patients (60%), of which 73 monoclonal and 2 biclonal. Of this series, 47 patients had centralized a BM sample for MRD analysis at end of induction. Median age of this subset was 59 years (33-81), 53% were female, all were Ann Arbor stage IV, 34% were FLIPI intermediate and 43% high risk, respectively. Moreover, 56% received R-CHOP and 44% BR as induction treatment, resulting in a 79% CR rate and 89% MRD negativity rate (in line with the results of the general trial population). PFS at 3 years for this series was 69%: however, the 5 patients who scored MRD positive (MRD+) showed statistically significant worse PFS at 3 years than MRD negative cases (22% vs 73%, respectively, p=0.0069, Figure 1). Notably, 4 out of 5 MRD+ patients experienced POD24. Focusing on biclonal cases, one presented superimposable MRD trends between clones (both achieving MRD negativity), while in the other both clones remained MRD positive but showing one Log difference between them. Interestingly, only one of the two clones showed a MRD increase in the following timepoint (6 months after end of induction), heralding the clinical relapse subsequently occurred at month 12 (Figure 2). On the other hand, no IGH clonotype was identified in the first 17 patients who were screened for a novel MRD marker despite the absence of BM infiltration. Conclusions. The first data on the large-scale application of NGS-based marker screening tools in FL patients lacking a conventional BCL2::IGH marker for MRD enrolled in a prospective clinical trial showed that the EC IGH-based NGS approach was able to provide a new molecular marker in 60% of patients with evidence of BM infiltration (that means about the 28% of “no marker” FL patients); importantly, this approach was predictive of worse PFS for MRD+ cases. Overall, these data are promising for the wide employment of novel, effective MRD monitoring tools, able to recover more than one fourth of FL patients lacking a conventional molecular marker and so, by combining conventional and NGS-based techniques, potentially rising the percentage of MRD evaluable cases up to at least 70% cases.

Improved MRD Negativity Rates in Adverse Genomic Risk B-ALL Patients with Chemotherapy / Blinatumomab Induction: Experience from the Australasian Leukaemia Lymphoma Group (ALLG) ALL06/09 Studies

Introduction: Despite the benefits conferred by pediatric treatment protocols, outcomes in adolescent and young adult (AYA) acute lymphoblastic leukemia (ALL) remain inferior to children. Poor outcomes in AYA ALL have been associated with reduced tolerability of intensive cytotoxics and higher incidence of adverse risk disease. Our earlier ALLG ALL06 study demonstrated intensive pediatric induction/consolidation could be delivered in a similar time frame as in children suggesting tolerability was not a major contributor to poor outcome in AYA ALL. ALL06 established post consolidation (TP2=d79) minimal residual disease (MRD) positivity (MRD+) and high BMI was predictive of inferior disease free- and overall survival (OS). In order to improve TP2 MRD negativity (MRD-) in B-cell disease, the ALLG ALL09 study substituted standard consolidation for blinatumomab (blin) in phase I and II of a BFM-style protocol . We present the impact of blin on outcomes in genomic cohorts from ALL09 and compare these to similar cohorts from ALL06 treated with standard chemotherapy. Methods: Between Sep 2020 and Apr 2022, 55 patientswere enrolled to ALL09. Of these, 45 had appropriate samples for genomic analyses and 39/45 had an MRD marker identified. These 39 patients formed the genomic cohort presented below. Genomic subtyping was performed using whole transcriptome sequencing, MLPA, karyotype and FISH analyses, and disease was classified into genomic adverse (AR) vs standard (SR) risk based on contemporary literature classifiers. Bone marrow MRD was assessed using ASO-PCR for IGH and TCR rearrangements, at d33 post induction (TP1), and TP2 post blin consolidation. Differences in TP2 MRD were assessed, with estimated 3 yr relapse free survival (RFS) and OS calculated using the Kaplan-Meier method. Results: In the ALL09 cohort, median follow up was 629 days with 3 yr RFS 84.5% and OS 87.1%. In the genomic cohort, median follow up was 602 days with 3 yr RFS 88.8% and OS 93.2%, respectively. These compared favorably to 3 yr RFS and OS (both 67.5%) in the B-cell genomic cohort from ALL06. Between ALL09 and ALL06 there were similar proportions of AR patients (73% and 62.5%, respectively). In ALL09, the most frequent subtypes were PAX5alt (18%) and PAX5 p.P80R (11%) versus KMT2Ar and DUX4r (both 17.5%) in ALL06. In ALL09, Ph-like disease represented 13% vs 12.5% in ALL06. In the ALL09 genomic cohort, the MRD+ rate reduced between TP1 and TP2 (p< 0.001), with the reduction in the rate of positivity being more pronounced in the AR cohort (AR MRD+ TP1=85% vs AR TP2=39%; p< 0.001). In contrast, in ALL06 MRD+ in the AR cohort reduced by only 28% between TP1 and TP2 (p >0.05). This suggests a positive impact of blin consolidation in AR patients in ALL09. Despite improved AR MRD+ rates overall, not all genomic subtypes in ALL09 followed this trend (Table 1). To date, there are 5 relapses in the ALL09 cohort, 4 from the genomic cohort. Of these, 1 was hypodiploid and strikingly the remaining 3 patients harbored a TCF3 rearrangement ( TCF3r). All 4 relapses were significantly associated with MRD+ at TP2 (RFS: MRD-, 100% (n=14) vs MRD+, 71% (n=25); p= 0.013), and all were observed in the AR genomic group. Of interest, the fifth case, outside of the genomic cohort, was classified as PAX5alt. In ALL09, high-risk (HR) patients proceeded to intensive HR block chemotherapy with the aim of eliminating MRD prior to allogeneic transplant (SCT). In the genomics cohort 15 patients proceeded to HR therapy, n=12 were considered AR. Sustained MRD+ through HR blocks 1 and 2 was noted in patients harboring PAX5 p.P80R (3/3), KMT2Ar (2/2) and TCF3r (2/3). Of the 9 patients who proceeded to SCT, 3 had PAX5 p.P80R 1 had PAX5alt, 2 were KMT2Ar and there was 1 each of DUX4r, hypodiploid and TCF3r. The 3 patients who remained MRD+ in the SR cohort had PAX5 p.P80R and all 3 proceeded to SCT. Conclusion: This is the first report of the impact of blin consolidation on genomic subsets of de novo AYA ALL. Our study suggests blin sensitivity as assessed by TP2 MRD response may be genomic subtype dependent, with TCF3r associated with sustained MRD+, leading to eventual relapse (3/3 patients). In addition, the PAX5 p.P80R subset was also associated with sustained high levels of TP2 MRD+ (3/4 with an MRD marker) and SCT (3/5 patients). These results may assist with identification of AYA ALL subsets that could benefit from alternative approaches as blin becomes increasingly incorporated into front-line protocols.

