Histone Deacetylase Inhibitors Treatment Research Articles

Lineage-specific proteome remodeling of diverse lung cancer cells by targeted epigenetic inhibitors.

Epigenetic inhibitors exhibit powerful antiproliferative and anticancer activities. However, cellular responses to small-molecule epigenetic inhibition are heterogenous and dependent on factors such as the genetic background, metabolic state, and on-/off-target engagement of individual small-molecule drugs. To determine the mechanisms that drive these heterogeneous cellular responses, we quantified chromatin, proteome, and transcriptome remodeling due to histone deacetylase inhibitor (HDACi) -treated cells derived from diverse genetic backgrounds. We utilized high-throughput sample multiplexed proteomics and integrated intelligent data acquisition methods to map proteomes of cancer cell lines in response to HDACi. We determined cell type-specific and ubiquitous cellular responses based on the quantification of 10,621 total proteins. We then established how coordinated remodeling of the proteome, transcriptome and chromatin state of HDACi treated cancer cells revealed convergent (JUN, MAP2K3, CDKN1A) and divergent (CCND3, ASF1B, BRD7) molecular phenotypes. HDACi-regulated proteins differ greatly across cell lines owing to heterogeneous molecular states of these cell lines. Finally, we demonstrated that HDACi treatment drove a highly cell-type specific response that may in part be explained by cell line-specific off-target drug engagement.

Laminin-associated integrins mediate Diffuse Intrinsic Pontine Glioma infiltration and therapy response within a neural assembloid model

Diffuse Intrinsic Pontine Glioma (DIPG) is a highly aggressive and fatal pediatric brain cancer. One pre-requisite for tumor cells to infiltrate is adhesion to extracellular matrix (ECM) components. However, it remains largely unknown which ECM proteins are critical in enabling DIPG adhesion and migration and which integrin receptors mediate these processes. Here, we identify laminin as a key ECM protein that supports robust DIPG cell adhesion and migration. To study DIPG infiltration, we developed a DIPG-neural assembloid model, which is composed of a DIPG spheroid fused to a human induced pluripotent stem cell-derived neural organoid. Using this assembloid model, we demonstrate that knockdown of laminin-associated integrins significantly impedes DIPG infiltration. Moreover, laminin-associated integrin knockdown improves DIPG response to radiation and HDAC inhibitor treatment within the DIPG-neural assembloids. These findings reveal the critical role of laminin-associated integrins in mediating DIPG progression and drug response. The results also provide evidence that disrupting integrin receptors may offer a novel therapeutic strategy to enhance DIPG treatment outcomes. Finally, these results establish DIPG-neural assembloid models as a powerful tool to study DIPG disease progression and enable drug discovery.

HDAC inhibitors activate lipid peroxidation and ferroptosis in gastric cancer

Gastric cancer remains among the deadliest neoplasms worldwide, with limited therapeutic options. Since efficacies of targeted therapies are unsatisfactory, drugs with broader mechanisms of action rather than a single oncogene inhibition are needed. Preclinical studies have identified histone deacetylases (HDAC) as potential therapeutic targets in gastric cancer. However, the mechanism(s) of action of HDAC inhibitors (HDACi) are only partially understood. This is particularly true with regard to ferroptosis as an emerging concept of cell death.In a panel of gastric cancer cell lines with different molecular characteristics, tumor cell inhibitory effects of different HDACi were studied. Lipid peroxidation levels were measured and proteome analysis was performed for the in-depth characterization of molecular alterations upon HDAC inhibition. HDACi effects on important ferroptosis genes were validated on the mRNA and protein level.Upon HDACi treatment, lipid peroxidation was found increased in all cell lines. Class I HDACi (VK1, entinostat) showed the same toxicity profile as the pan-HDACi vorinostat. Proteome analysis revealed significant and concordant alterations in the expression of proteins related to ferroptosis induction. Key enzymes like ACSL4, POR or SLC7A11 showed distinct alterations in their expression patterns, providing an explanation for the increased lipid peroxidation. Results were also confirmed in primary human gastric cancer tissue cultures as a relevant ex vivo model.We identify the induction of ferroptosis as new mechanism of action of class I HDACi in gastric cancer. Notably, these findings were independent of the genetic background of the cell lines, thus introducing HDAC inhibition as a more general therapeutic principle.

