Identification of concurrent STAT3::RARA and RARA::STAT5b fusions in a variant APL case.

Acute promyelocytic leukemia (APL) with typically PML::RARA fusion gene caused by t (15;17) (q22; q12) was distinguished from other types of acute myeloid leukemia. In a subset of patients with APL, t (15;17) (q22;q21) and PML::RARA fusion cannot be detected. In this report, we identified the coexistence of STAT3::RARA and RARA::STAT5b fusions for the first time in a variant APL patient lacking t (15;17)(q22;q21)/PML::RARA fusion. Then, this patient was resistant to all-trans retinoic acid combined arsenic trioxide chemotherapy. Accurate detection of RARA gene partners is crucial for variant APL, and effective therapeutic regime is urgently needed.

Molecular genetics and management of world health organization defined atypical chronic myeloid leukemia.

Atypical chronic myeloid leukemia (CML) is a rare BCR::ABL1-negative hematopoietic stem cell disease characterized by granulocytic proliferation and granulocytic dysplasia. Due to both the challenging diagnosis and the rarity of atypical CML, comprehensive molecular annotation-based analyses of this disease population have been scarce, and it is currently difficult to identify the optimal treatment for atypical CML. To explore atypical CML genomic landscape and treatment options, we performed a systematic retrospective of the clinical data and outcomes of 31 atypical CML patients. We observed that the molecular landscape of atypical CML was highly heterogeneous, with multiple molecular events driving its pathogenesis. Patients with atypical CML had a low response to current therapies, with an overall response rate (ORR) of 33.3% to hypomethylating agent (HMA)-based therapy. The current treatment strategies, including hematopoietic stem cell transplantation (HSCT), did not improve overall survival (OS) in atypical CML patients, with a median survival of 20 months. Thus, the benefits from HSCT and candidates for HSCT remain to be further evaluated. Acute myeloid leukemia (AML)-like chemotherapy followed by bridging allogeneic HSCT may be an ideal regimen for suitable individuals. The large-scale and prospective clinical studies will help to address the dilemma.

Integrative genomic and transcriptomic profiling reveals distinct molecular subsets in adult mixed phenotype acute leukemia.

Mixed phenotype acute leukemia (MPAL) is a subtype of leukemia in which lymphoid and myeloid markers are co-expressed. Knowledge regarding the genetic features of MPAL is lacking due to its rarity and heterogeneity. Here, we applied an integrated genomic and transcriptomic approach to explore the molecular characteristics of 176 adult patients with MPAL, including 86 patients with T-lymphoid/myeloid MPAL (T/My MPAL-NOS), 42 with Ph+ MPAL, 36 with B-lymphoid/myeloid MPAL (B/My MPAL-NOS), 4 with t(v;11q23), and 8 with MPAL, NOS, rare types. Genetically, T/My MPAL-NOS was similar to B/T MPAL-NOS but differed from Ph+ MPAL and B/My MPAL-NOS. T/My MPAL-NOS exhibited higher CEBPA, DNMT3A, and NOTCH1 mutations. Ph+ MPAL demonstrated higher RUNX1 mutations. B/T MPAL-NOS showed higher NOTCH1 mutations. By integrating next-generation sequencing and RNA sequencing data of 89 MPAL patients, we defined eight molecular subgroups (G1-G8) with distinct mutational and gene expression characteristics. G1 was associated with CEBPA mutations, G2 and G3 with NOTCH1 mutations, G4 with BCL11B rearrangement and FLT3 mutations, G5 and G8 with BCR::ABL1 fusion, G6 with KMT2A rearrangement/KMT2A rearrangement-like features, and G7 with ZNF384 rearrangement/ZNF384 rearrangement-like characteristics. Subsequently, we analyzed single-cell RNA sequencing data from five patients. Groups G1, G2, G3, and G4 exhibited overexpression of hematopoietic stem cell disease-like and common myeloid progenitor disease-like signatures, G5 and G6 had high expression of granulocyte-monocyte progenitor disease-like and monocyte disease-like signatures, and G7 and G8 had common lymphoid progenitor disease-like signatures. Collectively, our findings indicate that integrative genomic and transcriptomic profiling may facilitate more precise diagnosis and develop better treatment options for MPAL.