Prospective Evaluation of Minimal Residual Disease in Waldenström Macroglobulinemia across Different Tissues and Treatments: Results of the “BIO-WM” Trial of the Fondazione Italiana Linfomi (FIL)

Introduction. MYD88 L265P is the hallmark mutation in Waldenström Macroglobulinemia (WM) and is becoming increasingly important in the management of IgM-gammopathies, due to its role as prognostic and predictive biomarker. Recently, MYD88 L265P detection by allele-specific quantitative PCR was proposed as reliable minimal residual disease (MRD) marker in bone marrow (BM) samples (Varettoni, Hematol Oncol 2022). Novel, more sensitive, techniques as droplet digital PCR (ddPCR) might extend the feasibility of MRD detection in WM also in non-invasive tissues, as peripheral blood (PB) or plasmatic cell-free (cf) DNA. This was a secondary endpoint of the multicenter, observational “BIOWM” (NCT03521596) trial, sponsored by the Fondazione Italiana Linfomi (FIL) and the International WM Foundation/Leukemia and Lymphoma Society. From 2018 to 2020 this trial enrolled 300 consecutive patients with primary diagnosis of WM or IgM-MGUS and a systematic biobanking was performed. Here are presented the first results of the MRD study on the patient subset who received frontline treatment, with the aim of driving correlations with clinical response, type of therapy received and outcome prediction. Methods. Paired BM, PB and plasma samples were collected for each patient at baseline (T0) and follow-up and MYD88 L265P detection was performed by ddPCR in all specimens (Drandi, Haematologica 2018). As of today, 59/300 patients (58 WM and 1 IgM-MGUS) received frontline treatment and MRD levels after therapy (T2) were detected by ddPCR: actually, T0-T2 paired samples were available in 49 (BM), 56 (PB) and 48 (cfDNA) cases, respectively. Moreover, MRD was evaluated also by 8-color multiparametric flow cytometry (MFC) in 23 BM and 24 PB samples with a minimum of 10 million cells acquired. Results. Median age of the 59 treated patients was 68 years (24-85), 32% were female, median IgM value was 2420 mg/dL (101-8840), median Hb 10.5 g/dL (8 -17) and IPSS-WM was high in 35% and intermediate in 46%. MYD88 L265P mutation rate was 94% (46/49) in BM, 80% (45/56) in PB and 90% (43/48) in cfDNA samples, respectively. Treatment was started because of anemia in 32% cases, lymphadenopathy or splenomegaly in 34%, hyperviscosity in 20%, other reasons in 13%, and the only MGUS patient required treatment for anti-myelin-associated glycoprotein polyneuropathy. Thirty-one out of 59 patients received bendamustine-rituximab (BR), 23/59 dexamethasone, rituximab and cyclophosphamide (DRC) or DRC-like regimen and 5/59 a single agent therapy (namely, 3 rituximab, 1 cyclophosphamide, 1 ibrutinib). Overall response rate was 90% (87% for BR and 96% for DRC), with 19% CR, 29% VGPR and 35% PR in BR and 4% CR, 0% VGPR and 91% PR in DRC, respectively. Overall, MRD negativity after treatment was 30% (14/46) in BM, 89% (40/45) in PB and 54% (23/43) in cfDNA, respectively. Moreover, among patients still MRD positive after treatment, median MYD88 L265Ptumor burden shrinkage was about 2 Logs in BM (5.5E-03 vs 2.1E-01) and around 1 Log in cfDNA (5.9E-03 vs 3.7E-02) and in PB samples (3.6E-03 vs 2.5E-02), respectively (Figure 1). Interestingly, the MRD shrinkage in BM was deeper among patients receiving BR (48% MRD negative with a median MYD88 L265P decrease of more than 2 Logs among patients still MRD positive) vs patients receiving DRC (10% MRD negative with a median decrease of 1 Log), Figure 2. Similar trends in favor of BR in inducing MRD negativity were reported in PB (96% vs 88%) and cfDNA (70% vs 35%), respectively. MRD results by MFC were overall in line with ddPCR results: namely 27% of patients reached MRD negativity in BM and 69% in PB. The median follow-up for the whole series was 41 months, resulting in a 3-years PFS of 71% and 3-years OS of 89%, respectively. Interestingly, despite the current limited follow-up for such an indolent disease, MRD positivity by ddPCR in PB predicted a dismal clinical outcome if compared with MRD negative patients (3-years PFS 40% vs 73%, p=0.038). Conclusions. To the best of our knowledge, this is the first report of prospective MRD evaluation in a WM clinical trial. Our preliminary results suggest that MYD88 L265P monitoringby ddPCR is a suitable target for MRD analysis, in the contest of common immunochemotherapeutic schedules. Moreover, different levels of disease persistence were described across BM, PB and cfDNA, with MRD monitoring in non-invasive tissues showing promising predictive value for outcome discrimination.