The thiol methyltransferase activity of TMT1A (METTL7A) is conserved across species

Although few resistance mechanisms for histone deacetylase inhibitors (HDACis) have been described, we recently demonstrated that TMT1A (formerly METTL7A) and TMT1B (formerly METTL7B) can mediate resistance to HDACis with a thiol as the zinc-binding group by methylating and inactivating the drug. TMT1A and TMT1B are poorly characterized, and their normal physiological role has yet to be determined. As animal model systems are often used to determine the physiological function of proteins, we investigated whether the ability of these methyltransferases to methylate thiol-based HDACis is conserved across different species. We found that TMT1A was conserved across rats, mice, chickens, and zebrafish, displaying 85.7%, 84.8%, 60.7%, and 51.0% amino acid sequence identity, respectively, with human TMT1A. Because TMT1B was not found in the chicken or zebrafish, we focused our studies on the TMT1A homologs. HEK-293 cells were transfected to express mouse, rat, chicken, or zebrafish homologs of TMT1A and all conferred resistance to the thiol-based HDACIs NCH-51, KD-5170, and romidepsin compared to empty vector-transfected cells. Additionally, all homologs blunted the downstream effects of HDACi treatment such as increased p21 expression, increased acetylated histone H3, and cell cycle arrest. Increased levels of dimethylated romidepsin were also found in the culture medium of cells transfected to express any of the TMT1A homologs after a 24 h incubation with romidepsin compared to empty-vector transfected cells. Our results indicate that the ability of TMT1A to methylate molecules is conserved across species. Animal models may therefore be useful in elucidating the role of these enzymes in humans.

Class I HDAC inhibitors enhance antitumor efficacy and persistence of CAR-T cells by activation of the Wnt pathway

Epigenetic modification shapes differentiation trajectory and regulates the exhaustion state of chimeric antigen receptor T (CAR-T) cells. Limited efficacy induced by terminal exhaustion closely ties with intrinsic transcriptional regulation. However, the comprehensive regulatory mechanisms remain largely elusive. Here, we identify class I histone deacetylase inhibitors (HDACi) as boosters of CAR-T cell function by high-throughput screening of chromatin-modifying drugs, in which M344 and chidamide enhance memory maintenance and resistance to exhaustion of CAR-T cells that induce sustained antitumor efficacy both invitro and invivo. Mechanistically, HDACi decrease HDAC1 expression and enhance H3K27ac activity. Multi-omics analyses from RNA-seq, ATAC-seq, and H3K27ac CUT&Tag-seq show that HDACi upregulate expression of TCF4, LEF1, and CTNNB1, which subsequently activate the canonical Wnt/β-catenin pathway. Collectively, our findings elucidate the functional roles of class I HDACi in enhancing CAR-T cell function, which provides the basis and therapeutic targets for synergic combination of CAR-T cell therapy and HDACi treatment.

The impact of selective HDAC inhibitors on the transcriptome of early mouse embryos

BackgroundHistone acetylation, which is regulated by histone acetyltransferases (HATs) and histone deacetylases (HDACs), plays a crucial role in the control of gene expression. HDAC inhibitors (HDACi) have shown potential in cancer therapy; however, the specific roles of HDACs in early embryos remain unclear. Moreover, although some pan-HDACi have been used to maintain cellular undifferentiated states in early embryos, the specific mechanisms underlying their effects remain unknown. Thus, there remains a significant knowledge gap regarding the application of selective HDACi in early embryos.ResultsTo address this gap, we treated early embryos with two selective HDACi (MGCD0103 and T247). Subsequently, we collected and analyzed their transcriptome data at different developmental stages. Our findings unveiled a significant effect of HDACi treatment during the crucial 2-cell stage of zygotes, leading to a delay in embryonic development after T247 and an arrest at 2-cell stage after MGCD0103 administration. Furthermore, we elucidated the regulatory targets underlying this arrested embryonic development, which pinpointed the G2/M phase as the potential period of embryonic development arrest caused by MGCD0103. Moreover, our investigation provided a comprehensive profile of the biological processes that are affected by HDACi, with their main effects being predominantly localized in four aspects of zygotic gene activation (ZGA): RNA splicing, cell cycle regulation, autophagy, and transcription factor regulation. By exploring the transcriptional regulation and epigenetic features of the genes affected by HDACi, we made inferences regarding the potential main pathways via which HDACs affect gene expression in early embryos. Notably, Hdac7 exhibited a distinct response, highlighting its potential as a key player in early embryonic development.ConclusionsOur study conducted a comprehensive analysis of the effects of HDACi on early embryonic development at the transcriptional level. The results demonstrated that HDACi significantly affected ZGA in embryos, elucidated the distinct actions of various selective HDACi, and identified specific biological pathways and mechanisms via which these inhibitors modulated early embryonic development.

