Evidence Of Minimal Residual Disease Research Articles

NSABP C-14/Exact Sciences 16-002: CORRECT-MRD II—Second colorectal cancer clinical validation study to predict recurrence using a circulating tumor DNA assay to detect minimal residual disease.

TPS241 Background: Detectable circulating tumor DNA (ctDNA) after resection of early-stage solid tumors has been associated with very high risk of recurrence, suggesting ctDNA is evidence of minimal residual disease (MRD). Several studies are ongoing to investigate the role of ctDNA in the optimal management of patients (pts) with colorectal cancer using different assay technologies. Methods: This is a prospective, observational, multicenter study in the United States and Canada of 750 pts who have undergone complete surgical resection for stage II or III colorectal cancer, who have FFPE tissue available from the primary resection sufficient for a novel bespoke MRD assay and are willing to provide serial whole blood specimens for ctDNA analysis. Participants are asked to provide study specimens after definitive surgical resection, pre-recurrence follow-up, and clinical recurrence (if applicable). Recently amended eligibility criteria include inclusion of rectal cancer pts who have completed neo-adjuvant therapy and surgical resection, as well as enrollment of all stage II and III pts regardless of microsatellite stability status. The Oncotype Colon Recurrence Score® will be assessed on all pts from their surgical specimen, if criteria are met for this testing. ctDNA will be analyzed with an NGS-based tumor-informed MRD assay that identifies somatic genomic alterations from DNA derived from the patient’s tumor tissue, subtracts germline variants, and detects a selected subset of tumor-specific (bespoke) ctDNA in their blood. All primary tumor specimens will undergo full exome and transcriptome sequencing using the Onco ExTra assay. If there is evidence of disease recurrence, the metastatic tissue will also undergo Onco ExTra testing and results will be shared with participants. The primary objective is to validate the association of post-definitive therapy and pre-recurrence follow-up ctDNA positivity with recurrence-free interval (RFI). Further objectives are to assess the: sensitivity and specificity of ctDNA positivity for subsequent clinical recurrence; contribution of post-surgery baseline, post-adjuvant therapy, and pre-recurrence follow-up ctDNA results on RFI; time from positive ctDNA to clinical recurrence in participants who had a positive ctDNA result; and to compare the Oncotype Colon Recurrence Score estimate of 3-year recurrence risk with the observed 3-year recurrence rate. The primary analysis will use a Cox proportional hazards regression applied to the RFI with ctDNA result (positive or negative) measured after surgery, or end of adjuvant therapy if received, and serially after that as a single, time-dependent covariate. Enrollment has currently surpassed the 50% target. Protocol#: NSABP C-14 / Exact Sciences 16-002 NCT#: 05210283. Support: NSABP Foundation, ExactSciences. Clinical trial information: NCT05210283.

CtDNA guided adjuvant chemotherapy versus standard of care adjuvant chemotherapy after curative surgery in patients with high risk stage II or stage III colorectal cancer: a multi-centre, prospective, randomised control trial (TRACC Part C)

BackgroundCirculating tumour DNA (ctDNA) to detect minimal residual disease (MRD) is emerging as a biomarker to predict recurrence in patients with curatively treated early stage colorectal cancer (CRC). ctDNA risk stratifies patients to guide adjuvant treatment decisions. We are conducting the UK’s first multi-centre, prospective, randomised study to determine whether a de-escalation strategy using ctDNA to guide adjuvant chemotherapy (ACT) decisions is non-inferior to standard of care (SOC) chemotherapy, as measured by 3-year disease free survival (DFS) in patients with resected CRC with no evidence of MRD (ctDNA negative post-operatively). In doing so we may be able to spare patients unnecessary chemotherapy and associated toxicity and achieve significant cost savings for the National Health Service (NHS).MethodsWe are recruiting patients with fully resected high risk stage II and stage III CRC who are being considered for ACT into the study which uses results from a plasma-only ctDNA assay to guide treatment decisions. Eligible patients are randomised 1:1 to receive ctDNA-guided chemotherapy versus SOC chemotherapy. The primary endpoint is the difference in DFS at 3 years between the trial arms. Secondary endpoints include the proportion of patients in the ctDNA-guided arm who are ctDNA negative post-operatively and receive de-escalated ACT compared to the standard arm, the difference in overall survival (OS), neurotoxicity and quality of life between the arms, and the cost-effectiveness of ctDNA-guided therapy compared to SOC treatment. We hypothesise that using a ctDNA-guided approach to ACT decisions is non-inferior to SOC. Target accrual is 1621 patients over 4 years, which will provide a power of 80% with an alpha of 0.1 to demonstrate non-inferiority with a margin of 1.25 in survival of the ctDNA-guided approach compared to SOC. We anticipate approximately 50 UK centres will participate. The study opened with the Guardant Reveal plasma-only ctDNA assay in August 2022.DiscussionThe trial will determine whether ctDNA guided ACT is non-inferior to SOC ACT in patients with fully resected high risk stage II and stage III resected CRC, with the potential to significantly reduce unnecessary ACT and the toxicity associated with it.Trial registrationNCT04050345.

Safety and efficacy of a modified busulfan/cyclophosphamide conditioning regimen incorporating cladribine for autologous hematopoietic stem cell transplantation in acute myeloid leukemia.

This is a small phase I study examining the safety and efficacy of a cladribine (CLAD)-containing conditioning regimen prior to autologous hematopoietic stem cell transplantion (auto-HSCT) for patients with acute myeloid leukemia (AML). All patients, aged 15-54years (median 32years), had favorable/intermediate risk AML (n = 20) or acute promyelocytic leukemia (APL; n = 2) and no evidence of minimal residual disease (MRD) prior to transplantation. Fourteen of the 22 patients received the conditioning regimen as follows: busulfan (Bu) + cyclophosphamide (Cy) + CLAD + cytarabine (Ara-c) or idarubicin. The conditioning regimen of 8 patients was without Cy nor idarubicin to reducing adverse cardiac reaction: the regimen of Bu + CLAD+ Ara-c for 6 patients; and the regimen of Bu + melphalan + CLAD + Ara-c for the other 2 patients. All 22 AML patients received peripheral blood stem cell transplantation. The number of infused mononuclear cells and CD34+ cells was 10.00 (2.88-20.97) × 108/kg and 1.89 (1.52-10.44) × 106/kg, respectively. Hematopoietic reconstitution was achieved in all patients, with a median time of 13 (10-34) days for neutrophils and 28 (14-113) days for platelets. Two patients suffered from pulmonary infection, 4 patients suffered from septicemia during the neutropenic stage, and the others suffered from infection or gastrointestinal reaction without exceeding grade 3 after conditioning, which were all alleviated by anti-infection and other supportive treatment. None of the patients died of transplantation-related complications. At a median follow-up of 29.5 (ranging from 4.0 to 60.0) months, three patients relapsed after auto-HSCT at a median time of 6 (ranging from 0.5 to 10.0) months. One patient died due to relapse at 18months after auto-HSCT. The remaining 21 patients were all alive, including 19 patients with negative MRD. The other 2 patients achieved negative MRD after allogeneic HSCT or chemotherapy. The estimated 2-year survival, relapse, and Leukemia-free survival rates were 94.1 ± 5.7%, 14.7 ± 7.9% and 85.3 ± 7.9%, respectively. A CLAD-combination conditioning regimen is efficient and safe for auto-HSCT, indicating an effective approach for AML treatment.

NSABP C-14: CORRECT-MRD II—Second colorectal cancer clinical validation study to predict recurrence using a circulating tumor DNA assay to detect minimal residual disease.