Use of Circulating Extracellular Vesicles and CtDNA From Lymphatic Fluid Exudate as Markers for Minimal Residual Disease in Melanoma Patients

Use of Circulating Extracellular Vesicles and CtDNA From Lymphatic Fluid Exudate as Markers for Minimal Residual Disease in Melanoma Patients

Determination of the androgen receptor status of disseminated tumor cells in primary breast cancer patients

PurposeAndrogen receptor (AR) can serve as a new therapeutic target since it was shown to play a proliferative role in several breast cancer (BC) subtypes. Moreover, AR positivity has been suggested to reflect the metastatic potential of tumor cells in some BC subtypes. The aim of this study was to determine the AR expression on disseminated tumor cells (DTCs) as a surrogate marker of minimal residual disease (MRD) and potential precursor of metastasis in early BC.MethodsBone marrow (BM) aspirates from 62 DTC-positive early BC patients were included into this study and analyzed by immunofluorescence staining for the presence of AR-positive DTCs. CK-positive, CD45-negative cells containing an intact nucleus (DAPI positive) were identified as DTCs. AR expression of the primary tumor (PT) was assessed by immunohistochemistry on formalin-fixed, paraffin-embedded (FFPE) tumor sections from core biopsies and surgical specimens.ResultsAR status of DTCs could be determined in 21 patients. We detected AR-positive DTCs in nine samples (43%). AR expression of DTCs and corresponding PT showed a concordance rate of 33%. The DTC-AR status did not correlate with clinicopathological factors, nor did we observe a significant correlation between the AR status of the PT and other established prognostic factors for BC.ConclusionAR-positive DTCs can be detected in BM of early BC patients with a marked discordance of the AR status between DTCs and corresponding PTs. The clinical significance of these findings needs further investigation.

CELL‐FREE DNA KINETICS DECIPHER POTENTIAL MECHANISMS OF ACTION OF IBRUTINIB COMBINATION THERAPY IN MANTLE CELL LYMPHOMA (MCL)

Introduction: The European MCL Network trial TRIANGLE (NCT02858258) evaluated the addition of ibrutinib to standard treatment with autologous stem cell transplantation (ASCT) (arm A+I) in comparison to ASCT alone (arm A) and ibrutinib plus chemoimmunotherapy without ASCT (arm I) demonstrating significant and clinically relevant superiority of additional ibrutinib on outcome. Cell-free (cf)DNA captures tumor genetics and can be used to assess disease kinetics and minimal residual disease (MRD). We used capture based targeted sequencing for genotyping and MRD monitoring in cfDNA in the TRIANGLE trial to gain insight into the mechanisms of action of Ibrutinib and to identify patients (pts) with high risk of relapse. Methods: Peripheral blood (PB) and plasma samples from 57 pts with a PB infiltration of ≥10% at baseline (BL) and samples at mid induction (MI, n = 57) and end of induction (EoI, n = 53) were sequenced using the EuroClonality (EC)-NDC assay (Univ8® Genomics UK) to detect immunoglobulin and T-cell receptor (IG/TR) rearrangements, structural- (SV) and single nucleotide variants (SNVs) in 72 genes. Bioinformatics by ARResT/Interrogate with adaptations for cfDNA. Pts were considered MRD+ by detecting ≥1 read of a lymphoma-specific IG, SV or SNV. MRD in PB was analyzed by ASO-qPCR. Results: cfDNA levels were high at BL (mean 93 ng/mL) and MI (124 mg/mL) and decreased at EoI (12 ng/mL). BL cfDNA levels correlated with MIPI scores and identified pts with poor outcome. Genotyping by EC-NDC showed fully concordant IG clonotypes and SVs among 57 paired PB gDNA and cfDNA samples. 126 SNVs were identified (VAF 2.7%–75%, median 30%) in genes described as drivers in MCL, ATM (44%), TP53 (19%), KMT2D (16%), SAMHD1 (8%), CCND1 (8%) and 18 others. On average, 6 MRD markers were identified per patient (range 3-10). Copy number variation analysis revealed deletions in ATM (10%), CDKN2A (16%) and TP53 (5%) pts. TP53 mutations/deletions are a poor prognostic indicator in MCL, but therapy in arms A+I/I (n = 6) overcomes this poor outcome in comparison to arm A (n = 5). At MI, total cfDNA and ctDNA levels were significantly increased in arms A+I/I as compared to arm A (p = 0.049). 20/57 (35%) pts were MRD+ in PB and 35/55 (64%) in cfDNA. ctDNA was detected in 17/21 pts (81%) in Arm A and 18/34 pts (53%) in Arms A+I/I, suggesting effective disease clearance in the lymph nodes by ibrutinib. ctDNA detection at MI predicted outcome only in arm A. At EoI 8/53 (15%) pts were MRD+ in PB and 20/40 (50%) pts were MRD+ in cfDNA. ctDNA was detected in 12/18 pts (66%) in Arm A and 8/22 pts (36%) in Arms A+I/I. Similar to MI, ctDNA at EoI predicted outcome only in arm A. Keywords: combination therapies, liquid biopsy, minimal residual disease Conflicts of interests pertinent to the abstract S. Ferrero Consultant or advisory role: EUSA Pharma, Jannsen Honoraria: Servier Research funding: Gilead, Morphosys, Jannsen V. H. van der Velden Other remuneration: EuroFlow: Patents and Royalties R. García Sanz Honoraria: Janssen, BeiGene, Gilead, Astellas, Amgen, Takeda Research funding: Janssen, Gilead, Astellas, Takeda Educational grants: Janssen M. Dreyling Consultant or advisory role: Novartis, Lilly/Loxo, AstraZeneca, Roche, Janssen, Gilead/Kite, BMS/Celgene, Beigene Honoraria: Novartis, Lilly/Loxo, Amgen, AstraZeneca, Roche, Janssen, Gilead/Kite Research funding: Roche, Janssen, Bayer, AbbVie A. W. Langerak Research funding: Genentech, Roche Other remuneration: Spearkers Bureau: Janssen, Gilead D. Gonzalez de Castro Employment or leadership position: Founder and director of Univ8 Genomics Ltd.