Altered histone acetylation patterns in pancreatic cancer cell lines induce subtype‑specific transcriptomic and phenotypical changes.

Pancreatic ductal adenocarcinoma (PDAC) is often diagnosed at advanced tumor stages with chemotherapy as the only treatment option. Transcriptomic analysis has defined a classical and basal‑like PDAC subtype, which are regulated by epigenetic modification. The present study aimed to determine if drug‑induced epigenetic reprogramming of pancreatic cancer cells affects PDAC subtype identity and chemosensitivity. Classical and basal‑like PDAC cell lines PaTu‑S, Capan‑1, Capan‑2, Colo357, PaTu‑T, PANC‑1 and MIAPaCa‑2, were treated for a short (up to 96h) and long (up to 30weeks) period with histone acetyltransferase (HAT) and histone deacetylase (HDAC) inhibitors. The cells were analyzed using gene expression approaches, immunoblot analysis, and various cell assays to assess cell characteristics, such as proliferation, colony formation, cell migration and sensitivity to chemotherapeutic drugs. Classical and basal‑like PDAC cell lines showed pronounced epigenetic regulation of subtype‑specific genes through acetylation of lysine 27 on Histone H3 (H3K27ac). Moreover, classical cell lines revealed a significantly decreased expression of HDAC2 and increased total levels of H3K27ac in comparison with the basal‑like cell lines. Following HAT inhibitor treatment, classical cell lines exhibited a loss of epithelial marker gene expression, decreased chemotherapy response gene score and increased cell migration invitro, indicating a tumor‑promoting phenotype. HDAC inhibitor treatment, however, exerted minimal reprogramming effects in both subtypes. Epigenetic reprogramming of classical and basal‑like tumor cells did not have a major impact on gemcitabine response, although the gemcitabine transporter gene SLC29A1 (solute carrier family 29 member 1) was epigenetically regulated.

Synergistic integration of histone deacetylase inhibitors apparently enhances the cytokine-induced killer cell efficiency in multiple myeloma via the NKG2D pathway.

The rapid recognition of epigenetic manipulation's potential in restricting cancer cell capabilities spurred translational initiatives, including histone deacetylase inhibitors (HDACis). Clinical trials on multiple myeloma (MM) demonstrated substantial benefits of HDACis, coupled with promising outcomes from cytokine-induced killer cell (CIK) immunotherapy. Intriguingly, the unexplored synergy of HDACis and CIK cell immunotherapy in MM prompted our study. We examined clinically relevant HDACis (panobinostat/LBH589 and romidepsin) alongside CIK cells derived from peripheral blood mononuclear cells across diverse MM cell lines (U266, RPMI8226, OPM-2 and NCI-H929). Utilising various in vitro methodologies, we investigated how HDACis enhance CIK cell lysis of myeloma cells through NKG2D/NKG2D ligand interactions. The results of our analysis indicated several key findings. (1) Enhanced cytotoxicity of CIK cells in MM cells when combined with HDACis. (2) Significant increase in apoptosis, suggesting HDACis and CIK may together enhance apoptotic effects in specific MM cell lines. (3) Elevated IFN-γ secretion and alterations in granzyme Bsecretion because of the independent activity of HDACis. (4) Notably, HDACis increased the expression of MICA/B and ULBP2, crucial for inducing antitumor cytotoxicity of NKTcells. Validation through NKG2D receptor blocking in CIK cells with a purified mouse antihuman NKG2D antibody further supported our findings. Our analyses provide sufficient evidence to consider this clinically forgotten instance (HDACis-CIK cell combination) as a therapeutic priority for MM treatment. Furthermore, we suggest that NKG2D/NKG2D-ligand interactions activating NK/NKTcells may contribute to enhanced myeloma cell lysis in response to HDACis treatment by CIK cells.

Underlying anti-cancer mechanisms of histone deacetylase (HDAC) inhibitors in tamoxifen-resistant breast cancer cells.