TPS284 Background: Detectable ctDNA after resection of early-stage solid tumors has been associated with very high risk of recurrence, suggesting ctDNA is evidence of minimal residual disease (MRD). Several studies are ongoing to investigate the role of ctDNA in the optimal management of pts with colorectal cancer using different assay technologies. Methods: This is a prospective, observational, multicenter study in the United States and Canada of 750 patients who have undergone complete surgical resection for stage II or III colorectal cancer, who have FFPE tissue available from the primary resection sufficient for a novel bespoke MRD assay and are willing to provide serial whole blood specimens for ctDNA analysis. Participants are asked to provide study specimens after definitive surgical resection, pre-recurrence follow-up, and clinical recurrence (if applicable). Recently amended eligibility criteria include inclusion of rectal cancer patients who have completed neo-adjuvant therapy and surgical resection, as well as enrollment of all stage II and III patients regardless of microsatellite stability status. The Oncotype Colon Recurrence Score will be assessed on all patients from their surgical specimen if criteria are met for this testing. ctDNA will be analyzed with an NGS-based tumor-informed MRD assay that identifies somatic genomic alterations from DNA derived from the patient’s tumor tissue, subtracts germline variants, and detects a selected subset of tumor-specific (bespoke) ctDNA in their blood. All primary tumor specimens will undergo full exome and transcriptome sequencing using the Oncomap ExTra assay. If there is evidence of disease recurrence, the metastatic tissue will also undergo Oncomap ExTra testing, which will be shared with participants. The primary objective is to validate the association of post-definitive therapy and pre-recurrence follow-up ctDNA positivity with recurrence-free interval (RFI). Further objectives are to assess the: sensitivity and specificity of ctDNA positivity for subsequent clinical recurrence; contribution of post-surgery baseline, post-adjuvant therapy, and pre-recurrence follow-up ctDNA results on RFI; time from positive ctDNA to clinical recurrence in participants who had a positive ctDNA result; and compare the Oncotype Colon Recurrence Score estimate of 3-year recurrence risk with the observed 3-year recurrence rate. The primary analysis will use a Cox proportional hazards regression applied to the RFI with ctDNA result (positive or negative) measured at post-surgical baseline (or end of adjuvant therapy if used) and serially after that as a single, time-dependent covariate. Protocol#: NSABP C-14 / ES 16-002. Support: NSABP Foundation, ExactSciences Clinical trial information: 05210283 .

Pediatric Acute Myeloid Leukemia with Co-Occurring BCR::ABL and CBFA2T3::GLIS2 Dual Fusion with Deep Response to FOLR1-Targeting Antibody Drug Conjugate Stro-002 and Tyrosine Kinase Inhibitor

Introduction: Pediatric acute myeloid leukemia (AML) is characterized by significant cytogenetic and molecular heterogeneity. AML with CBFA2T3::GLIS2 fusion (CBF::GLIS) involving infants and very young children is a rare but uniformly refractory disease with high mortality. Laboratory data demonstrates that this fusion is sufficient for leukemic transformation without recurrent secondary variants. These studies have also linked CBF::GLIS fusion to induction of folate receptor alpha (FOLR1) expression in transformed cells (Meshinchi et al, 2020). Incidence of Ph+ de novo (BCR::ABL minor breakpoint, p190 hybrid) disease ranges from 0.5-3% of all AML cases across the age spectrum (Soupir et al, 2017). We, for the first-time, report dual existence of both fusions in a child with AML and discuss novel interventions including combination of a BCR::ABL tyrosine kinase inhibitor (TKI) and antifolate therapy consisting of methotrexate (MTX) and FLOR1-specific antibody-drug conjugate (ADC) STRO-002 (Sutro Biopharma) based on available data regarding its in-vitro preclinical activity in CBF::GLIS disease (Meshinchi et al, ASH abstract, 2021). Case: A 2-year-old girl of Vietnamese descent received a bone marrow examination for bicytopenia and macrocytosis. Flow cytometry confirmed the poor prognostic RAM immunophenotype AML (CD45 dim, HLA-DR negative, CD38 dim, CD56 bright) and FOLR1 expression. Marrow aspirate smears showed numerous hypercellular spicules and blasts with scant cytoplasm and some blebbing but no granules or Auer rods. Marrow exhibited 100% cellularity. CNS was not involved. Review of diagnostic genomic profile mid induction showed a BCR::ABL minor breakpoint fusion as well as CBF::GLIS fusion. The child began AML induction on COG AAML1831 trial randomized to the experimental arm with CPX-351 and gemtuzumab ozogamicin (GO). Due to lack of response to AML therapy at end of Induction (EOI) I, she was taken off protocol and started on modified ALL regimen for Induction II consisting of cytarabine, low dose weekly methotrexate and bimonthly peg-asparaginase with addition of an oral TKI, dasatinib. EOI II marrow examination showed improvement with reduction in blasts (11%) and detectable BCR::ABL but undetectable CBF::GLIS by Q-RT-PCR. Induction III included low dose weekly methotrexate, cytarabine, GO, and dasatinib. EOI III marrow examination showed complete morphologic remission (CR) with no evidence of minimal residual disease (MRD) or fusion transcripts of either clone. Due to availability of a matched sibling donor, the child underwent hematopoietic stem cell transplantation (HSCT); however, was noted to have 3% blasts on day +83 marrow examination despite previously commencing dasatinib post-transplant prophylaxis. BCR::ABL and CBF::GLIS transcripts were again detectable at low levels. Dasatinib dosage was maximized and weekly MTX and bimonthly peg-asparaginase were reintroduced, but the marrow disease burden increased significantly to 78% blasts positive only for CBF::GLIS fusion. Given availability of FOLR1 directed ADC on single-patient compassionate use basis, patient received single agent STRO-001 on a bi-monthly basis. With rising ANC, evaluation of marrow after 2nd STRO-002 dose remarkably showed CR with MRD negative remission by flow and molecular studies. This time remission was consolidated with monthly STRO-002 and donor lymphocyte infusions (DLI). Currently, patient remains in MRD-negative remission after the 3 of 4 planned STRO-002/DLI maintenance. Patient has maintained a 100% Lansky play score and tolerated the regimen with only prolonged initial cytopenia requiring supportive care as a toxicity. Plan is to continue TKI and monthly STRO-002 as maintenance therapy. Conclusion: Complete genomic sequencing has revolutionized pediatric AML categorization. It is quite conceivable that (BCR::ABL, CBF::GLIS) dual fusion disease will become a more common finding in future. Although CBF::GLIS AML is uniformly refractory to conventional therapy, targeted therapy with FOLR1-directed STRO-002 was highly effective in eliminating CBF::GLIS clone in this patient, and combination with DLI appears to be feasible. Long-term follow-up is required to assess durability of remission. Additional testing of this approach in a larger patient population is needed to determine the role of STRO-002 in this high-risk pediatric AML population. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

MRD Status By Clonoseq ® Is a Poor Predictor of Long-Term Outcomes after Bispecific LV20.19 CAR T-Cell Therapy for Relapsed, Refractory B-Cell NHL