Anti-CD19 and anti-BCMA CAR T cell therapy followed by lenalidomide maintenance after autologous stem-cell transplantation for high-risk newly diagnosed multiple myeloma.

Few prospective studies have examined posttransplant chimeric antigen receptor (CAR) T cell infusion as candidates for front-line consolidation therapy for high-risk multiple myeloma (MM) patients. This single-arm exploratory clinical trial is the first to evaluate the safety and efficacy of sequential anti-CD19 and anti-BCMA CAR-T cell infusion, followed by lenalidomide maintenance after autologous stem cell transplantation (ASCT), in 10 high-risk newly diagnosed multiple myeloma (NDMM) patients. The treatment was generally well tolerated, with hematologic toxicities being the most common grade 3 or higher adverse events. All patients had cytokine release syndrome (CRS), which was grade 1 in 5 patients (50%) and grade 2 in 5 patients (50%). No neurotoxicity was observed after CAR-T cell infusion. The overall response rate was 100%, with the best response being 90% for a stringent complete response (sCR), and 10% for a complete response (CR). At a median follow-up of 42 (36-49) months, seven (70%) of 10 patients showed sustained minimal residual disease (MRD) negativity for more than 2 years. The median progression-free survival (PFS) and overall survival (OS) were not reached. Although the sample size was small and there was a lack of control in this single-arm study, the clinical benefits observed warrant ongoing randomized controlled trials.

Akt-mediated platelet apoptosis and its therapeutic implications in immune thrombocytopenia.

Immune thrombocytopenia (ITP) is an autoimmune disorder characterized by low platelet count which can cause fatal hemorrhage. ITP patients with antiplatelet glycoprotein (GP) Ib-IX autoantibodies appear refractory to conventional treatments, and the mechanism remains elusive. Here we show that the platelets undergo apoptosis in ITP patients with anti-GPIbα autoantibodies. Consistent with these findings, the anti-GPIbα monoclonal antibodies AN51 and SZ2 induce platelet apoptosis in vitro. We demonstrate that anti-GPIbα antibody binding activates Akt, which elicits platelet apoptosis through activation of phosphodiesterase (PDE3A) and PDE3A-mediated PKA inhibition. Genetic ablation or chemical inhibition of Akt or blocking of Akt signaling abolishes anti-GPIbα antibody-induced platelet apoptosis. We further demonstrate that the antibody-bound platelets are removed in vivo through an apoptosis-dependent manner. Phosphatidylserine (PS) exposure on apoptotic platelets results in phagocytosis of platelets by macrophages in the liver. Notably, inhibition or genetic ablation of Akt or Akt-regulated apoptotic signaling or blockage of PS exposure protects the platelets from clearance. Therefore, our findings reveal pathogenic mechanisms of ITP with anti-GPIbα autoantibodies and, more importantly, suggest therapeutic strategies for thrombocytopenia caused by autoantibodies or other pathogenic factors.

HLA-mismatched stem cell microtransplantation compared to matched-sibling donor transplantation for intermediate/high-risk acute myeloid leukemia.

