CRLF2 overexpression

The utilization of RNA-seq enables the identification of clinically significant genetic abnormalities that may go undetected through conventional detection methods. Its robust analytical performance might bring great application value for clinical diagnosis, prognosis, and therapy in ALL.

High Resolution Melting analysis unveiled concurrent CRLF2 or JAK2 variants in 8.19% of samples, as well as a dynamic nature of CRLF2 alterations during disease progression. Overall, this approach provides an accurate identification of CRLF2 alterations, enabling improved diagnostic and facilitating therapeutic decision-making.

Such heterogeneity was attributed to the different molecular mechanisms leading to CLRF2 upregulation, where the CRLF2 overexpression level was high in Ph-like B-ALL and medium in high hyperdiploid B-ALL. This study highlights the importance of the molecular mechanisms of the upregulation of CRLF2 expression in predicting the prognosis of pediatric B-ALL patients.

Pts received hyper-CVAD alternating with high-dose methotrexate (MTX) and cytarabine (Ara-C) for up to 4 cycles, followed by 4 cycles of blinatumomab at standard doses. Pts with CD20+ disease (≥1% cells) received 8 doses of ofatumumab (2000 mg) or rituximab (375 mg/m2)...One pt discontinued blinatumomab due to a related adverse event (grade 2 encephalopathy and dysphasia). No pts discontinued INO due to toxicity, and no cases of veno-occlusive disease have been observed.

Cell apoptosis assays demonstrated the CRLF2-dependent potency of CRLF2-DM1 LIP in Ph-like ALL cell lines. This study is the first to highlight the therapeutic potential of a CRLF2-directed scFv-Fc-liposomal conjugate for targeting Ph-like ALL.

Patients with DS-ALL exhibit an increased rate of relapse and particularly of treatment-related mortality. Novel, less-toxic therapeutic strategies are needed to improve outcomes.

Furthermore, concomitant CNA of IKZF1 in CRLF2 positive patients was associated with a greater hazard for poor overall and event free survival, compared to patients without these alterations or presence of any one of them. Our findings demonstrate that the surface CRLF2 expression in association with IKZF1 copy number alteration can be used to risk stratify pediatric B-ALL patients.

With this practical approach, we reported a high incidence of BCR::ABL1-like ALLs, and a lower frequency of CRLF2 alteration & associated CGFs. Recognising this entity, early at diagnosis is crucial to optimise personalised treatment strategies.

However, long-term efficacy remains to be determined and it is under evaluation in clinical trials.1,22,23 Preclinical and more recent clinical studies have shown variable activity of JAK inhibitors in JAK-STAT activating ALL17,24 and the need to simultaneously inhibit multiple pathways, including phosphoinositide 3-kinase (PI3K)/mTOR or mitogen-activated protein kinase (MEK)/ receptor tyrosine kinases (e.g. FLT3) to block the growth of leukemic blasts.25–27 Promising alternative approaches to TKIs are represented by antibody-based (e.g. blinatumomab or inozutumab) and cellular (CAR T-cell therapy) immunotherapy which have shown discernable efficacy in BCR::ABL1-like ALL among other subtypes.28–30 Moreover, the efficacy of proteolysis-targeting chimeras (PROTACs) directed against JAK231 or GSPT1/2 degraders32 has been recently reported in preclinical models of CRLF2-rearranged and other JAK-activated BCR::ABL1-like ALL. Conclusions BCR::ABL1-like ALL is driven by numerous targetable kinase fusions whose diagnosis requires the use of comprehensive assays. The incorporation of TKIs and/or different immunotherapies is reversing the historically poor outcome of this ALL subtype.

Notably, ex vivo drug screening revealed sensitivity of IKZF1plus blasts for drugs active against Ph-like ALL such as Birinapant and histone deacetylase inhibitors. We provided data in a large setting of a rare condition (DS-ALL) supporting that these patients, not associated with other high-risk features, need tailored therapeutic strategies.

These findings indicate P2RY8-CRLF2 identifies a subset of patients with specific features and adverse outcomes that could be improved by risk-directed treatment.

In developing countries, detecting Philadelphia (Ph)-like B-lineage acute lymphoblastic leukemia is complicated and challenging due to its diverse genetic landscape. There is no well-defined and cost-effective methodology for its detection. The incidence of this high-risk subtype is very high in adult cases, and there is an urgent need for its accurate detection. We have developed an online PHi-RACE classifier for its rapid detection, followed by delineating the genomic and proteomic landscape of Ph-like acute lymphoblastic leukemias for the first time in Indian patients.

P1/2, N=26, Not yet recruiting, Princess Maxima Center for Pediatric Oncology

Mechanistically, we showed that high doses of TSLP induced loss of its receptor and loss of CRLF2 signals in vitro. These results suggest that high doses of TSLP could be further investigated as a potential therapy for the treatment of CRLF2 B-ALL.

The presence of CRLF2 rearrangements within the context of a complex karyotype is often associated with CRLF2 overexpression and poor prognosis. The heterogeneity of B-ALL and the variability in the outcomes of patients that lack characteristic genetic abnormalities highlight the importance of profiling unusual genetic cases such as this one and continuing research to understand the molecular mechanisms of rarer mutations.

Performing an ex-vivo drug screening with 174 drugs in early to late clinical trials on blasts of 3 IKZF1plus DS-ALL patients and on 14 controls (5 B-cell lymphoblastoid cell lines, 3 PBMCs, 3 T-cells and 3 CD34+ cells, all derived from healthy donors) we observed the efficacy of drugs known to be effective in Ph-like patients such as Birinapant, a SMAC mimetic, and HDAC inhibitors...Ph-like signature and IKZF1 deletions were associated with poor outcome, with the risk of relapse further increased for IKZF1plus patients. These alterations characterize subgroups of DS-ALL patients who need tailored therapeutic strategies.

The GIMEMA LAL2317 is a clinical trial designed for newly diagnosed adult B-lineage Ph-negative ALL that includes two cycles of blinatumomab in the consolidation phase...1. Chiaretti S et al., BJH 2018

We demonstrate the use of a single-tube multimodal NGS assay for comprehensive genomics profiling that simultaneously screens DNA and RNA for expression and variants. It is a powerful and cost-effective tool to help classify PhL B-ALL subgroup for clinical management and guiding therapeutic intervention.

Therapy with ruxolitinib led to complete (n = 7) and partial (n = 2) remission, in three individuals no information was found. Based on the limited number of studies describing the efficacy of ruxolitinib as an additional compound administrated with standard ALL therapy, we conclude that this approach can be considered in patients with aberrations activating JAK-STAT pathway.

The identification of the CRLF2 antigen using the MFC, based on the percentage of positivity and MFI values, is a useful tool for predicting JAK2 mutations and CRLF2-r.

Mucositis, infections, and hyperglycemia were significantly more frequent in all patients with DS, while seizures were more frequent in patients with DS on HR trials (4.1% vs 1.7%, p = 0.001).ConclusionsPatients with DS B-ALL exhibit increased rates of treatment toxicity, death during treatment, and relapse. Novel therapeutic strategies are needed to improve these outcomes.

In conclusion, in our cohort, a high occurrence of CRLF2 abnormalities was documented, particularly the P2RY8-CRLF2 rearrangement, which might represent a characteristic of the Mexican population. Targeted therapy to treat this group of patients could improve OS.

Combining miRNA and mRNA expression, DNA copy number, and methylation status into one multidimensional survival analysis, we found a significant association between greater numbers of alterations and poorer overall survival for multiple component genes. While the intricacies of CRL2 complex gene regulation require additional research, it is evident that multiple causes for the deregulation of these genes must be considered in HCC, including non-traditional mechanisms.

We identified that IKZF1 transcripts with intron retention were upregulated in the CRLF2-high subgroup. Here, we delineated the gene expression profile of CRLF2-high T-ALL samples and unravelled the crucial role of PRC2 in CRLF2 regulation in ETP-ALL.

Thus, CRLF2 overexpression results in reduced B cell differentiation and enhanced E2F signaling in Dp16 B-progenitor cells and DS-ALL patient samples. These findings suggest a functional basis for the high frequency of CRLF2-R in DS-ALL as well as a potential therapeutically targetable pathway.

These genetic alterations induce constitutive JAK/STAT and other kinase signaling that may be targetable by the JAK1/2 inhibitor ruxolitinib...These results suggest potential for both anti-ALL efficacy and CRS dampening when JAK inhibition and TSLPRCART are properly time-sequenced and may have broader applicability for toxicity mitigation with other cellular immunotherapies. Studies of CAR T cell immunotherapy and other kinase inhibitors in additional CRLF2 -rearranged and non-rearranged DS-ALL models are ongoing.