Breast cancer is an important women's malignancy with high cancer-related deaths worldwide. Drug resistance lowers the treatment efficacy in this malignancy. This study aimed to explore the underlying mechanisms of histone deacetylase (HDAC) inhibitor trichostatin A (TSA) to overcome resistance to tamoxifen in breast cancer cells. Tamoxifen-resistance in MCF-7 breast cancer cells was simulated. MTT assay was used to detect the cytotoxic effects of HDAC inhibitor and PI3K inhibitor on the cancer cells. Trans-well assay was applied to evaluate the invasion and migration of the treated cancer cells. Flow cytometer assay was also applied to evaluate cell cycle phases in the treated cancer cells. Finally, expression of vascular endothelial growth factor (VEGF), E-cadherin, Vimentin, phosphorylated phosphatidylinositol kinase (p-PI3k), phosphorylated protein kinase B (p-AKT), and phosphorylated mammalian target protein of rapamycin (p-mTOR) was evaluated by western blotting. The obtained results indicated that HDAC inhibitor treatments significantly decreased viability, migration, and invasion in the cancer cells. Furthermore, the frequency of the treated cancer cells significantly increased in the S phase as well as significantly decreasing in the G2/M phase of the cell cycle. Moreover, HDAC inhibitor modified levels of VEGF, E-cadherin, Vimentin, p-PI3k, p-AKT, and p-mTOR proteins. However, HDAC inhibitor combined with PI3K inhibitor exerts more profound effects on the cancer cells as compared to HDAC inhibitor monotherapy. HDAC inhibitors inhibited the survival of breast cancer drug-resistant cells, invasion, migration, and angiogenesis by inhibiting the PI3k/Akt/mTOR signaling pathway.

Epigenetic modifications regulate cultivar-specific root development and metabolic adaptation to nitrogen availability in wheat

The breeding of crops with improved nitrogen use efficiency (NUE) is crucial for sustainable agriculture, but the involvement of epigenetic modifications remains unexplored. Here, we analyze the chromatin landscapes of two wheat cultivars (KN9204 and J411) that differ in NUE under varied nitrogen conditions. The expression of nitrogen metabolism genes is closely linked to variation in histone modification instead of differences in DNA sequence. Epigenetic modifications exhibit clear cultivar-specificity, which likely contributes to distinct agronomic traits. Additionally, low nitrogen (LN) induces H3K27ac and H3K27me3 to significantly enhance root growth in KN9204, while remarkably inducing NRT2 in J411. Evidence from histone deacetylase inhibitor treatment and transgenic plants with loss function of H3K27me3 methyltransferase shows that changes in epigenetic modifications could alter the strategy preference for root development or nitrogen uptake in response to LN. Here, we show the importance of epigenetic regulation in mediating cultivar-specific adaptation to LN in wheat.

HDAC2-miRNA183 Epigenetic Circuit Contributes to the Growth of Philadelphia-Positive Acute Lymphoblastic Leukemia Via BCR-ABL/PTEN/p-AKT and c-MYC Signaling Pathways