HLA-mismatched stem cell microtransplantation is a new form of transplantation reported in recent years. We compared 59 patients undergoing microtransplantation to 66 patients undergoing HLA-matched sibling donor (MSD) transplantation at the same period from April 2012 to December 2016, who all suffered from intermediate/high-risk acute myelogenous leukemia (AML) in first complete remission (CR1). The estimated overall survival (OS) at 2 years was 74.1% ± 6.2% and 34.3% ± 7.9% in MSD and microtransplantation group, respectively (P = 0.001). The estimated leukemia-free survival (LFS) at 2 years was 73.3% ± 6.1% in the MSD group and 31.6% ± 7.6% in the microtransplantation group (P = 0.000). The 2-year cumulative incidence of relapse was 17.6% and 62.3% in the MSD and microtransplantation groups, respectively (P < 0.0001). The 2-year cumulative incidence of nonrelapse mortality was 10.9% in MSD group and 4.2% in the microtransplantation group (P = 0.251). Hematopoietic recovery time was shorter in the microtransplantation group than in the MSD group (P < 0.05). The infection rate was higher in the MSD group than in the microtransplantation group (P = 0.012). The preliminary results suggested that OS and LFS of microtransplantation were inferior to MSD transplantation for intermediate/high-risk AML in CR1.

Characterizing the Tumor Suppressor Role of CEACAM1 in Multiple Myeloma.

BACKGROUND/AIMS: Carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), also known as CD66a, is a member of the immunoglobulin (Ig) superfamily that belongs to the carcinoembryonic antigen (CEA) family which plays a dual role in cancer. Previous studies showed high expression of CEACAM1 in multiple myeloma (MM). The aim of this study was to investigate the biological consequences of CEACAM1 overexpression in MM. METHODS: pEGFP-N1-CEACAM1 and pcDNA3.1-CEACAM1 expression plasmids were transfected into U-266 and RPMI8266 cell lines . Effect of CEACAM1 overexpression on the proliferation of two cell lines were tested by the CCK8 assay. Cell cycle and Apoptotic changes after CEACAM1 transfection were examined with AnnexinV-FITC/PI by flow cytometry. Hochest staining assay was used to confirm the apoptotic changes. Caspase-3 activity was examined by Western blotting. The cell invasion and migration activity change after CEACAM1 transfection were performed by well chamber assays and a wound healing, respectively. MMP-2 and MMP-9 proteins expression were detected by Western blotting. Flow cytometry immunophenotyping was be evaluated on myeloma cells from bone marrow taken from 50 patients with symptomatic MM newly diagnosed. The correlations between CEACAM1 expression levels and the clinical features across all groups were investigated. RESULTS: CEACAM1 overexpression significantly suppressed MM cell proliferation, induced cell apoptosis, and inhibited cell invasion and migration possibly through activation of caspase-3 and downregulation of MMP-2 and MMP-9. CEACAM1 expression in patients with DS stage I was more frequent (61.5%) than those with DS stage II (21.1%) or III (22.2%). Furthermore, patients with ß2-microglobulin levels equal to or less than 3.5 mg/L had higher CEACAM1 expression than those with ß2-microglobulin levels greater than 3.5 mg/L. CONCLUSION: Our findings suggest that CEACAM1 may act as a tumor suppressor in MM.

Haploidentical hematopoietic cell transplantation with or without an unrelated cord blood unit for adult acute myeloid leukemia: a multicenter, randomized, open-label, phase 3 trial.

Coinfusion of unrelated cord blood (UCB) units in haploidentical hematopoietic cell transplantation (haplo-HCT) (haplo-cord HCT) for hematopoietic malignancies showed promising results in previous reports, but the efficiency of haplo-cord HCT in acute myeloid leukemia (AML) still lacks sufficient evidence. This multicenter, randomized, phase 3 trial (ClinicalTrials.gov NCT03719534) aimed to assess the efficacy and safety of haplo-cord HCT in AML patients. A total of 268 eligible patients aged 18-60 years, diagnosed with measurable residual disease in AML (excluding acute promyelocytic leukemia), with available haploidentical donors and suitable for allotransplantation, were randomly allocated (1:1) to receive haplo-cord HCT (n = 134) or haplo-HCT (n = 134). The 3-year overall survival (OS) was the primary endpoint in this study. Overall median follow-up was 36.50 months (IQR 24.75-46.50). The 3-year OS of Haplo-cord HCT group was better than haplo-HCT group (80.5%, 95% confidence interval [CI]: 73.7-87.9 vs. 67.8% 95% CI 60.0-76.5, p = 0.013). Favorable progression-free survival (70.3%, 95% CI 62.6-78.8 vs. 57.6%, 95% CI 49.6-67.0, p = 0.012) and cumulative incidence of relapse (12.1%, 95% CI 12.0-12.2 vs. 30.3%, 95% CI 30.1-30.4, p = 0.024) were observed in haplo-cord HCT group. Grade 3-4 adverse events (AEs) within two years posttransplantation in the two groups were similar. Haplo-cord HCT patients exhibited a faster cumulative incidence of neutrophil recovery (p = 0.026) and increased T-cell reconstitution in the early period posttransplantation. Haplo-cord HCT can improve OS in AML patients without excessive AEs, which may exert additional benefits for recipients of haplo-HCT.