COMBINED USE OF MINIMAL RESIDUAL DISEASE MONITORING AND FDG‐PET FOR OUTCOME PREDICTION IN FOLLICULAR LYMPHOMA: RESULTS FROM THE FONDAZIONE ITALIANA LINFOMI (FIL) FOLL12 TRIAL

Introduction: In follicular lymphoma (FL), FDG-PET is routinely employed to evaluate response at the end of induction (EoI), while minimal residual disease (MRD) analysis is highly reliable in predicting relapse over the entire disease history. Nonetheless, scant data are available describing the integration of these powerful prognostic tools for response evaluation in FL. This issue was investigated in the phase III FIL “FOLL12” trial [Luminari, JCO 2022]. Methods: Only patients with an available molecular marker for MRD and centrally reviewed EoI PET were included. MRD was systematically assessed by RQ-PCR with BCL2::IGH consensus primers in peripheral blood (PB) and bone marrow (BM), as previously reported with inferior follow-up [Ladetto ASH 2021]. PET scans were classified according to Deauville score (DS) criteria: DS1–3 was negative and DS4–5 positive. Results: Overall, 394 patients out of 780 were included (51%), as expected based on the MRD marker availability. These patients had a slightly better outcome than excluded cases (CR rate 82% vs. 73%, p = 0.003, 5y PFS 66% vs. 61%, p = 0.046). At EoI, both BM MRD and PET positivity were independent predictors of poor PFS in multivariate analysis (HR 1.66, CI: 1.01–2.71 and HR 2.03, CI: 1.30–3.18). The 68 discordant cases (i.e., MRD+/PET− or MRD−/PET+) showed an intermediate outcome (PFS HR 1.93, CI: 1.28–2.92) between the 314 double negative (reference) and the 10 double positive ones (HR 6.22, CI: 2.99–12.9), Figure 1A. Moreover, both EoI PET+ and the persistence/reappearance of MRD positivity in BM during the 18 months of rituximab maintenance/observation were independently associated to a higher risk of POD24 (HR 5.61, CI: 2.59–12.1, and HR 2.37, CI: 1.15–4.84, respectively). On the other hand, MRD kinetic analysis by non-invasive PB sampling during the rituximab maintenance/observation period substantially updated the risk status over PET results. Indeed, the marginal PFS HR for an MRD+ after an EoI PET negative result increased from 1.36 (CI: 0.47–3.96) at 6 months, to 2.09 (CI: 1.01–4.18) at 12 months, up to 3.60 (CI: 2.30–5.64) at 24 months (Figure 1B, blue curve). This means that patients scoring PET- at EoI but showing a persistent or reappearing MRD+ signal during the following 24 months had a dismal PFS, superimposable to that of PET+ patients (Figure 1C). Accordingly, in landmark multivariate analysis PB MRD+ either at 12 or 24 months predicted adverse PFS (HR 3.79, CI: 1.60–8.96 and HR 5.21, CI: 1.88–14.4, respectively), independently from EoI PET result. Keywords: indolent non-Hodgkin lymphoma, minimal residual disease, PET-CT Conflicts of interests pertinent to the abstract S. Ferrero Consultant or advisory role: Janssen, EUSA Pharma, Abbvie, Incyte, Clinigen, Sandoz, Italfarmaco Honoraria: Janssen, EUSA Pharma, Servier, Gentili Research funding: Morphosys, Janssen, Gilead, Beigene Educational grants: Takeda, Janssen, EUSA Pharma

Liquid biopsies and minimal residual disease in lymphoid malignancies.

Minimal residual disease (MRD) assessment using peripheral blood instead of bone marrow aspirate/biopsy specimen or the biopsy of the cancerous infiltrated by lymphoid malignancies is an emerging technique with enormous interest of research and technological innovation at the current time. In some lymphoid malignancies (particularly ALL), Studies have shown that MRD monitoring of the peripheral blood may be an adequate alternative to frequent BM aspirations. However, additional studies investigating the biology of liquid biopsies in ALL and its potential as an MRD marker in larger patient cohorts in treatment protocols are warranted. Despite the promising data, there are still limitations in liquid biopsies in lymphoid malignancies, such as standardization of the sample collection and processing, determination of timing and duration for liquid biopsy analysis, and definition of the biological characteristics and specificity of the techniques evaluated such as flow cytometry, molecular techniques, and next generation sequencies. The use of liquid biopsy for detection of minimal residual disease in T-cell lymphoma is still experimental but it has made significant progress in multiple myeloma for example. Recent attempt to use artificial intelligence may help simplify the algorithm for testing and may help avoid inter-observer variation and operator dependency in these highly technically demanding testing process.

