The orally bioavailable GSPT1/2 degrader SJ6986 exhibits in vivo efficacy in acute lymphoblastic leukemia.

Advancing cure rates for high-risk acute lymphoblastic leukemia (ALL) has been limited by the lack of agents that effectively kill leukemic cells, sparing normal hematopoietic tissue. Molecular glues direct the ubiquitin ligase cellular machinery to target neosubstrates for protein degradation. We developed a novel cereblon modulator, SJ6986, that exhibits potent and selective degradation of GSPT1 and GSPT2 and cytotoxic activity against childhood cancer cell lines. Here, we report in vitro and in vivo testing of the activity of this agent in a panel of ALL cell lines and xenografts. SJ6986 exhibited similar cytotoxicity to the previously described GSPT1 degrader CC-90009 in a panel of leukemia cell lines in vitro, resulting in apoptosis and perturbation of cell cycle progression. SJ6986 was more effective than CC-90009 in suppressing leukemic cell growth in vivo, partly attributable to favorable pharmacokinetic properties, and did not significantly impair differentiation of human CD34+ cells ex vivo. Genome-wide CRISPR/Cas9 screening of ALL cell lines treated with SJ6986 confirmed that components of the CRL4CRBN complex, associated adaptors, regulators, and effectors were integral in mediating the action of SJ6986. SJ6986 is a potent, selective, orally bioavailable GSPT1/2 degrader that shows broad antileukemic activity and has potential for clinical development.

Acquired copy number amplification at the MYC enhancer in human B-precursor acute lymphoblastic leukemia cell lines.

Our study highlights the discovery of recurrent copy number alterations in noncoding regions, specifically blood enhancer cluster (BENC-CNA), in B-precursor acute lymphoblastic leukemia (BCP-ALL) cell lines. We demonstrate that BENC-CNA acts as a super-enhancer, driving MYC expression and possibly contributing to the immortalization and proliferative advantage of BCP-ALL cells in vitro.

Outcomes in children with first-relapsed acute lymphoblastic leukemia in Japan: Results from JCCG Study JPLSG-ALL-R08.

BACKGROUND/OBJECTIVES: The Berlin-Frankfurt-Münster (BFM)-S classification is a crucial prognostic indicator in children experiencing first-relapsed acute lymphoblastic leukemia (ALL). Early molecular response to therapy, evaluated by measurable/minimal residual disease (MRD), has a significant impact on the survival of patients with childhood ALL. Applying risk stratification based on the BFM-S classification and MRD response after induction, the first nationwide prospective multicenter study, ALL-R08, was conducted in children with first-relapsed ALL in Japan. METHODS: The ALL-R08 study comprised two parts: ALL-R08-I, an observational study aimed at obtaining an overall picture of outcomes in first-relapsed childhood ALL, and ALL-R08-II, a clinical trial for the non-T-ALL S2 risk group. In ALL-R08-II, patients with an MRD level of ≥10-3 at the end of induction therapy were assigned to undergo allogeneic hematopoietic stem cell transplantation (allo-HCT), whereas those with an MRD level less than 10-3 and isolated extramedullary relapse continued to receive chemotherapy. RESULTS: In total, 163 patients were enrolled in the ALL-R08 study, and 82 and 81 patients were enrolled in the ALL-R08-I and the ALL-R08-II, respectively. In ALL-R08-I, the probability of 3-year event-free survival (EFS) for patients with S1, S2, S3, S4, and post-HCT groups was 83% ± 15%, 37% ± 11%, 28% ± 8%, 14% ± 7%, and 0%, respectively. In the ALL-R08-II trial, 3-year EFS in patients with post-induction MRD less than 10-3 and ≥10-3 was 70% ± 9% (n = 27) and 68% ± 8% (n = 31) (p = .591), respectively. CONCLUSIONS: ALL-REZ BFM-type treatment is equally effective for children with first-relapsed ALL treated according to the Japanese frontline protocols and for children with first-relapsed ALL treated according to the BFM-type frontline protocols.

Metronomic Chemotherapy for Pediatric Refractory Solid Tumors: A Retrospective Single-center Study.

Metronomic chemotherapy (MC) is based on chronic administration of chemotherapeutic agents at minimally toxic doses without prolonged drug-free breaks, that inhibits tumor angiogenesis and induces tumor dormancy. This study aimed to determine the efficacy of MC for pediatric refractory solid tumors. We retrospectively analyzed the data of pediatric patients with relapsed/refractory solid tumors who received treatment, including low-dose continuous administration of anticancer drugs, at our institute. Of the 18 patients, the disease statuses at the initiation of MC were complete remission (n=2), partial remission/stable disease (n=5), and progressive disease (n=11). The overall survival rate was 61% at 12 months and 34% at 24 months, and the progression-free survival rate was 21% at 12 and 24 months. Although only 5 of the 18 patients showed certain tumor regression or maintained remission, tumors that stabilized, maintained remission/stable disease, and showed certain advantages in terms of overall survival rate, even if limited to progressive disease. Approximately half of the patients demonstrated temporal tumor stabilization and improved survival time. Overall, previous reports and the present study support the conclusion that MC has the potential to play an important role in pediatric cancer treatment during the advanced stage.