INTRODUCTION: The clonoSEQ® Assay (Adaptive Biotechnologies Corporation, Seattle, USA) uses next-generation sequencing (NGS) based testing to identify disease-specific immunoglobulin sequences and detect evidence of minimal residual disease (MRD). MRD assessments have been shown to predict progression-free survival and overall survival in non-Hodgkin's lymphoma (NHL) patients. We report MRD results via the commercially available clonoSEQ® assay and its association with durability of response in patients treated with bispecific LV20.19 CAR in our phase 1/2 trial (NCT04186520). METHODS: We conducted a phase 1/2 single-center prospective trial (NCT04186520) to evaluate the efficacy and safety of LV20.19 CAR T-cells in patients with NHL at 2.5x106 cells/kg dose. LV20.19 CAR T-cells were manufactured onsite at varying lengths of manufacturing (8 vs 12 days) in the CliniMACS Prodigy device with IL7+15 for cell expansion, with the goal of fresh infusion. The commercial clonoSEQ® assay (by Adaptive Biotechnologies) was used to identify and track tumor clonotypes in responding patients at first assessment performed between Day 28-60. We evaluated associations between MRD status and long-term response post LV20.19 CAR-T cell therapy. Descriptive statistics were utilized for baseline characteristics. Survival analyses were calculated utilizing the Kaplan-Meier method and differences between groups were evaluated via the log-rank test. RESULTS: Among the 32 patients enrolled in the trial, there were 26 responders on Day 28. Of these responders, 23 patients had clonoSEQ® B-cell MRD assay done between Day 28-60 post CAR infusion (Table 1). MRD was performed in 11 patients with Diffuse Large B-cell Lymphoma (DLBCL), 3 patients with Follicular lymphoma (FL), and 9 patients with Mantle Cell Lymphoma (MCL). The median age was 64 years (45-74) and the median lines of prior therapy were 4 (2-11). Of the 23 patients with early MRD status available, the day 28 objective response rate (ORR) was 100% (complete response=57%, n=13 and partial response=43%, n=10). Eighteen patients (78%) were MRD negative at first assessment between days 28-60 and 5 (22%) were MRD positive. Of the 5 patients who were MRD positive, 3 became MRD negative at day 90, 1 patient was non-evaluable due to a non-relapse mortality event, and the remaining patient remains positive but with decreasing MRD status at Day 90. None of the patients with MRD positivity at day 28 have relapsed to date. Among patients who achieved early MRD negativity (n=18), 7 (39%) have relapsed, and all were patients with either DLBCL or FL. No MRD-negative MCL patient has relapsed to date. The duration of response in MRD-negative patients was significantly improved in MCL patients compared to DLBCL/FL patients (Figure 1), p=0.02. Among the 7 MRD negative patients who relapsed, only 1 patient was detected to have disease relapse by MRD positivity, 2 were detected simultaneously on MRD testing and imaging, while 3 were detected by physical exam or imaging findings and did not have MRD testing. Interestingly, 1 patient had a central nervous system relapse while being MRD negative. CONCLUSION: Relapse after CAR T-cell therapy remains a significant clinical challenge. New predictors are indicated to determine which responding patients after CAR-T cell therapy are more likely to relapse. Utilizing the commercially available Adaptive clonoSEQ® assay, early MRD assessment was not predictive of long-term clinical outcomes in DLBCL and FL. MRD positive patients maintained durable remission and converted to MRD negativity, while early MRD negative patients continued to relapse. In contrast, MRD negativity post-CAR-T therapy did appear to predict durable clinical remission in MCL patients. Newer assays, including cell-free technologies, may be more informative for non-MCL histologies. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

NSAB C-14: CORRECT-MRD II—Second colorectal cancer clinical validation study to predict recurrence using a circulating tumor DNA assay to detect minimal residual disease.

TPS3632 Background: Patients (pts) with stage II and III colon cancer (CC) have unique post-operative decisions regarding adjuvant chemotherapy (ACT). There is a subset of stage II pts with defined clinicopathologic features associated with poor prognosis who may benefit from ACT, although more discriminating and objective predictors of benefit are needed. In addition, there may be a subset of Stage III CC pts who could tolerate a de-escalation of ACT or who may require intensification of ACT to improve clinical outcome. Detectable ctDNA after resection of early-stage solid tumors has been associated with very high risk of recurrence, suggesting ctDNA is evidence of minimal residual disease (MRD). Several studies are ongoing to investigate the role of ctDNA in the optimal management of pts with CC using different assay technologies. Methods: This is a prospective, observational, multicenter study in the United States and Canada of 750 pts who have undergone complete surgical resection for stage II or III CC, have FFPE tissue available from the primary resection sufficient for a novel bespoke MRD assay, and are willing to provide serial whole blood specimens for ctDNA analysis. Subjects are asked to provide study specimens at baseline, pre-recurrence follow-up, and clinical recurrence (if applicable) study visits. ctDNA will be analyzed with an NGS-based MRD assay that identifies somatic genetic alterations from DNA derived from the pt’s tumor tissue, subtracts germline variants, and detects a subset of these tumor-specific (bespoke) ctDNA in the pt’s blood. The primary objective is to validate the association of post-definitive therapy and pre-recurrence follow-up ctDNA positivity with recurrence-free interval (RFI). Further objectives are to assess the: sensitivity and specificity of ctDNA positivity for subsequent clinical recurrence; contribution of post-surgery baseline, post-adjuvant therapy, and pre-recurrence follow-up ctDNA results on RFI; time from positive ctDNA to clinical recurrence in subjects who had a positive ctDNA result; and compare the Oncotype Colon Recurrence Score estimate of 3 yr recurrence risk with the observed 3 yr recurrence rate. The primary analysis will use a Cox proportional hazards regression applied to the RFI with ctDNA result (positive or negative) measured at post-surgical baseline (or end of ACT if ACT was used) and serially after that as a single, time-dependent covariate. Protocol: 16-002/NSABP C-14. Support: NSABP Foundation. Clinical trial information: 05210283.

90-yttrium-ibritumomab tiuxetan as first-line treatment for follicular lymphoma: updated efficacy and safety results at an extended median follow-up of 9.6\xa0years

Radioimmunotherapy with 90-yttrium-ibritumomab tiuxetan (90Y-IT) as first-line treatment in patients with follicular lymphoma (FL) demonstrated promising results with a complete remission (CR) rate of 56% and a median progression-free survival (PFS) of 26 months, when initially analyzed after a median follow-up of 30.6 months. The aim of this long-term follow-up was to investigate whether clinical benefits were maintained and new safety signals appeared. Fifty-nine patients, aged ≥ 50 years, with FL grade 1 to 3A in stages II to IV were treated with 90Y-IT as first-line therapy. If CR without evidence of minimal residual disease (MRD), partial response or stable disease was achieved 6 months after treatment, patients were observed without further treatment. Patients with CR but persisting MRD received consolidation therapy with rituximab. The primary endpoint was the clinical response rate. Secondary endpoints were time to progression, safety, and tolerability. After a median follow-up of 9.6 years, median PFS was 3.6 years, and 8-year PFS was 38.3%. Median overall survival (OS) was not reached during the extended follow-up, and 8-year OS amounted to 69.2%. Age 65 years and above or disease progression within 24 months of treatment were significantly associated with shorter OS. An important finding was the lack of new safety signals. In particular, no increase in secondary malignancies or transformation into aggressive lymphoma was observed compared to trials with a similar follow-up. In summary, 90Y-IT as first-line treatment demonstrates a favorable safety profile and long-term clinical activity in a substantial fraction of FL patients in need of therapy. ClinicalTrials.gov Identifier: NCT00772655.