Urine Protein Immunofixation Electrophoresis: Free Light Chain Urine Immunofixation Electrophoresis Is More Sensitive than Conventional Assays for Detecting Monoclonal Light Chains and Could Serve as a Marker of Minimal Residual Disease.

Immunoglobulin monoclonal light chains (MLCs) in serum and urine are markers for monoclonal gammopathy and could serve as markers of minimal residual disease (MRD) in multiple myeloma (MM). Excretion of MLCs in urine is known to result in renal damage and shorter survival in patients with LC-predominantMM. Retrospective review of urine immunofixation in 1738 specimens at 3 medical centers was conducted to assess the utility ofurinalysis for diagnosis and monitoring of monoclonal gammopathy. We tested 228 stored urine specimens via the modified urine immunofixation method, using antisera to assay free LCs (FLCs). Our review of urine immunofixation results and medical records validated the theory that the only meaningful value-added finding was detection of monoclonal free light chains. Examination of 228 urine specimens using our novel method revealed 18.4% additional positive results. The rate of incremental findings for lambda LCs was nearly 3-fold higher than for kappa LCs. The new method of urine immunofixation is significantly more sensitive and more efficient than the conventional method for detecting MLCs in urine. The new assay appears to be sensitive enough to prove that MLCs serve as a marker of MRD in MM.

Circulating tumour DNA in the evolving treatment landscape of locally advanced rectal cancer: where does it fit in?

The management of locally advanced rectal cancer (LARC) requires multimodality treatment, typically with neoadjuvant chemoradiotherapy (CRT) followed by total mesorectal excision. However, the treatment landscape is rapidly evolving with total neoadjuvant therapy and non-operative management for selected patients emerging as other novel treatment approaches. With so many treatment options, there is a need for biomarkers to direct a more personalised treatment strategy for patients with LARC. In this review, we summarise the available data regarding the use of circulating tumour DNA (ctDNA) in patients with LARC, as both a marker of treatment response to neoadjuvant therapy and as a marker of minimal residual disease (MRD) after patients have completed definitive local treatment. To date, the ability of ctDNA status to predict for pathologic complete response at any timepoint during multimodality treatment has been variably reported. The most consistent finding across available studies is the ability of ctDNA to detect MRD after CRT and surgery, the presence of which confers a significantly poor prognosis, with increased risk of cancer recurrence and worse overall survival. It is yet to be determined if providing additional therapies to patients with MRD improves outcomes. The available studies assessing the potential utility of ctDNA in LARC are limited by significant heterogeneity in the choice of ctDNA assay, timepoint at which ctDNA was collected, treatment that patients received and length of follow-up, leading to uncertainties about how to implement it into daily clinical practice. As the treatment landscape evolves, larger randomised trials assessing the role of ctDNA in LARC are needed.

Prognostic impact of postoperative circulating tumor DNA as a molecular minimal residual disease marker in patients with pancreatic cancer undergoing surgical resection.

We aimed to clarify the prognostic impact of postoperative circulating tumor DNA (ctDNA) shortly after pancreatectomy in patients with pancreatic ductal adenocarcinoma (PDAC). Preoperative and paired postoperative blood samples were obtained from 66 patients in patients with PDAC. Cell-free DNA was extracted from the plasma, and KRAS mutations, as a benchmark of ctDNA, were examined using droplet digital PCR. Disease-free survival (DFS) and overall survival (OS) were compared between patients with presence and absence of ctDNA. In univariate analysis, patients with detectable postoperative ctDNA showed worse survival than those with undetectable in both DFS (P=.034) and OS (P=.022). Multivariate analysis also revealed that the presence of postoperative ctDNA was an independent risk factor for recurrence (hazard ratio: 2.677, P=.011). In contrast, preoperative ctDNA detection did not affect long-term outcomes. These trends persisted in 34 patients with resectable PDAC who underwent resection after neoadjuvant chemotherapy. Patients with detectable postoperative ctDNA were more prone to developing hepatic recurrence than those with undetectable postoperative ctDNA (P=.039). Postoperative ctDNA, as a minimal residual marker, can be useful for predicting the risk of recurrence in patients with PDAC even after curative resection.

Prophylactic Donor Lymphocyte Infusion in Patients after Allogeneic Stem Cell Transplantation with Post-Transplant Cyclophosphamide Is Associated with Low Relapse Risk and Excellent Survival in Patients below 65 Years with Acute Myeloid Leukemia and High-Risk Myelodysplasia