Evaluation of next-generation sequencing versus next-generation flow cytometry for minimal-residual-disease detection in Chinese patients with multiple myeloma.

PURPOSE: To evaluate the efficacy of next-generation sequencing (NGS) in minimal-residual-disease (MRD) monitoring in Chinese patients with multiple myeloma (MM). METHODS: This study analyzed 60 Chinese MM patients. During MRD monitoring in these patients' post-therapy, clonal immunoglobulin heavy chain (IGH) rearrangements were detected via NGS using LymphoTrack assays. MRD monitoring was performed using NGS or next-generation flow cytometry (NGF), and the results were compared. Additionally, the sensitivity and reproducibility of the NGS method were assessed. RESULTS: The MRD detection range of the NGS method was 10-6-10-1, which suggested good linearity, with a Pearson correlation coefficient of 0.985 and a limit of detection of 10-6. Intra- and inter-assay reproducibility analyses showed that NGS exhibited 100% reproducibility with low variability in clonal cells. At diagnosis, unique clones were found in 42 patients (70.0%) with clonal IGH rearrangements, which were used as clonality markers for MRD monitoring post-therapy. Comparison of NGS and NGF for MRD monitoring showed 79.1% concordance. No samples that tested MRD-positive via NGF were found negative via NGS, indicating the higher sensitivity of NGS. MRD could be detected using NGS in 6 of 7 samples before autologous hematopoietic stem-cell transplantation, and 5 of them tested negative post-transplantation. In contrast, the NGF method could detect MRD in only 1 sample pre-transplantation. CONCLUSION: Compared with NGF, NGS exhibits higher sensitivity and reproducibility in MRD detection and can be an effective strategy for MRD monitoring in Chinese MM patients.

Effects of rapamycin combined with low dose prednisone in patients with chronic immune thrombocytopenia.

We conducted this randomized trial to investigate the efficacy and safety of rapamycin treatment in adults with chronic immune thrombocytopenia (ITP). Eighty-eight patients were separated into the control (cyclosporine A plus prednisone) and experimental (rapamycin plus prednisone) groups. The CD4⁺CD25⁺CD127(low) regulatory T (Treg) cells level, Foxp3 mRNA expression, and the relevant cytokines levels were measured before and after treatment. The overall response (OR) was similar in both groups (experimental group versus control group: 58% versus 62%, P = 0.70). However, sustained response (SR) was more pronounced in the experimental group than in the control group (68% versus 39%, P < 0.05). Both groups showed similar incidence of adverse events (7% versus 11%, P = 0.51). As expected, the low pretreatment baseline level of Treg cells was seen in all patients (P < 0.001); however, the experimental group experienced a significant rise in Treg cell level, and there was a strong correlation between the levels of Treg cells and TGF-beta after the treatment. In addition, the upregulation maintained a stable level during the follow-up phase. Thus, rapamycin plus low dose prednisone could provide a new promising option for therapy of ITP.

[The variation and clinical significance of paroxysmal nocturnal hemoglobinuria clone in patients with aplastic anemia before and after immunosuppressive therapy].

OBJECTIVE: To evaluate the evolution of paroxysmal nocturnal hemoglobinuria (PNH) clone and its clinical significance before and after immunosuppressive therapy (IST) in patients with aplastic anemia (AA). METHODS: A total of 186 patients diagnosed as AA were enrolled in this study. Among them, 55 patients were diagnosed as severe AA (SAA) and treated with cyclosporine (CsA) plus anti-thymocyte globulin (ATG), 131 were diagnosed as non SAA (NSAA) and treated with CsA alone. All patients were screened for PNH clone by flow cytometry before treatment and followed up for 18-76 months, with a median time of 22 months. RESULTS: Positive PNH clones were detected in 10 SAA (18.9%) patients, significantly more than that of NSAA group [9 patients (7.4%), t = 5.041, P = 0.025]. The proportions of PNH clones in SAA group at 6, 12, 24 and > 24 months were 13.38%, 14.88%, 20.00% and 18.85%, respectively, also significantly higher than those of NSAA patients (5.67%, 5.31%, 5.47% and 9.08%, all P values < 0.05). Clinical response rates were comparable in both ATG+CsA or CsA alone groups no matter PNH clone was positive or negative. CONCLUSIONS: PNH clone are detectable in AA patients either treated with ATG plus CsA or CsA alone, and more significant by ATG plus CsA. Whether PNH clone occurred before or after IST does not affect the therapeutic efficacy.

[Changes of transcription factors Bcl-6, Foxp3 and RORγt in CD4(+) cells in bone marrow of immuno-related hematocytopenia].

OBJECTIVE: To evaluate the possible mechanism of transcription factors B cell lymphoma 6 (Bcl-6) , forkhead/winged helix transcription factor 3 (Foxp3) and retinoic acid related orphan receptor (RORγt) in CD4(+) T cells for immuno-related hematocytopenia (IRH). METHODS: CD4(+) T cells were harvested from 40 IRH patients, 38 aplastic anemia subjects and 25 normal controls and separated by magnetic activated cell sorting (MACS). Then the expressions of transcription factors of Foxp3, RORγ and Bcl-6 in CD4(+) T cells were measured by real time fluorescent quantitative-polymerase chain reaction (QRT-PCR). RESULTS: Auto-antibody was detected on CD34(+) cells (67.5% (27/40) ), CD15(+) cells (65.0% (26/40)), GlyA(+) cells (75.0% (30/40) ), auto-antibody involving three, two or one myeloid cell were detected in 27.5% (11/40), 52.5% (21/40), 20.0% (8/40) of IRH patients. Compensatory increase of Foxp3 mRNA was found in IRH (0.124 (0.073-0.198) vs 0.071 (0.046-0.118), P < 0.05). The expression of Bcl-6 was higher (2.243 (0.854-4.544) vs 1.211 (0.131-2.816), P < 0.05). Compared to aplastic anemia, the expression of RORγt was lower in IRH (0.133 (0.068-0.189) vs 0.290 (0.138-0.480), P < 0.01) and the ratio of Treg/Th17 shifted to Th17 in patients with aplastic anemia (Foxp3/RORγt ratio,0.500 (0.240-0.795) vs 0.975 (0.483-1.416), P < 0.01). CONCLUSION: As one kind of bone marrow failures caused by autoantibody to bone marrow cells, IRH may occur due to a high expression of Bcl-6 in CD4(+) T cells, its immunopathogenesis is different from that of aplastic anemia.

Platelet activation is upregulated in cirrhotic patients with portal vein thrombosis.