Liquid BIOpsy for MiNimal RESidual DiSease Detection in Head and Neck Squamous Cell Carcinoma (LIONESS)\u2014a personalised circulating tumour DNA analysis in head and neck squamous cell carcinoma

BackgroundHead and neck squamous cell carcinoma (HNSCC) remain a substantial burden to global health. Cell-free circulating tumour DNA (ctDNA) is an emerging biomarker but has not been studied sufficiently in HNSCC.MethodsWe conducted a single-centre prospective cohort study to investigate ctDNA in patients with p16-negative HNSCC who received curative-intent primary surgical treatment. Whole-exome sequencing was performed on formalin-fixed paraffin-embedded (FFPE) tumour tissue. We utilised RaDaRTM, a highly sensitive personalised assay using deep sequencing for tumour-specific variants, to analyse serial pre- and post-operative plasma samples for evidence of minimal residual disease and recurrence.ResultsIn 17 patients analysed, personalised panels were designed to detect 34 to 52 somatic variants. Data show ctDNA detection in baseline samples taken prior to surgery in 17 of 17 patients. In post-surgery samples, ctDNA could be detected at levels as low as 0.0006% variant allele frequency. In all cases with clinical recurrence to date, ctDNA was detected prior to progression, with lead times ranging from 108 to 253 days.ConclusionsThis study illustrates the potential of ctDNA as a biomarker for detecting minimal residual disease and recurrence in HNSCC and demonstrates the feasibility of personalised ctDNA assays for the detection of disease prior to clinical recurrence.

Liposomal Cytarabine and Daunorubicin (CPX-351) in Combination with Gemtuzumab Ozogamicin (GO) in Relapsed Refractory (R/R) Acute Myeloid Leukemia (AML) and Post-Hypomethylating Agent (Post-HMA) Failure High-Risk Myelodysplastic Syndrome (HR-MDS)

Liposomal Cytarabine and Daunorubicin (CPX-351) in Combination with Gemtuzumab Ozogamicin (GO) in Relapsed Refractory (R/R) Acute Myeloid Leukemia (AML) and Post-Hypomethylating Agent (Post-HMA) Failure High-Risk Myelodysplastic Syndrome (HR-MDS)

Abstract 553: Personalized circulating tumor DNA analysis in head and neck squamous cell carcinoma: Preliminary results of the Liquid BIOpsy for MiNimal RESidual DiSease Detection in Head and NeckSquamous Cell Carcinoma (LIONESS) study

Abstract Introduction Head and neck squamous cell carcinoma (HNSCC) remains a substantial burden to global health. Despite evolving therapies, 5-year survival is less than 50% and unlike other cancers, reliable biomarkers to monitor treatment response do not exist. Cell-free circulating tumor DNA (ctDNA) is an emerging biomarker but has not yet been studied sufficiently for HNSCC. The detection of ctDNA as a marker of minimal residual disease following curative-intent treatment holds promise for identifying patients at an increased risk of relapse, who may benefit from adjuvant radio(chemo)therapy or facilitate close monitoring with repeat resection if needed. Methods We conducted a single-center prospective experimental evidence-generating cohort study to assess ctDNA in 30 patients with p16-negative HNSCC (stages I-IVB) who received primary surgical treatment with curative intent at our institution. Whole exome sequencing (WES) was performed on formalin-fixed paraffin-embedded tumor tissue to a median depth of 250x. For each patient, we selected up to 48 somatic variants for personalized ctDNA assay design. We used the RaDaRTM assay to analyze serial pre- and post-operative plasma samples (range 2-6) for evidence of minimal residual disease or recurrence. Results In a subset of patients analyzed to evaluate the performance of RaDaR, personalized panels were designed with between 34 and 48 somatic variants (median 48). Preliminary data shows 100% ctDNA detection in baseline samples taken prior to surgery at tumor fractions ranging from 312 ppm (equivalent to 0.03% AF) to 7579 ppm (equivalent to 0.76% AF). In post-surgery samples, ctDNA could be detected at levels as low as 26 ppm (equivalent to 0.0026% AF). Analysis of follow-up plasma samples will be presented along with data from the full patient cohort. Conclusions This study illustrates the potential of ctDNA as a biomarker in HNSCC and demonstrates the feasibility of personalized ctDNA assays for the detection of minimal residual disease post-treatment and for monitoring for early detection of relapse. Citation Format: Susanne Flach, Karen Howarth, Sophie Hackinger, Christodoulos Pipinikas, Kirsten McLay, Giovanni Marsico, Christoph Walz, Olivier Gires, Martin Canis, Philipp Baumeister. Personalized circulating tumor DNA analysis in head and neck squamous cell carcinoma: Preliminary results of the Liquid BIOpsy for MiNimal RESidual DiSease Detection in Head and NeckSquamous Cell Carcinoma (LIONESS) study [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 553.

Liposomal Cytarabine and Daunorubicin (CPX-351) in Combination with Gemtuzumab Ozogamicin (GO) in Relapsed Refractory (R/R) Patients with Acute Myeloid Leukemia (AML) and Post-Hypomethylating Agent (Post-HMA) Failure High-Risk Myelodysplastic Syndrome (HR-MDS)

Liposomal Cytarabine and Daunorubicin (CPX-351) in Combination with Gemtuzumab Ozogamicin (GO) in Relapsed Refractory (R/R) Patients with Acute Myeloid Leukemia (AML) and Post-Hypomethylating Agent (Post-HMA) Failure High-Risk Myelodysplastic Syndrome (HR-MDS)

Significance of minimal residual disease monitoring by real-time quantitative polymerase chain reaction in core binding factor acute myeloid leukemia for transplantation outcomes.

Despite the well-defined role of minimal residual disease (MRD) monitoring by real-time quantitative polymerase chain reaction (RT-PCR) for RUNX1/RUNX1T1 and CBFB-MYH11 transcripts in core binding factor (CBF) acute myeloid leukemia (AML) after intensive chemotherapy, there has been a paucity of data assessing the utility of MRD monitoring at and after allogeneic hematopoietic stem cell transplantation (HSCT). Patients with CBF AML who underwent HSCT in complete remission (first or second) from January 2007 through December 2018 were included in this analysis. MRD by polymerase chain reaction at HSCT was assessed in 50 of 76 patients, and 44 (88%) had evidence of MRD (MRDpos). MRDpos patients had 3-year overall survival (OS) and leukemia-free survival (LFS) rates of 69.3% and 66.3%, respectively. Six MRD-negative patients had 3-year OS and LFS rates of 100% and 100%, respectively. Thirty-five of the 70 evaluable patients (50%) had a day +100 MRD assessment by RT-PCR, and 14 (40%) were MRDpos. The presence of MRD by RT-PCR on day +100 was not associated with lower estimates of LFS (75% vs 82.2%; P=.3) but was associated with a higher relapse incidence, although the difference did not reach statistical significance (27.6% vs 9.7%; P=.2). Durable complete remissions can be achieved in patients with CBF AML with HSCT even if they are MRDpos by RT-PCRat HSCT. The clinical impact of frequent MRD monitoring for identifying a group at high risk for early relapse and then for determining the best time point for therapeutic interventions to prevent impending relapse warrants investigation in prospectively designed clinical trials.