BackgroundPost-transplant cyclophosphamide (PTCy) effectively prevents graft-versus-host-disease (GVHD) in allogeneic stem cell transplantation (alloSCT) with matched related and matched unrelated donors resulting in low risk of non-relapse mortality (NRM) (<10%). However, PTCy-alloSCT is associated with high relapse rates of approximately 20-30% after 1 year and 30-40% after 2 years (Gooptu et al. Blood 2021, Ruggeri et al. J Hemat Oncol 2018). Prophylactic donor lymphocyte infusion (DLI) may reduce relapse, although it may increase the risk of GVHD. AimTo provide evidence for reduction of relapse and improvement of survival by combining allogeneic SCT with PTCy and subsequent standardized prophylactic donor lymphocyte infusions (DLI) in a cohort of patients with acute myeloid leukemia (AML) and high risk myelodysplasia (MDS). Patients and MethodsSixty-five patients with AML (n=50) or high risk MDS (n=15) received myeloablative (n=15) or reduced-intensity (n=50) alloSCT with a matched related (n=8) or 9/10 (n=19) or 10/10 (n=38) matched unrelated donor. Based on modified ELN criteria, 10 AML patients were intermediate risk and 40 poor risk. GVHD prophylaxis consisted of PTCy, mycophenolate until day +30, and tacrolimus tapered from +70 (related or 10/10 unrelated) or day +90 (9/10 unrelated). All patients were planned to receive DLI at 6 months after transplantation at a dose of 3 x 10^6 CD3+ cells/kg in related and 1.5 x 10^6 CD3+ cells/kg in unrelated alloSCT. In poor risk patients (poor risk AML and high risk MDS) additional low dose DLI (0.3 or 0.15 x 10^6 CD3+ cells/kg in related and unrelated alloSCT, respectively) was planned one month after tacrolimus tapering (i.e. 3-4 months after alloSCT). In poor risk patients with a sensitive molecular or immunophenotypical minimal residual disease (MRD) marker (<0.1%) low dose DLI was not administered but MRD monitored; only in case of MRD positivity at 2 or 4 months additional low dose DLI was given. ResultsOf the 65 patients, 49 received standard dose DLI at 6 months after alloSCT (75%), either as a first dose at 6 months (40%), or as a second DLI after a low dose DLI at 3-4 months after alloSCT (35%). Reasons for not receiving standard dose DLI were NRM (5%), GVHD (9%), relapse (9%) and donor-related (2%). Of the 22 poor risk patients with a sensitive MRD marker, 15 received the first DLI at 6 months; 6 received both low dose DLI and 6 month DLI because of MRD positivity at 2 or 4 months after alloSCT; one patient did not receive prophylactic DLI because of systemic relapse at day 105. With a median follow up of 400 days (range 183-814), 12 patients developed systemic relapse. Most relapses (67%) occurred very early after transplant (median 110 days, range 63 to 165 days), before low dose DLI could be effectively given. These early relapses occurred significantly more in patients >= 65 years (88%) than <65 years (p=0.01 Fisher's exact). Overall survival at 1 year was 95% (95% CI 89-100%) in patients <65 (n=42), significantly higher than the 68% (95% CI 49-95%) in patients >=65 (n=23) (p=0.01). Relapse rate at 1 year was 5% (95% CI 1-15%) in patients <65, significantly lower than the 37% (95% CI 25-58%) in patients >=65 (p=0.001). Grade 2-4 acute and chronic GVHD incidence at 1 year was low (6% (95% CI 2-14%) and 11% (2.5-25%), respectively). Survival at 1 year was similar (p=0.35) in poor risk patients without MRD marker (80%, 95% CI 67-96%), poor risk with MRD marker (95%, 95% CI 87-100%) and intermediate risk (86%, 95% CI 63-100%). For these groups relapse risk was also similar: 18% (95% CI 7-33%), 16% (95% CI 3-36%) and 11% (95% CI 5-41%), respectively. ConclusionsThis study is the first to demonstrate feasibility of prophylactic DLI after PTCy-alloSCT with a low associated risk of GVHD. The observed low incidence of relapse at 1 year after transplantation (5%) of patients aged <65 years appears to be substantially less than the reported risk without pre-planned DLI (20-30%). Although the value of early DLI administration (around 3-4 months) is still unclear, standard dose DLI administration at 6 months appears feasible in intermediate risk patients and poor risk patients with sensitive MRD markers. Early relapses after alloSCT, at a time point before early DLI could be effective, caused the inferior survival of patients aged >= 65 years. The results of this study can serve as a reference for designing future prophylactic cellular interventions in the PTCy-alloSCT setting. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

CD58 Could be As a Leukemic Marker in Relapsed/Refractory B-ALL Patients after Multiline Therapies

It has been verified that CD58, a member of the immunoglobulin superfamily, is expressed in B-cell lymphoblastic leukemia (B-ALL) at a higher level and is a marker of minimal residual disease in B-ALL. A study reported that CD58 was expressed in 99.4% of B-ALL cases at diagnosis. Nevertheless, little is known regarding CD58 expression in relapsed/refractory (r/r) B-ALL patients having received multiline therapies especially chimeric antigen receptor (CAR) T-cells. Although CD19-specific CAR-T therapy resulted in high complete remission (CR) rates, some patients relapsed after CAR-T and part of them showed CD19 loss, under this circumstance, additional B-cell antigen specifically expressed on the B-ALL cells could help us to identify the leukemia cells. In this study, we investigated the CD58 expression in r/r B-ALL patients before CAR-T, and the influence of CAR-T cells on CD58 antigen of lymphoblasts in patients failed or relapsed after CAR-T. CD58 on leukemic cells of r/r B-ALL patients has been routinely detected by flow cytometry in our center. Data of r/r B-ALL patients were collected from May 2019 through May 31, 2022. Samples were detected on an 8-color BD FACSCanto II flow cytometer, and analyzed by FACS Diva software from BD or Kaluza software from Beckman Coulter. Antigen full expression, partial expression and no expression were defined as >80%, 20%-80% and <20% of gated cells displaying the antigen of interest, respectively. A total of 274 r/r B-ALL patients planned to be administered CD19 CAR-T cells were assayed CD58 antigen before CAR-T, all of them had received multiline chemotherapies, 61 (22.3%) underwent allogeneic hematopoietic cell transplant (allo-HCT) and 6 cases had a history of antibody (CD20, CD19 or CD22) treatment. The median age was 12 years (range, 1-75 years), including 97 adults and 177 children younger than 18 years. Among all 274 patients, 228 (83.2%, 228/274) showed CD58 expression, consisting of 180 (78.9%, 180/228) cases with full expression and 48 (21.1%, 48/228) with partial expression; whereas 46 (16.8%, 46/274) patients were CD58-negative, demonstrating that CD58 was expressed on the majority (83.3%) of r/r B-ALL patients after treated by multiple intensive chemo drugs and even allo-HCT. Then, we observed the influence of CAR-T cells on the expression of CD58 antigen of lymphoblasts. We followed-up 57 patients who had CD58 expression before CD19 CAR-T but failed or relapsed after CAR-T. Of them, 45.6% (26/57) cases received the further treatment as a consolidation following CD19 CAR-T (CD22 CAR-T, 14; allo-HCT, 12). Among 57 patients, 10 were CD58-negative while 47 cases still showed CD58-positive, the frequency of CD58 expression was 82.5% (47/57) after treated by CAR-T cells and allo-HCT. When we excluded the 26 patients who received following CD22 CAR-T or allo-HCT, and only considered the 31 patients exposed to one-time CD19 CAR-T cells, the CD58 expression rate was 87.1% (27/31). It was reported that CD58 loss in tumor cells induced resistance to CAR-T therapy in cell lines and mouse models. We therefore analyzed the influence of CD58 expression on the treatment response of CAR-T cells. Among 274 r/r B-ALL patients received CD19 CAR-T, 6 were unable to evaluate the treatment response due to death or hospital discharge within 15 days after cell infusion, the remaining 268 patients had evaluation data including 223 cases with CD58 expression (full or partial expression) and 45 without CD58 expression. After CD19 CAR-T therapy, the CR (including MRD+CR) rate in patients with CD58 expression was 91.9% (205/223) while it was 95.6% (43/45) in patients without CD58 expression, indicating that CD58 negativity was irrelevant to CAR-T treatment failure in r/r B-ALL. In conclusion, CD58 antigen was expressed in the majority (83.2%) of r/r B-ALL cells without exposing to CAR-T treatment, and still expressed on the lymphoblasts of most patients (>80%) failed or relapsed after CAR-T, therefore, it could be as a leukemic marker for identifying tumor cells in r/r B-ALL patients who have received multiline therapies including allo-HCT and CAR-T. Additionally, CD58-negative patients achieved a similar high CR rate as CD58-positive patients did after CD19 CAR-T.

Time-Sequential Change of the Predictive Power of Peripheral Blood WT1mRNA Levels after Allogeneic Stem Cell Transplantation for Myeloid Neoplasm Relapse and Development of a Dynamic Relapse Prediction Model

Introduction:Wilms tumor 1 messenger RNA (WT1mRNA) is detected in patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS), and is reportedly a useful minimal residual disease (MRD) marker. Allogeneic stem cell transplantation (allo-SCT) is the established cellular immunotherapy with a graft-versus-leukemia (GVL) effect and eliminates residual leukemia cells via the alloimmune response. The GVL effect is generally considered to occur after immune reconstruction. Therefore, the predictive power of WT1mRNA as an MRD marker for relapse may differ between the early and late periods after allo-SCT because the GVL effect is not expected in the early period without enough immune reconstruction. However, no studies have reported whether the predictive power of WT1mRNA for relapse changes in a time-sequential manner after allo-SCT. Hence, we investigated the relationship between WT1mRNA as a time-dependent variable and days after allo-SCT for AML/MDS relapse using a dynamic landmarking approach, and developed a dynamic relapse prediction model using the WT1mRNA level and measurement timepoint after allo-SCT. Methods: We retrospectively analyzed AML and MDS patients who received allo-SCT at Osaka Metropolitan University Hospital, and whose WT1mRNA levels were measured at least once from January 2008 to April 2021. WT1mRNA levels in peripheral blood were measured using an "Otsuka" WT1mRNA assay kit (Otsuka Pharmaceutical Co., Ltd.). Relapse was defined as hematological relapse or salvage chemotherapy intervention and/or donor lymphocyte infusion. To evaluate time-sequential changes in the predictive power of WT1mRNA for relapse and to develop a dynamic relapse prediction model, dynamic landmarking analysis was performed, which was reported by Hans C. van Houwelingen et al. The probabilities of the 1 year cumulative incidence of relapse (CIR) post-landmark timepoint were estimated with the Kaplan-Meier method. The Cox proportional hazard regression model was used to calculate the time-sequential hazard ratio for 1 year CIR not only post-landmark but also post-any timepoints after allo-SCT, and to develop a dynamic relapse prediction model. p < 0.05 was considered significant. Results: We analyzed 265 patients (AML: 197 and MDS: 68) and 2,539 WT1mRNA tests. Median patient age was 46 years, 55% of patients achieved complete remission at the time of allo-SCT, and the 2 year relapse-free survival rate was 46%. First, patients were divided into two groups, namely, those with WT1mRNA levels higher and lower than 200 copies/μg RNA, which is a previously reported cutoff value. Figure 1 shows a summary plot of the 1 year CIR of the two groups post-landmark timepoint. The 1 year CIR decreased over time after allo-SCT in the lower WT1mRNA group but not in the higher WT1mRNA group. The difference in 1 year CIR between the two groups increased over time after allo-SCT. Moreover, the hazard ratio of log WT1mRNA levels for 1 year CIR post landmark timepoints after allo SCT increased over time, and there was a significant interaction between WT1mRNA levels and landmark time (p < 0.001). These findings indicate that the predictive power of WT1mRNA levels for relapse increases over time after allo-SCT. Second, we developed a dynamic relapse prediction model that can predict the 1 year CIR post-any timepoint after allo-SCT based on individual post-transplant WT1mRNA levels and measurement timepoints. Figure 2 shows a time series of predictive values for 1 year relapse post-any timepoints derived from this prediction model, which assumed that WT1mRNA levels remain constant over time after allo-SCT. Conclusions: We demonstrated that the predictive power of peripheral blood WT1mRNA for relapse increases after allo-SCT over time. This finding implies that MRD in the early phase after allo-SCT is more likely to disappear owing to the GVL effect after immune reconstruction compared to one in the late phase. Transplant clinicians should pay attention to the difference in predictive power for relapse depending on the period when they observe elevated WT1mRNA levels. Dynamic landmark analysis also enabled prediction of continuous 1 year CIR post-any timepoint after allo-SCT by inputting the individual WT1mRNA level and measurement timepoint. Dynamic relapse prediction will offer useful information for clinical decision making. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Circulating Tumor DNA as a Marker of Minimal Residual Disease After Radical Resection of Colorectal Liver Metastases.