Purpose: To characterize the expression and function of leukemia related/specific microRNAs (miRNAs) in Philadelphia-positive Acute Lymphoblastic Leukemia (Ph-positive ALL), and to identify the mechanistic signaling pathways in Ph-positive ALL. Method: Differentially expressed miRNAs in Ph-positive ALL patient samples and cell line were identified using RNA-seq and qPCR. Cell proliferation and apoptosis were detected with CCK-8 and flow cytometry assays, respectively. Bioinformatics software predicted the upstream and downstream regulatory pathway of miRNA183; the dual luciferase reporter gene analysis verified the regulatory effect of miRNA183 on the target gene BCR-ABL; ChIP experiment detected the recruitment of HDACs in the miRNA183 promoter region, and western blot detected the expression of related protein in the miRNA183 related pathway. Results: We identified thirty-one differentially expressed miRNAs in three Ph-positive ALL samples vs. normal karyotype B-ALL samples. Among these alternately expressed miRNAs, we found miRNA183 was significantly downregulated in Ph-positive ALL. qPCR verified the down-expression of miRNA183 in Ph-positive ALLcasesamples and Ph-positive ALL cell line SUP-B15. Forced over-expression of miRNA183 inhibited proliferation of SUP-B15 cells, but had no effect on NALM-6 cells (normal karyotype B-ALL). Furthermore, either forced over-expression of miRNA183 or silence down of BCR-ABL1 promoted SUP-B15 cell apoptosis.TargetScan software prediction found that miRNA183 and BCR-ABL (e1a2) had potential binding sites, dual luciferase reporter gene experiments further proved that miRNA183 could direct bind to BCR-ABL gene; qPCR and western blot further showed that miRNA183 inhibited the expression of P190. Our results showed that BCR-ABL (e1a2) is the target gene of miRNA183. In addition, forced over-expression of miRNA183 and silence down of BCR-ABL1 regulated the downstream proteins including PTEN, PARP, p-AKT and c-MYC with the same change trend. In complementary loss-of-function studies, inhibition of miRNA183 restored the expression of these proteins.Treatment of SUP-B15 cells with decitabine (DAC) and histone deacetylase inhibitor (HDACi), respectively, resulted in decrease of cell viability in a dose-dependent manner. Surprisingly, miRNA183 levels were significantly increased in HDACi exposure but not in DAC exposure. Furthermore, miRNA183 inhibitor reduced HDACi-induced upregulation of miRNA183 expression and attenuated HDACi-mediated apoptosis in SUP-B15 cells. Biology software prediction showed the presence of a transcription start site (TSS) 5 kb upstream of miRNA183. ChIP assay further demonstrated that HDAC2 but not HDAC1 was specifically recruited to the miRNA183 promoter region. Additionally, after HDACi treatment, HDACi-induced H3K27ac was recruited and accumulated at the miRNA183 promoter region. Western blotassay further proved the changes of the related proteins. Conclusion :Our results indicate that miRNA183 is a potential biomarker for Ph-positive ALL and suggest that a novel “HDAC2-miRNA183 Epigenetic Circuit” may be involved in the growth and progression of Ph-positive ALL by PTEN/p-AKT and c-MYC signaling pathways. Taken together, this study may provide novel approaches to the design of targeted drugs for Ph-positive ALL.

Applying HDACis to increase SSTR2 expression and radiolabeled DOTA-TATE uptake: from cells to mice

AimsThe aim of our study was to determine the effect of histone deacetylase (HDAC) inhibitors (HDACis) on somatostatin type-2 receptor (SSTR2) expression and [111In]In-/[177Lu]Lu-DOTA-TATE uptake in vitro and in vivo. Materials and methodsThe human cell lines NCI-H69 (small-cell lung carcinoma) and BON-1 (pancreatic neuroendocrine tumor) were treated with HDACis (i.e. entinostat, mocetinostat (MOC), LMK-235, CI-994 or panobinostat (PAN)), and SSTR2 mRNA expression levels and [111In]In-DOTA-TATE uptake were measured. Furthermore, vehicle- and HDACi-treated NCI-H69 and BON-1 tumor-bearing mice were injected with radiolabeled DOTA-TATE followed by biodistribution studies. Additionally, SSTR2 and HDAC mRNA expression of xenografts, and of NCI-H69, BON-1, NCI-H727 (human pulmonary carcinoid) and GOT1 (human midgut neuroendocrine tumor) cells were determined. Key findingsHDACi treatment resulted in the desired effects in vitro. However, no significant increase in tumoral DOTA-TATE uptake was observed after HDACi treatment in NCI-H69 tumor-bearing animals, whereas tumoral SSTR2 mRNA and/or protein expression levels were significantly upregulated after treatment with MOC, CI-994 and PAN, i.e. a maximum of 2.1- and 1.3-fold, respectively. Analysis of PAN-treated BON-1 xenografts solely demonstrated increased SSTR2 mRNA expression levels. Comparison of HDACs and SSTR2 expression in BON-1 and NCI-H69 xenografts showed a significantly higher expression of 6/11 HDACs in BON-1 xenografts. Of these HDACs, a significant inverse correlation was found between HDAC3 and SSTR2 expression (Pearson r = −0.92) in the studied cell lines. SignificanceTo conclude, tumoral uptake levels of radiolabeled DOTA-TATE were not enhanced after HDACi treatment in vivo, but, depending on the applied inhibitor, increased SSTR2 expression levels were observed.