Phased Variant Enrichment for Enhanced Minimal Residual Disease Detection from Cell-Free DNA

Background: Circulating tumor DNA (ctDNA) is an emerging biomarker in non-Hodgkin lymphomas (NHLs). Current methods for ctDNA minimal residual disease (MRD) are limited by two factors - low input DNA amounts and high background error rates. VDJ sequencing (i.e., IgHTS) has low background but is limited by low cell-free DNA (cfDNA). Tracking multiple mutations via CAPP-Seq improves sensitivity, but detection is limited by background errors. Clustered mutations have been described in multiple cancers including NHLs and potentially have lower error rates. We explored clustered mutations from whole-genome sequencing (WGS) to identify 'phased variants' (PVs), defined as multiple mutations on a single DNA molecule (Fig 1A). We designed a method to capture PVs for improved ctDNA detection and explored its utility for MRD in DLBCL. Methods: We reanalyzed WGS from 1455 tumors across 11 cancer types. We identified genomic regions recurrently containing PVs and designed an assay for deep cfDNA sequencing. We applied this assay to 171 patients with large B-cell lymphomas. We compared the performance of PVs for disease detection to current ctDNA techniques, including CAPP-Seq and duplex sequencing. Results: To utilize PVs, mutations must occur within a typical cfDNA strand (~170bp). We measured the frequency of putative PVs in WGS, focusing on pairs of mutations occurring within <170bp. PVs were more frequent in NHLs than any other histology (median: DLBCL, 642; FL, 307; Burkitt, 89.5; CLL, 34; breast, 46; lung, colorectal, melanoma, bladder, cervical, head & neck < 10 per case; P < 0.001 for NHLs vs others). PVs in NHLs were enriched in single base substitution mutational signatures associated with activation-induced cytidine deaminase (AID) (SBS84 & 85). PVs in NHLs occurred in stereotyped regions, including canonical AID targets such as IGH, IGK, and IGL, as well as 44 other AID targets (Schmitz, NEJM 2018) (Fig 1B). We additionally identified novel regions not previously implicated as targets of AID, including LPP, XBP1, BZRAP1, and HLA-DQ. We designed an approach for enriching PVs from ~115kb (Phased variant Enrichment Sequencing, PhasE-Seq) and other regions recurrently mutated in B-NHLs. We compared PhasE-Seq and CAPP-Seq using tumor and plasma samples from 16 patients. Compared to CAPP-Seq, PhasE-Seq yielded more SNVs and PVs per case (median SNVs: 331 vs 114, P<0.001; PVs: 729 vs 222.5, P <0.001). We next applied PhasE-Seq to 171 patients with untreated lymphomas (DLBCL, 148; primary mediastinal B-cell lymphoma, PMBCL, 23) profiling 58 tumor and 171 plasma samples with matched germline. We observed significant differences in the distribution of PVs between subtypes - for example, GCB-DLBCL had more PVs in BCL2, MYC, and SGK1, while ABC-DLBCL had more in PIM1 and IGHV4-34 (Fig 1C). Similarly, we noted enrichment in PVs in PMBCL in CIITA, SOCS1, CD83, and ITPKB. We then compared PhasE-Seq to alternative methods for MRD detection. We used limiting dilutions of patient ctDNA down to 1:1,000,000 to establish the detection limit (LOD, Fig 1D). PhasE-Seq outperformed CAPP-Seq and duplex sequencing for recovery of expected tumor content, with a high degree of linearity down to ~1:1,000,000. We applied standard CAPP-Seq and PhasE-Seq to patient cfDNA samples after two cycles of front-line therapy (n=92). We previously reported a 2.5-log reduction in ctDNA as prognostic at this time-point (Kurtz, JCO 2018). Using CAPP-Seq, 58% (53/92) of samples were undetectable. Using PhasE-Seq, 30% (16/53) of samples not detected by CAPP-Seq had evidence of MRD, with levels as low as 2:1,000,000. In patients with ctDNA undetected by CAPP-Seq, detection by PhasE-Seq significantly stratified outcomes (Fig 1E). Conclusions: PVs are frequent in NHLs, likely due to AID, and correlate with disease biology. PhasE-Seq allows for superior detection of ctDNA, including MRD detection in the majority of patients after 2 cycles. Targeted sequencing of ctDNA should consider PVs to maximize detection and guide precision approaches. Figure 1: A) Structure of phased variants B) Distribution of putative PVs from WGS data C) Genomic enrichment in PVs in lymphoma subtypes D) Dilution series comparing PhasE-Seq, CAPP-Seq, and duplex sequencing E) Waterfall plot showing ctDNA level vs outcome; undetectable ctDNA by CAPP-Seq is highlighted F) EFS of patients with undetectable ctDNA by CAPP-Seq after 2 cycles, stratified by PhasE-Seq Disclosures Kurtz: Roche: Consultancy. Diehn:Roche: Consultancy; AstraZeneca: Consultancy; Novartis: Consultancy; BioNTech: Consultancy; Quanticell: Consultancy. Alizadeh:Pharmacyclics: Consultancy; Janssen: Consultancy; Genentech: Consultancy; Roche: Consultancy; Gilead: Consultancy; Celgene: Consultancy; Chugai: Consultancy; Pfizer: Research Funding.

Monitoring for Minimal Residual Disease before and after Allogeneic Hematopoietic Stem Cell Transplantation in Newly Diagnosed Acute Leukemia

Objective:To assess the relevance between minimal residual disease (MRD) levels and disease recurrence and prognosis in patients with acute leukemia before and after hematopoietic stem cell transplantation (HSCT), and further to explore potential benefit based on pre-transplant MRD stratification. Methods: A total of 113 patients with newly diagnosed acute leukemia in Fujian Union hospital from April 2013 to April 2019 were retrospectively analyzed. 94 patients were complete remission (CR)( 50 cases of AML and 44 cases of ALL). 19 patients didn't achieve CR( 10 patients with AML and 9 patients with ALL). The median age was 26(1-56) years. 67 patients were male and 46 patients were female. The sources of hematopoietic stem cell include bone marrow, cord blood and peripheral blood stem cell. All patients underwent myeloablative conditioning before transplantation. Standard graft-versus-host disease (GVHD) prophylaxis was applied in all patients. MRD was measured by flow cytometry and real-time quantitative polymerase chain reaction(PCR). MRD values >10-4 or BCR-ABL were considered positive for minimal residual disease and compared with bone marrow morphological findings. Median follow time was 15.2 (2.2 to 70.5) months. Results: Among patients in CR, achieving pre-MRD negativity was associated with longer 2-year relapse-free survival (2-year RFS; 81.4% vs 49.1%; P=0.003) and 2-year overall survival (2-year OS; 84% vs 50.6%; P=0.012). Compared to pre-MRDneg patients, pre-MRDpos patients had a higher incidence of relapse (31.2% vs. 5.1%, P=0.001). There was no significant difference between OS and MRD level after transplantation (OS; post-MRDneg 82.9% vs. post-MRDpos 66.7%, P =0.468). In the second set of analyses, CR patients were classified into the MRDpos /MRDpos group, the MRDpos /MRDneg group, the MRDneg /MRDpos group, and the MRDneg /MRDneg group according to MRD dynamics. Compared to the other three groups, patients from the MRDpos /MRDpos group had higher cumulative incidences of relapse (MRDpos /MRDpos, 48.3%; MRDpos /MRDneg, 22.2%, MRDneg /MRDpos,40.0%; MRDneg /MRDneg, 2.7%; P<0.000) and worse 2-year RFS(MRDpos /MRDpos, 34.3%; MRDpos /MRDneg, 60.0%, MRDneg /MRDpos,60.0%; MRDneg /MRDneg, 76.6%; P=0.012). 1-year RFS and 1-year OS were similar in patients who didn't achieve CR, regardless of the post-MRD response(1-year RFS; 50.0% vs. 14.3%, P=0.063 and 1-year OS; 50.0% vs. 14.3%, P=0.184).In this group, 7 patients with MRD-positive after transplantation did not survive, but 12 post-HSCT MRD-negative patients had not relapsed. Conclusion: MRD can be used as a sensitive indicator to evaluate disease prognosis. Patients who are in morphologic remission and have no evidence of MRD before the HSCT come to encouraging leukemia-free survival. Continuous regular dynamic observation is important for guiding disease recurrence and prognosis. Disclosures No relevant conflicts of interest to declare.