Prognostic tools to estimate the risk of relapse for patients with liver-limited metastatic colorectal cancer (LL-mCRC) undergoing resection with curative intent are needed. Circulating tumor DNA (ctDNA) as a surrogate of postsurgical minimal residual disease is a promising marker in localized CRC. We explored the role of postoperative ctDNA as a marker of minimal residual disease in patients with radically resected LL-mCRC. Seventy-six patients with LL-mCRC were retrospectively included. DNA from tumor tissue was sequenced, and one somatic mutation was then assessed by digital droplet polymerase chain reaction in plasma samples collected after surgery to identify the persistence of ctDNA. Relapse-free survival and postresection overall survival were compared between patients with positive vs negative postoperative ctDNA. ctDNA was found in 39 (51%) of 76 patients with LL-mCRC. At a median follow-up of 77 months, 33 of 39 ctDNA-positive patients and 20 of 37 ctDNA-negative patients experienced disease relapse (P = .008). ctDNA-positive patients reported significantly shorter RFS than ctDNA-negative ones (median RFS 12.7 v 27.4 months hazard ratio, 2.09, P = .008). In the multivariable model including other prognostic covariates, this association was still significant (P = .046) and a trend toward shorter overall survival among ctDNA-positive patients was reported (hazard ratio, 1.65, P = .183). The detection of postsurgical ctDNA is an independent negative prognostic marker and identifies patients at high risk of relapse after liver metastases resection.

The Clinical Significance of Light Chain Predominant Multiple Myeloma

Abstract Multiple Myeloma (MM) is a neoplastic clonal proliferation of terminally differentiated B- lymphocytes. It accounts for 1.6% of all cancers and about 10-15% of all hematological malignancies in the United States. Monoclonal immunoglobulins provide an indication of tumor burden in patients with plasma cell neoplasm. Recent development has shown that a proportion of intact immunoglobulin producing multiple myeloma secrete much higher amounts of free light chains than usual; i.e. light chain predominant multiple myeloma (LCPMM). About 18% of multiple myeloma secreting intact immunoglobulins secrete higher amount of free light compared to other intact immunoglobulin secreting myelomas and are associated with significantly poorer clinical outcomes. In this study, a retrospective investigation comparing the laboratory and clinical findings in conventional MM and LCPMM was undertaken. Three hundred and sixteen patients with conventional MM and seventy-one patients with LCPMM were studied. The results were remarkable for marked increase in death rate in the LCPMM group compared to the conventional MM group, 35.2% and 13.6% respectively; P-value .00001. The occurrence of dialysis was also markedly increased in the LCPMM group compared with the conventional MM; 18.3% vs. 3.5% P-value .00001. A markedly lower eGFR was observed in patients with LCPMM group compared to patients with conventional MM with an average value of 46.3 versus 69.4 ml respectively P value .0000003. In conclusion, light chain predominant plasma cell myeloma confers a significant disease burden, poor clinical outcomes and shorter survival. Further prospective trial will be helpful in understanding the pathogenesis, the course, and management of the disease. An assay for serum free monoclonal light chains has been described and could serve as a marker for minimal residual disease in such patients. We aim to further study the clinical impact of light chain predominant plasma cell myeloma (LCPMM) to increase the current knowledge of the disease and investigate therapeutic interventions.

Significance of Gene Diagnosis in Acute Myeloid Leukemia with the Emergence of New Molecular Target Drug Treatment

Acute myeloid leukemia (AML) is a heterogeneous hematopoietic malignancy accompanied by impaired differentiation and autonomous proliferation of hematopoietic stem cells. Standard induction therapy results in first complete remission among 70% of patients with AML; however, approximately half of these patients relapse and become refractory. Allogeneic hematopoietic cell transplantation is a useful treatment for relapsed and refractory cases. However, transplantation-related mortality is approximately 20%, which is not a low value, and quality of life after transplantation decreases. Therefore, there is a need to stratify the prognosis of each patient and implement this treatment appropriately. Owing to recent advances in genome analysis technology, many gene mutations involved in onset and recurrence of AML have been discovered. These abnormalities and mutations not only have clinical application as prognostic factors and minimal residual disease markers, but they may also contribute to novel molecular targeted drug development. Many new drugs such as first-generation FMS-like tyrosine kinase 3 (FLT3), isocitrate dehydrogenase 1 and 2 (IDH1/2), and B cell lymphoma 2 (BCL2) inhibitors have been developed in the West. In addition, the second-generation FLT3 inhibitors gilteritinib and quizartinib were developed in Japan, and treatment outcomes for patients with AML have improved. However, there is still a large disparity in drug availability between the West, and Japan. As a result, treatment guidelines in the West cannot be applied in the clinical setting in Japan. In this study, we assessed the molecular target drug treatment by gene diagnosis for treatment of AML patients.

Circulating Tumor DNA as a Marker of Minimal Residual Disease.

Circulating Tumor DNA as a Marker of Minimal Residual Disease.