Rare TCF3 variants associated with pediatric B cell acute lymphoblastic leukemia.

Germline genetic variants influence development of pediatric B cell acute lymphoblastic leukemia (B-ALL). Genome-wide association studies (GWAS) have identified several pediatric B-ALL susceptibility loci. IKZF1 and PAX5, transcription factors involved in B cell development, have been reported as susceptibility genes for B-ALL development. Therefore, we hypothesized that rare variants of genes involved in B cell development would be candidate susceptibility loci for pediatric B-ALL. Thus, we sequenced TCF3, a key transcription factor gene involving in B cell development. Saliva DNA from 527 pediatric patients with pediatric B-ALL in remission who were registered with the Tokyo Children's Cancer Study Group (TCCSG) were examined. As a TCF3 gene-based evaluation, the numbers of rare deleterious germline TCF3 sequence variants in patients with pediatric B-ALL were compared with those in cancer-free individuals using data in public databases. As a TCF3 single-variant evaluation, the frequencies of rare deleterious germline TCF3 sequence variants in patients with pediatric B-ALL were also compared with those in control data. TCF3 gene-based analysis revealed significant associations between rare deleterious variants and pediatric B-ALL development. In addition, TCF3 variant-based analysis showed particularly strong association between variant rs372168347 (three in 521 TCCSG and three in the 15780 gnomAD whole genome analysis cohort, p = 0.0006) and pediatric B-ALL development. TCF3 variants are known to influence B cell maturation and may increase the risk of preleukemic clone emergence.

[Experience with an international study on standard-risk relapsed childhood acute lymphoblastic leukemia (IntReALL SR 2010 study) in Japan].

Many effective new agents for relapsed childhood acute lymphoblastic leukemia (ALL) are now becoming available, and international standard chemotherapy should be developed to optimize use of these agents. Randomized controlled trials (RCTs) are needed to establish a standard treatment, but few have been conducted for relapsed childhood ALL in Japan due to the small patient population. Participation in international RCTs is necessary to access sufficient patients for informative study results, but differences in approved drugs and healthcare systems between countries make this challenging. In 2014, the Japanese Pediatric Leukemia/Lymphoma Study Group (JPLSG) participated in an international study on standard-risk relapsed childhood ALL (IntReALL SR 2010) involving two RCTs and multiple drugs not approved in Japan, which was addressed by replacing the unapproved drugs with alternative approved drugs with the same or similar efficacy. This article discusses the historical background of treatment development for relapsed childhood ALL, our experience in participating in the IntReALL SR 2010 trial, and prospects for treating relapsed childhood ALL.

Tumor-intrinsic and -extrinsic determinants of response to blinatumomab in adults with B-ALL.

Blinatumomab, a bispecific antibody that directs CD3+ T cells to CD19+ tumor cells, shows variable efficacy in B-progenitor acute lymphoblastic leukemia (B-ALL). To determine tumor-intrinsic and -extrinsic determinants of response, we studied 44 adults with relapsed or refractory B-ALL (including 2 minimal residual disease positive) treated with blinatumomab using bulk tumor and single-cell sequencing. The overall response rate in patients with hematological disease was 55%, with a high response rate in those with CRLF2-rearranged Philadelphia chromosome-like ALL (12 [75%] of 16). Pretreatment samples of responders exhibited a tumor-intrinsic transcriptomic signature of heightened immune response. Multiple mechanisms resulted in loss of CD19 expression, including CD19 mutations, CD19-mutant allele-specific expression, low CD19 RNA expression, and mutations in CD19 signaling complex member CD81. Patients with low hypodiploid ALL were prone to CD19- relapse resulting from aneuploidy-mediated loss of the nonmutated CD19 allele. Increased expression of a CD19 isoform with intraexonic splicing of exon 2, CD19 ex2part, at baseline or during therapy was associated with treatment failure. These analyses demonstrate both tumor-intrinsic and -extrinsic factors influence blinatumomab response. We show that CD19 mutations are commonly detected in CD19- relapse during blinatumomab treatment. Identification of the CD19 ex2part splice variant represents a new biomarker predictive of blinatumomab therapy failure.

Degradation of Janus kinases in CRLF2-rearranged acute lymphoblastic leukemia.