T Cells Expressing an Anti-B-Cell Maturation Antigen (BCMA) Chimeric Antigen Receptor with a Fully-Human Heavy-Chain-Only Antigen Recognition Domain Induce Remissions in Patients with Relapsed Multiple Myeloma

Chimeric antigen receptor (CAR) T cells expressing B-cell maturation antigen (BCMA) can target and kill multiple myeloma (MM). BCMA was chosen as a target for MM because it is expressed by almost all cases of MM but has a restricted expression pattern on normal cells. CAR antigen-recognition domains made up of monoclonal antibody-derived, single-chain-variable fragments (scFv) are potentially immunogenic. To reduce the risk of recipient immune responses against CAR T cells, we used the sequence of a novel anti-BCMA, fully-human, heavy-chain-only binding domain designated FHVH33. The FHVH33 binding domain sequence was from TeneoBio, Inc. FHVH33 is smaller than a scFv. FHVH33 lacks the light chain, artificial linker sequence, and 2 associated junctions of a scFv, so it is predicted to be less immunogenic than a scFv, especially murine-derived scFvs. We constructed a CAR incorporating FHVH33, CD8α hinge and transmembrane domains, a 4-1BB costimulatory domain, and a CD3ζ T-cell activation domain. The CAR, FHVH33-CD8BBZ, is encoded by a γ-retroviral vector. FHVH33-CD8BBZ-expressing T cells (FHVH-BCMA-T) exhibited a full range of T-cell functions in vitro and eliminated tumors and disseminated malignancy in mice (Lam et al, Blood (ASH abstract) 2017 vol 130: 504). We are conducting the first clinical trial of FHVH-BCMA-T. Patients receive conditioning chemotherapy on days -5 to -3 with 300 mg/m2 of cyclophosphamide and 30 mg/m2 of fludarabine followed by infusion of FHVH-BCMA-T on day 0. This dose-escalation trial has 5 planned dose levels (DL). Twelve patients have received FHVH-BCMA-T on 3 DLs, 0.75x106, 1.5x106 and 3x106 CAR+ T cells/kg of bodyweight. Three patients were enrolled on the trial but not treated. The median age of patients enrolled was 63 (range 52-70); patients received a median of 6 lines of anti-myeloma therapy (range 3-10) prior to treatment with FHVH-BCMA-T. Ten patients out of 12 patients have achieved objective responses (OR). Five patients have obtained CRs or VGPRs to date. One patient achieved a partial remission (PR) 26 weeks after FHVH-BCMA-T infusion through a continued decrease in a measurable plasmacytoma. Five out of 7 patients who had myeloma with high-risk cytogenetics had an OR (Table). ORs occurred in patients with large soft-tissue plasmacytomas. Loss of BCMA expression on myeloma cells after treatment was documented in 2 patients. Two patients who developed progressive MM after CAR T-cell infusion had evidence of minimal residual disease in bone marrow 1-2 months post infusion of CAR T cells (patients 7,8). Eleven out of 12 patients had cytokine release syndrome (CRS); CRS grades ranged from 1-3 (Lee et al. Biol Blood Marrow Transplant 25 (2019) 625-638). The median peak C reactive protein (CRP) of the patients with CRS was 156.3 mg/L. Of 12 patients, 1 received the interleukin-6-receptor antagonist tocilizumab on day +6 to treat grade 3 CRS with hypotension requiring low-dose pressor therapy, grade 2 ejection fraction (EF) decrease and elevation of creatinine kinase (CK). All parameters returned to baseline by day +10. Patient 12 had a grade 3 decrease in EF which resolved by day +29. Two patients had grade 2 neurotoxicity that resolved without intervention: patient 3 had headaches, dysarthria and word-finding difficulties that resolved after 6 days while patient 6 had headaches on day +4. Patient 12 had grade 3 neurotoxicity with confusion on day +2; she was given dexamethasone with improvement in mental status the same day. After attaining a response, patient 6 died from influenza complications 6 weeks after FHVH-BCMA-T infusion. A median of 10.6% (range 1.1-46) of bone marrow T cells were CAR+ when assessed 14 days after FHVH-BCMA-T infusion. We assessed blood CAR+ cells by quantitative PCR. The median peak level of CAR+ cells was 76.5 cells/µl (range 3-347 cells/µl) and the median day post-infusion of peak blood CAR+ cell levels was 13 (range 9-14). The results from this phase 1 trial demonstrate that FHVH-BCMA-T cells can induce responses at low dose levels. Patients who had no CRS or low-grade CRS achieved objective responses. Toxicity was limited and reversible. Accrual to this trial continues. A maximum tolerated dose has not been determined yet. These results encourage further development of FHVH CAR-T. Table Disclosures Manasanch: Janssen: Honoraria; Sanofi: Honoraria; Takeda: Honoraria; Merck: Research Funding; Skyline Diagnostics: Research Funding; Sanofi: Research Funding; Quest Diagnostics: Research Funding; Celgene: Honoraria. Trinklein:Teneobio, Inc.: Employment, Equity Ownership. Buelow:Teneobio, Inc.: Employment, Equity Ownership. Kochenderfer:Kite and Celgene: Research Funding; Bluebird and CRISPR Therapeutics: Other: received royalties on licensing of his inventions. OffLabel Disclosure: Cyclophosphamide and fludarabine are used in combination for conditioning chemotherapy prior to CAR T-cell infusion

Final Clinical Results of a Phase II Study of High Dose Cytarabine Followed By Pembrolizumab in Relapsed/Refractory AML