Peripheral blood DNA methylation and neuroanatomical responses to HDACi treatment that rescues neurological deficits in a Kabuki syndrome mouse model

BackgroundRecent findings from studies of mouse models of Mendelian disorders of epigenetic machinery strongly support the potential for postnatal therapies to improve neurobehavioral and cognitive deficits. As several of these therapies move into human clinical trials, the search for biomarkers of treatment efficacy is a priority. A potential postnatal treatment of Kabuki syndrome type 1 (KS1), caused by pathogenic variants in KMT2D encoding a histone-lysine methyltransferase, has emerged using a mouse model of KS1 (Kmt2d+/βGeo). In this mouse model, hippocampal memory deficits are ameliorated following treatment with the histone deacetylase inhibitor (HDACi), AR-42. Here, we investigate the effect of both Kmt2d+/βGeo genotype and AR-42 treatment on neuroanatomy and on DNA methylation (DNAm) in peripheral blood. While peripheral blood may not be considered a “primary tissue” with respect to understanding the pathophysiology of neurodevelopmental disorders, it has the potential to serve as an accessible biomarker of disease- and treatment-related changes in the brain.MethodsHalf of the KS1 and wildtype mice were treated with 14 days of AR-42. Following treatment, fixed brain samples were imaged using MRI to calculate regional volumes. Blood was assayed for genome-wide DNAm at over 285,000 CpG sites using the Illumina Infinium Mouse Methylation array. DNAm patterns and brain volumes were analyzed in the four groups of animals: wildtype untreated, wildtype AR-42 treated, KS1 untreated and KS1 AR-42 treated.ResultsWe defined a DNAm signature in the blood of KS1 mice, that overlapped with the human KS1 DNAm signature. We also found a striking 10% decrease in total brain volume in untreated KS1 mice compared to untreated wildtype, which correlated with DNAm levels in a subset KS1 signature sites, suggesting that disease severity may be reflected in blood DNAm. Treatment with AR-42 ameliorated DNAm aberrations in KS1 mice at a small number of signature sites.ConclusionsAs this treatment impacts both neurological deficits and blood DNAm in mice, future KS clinical trials in humans could be used to assess blood DNAm as an early biomarker of therapeutic efficacy.

Targeting the Clear Cell Sarcoma Oncogenic Driver Fusion Gene EWSR1::ATF1 by HDAC Inhibition.

This study reveals the epigenetic and transcriptional suppression mechanism of the fusion oncogene EWSR1::ATF1 in clear cell sarcoma by histone deacetylase inhibitor treatment as well as identifying SOX10 as a transcription factor that regulates EWSR1::ATF1 expression.

In the quest for histone deacetylase inhibitors: current trends in the application of multilayered computational methods.

Histone deacetylase (HDAC) inhibitors have gained attention over the past three decades because of their potential in the treatment of different diseases including various forms of cancers, neurodegenerative disorders, autoimmune, inflammatory diseases, and other metabolic disorders. To date, 5 HDAC inhibitor drugs are marketed for the treatment of hematological malignancies and several drug-candidate HDAC inhibitors are at different stages of clinical trials. However, due to the toxic side effects of these drugs resulting from the lack of target selectivity, active studies are ongoing to design and develop either class-selective or isoform-selective inhibitors. Computational methods have aided the discovery of HDAC inhibitors with the desired potency and/or selectivity. These methods include ligand-based approaches such as scaffold hopping, pharmacophore modeling, three-dimensional quantitative structure-activity relationships (3D-QSAR); and structure-based virtual screening (molecular docking). The current trends involve the application of the combination of these methods and incorporating molecular dynamics simulations coupled with Poisson-Boltzmann/molecular mechanics generalized Born surface area (MM-PBSA/MM-GBSA) to improve the prediction of ligand binding affinity.This review aimed at understanding the current trends in applying these multilayered strategies and their contribution to the design/identification of HDAC inhibitors.

GOLIDOCITINIB IN TREATING REFRACTORY OR RELAPSED PERIPHERAL T‐CELL LYMPHOMA: PRIMARY ANALYSIS OF THE MULTINATIONAL PIVOTAL STUDY RESULTS (JACKPOT8)

Introduction: Currently there is no consensus on the treatment of relapsed/refractory (r/r) PTCL, and patient prognosis was poor. Golidocitinib is an orally available, JAK1-selective inhibitor currently being evaluated in a multinational, pivotal study in r/r PTCL (JACKPOT8 Part B, NCT04105010). Here we reported the primary analysis of efficacy and safety results of this study. Methods: PTCL patients who had relapsed from or had been refractory/intolerant to ≥1 (but ≤3) prior systemic therapy(ies) were enrolled to receive golidocitinib at 150 mg once daily until disease progression or pre-defined discontinuation criteria were met. The primary endpoint was CT-based objective response rate (ORR) assessed by an independent review committee (IRC) according to Lugano 2014 classification. The efficacy analysis set included patients whose pathological diagnosis of PTCL had been retrospectively confirmed by a central laboratory and who had at least one measurable lesions at baseline. The safety analysis set included all dosed patients. Results: As of 30 November 2022, a total of 104 patients with r/r PTCL were enrolled and dosed with golidocitinib. Baseline characteristics: median age was 58 yrs; 64.4% were male; 20.2% had baseline bone marrow involvement. Major subtypes included NOS (46.2%), AITL (15.4%) and ALCL (9.6%). The median prior lines of therapies were two. All of the patients had been treated with chemotherapies, and 48.1% had been treated with histone deacetylase inhibitors. By the data cut-off (DCO) date, a total of 80 patients had both IRC assessment and central pathology review results available, and thus were included in the efficacy analysis. Among them, 35 patients achieved tumor response (ORR = 43.8%), including 20 patients (25.0%) achieved complete responses. Tumor response was observed in various subtypes, including AITL (56.3%), NOS (45.7%), ALCL (11.1%) and others (44.4%). In patients who relapsed from HDAC inhibitor treatment, 54.8% achieved tumor response. With a median follow-up of 5.5 months for responders, the median duration of response (DoR) has not been reached. Golidocitinib was tolerated in patients with r/r PTCL. By the DCO date, the longest duration on treatment was 15.7 months (still responding). The most common ≥ grade 3 treatment-related adverse events (TRAEs) were hematological in nature, including neutrophil count decreased (26.0%), white blood cell count decreased (25.0%), platelet count decreased (16.3%) and lymphocyte count decreased (16.3%). TRAEs leading to dose interruption, reduction, and discontinuation were 37.5%, 5.8%, and 7.7%, respectively. The majority of TRAEs were reversible or clinically manageable. Conclusions: Golidocitinib demonstrated its potential as a novel targeted therapy for the treatment of r/r PTCL. The updated data will be presented at the conference. Encore Abstract—previously submitted to ASCO 2023 Keywords: aggressive T-cell non-Hodgkin lymphoma, molecular targeted therapies Conflicts of interests pertinent to the abstract W. Yu Employment or leadership position: Employed by Dizal Pharmaceutical

Golidocitinib in treating refractory or relapsed peripheral T-cell lymphoma: Primary analysis of the multinational pivotal study results (JACKPOT8).

7503 Background: Currently there is no consensus on the treatment of relapsed/refractory (r/r) PTCL, and patient prognosis was poor. Golidocitinib is an orally available, JAK1-selective inhibitor currently being evaluated in a multinational, pivotal study in r/r PTCL (JACKPOT8 Part B, NCT04105010). Here we reported the primary analysis of efficacy and safety results of this study. Methods: PTCL patients who had relapsed from or had been refractory/intolerant to ≥ 1 (but ≤ 3) prior systemic therapy(ies) were enrolled to receive golidocitinib at 150 mg once daily until disease progression or pre-defined discontinuation criteria were met. The primary endpoint was CT-based objective response rate (ORR) assessed by an independent review committee (IRC) according to Lugano 2014 classification. The efficacy analysis set included patients whose pathological diagnosis of PTCL had been retrospectively confirmed by a central laboratory and who had at least one measurable lesions at baseline. The safety analysis set included all dosed patients. Results: As of November 30, 2022, a total of 104 patients with r/r PTCL were enrolled and dosed with golidocitinib. Baseline characteristics: median age was 58 years old; 64.4% were male; 20.2% had baseline bone marrow involvement. Major subtypes included NOS (46.2%), AITL (15.4%) and ALCL (9.6%). The median prior lines of therapies were two. All of the patients had been treated with chemotherapies, and 48.1% had also been treated with histone deacetylase inhibitors. By the data cut-off (DCO) date, a total of 80 patients had both IRC assessment and central pathology review results available, and thus were included in the efficacy analysis. Among them, 35 patients achieved tumor response (ORR = 43.8%), including 20 patients (25.0%) achieved complete responses. Tumor response was observed in various subtypes, including AITL (56.3%), NOS (45.7%), ALCL (11.1%) and others (44.4%). In patients who relapsed from HDAC inhibitor treatment, 54.8% achieved tumor response. With a median follow-up of 5.5 months for responders, the median duration of response (DoR) has not been reached. Golidocitinib was tolerated in patients with r/r PTCL. By the DCO date, the longest duration on treatment was 15.7 months (still responding). The most common ≥ grade 3 treatment-related adverse events (TRAEs) were hematological in nature, including neutrophil count decreased (26.0%), white blood cell count decreased (25.0%), platelet count decreased (16.3%) and lymphocyte count decreased (16.3%). TRAEs leading to dose interruption, reduction, and discontinuation were 37.5%, 5.8%, and 7.7%, respectively. The majority of TRAEs were reversible or clinically manageable. Conclusions: Golidocitinib demonstrated its potential as a novel targeted therapy for the treatment of r/r PTCL. The updated data will be presented at the conference. Clinical trial information: NCT04105010 .