CRLF2-rearranged (CRLF2r) acute lymphoblastic leukemia (ALL) accounts for more than half of Philadelphia chromosome-like (Ph-like) ALL and is associated with a poor outcome in children and adults. Overexpression of CRLF2 results in activation of Janus kinase (JAK)-STAT and parallel signaling pathways in experimental models, but existing small molecule inhibitors of JAKs show variable and limited efficacy. Here, we evaluated the efficacy of proteolysis-targeting chimeras (PROTACs) directed against JAKs. Solving the structure of type I JAK inhibitors ruxolitinib and baricitinib bound to the JAK2 tyrosine kinase domain enabled the rational design and optimization of a series of cereblon (CRBN)-directed JAK PROTACs utilizing derivatives of JAK inhibitors, linkers, and CRBN-specific molecular glues. The resulting JAK PROTACs were evaluated for target degradation, and activity was tested in a panel of leukemia/lymphoma cell lines and xenograft models of kinase-driven ALL. Multiple PROTACs were developed that degraded JAKs and potently killed CRLF2r cell lines, the most active of which also degraded the known CRBN neosubstrate GSPT1 and suppressed proliferation of CRLF2r ALL in vivo, e.g. compound 7 (SJ988497). Although dual JAK/GSPT1-degrading PROTACs were the most potent, the development and evaluation of multiple PROTACs in an extended panel of xenografts identified a potent JAK2-degrading, GSPT1-sparing PROTAC that demonstrated efficacy in the majority of kinase-driven xenografts that were otherwise unresponsive to type I JAK inhibitors, e.g. compound 8 (SJ1008030). Together, these data show the potential of JAK-directed protein degradation as a therapeutic approach in JAK-STAT-driven ALL and highlight the interplay of JAK and GSPT1 degradation activity in this context.

BCL6 inhibition ameliorates resistance to ruxolitinib in CRLF2-rearranged acute lymphoblastic leukemia.

Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) is an intractable disease and most cases harbor genetic alterations that activate JAK or ABL signaling. The commonest subtype of Ph-like ALL exhibits a CRLF2 gene rearrangement that brings about JAK1/2-STAT5 pathway activation. However, JAK1/2 inhibition alone is insufficient as a treatment, so combinatorial therapies targeting multiple signals are needed. To better understand the mechanisms underlying the insufficient efficacy of JAK inhibition, we explored gene expression changes upon treatment with a JAK1/2 inhibitor (ruxolitinib) and found that elevated BCL6 expression was one such mechanism. Upregulated BCL6 suppressed the expression of TP53 along with its downstream cell cycle inhibitor p21 (CDKN2A) and pro-apoptotic molecules, such as FAS, TNFRSF10B, BID, BAX, BAK, PUMA, and NOXA, conferring cells some degree of resistance to therapy. BCL6 inhibition (with FX1) alone was able to upregulate TP53 and restore the TP53 expression that ruxolitinib had diminished. In addition, ruxolitinib and FX1 concertedly downregulated MYC. As a result, FX1 treatment alone had growth-inhibitory and apoptosis- sensitizing effects, but the combination of ruxolitinib and FX1 more potently inhibited leukemia cell growth, enhanced apoptosis sensitivity, and prolonged the survival of xenografted mice. These findings provide one mechanism for the insufficiency of JAK inhibition for the treatment of CRLF2-rearranged ALL and indicate BCL6 inhibition as a potentially helpful adjunctive therapy combined with JAK inhibition.

Common Blood Test Indices for Predicting Transient Abnormal Myelopoiesis-Related Mortality in Infants with Down Syndrome.

Transient abnormal myelopoiesis (TAM) can cause early death in children with Down syndrome, and liver failure is the most common cause of death. The aim of this single-center retrospective study was to identify a quantitative index for predicting TAM-related mortality at the time of diagnosis. Of the 462 children with Down syndrome admitted to our hospital from 1992 to 2021, we studied 12 infants with TAM-related death and 31 survivors who were diagnosed with TAM. In the death and survival groups, the median gestational ages were 34.9 and 37.1 weeks, respectively (p = 0.12). At diagnosis, the white blood cell (WBC) counts were 99.2 and 36.2 × 109/L (p = 0.011), the hemoglobin concentrations were 131 and 159 g/L (p = 0.009), and the serum albumin concentrations were 23 and 31 g/L (p < 0.001), respectively. The areas under the receiver operating characteristic curve for the abilities of the WBC count, hemoglobin, and serum albumin at diagnosis to predict survival were 0.75, 0.76, and 0.85, respectively. The serum albumin concentration threshold of 28 g/L at diagnosis had sensitivity of 0.79 and specificity of 0.82. Gestational age and serum albumin concentration were entered into a logistic regression model. The serum albumin concentration was an independent indicator of TAM-related death (adjusted odds ratio, 0.78; 95% confidence interval, 0.65-0.93; p = 0.005). In conclusion, a low serum albumin concentration at diagnosis may be a good predictor of TAM-related death.