OBJECTIVE: The alteration of platelet function plays a key role in thrombosis formation. This study aimed to explore platelet function alterations in the formation of portal vein thrombosis (PVT) in cirrhosis. METHODS: Cirrhotic patients admitted to hospital between October 2021 and April 2023 were recruited and divided into PVT and non-PVT groups. Flow cytometry was used to detect the expression of CD62p, CD63, monocyte-platelet aggregates (MPAs), neutrophil-platelet aggregates (NPAs) and vWF-Ag to evaluate platelet activation and adhesion function. RESULTS: A total of 145 subjects were enrolled in our study including 60 cirrhotic PVT patients, 60 cirrhotic non-PVT patients and 25 healthy volunteers. The expression of CD41+CD62p+ and CD41+CD63+ platelets in the PVT group was significantly elevated compared with that in the non-PVT group(P＜0.05). Subgroup analysis showed that the mean fluorescent intensity (MFI) of CD62p and CD63 was associated with portal hypertension-related complications (P＜0.05), and CD63 MFI was significantly associated with thrombosis burden (P=0.019). CD41+CD62p+ and CD41+CD63+ platelets as well as MPAs and NPAs were highly expressed in the splenectomy group (P＜0.05). Positive correlations were found between CD62p and CD63 MFI, MPAs and NPAs(P＜0.05). In addition, platelet counts were also correlated with MPAs (r=0.556, P＜0.001) and NPAs (r=0.467, P＜0.001). Cirrhotic patients with PVT had higher mortality and were more likely to experience portal hypertension-related complications (P＜0.05). CONCLUSION: Highly activated platelet function exists in patients with cirrhosis, and platelet activation was elevated during PVT formation, suggesting that activated platelets may participate in the formation of PVT in patients with cirrhosis.

Human corin isoforms with different cytoplasmic tails that alter cell surface targeting.

Corin is a cardiac serine protease that activates natriuretic peptides. It consists of an N-terminal cytoplasmic tail, a transmembrane domain, and an extracellular region with a C-terminal trypsin-like protease domain. The transmembrane domain anchors corin on the surface of cardiomyocytes. To date, the function of the corin cytoplasmic tail remains unknown. By examining the difference between human and mouse corin cytoplasmic tails, analyzing their gene sequences, and verifying mRNA expression in hearts, we show that both human and mouse corin genes have alternative exons encoding different cytoplasmic tails. Human corin isoforms E1 and E1a have 45 and 15 amino acids, respectively, in their cytoplasmic tails. In transfected HEK 293 cells and HL-1 cardiomyocytes, corin isoforms E1 and E1a were expressed at similar levels. Compared with isoform E1a, however, isoform E1 was more active in processing natriuretic peptides. By cell surface labeling, glycosidase digestion, Western blotting, and flow cytometry, we found that corin isoform E1 was activated more readily as a result of more efficient cell surface targeting. By mutagenesis, we identified a DDNN motif in the cytoplasmic tail of isoform E1 (which is absent in isoform E1a) that promotes corin surface targeting in both HEK 293 and HL-1 cells. Our data indicate that the sequence in the cytoplasmic tail plays an important role in corin cell surface targeting and zymogen activation.

Clinical, immunophenotypic, cytogenetic, and molecular genetic features in 117 adult patients with mixed-phenotype acute leukemia defined by WHO-2008 classification.

Among 4,780 consecutive adult acute lymphoblastic/myeloblastic leukemia patients, we identified 117 (2.4%) patients with mixed-phenotype acute leukemia fulfilling WHO 2008 criteria; these were classified as: Blymphoid+ myeloid (n=64), T-lymphoid+myeloid (n=38), B+T-lymphoid (n=14) and trilineage (n=1). Of 92 patients karyotyped, 59 were abnormal and were classified as: complex (22 of 92), t(9;22)(q34;q11) (14 of 92), monosomy 7 (7 of 92), polysomy 21 (7 of 92), t(v;11q23) (4 of 92), t(10;11)(p15;q21) (3 of 92), while STIL-TAL1 fusion was detected in one (T+My) patient. After investigating common acute leukemia-related mutations in 17 genes, 12 of 31 (39%) patients were found to have at least one mutation, classified with: IKZF1 deletion (4 of 31), and EZH2 (3 of 31), ASXL1 (3 of 31), ETV6 (2 of 31), NOTCH1 (1 of 31), and TET2 (1 of 31) mutations. Array-CGH revealed genomic deletions of CDKN2A (4 of 12), IKZF1 (3 of 12), MEF2C (2 of 12), BTG1 (2 of 12), together with BCOR, EBF1, K-RAS, LEF1, MBNL1, PBX3, and RUNX1 (one of 12 each). Our results indicate that mixed-phenotype acute leukemia is a complex entity with heterogeneous clinical, immunophenotypic, cytogenetic, and molecular genetic features.