Background: Despite recent advances in the therapeutic armamentarium for AML, outcomes remain dismal for patients (pts) with relapsed/refractory (R/R) AML. Response rates with high dose cytarabine (HiDAC) salvage chemotherapy are approximately 20%. Multiple immune aberrations in AML lead to immune suppression, exhaustion, and senescence. Programmed Death-1 (PD-1), a co-inhibitory receptor (IR) on immune cells, suppresses immune activation and is exploited by leukemic cells to evade immune surveillance. PD-1 and other IRs are up-regulated during disease progression. We hypothesized that pembrolizumab, a monoclonal antibody targeting PD-1, after HiDAC would stimulate a T-cell mediated anti-leukemic immune response. Methods: Eligibility for this study included R/R AML 18-70 years, ECOG PS 0-1 and adequate organ function. Treatment consisted of HiDAC (<60 years: 2 gm/m2 IV Q12hours days 1-5; >60 years: 1.5 gm/m2 IV Q12hours days 1-5) followed by pembrolizumab 200 mg IV on day 14. The primary objective of this study was to estimate the overall complete remission (CR + CRi) rate. Secondary objectives included assessment of safety, durability of CR, overall survival (OS) and biomarker correlates of response. Overall responders were eligible to receive maintenance phase pembrolizumab 200 mg IV Q3weeks for up to 2 years until progression. Allogeneic stem cell transplant (alloSCT) was permissible before or after maintenance phase. Results: Thirty-seven pts were enrolled and evaluable (Table 1). Sixteen (43%) pts had refractory disease and 16 (43%) pts had relapsed AML with CR1 duration <1 year. Excluding electrolyte abnormalities, the most common pembrolizumab-related toxicities were febrile neutropenia (57%), ALT elevation (43%; Grade >3: n=1), AST elevation (32%; Grade >3: n=1), fatigue (27%), alkaline phosphatase elevation (24%), and maculopapular rash (19%; Grade >3: n=2). Grade >3 immune-related adverse events (iRAE) were rare (maculopapular rash: n=2, AST/ALT increase: n=2, right upper quadrant pain with lymphocytic infiltrate in liver: n=1) and self-limiting. Five (14%) pts required steroid administration for grade 2 hyperbilirubinemia (n=1), grade 3 ALT elevation (n=1), grade 3 AST elevation with liver biopsy revealing no evidence of iRAE (n=1), grade 3 bilirubin subsequently deemed to be a delayed hemolytic transfusion reaction (n=1), and grade 3 systolic dysfunction without evidence of myocarditis by endomyocardial biopsy or cardiac MRI (n=1). Sixty-day mortality was 3% (1/37) due to progressive AML. Median time to full neutrophil (>1x109/L) and platelet (>100x109/L) recovery was 32 and 31 days, respectively. The overall response (ORR: CR+CRi+PR+MLFS) and composite CR (CR+CRi) rates were 46% [29%,63%] and 38% [22%,55%], respectively, meeting the primary endpoint of the study. Notably, 13/28 (46%) pts receiving HiDAC + pembrolizumab as their first salvage regimen achieved CR/CRi. Two pts refractory to HiDAC (administered within past 6 months) achieved CR including one pt who was refractory to HiDAC salvage 1 month prior to enrollment and ultimately achieved CR without evidence of minimal residual disease. Nine (24%) pts received an alloSCT. There were no instances of Grade >3 acute GVHD or veno-occlusive disease post-alloSCT. Nine (24%) pts received maintenance phase pembrolizumab (median # of cycles = 3; range: 1-12) for CR (n=8) or PR (n=1). Seven out of 9 pts relapsed/progressed after maintenance phase. Median follow-up among survivors, and median OS, event-free survival and disease-free survival was 7.8 months, 8.9 months [6.0,13.1], 6.9 months [4.2,11.5], and 5.7 months [1.9,7.3], respectively. Conclusions: Pembrolizumab can be safely administered after HiDAC salvage in R/R AML. Severe iRAE's were uncommon despite administration after cytotoxic chemotherapy. The addition of pembrolizumab to HiDAC led to an encouraging overall CR rate meeting the primary endpoint of the study. Immunogenomic biomarker analyses consisting of B cell receptor amplicon sequencing, RNA-seq of blasts and CD8+ T cells, CD8+ T cell receptor repertoire, whole exome sequencing and flow cytometry analyses are ongoing to determine predictors of response. These results warrant further investigation of IR blockade and other immunomodulatory therapeutic strategies after intensive cytotoxic chemotherapy in AML. Disclosures Zeidner: Takeda: Research Funding; Merck: Research Funding; AsystBio Laboratories: Consultancy; Pfizer: Honoraria; Tolero: Honoraria, Research Funding; Daiichi Sankyo: Honoraria; Celgene: Consultancy, Honoraria, Research Funding; Agios: Honoraria; AbbVie: Honoraria. Vincent:Pharmacyclics: Research Funding; Merck: Research Funding. Foster:Bellicum Pharmaceuticals: Research Funding; Macrogenics: Research Funding; Celgene: Research Funding; Daiichi Sankyo: Consultancy. Coombs:Dedham Group: Consultancy; Covance: Consultancy; Cowen & Co.: Consultancy; Octopharma: Honoraria; H3 Biomedicine: Honoraria; Loxo: Honoraria; Pharmacyclics: Honoraria; Medscape: Honoraria. Webster:Pfizer: Consultancy; Amgen: Consultancy; Genentech: Research Funding. DeZern:Astex Pharmaceuticals, Inc.: Consultancy; Celgene: Consultancy. Smith:Jazz: Consultancy; Pfizer: Consultancy; Novartis: Consultancy; Agios: Consultancy. Levis:Amgen: Consultancy, Honoraria; Astellas: Consultancy, Research Funding; FUJIFILM: Consultancy, Research Funding; Menarini: Consultancy, Honoraria; Novartis: Consultancy, Research Funding; Daiichi Sankyo Inc: Consultancy, Honoraria; Agios: Consultancy, Honoraria. Luznik:Merck: Research Funding, Speakers Bureau; Genentech: Research Funding; AbbVie: Consultancy; WindMiL Therapeutics: Patents & Royalties: Patent holder. Serody:Merck: Research Funding; GlaxoSmithKline: Research Funding. Gojo:Amphivena: Research Funding; Amgen Inc: Consultancy, Honoraria, Research Funding; Juno: Research Funding; Merck: Research Funding; Jazz: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Abbvie: Consultancy, Honoraria. OffLabel Disclosure: Pembrolizumab is investigational for AML.

Abstract 1387: Tracking minimal residual disease in post-operative cell-free DNA using MSK-ACCESS

Abstract A noted clinical application for liquid biopsy is as a non-invasive method of detecting and monitoring of minimal residual disease (MRD) in patients with cancer. The low concentration of circulating tumor DNA in blood, especially in early stage cancers, however, complicates the detection of tumor-derived cell-free DNA. To address this challenge, we have developed MSK-ACCESS (Analysis of Circulating cfDNA to Examine Somatic Status), a custom NGS assay covering selected exons from 129 cancer related genes for high-sensitivity detection of somatic mutations from plasma. Using ultra-high depth sequencing, with duplex unique molecular indexing (UMI), unique dual sample barcodes, and background error suppression, MSK-ACCESS is able to detect low-frequency (0.1%) variants with high confidence. The design of MSK-ACCESS leverages our dataset of more than 30,000 tumors profiled by our institutional tumor sequencing assay, MSK-IMPACT, ensuring that the majority of patients harbor multiple mutations that can be tracked in plasma. We have validated MSK-ACCESS using plasma samples collected from 40 healthy individuals and 70 cancer patients harboring a range of somatic mutations in 11 genes. Greater than 95% of mutations at allele fractions >0.1% were empirically detected, and we established the performance characteristics of the assay through intra- and inter-assay reproducibility tests and dilution experiments. We have initiated clinical trials in multiple tumor types to evaluate the benefit of early therapeutic intervention in patients where MSK-ACCESS can detect circulating tumor DNA following surgery. Tumor mutations revealed by MSK-IMPACT in surgically resected specimens will be monitored at regular intervals as evidence of MRD. As a proof of concept, we have applied MSK-ACCESS to monitor variants known from tissue tumor sequencing in pre- and post-surgical cfDNA samples from 9 colon adenocarcinoma patients. All samples were sequenced to an average total depth of approximately 20,000X coverage and subsequently collapsed to consensus sequences exhibiting an average noise level less than 0.0006%. Circulating tumor DNA was detected in 66%(6/9) of the pre-surgical samples. Of these samples, ctDNA was also detected in 50% (3/6) of the post-surgical samples. Overall, this study shows that MSK-ACCESS can be used to successfully detect MRD. Citation Format: Maysun M. Hasan, Juber Patel, Ian Johnson, Fanli Meng, Grittney K. Tam, Xiaohong Jing, Julie L. Yang, A. Rose Brannon, Jayakumaran Gowtham, Dennis P. Stephens, Monica Diosdado, Ryma Benayed, Ahmet Zehir, Chin-Tung Chen, Martin R. Weiser, Dana Tsui, Brian Houck-Loomis, Michael Berger. Tracking minimal residual disease in post-operative cell-free DNA using MSK-ACCESS [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1387.