Class I HDAC inhibitor entinostat synergizes with PLK1 inhibitors in MYC-amplified medulloblastoma cells

PurposeWe and others have demonstrated that MYC-amplified medulloblastoma (MB) cells are susceptible to class I histone deacetylase inhibitor (HDACi) treatment. However, single drug treatment with HDACi has shown limited clinical efficacy. We hypothesized that addition of a second compound acting synergistically with HDACi may enhance efficacy.MethodsWe used a gene expression dataset to identify PLK1 as a second target in MB cells and validated the relevance of PLK1 in MB. We measured cell metabolic activity, viability, and cycle progression in MB cells after treatment with PLK1-specific inhibitors (PLK1i). Chou–Talalay synergy calculations were used to determine the nature of class I HDACi entinostat and PLK1i interaction which was validated. Finally, the clinical potential of the combination was assessed in the in vivo experiment.ResultsMYC-amplified tumor cells are highly sensitive towards treatment with ATP-competitive PLK1i as a monotherapy. Entinostat and PLK1i in combination act synergistically in MYC-driven MB cells, exerting cytotoxic effects at clinically relevant concentrations. The downstream effect is exerted via MYC-related pathways, pointing out the potential of MYC amplification as a clinically feasible predictive biomarker for patient selection. While entinostat significantly extended survival of mice implanted with orthotopic MYC-amplified MB PDX, there was no evidence of the improvement of survival when treating the animals with the combination.ConclusionThe combination of entinostat and PLK1i showed synergistic interaction in vitro, but not in vivo. Therefore, further screening of blood–brain barrier penetrating PLK1i is warranted to determine the true potential of the combination as no on-target activity was observed after PLK1i volasertib treatment in vivo.

Effects of HDAC inhibitors on neuroblastoma SH-SY5Y cell differentiation into mature neurons via the Wnt signaling pathway

Histone deacetylase (HDAC) inhibitors affect cell homeostasis, gene expression, and cell cycle progression and promote cell terminal differentiation or apoptosis. However, the effect of HDAC inhibition on SH-SY5Y cells, which are neuroblastoma cells capable of differentiating into neurons under specific conditions, such as in the presence of retinoic acid (RA), is unknown. In this study, we hypothesized that HDAC inhibitors induced the neuronal differentiation of SH-SY5Y cells. To test this hypothesis, we used phase contrast microscopy, immunocytochemistry (ICC), qPCR, and western blotting analysis. MS-275 and valproic acid (VPA), two HDAC inhibitors, were selected to evaluate neuronal differentiation. It was confirmed that cells treated with MS-275 or VPA differentiated into mature neurons, which were distinguished by bipolar or multipolar morphologies with elongated branches. In addition, the mRNA expression of neuronal markers (Tuj1 and NEFH) and the oligodendrocyte marker (CNP) was significantly increased with MS-275 or VPA treatment compared to that with RA treatment. In addition, the protein expression of the other neuronal markers, Tuj1 and NeuN, was highly increased with HDAC inhibitor treatments compared to that with RA treatment. Furthermore, we confirmed that noncanonical Wnt signaling was upregulated by HDAC inhibitors via MAPK signaling and the Wnt/JNK pathway. Therefore, both MS-275 and VPA promoted the differentiation of SH-SY5Y cells into mature neurons via the Wnt signaling pathway.

Research Highlights.

Research Highlights.