Case report: CD38-directed CAR-T cell therapy: A novel immunotherapy targeting CD38- positive blasts overcomes TKI and chemotherapy resistance of myeloid chronic myeloid leukemia in blastic phase.

Resistance to tyrosine kinase inhibitor (TKI) is a tough problem in the treatment of chronic myeloid leukemia in blastic phase (CML-BP), which was often associated with acquired mutations in the kinase domain and not eliminating the leukemic stem cells. The efficacy of TKI or combination with chemotherapy in CML-BP remains unsatisfactory. Chimeric antigen receptor T (CAR-T) cell immunotherapy may overcome TKI and chemotherapy resistance. However, lack of ideal targetable antigens is a major obstacle for treating patients with myeloid malignancies. CD38 is known to be expressed on most (acute myeloid leukemia) AML cells, and its lack of expression on hematopoietic stem cells renders it as a potential therapeutic target for myeloid CML-BP. We develop a CD38-directed CAR-T cell therapy for AML, and two patients with myeloid CML-BP were enrolled (NCT04351022). Two patients, harboring E255K and T315I mutation in the ABL kinase domain, respectively, were resistant to multiple TKIs (imatinib, dasatinib, nilotinib, and ponatinib) and intensive chemotherapy. The blasts in the bone marrow of two patients exhibited high expression of CD38. After tumor reduction chemotherapy and lymphodepletion chemotherapy, 1 × 107 CAR-T-38 cells per kilogram of body weight were administered. They achieved minimal residual disease-negative and BCR::ABL1-negative complete remission and experienced grade II cytokine release syndrome manifesting as fever. Our data highlighted that CAR-T-38 cell therapy may overcome TKI and chemotherapy resistance in patients with myeloid CML-BP.

Successful application of PD-1 knockdown CLL-1 CAR-T therapy in two AML patients with post-transplant relapse and failure of anti-CD38 CAR-T cell treatment.

Patients with relapsed/refractory acute myeloid leukemia (R/R AML) often show resistance to chemotherapy and have dismal outcomes. Therefore, it is urgent to develop new treatment strategies to address this problem. With tremendous achievement of chimeric antigen receptor T cells (CAR-T) therapy against B-cell malignancies, many efforts have been devoted to developing CAR-T therapy for R/R AML but with limited success, in part owing to a lack of specific targets. C-type lectin-like molecule-1 (CLL-1) is highly expressed on AML blasts with no expression on normal hematopoietic stem cells, which makes it an ideal target of immunotherapy for AML. Here, we report 2 R/R AML patients who relapsed after allogeneic stem cell transplantation and failed multiline salvage therapies including anti-CD38 CAR-T therapy, but were successfully treated with PD-1 silenced anti-CLL-1 CAR-T therapy. Both patients achieved molecular complete remission with incomplete hematologic recovery at 28 days of evaluation after CLL-1 CAR-T cell infusion. Cytokine release syndrome in cases 1 and 2 were grade 1 and 2, respectively. At the last follow-up, cases 1 and 2 had maintained continuous remission for 8 and 3 months, respectively. Our results demonstrated that CLL-1 CAR-T cells might be an effective and safe salvage therapy for AML patients with posttransplant relapse.