Antibody-based therapies in patients with acute lymphoblastic leukemia.

The use of multiagent combination chemotherapy regimens results in cure rates of >90% for children and ∼40% for adults with acute lymphoblastic leukemia (ALL) but is associated with extensive toxicity and disappointingly low efficacy in relapsed patients. ALL blast cells express several surface antigens, including CD20, CD22, and CD19, which represent valuable targets for immunotherapy. Monoclonal antibodies, antibody-drug conjugates, and bispecific T-cell-engaging antibodies targeting these antigens offer novel mechanisms of action. Within the last several years, the anti-CD20 antibody rituximab has been added to chemotherapy for newly diagnosed patients <60 years with CD20+ pre-B ALL and significantly improved the 2-year event-free survival from 52% to 65%. In adults with relapsed or refractory CD22+ ALL, the antibody-drug conjugate inotuzumab ozogamicin resulted in a complete response rate of 81% and median overall survival of 7.7 months with reduced toxicity compared with standard chemotherapy. Similarly, the bispecific T-cell-engaging antibody blinatumomab yielded a complete response rate of 44% and a median overall survival of 7.7 months in an extensively treated ALL population. Moreover, ∼80% of ALL patients in complete remission with evidence of minimal residual disease (MRD) achieved a complete MRD response following treatment with blinatumomab. These results highlight the tremendous promise of antibody-based treatment approaches for ALL. Ongoing and future research is critical to further define the role of the various immunotherapies in the frontline treatment of ALL. Additional challenges include the optimal sequencing of the available antibodies in the relapsed setting as well as their integration with stem cell transplant and chimeric antigen receptor T-cell therapy.

Effective Immunomodulation with Pomalidomide Beginning at Early Lymphocyte Recovery during Induction Timed Sequential Therapy (TST) for Acute Myeloid Leukemia (AML) and High-Risk Myelodysplasia (HR-MDS)

Introduction: Adults with AML have immune aberrations leading to immune suppression, exhaustion, evasion, and senescence. Early lymphocyte recovery (ELR) after induction TST is dominated by an expansion of oligoclonal peripherally derived regulatory T cells (Tregs). Pomalidomide (Pom), a small molecule immunomodulatory agent (IMiD), leads to the selective ubiquitination of Aiolos and Ikaros by cereblon; this ubiquitination increases IL-2 production, inhibits Tregs, and inhibits angiogenesis. We hypothesize that Pom administration after induction TST may enhance anti-leukemic activity through immune modulation.Methods: A multicenter phase 1 dose escalation study was conducted to determine the safety and tolerability of Pom administration after induction TST in newly diagnosed AML and HR-MDS patients 18-65 years. Favorable-risk cytogenetics were excluded. All patients received induction TST with AcDVP16: Cytarabine 667 mg/m2/day continuous IV days 1-3, daunorubicin 45 mg/m2 IV days 1-3 (or idarubicin 8-12 mg/m2 IV days 1-3 in daunorubicin shortage), etoposide 400 mg/m2 IV days 8-10, followed by Pom administration at ELR (after day 14 and within 3 days of the total white blood cell (WBC) reaching ≥0.2x109/L above nadir, but no later than day 30). Pom dose escalation occurred in 2 cohorts: 10 days versus 21 days of administration, in a traditional 3+3 design.Results: A total of 51 patients were enrolled and 43 (AML: n=39, HR-MDS: n=4) received Pom across 3 institutions (Table 1). Eight patients did not receive Pom due to no ELR by day 30 (n=3), sepsis (n=3), noncompliance with treatment (n=1), and death prior to ELR due to acute respiratory distress syndrome (n=1). Median time for Pom initiation after AcDVP16 induction was day 21 (range: 15-30 days). Pom maximal tolerated dose (MTD) was 4 mg for 21 consecutive days at ELR. The most common non-hematologic grade ≥3 toxicities related to Pom were febrile neutropenia (30%), maculopapular rash (14%), and aminotransferase elevation (5%). Dose-limiting toxicities (DLTs) of Pom at 8 mg for 21 days were grade 3 ALT/AST elevation and grade 4 respiratory failure, respectively. Pom was discontinued early (median duration = 7 days; range: 3-20 days) in 14 (33%) patients due to disease progression (n=4), grade 3 rash (n=3), DLT (n=2), patient decision (n=2), progressive cytopenias (n=1), and grade 4 acute kidney injury (n=1).Overall, 32/43 (74%) achieved CRc (CR: n= 30, CRi: n=2). Among the 4 HR-MDS patients, 2 (50%) achieved CRc, whereas 77% (30/39) of AML patients achieved CRc. Of the 32 CRc patients, 19 (59%) had no evidence of minimal residual disease (MRD) by standard testing (flow cytometry, FISH, cytogenetics, and/or molecular PCR). Encouraging CR rates were seen in poor-risk subsets: Age ≥60 years: 75% CRc (9/12); Secondary AML: 71% CRc (10/14); Adverse-risk cytogenetics: 82% CRc (14/17). Thirty- and 60-day mortality were 0 and 2%, respectively. Median time to neutrophil (≥1.0x109/L) and platelet (≥100x109/L) recovery was 38 (range: 28-86) and 33 days (range: 24-75), respectively. With a median follow-up of 23.9 months, median overall survival, disease-free survival and event-free survival were 33.8 months, 20.3 months, and 8.7 months, respectively.Pharmacodynamic biomarker studies revealed a significant decrease in the expression of Aiolos in peripheral blood CD4+ and CD8+ T cells when compared with AML controls who received AcDVP16 induction but did not receive pomalidomide (p <0.05; Figure 1). Moreover, these findings were corroborated in the bone marrow where Aiolos expression in CD4+ and CD8+ T cells substantially decreased after pomalidomide initiation.Conclusions: Pom can be added at the time of ELR after induction TST without increased toxicity and CR rates that compare favorably with historical controls of TST and other standard induction therapies. Despite administration at the time of profound cytopenias, Pom was well tolerated and did not significantly prolong hematologic recovery. Correlates suggest that Aiolos expression may be a pharmacodynamic biomarker of activity. Identification of immune biomarkers to predict for response and duration of response are ongoing. Further exploration of Pom both at the time of induction TST and during CR are warranted. [Display omitted] DisclosuresZeidner:Rafael Pharmaceuticals: Other: Travel Fees; Merck: Research Funding; Asystbio Laboratories: Consultancy; Takeda: Other: Travel fees, Research Funding; Tolero: Honoraria, Other: Travel Fees, Research Funding; Celgene: Honoraria. Zeidan:Agios: Consultancy; Incyte: Employment; Novartis: Consultancy; Celgene: Consultancy; Ariad: Consultancy, Speakers Bureau; Gilead: Consultancy; Pfizer: Consultancy; Abbvie: Consultancy. Pratz:Astellas: Consultancy, Research Funding; Agios: Research Funding; Boston Scientific: Consultancy; AbbVie: Consultancy, Research Funding; Millenium/Takeda: Research Funding. Foster:Shire: Honoraria; Pfizer: Research Funding; Macrogenics: Research Funding; Celgene: Research Funding. Coombs:H3 Biomedicine: Honoraria; AROG: Other: Travel fees; DAVA Oncology: Honoraria; Abbvie: Consultancy; Incyte: Other: Travel fees. Luznik:WIndMIL Therapeutics: Equity Ownership, Patents & Royalties. Gojo:Amgen: Membership on an entity's Board of Directors or advisory committees; Jazz Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; Merck inc: Research Funding; Amgen: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees.