Telomerase Inhibition Research Articles

Telomerase: a nexus between cancer nanotherapy and circadian rhythm.

Cancer represents a complex disease category defined by the unregulated proliferation and dissemination of anomalous cells within the human body. According to the GLOBOCAN 2020 report, the year 2020 witnessed the diagnosis of approximately 19.3 million new cases of cancer and 10.0 million individuals succumbed to the disease. A typical cell eventually becomes cancerous because of a long-term buildup of genetic instability and replicative immortality. Telomerase is a crucial regulator of cancer progression as it induces replicative immortality. In cancer cells, telomerase inhibits apoptosis by elongating the length of the telomeric region, which usually protects the genome from shortening. Many nanoparticles are documented as being available for detecting the presence of telomerase, and many were used as delivery systems to transport drugs. Furthermore, telomere homeostasis is regulated by the circadian time-keeping machinery, leading to 24-hour rhythms in telomerase activity and TERT mRNA expression in mammals. This review provides a comprehensive discussion of various kinds of nanoparticles used in telomerase detection, inhibition, and multiple drug-related pathways, as well as enlightens an imperative association between circadian rhythm and telomerase activity from the perspective of nanoparticle-based anticancer therapeutics.

Obesity-Senescence-Breast Cancer: Clinical Presentation of a Common Unfortunate Cycle.

There are few convincing studies establishing the relationship between endogenous factors that cause obesity, cellular aging, and telomere shortening. Without a functional telomerase, a cell undergoing cell division has progressive telomere shortening. While obesity influences health and longevity as well as telomere dynamics, cellular senescence is one of the major drivers of the aging process and of age-related disorders. Oxidative stress induces telomere shortening, while decreasing telomerase activity. When progressive shortening of telomere length reaches a critical point, it triggers cell cycle arrest leading to senescence or apoptotic cell death. Telomerase activity cannot be detected in normal breast tissue. By contrast, maintenance of telomere length as a function of human telomerase is crucial for the survival of breast cancer cells and invasion. Approximately three-quarters of breast cancers in the general population are hormone-dependent and overexpression of estrogen receptors is crucial for their continued growth. In obesity, increasing leptin levels enhance aromatase messenger ribonucleic acid (mRNA) expression, aromatase content, and its enzymatic activity on breast cancer cells, simultaneously activating telomerase in a dose-dependent manner. Meanwhile, applied anti-estrogen therapy increases serum leptin levels and thus enhances leptin resistance in obese postmenopausal breast cancer patients. Many studies revealed that shorter telomeres of postmenopausal breast cancer have higher local recurrence rates and higher tumor grade. In this review, interlinked molecular mechanisms are looked over between the telomere length, lipotoxicity/glycolipotoxicity, and cellular senescence in the context of estrogen receptor alpha-positive (ERα+) postmenopausal breast cancers in obese women. Furthermore, the effect of the potential drugs, which are used for direct inhibition of telomerase and the inhibition of human telomerase reverse transcriptase (hTERT) or human telomerase RNA promoters as well as approved adjuvant endocrine therapies, the selective estrogen receptor modulator and selective estrogen receptor down-regulators are discussed.

Telomerase inhibitor imetelstat kills AML cells via lipid ROS and ferroptosis.

Telomerase inhibitor imetelstat kills AML cells via lipid ROS and ferroptosis.

Telomerase and mitochondria inhibition promote apoptosis and TET2 and ANMT3a expression in triple negative breast cancer cell lines.

High metastasis, resistance to common treatments, and high mortality rate, has made triple-negative breast cancer (TNBC) to be the most invasive type of breast cancer. High telomerase activity and mitochondrial biogenesis are involved in breast cancer tumorigenesis. The catalytic subunit of telomerase, telomerase reverse transcriptase (hTERT), plays a role in telomere lengthening and extra-biological functions such as gene expression, mitochondria function, and apoptosis. In this study, it has been aimed to evaluate intrinsic-, extrinsic-apoptosis and DNMT3a and TET2 expression following the inhibition of telomerase and mitochondria respiration in TNBC cell lines. TNBC cells were treated with IC50 levels of BIBR1532, tigecycline, and also their combination. Then, telomere length, and DNMT3a, TET2, and hTERT expression were evaluated. Finally, apoptosis rate, apoptosis-related proteins, and genes were analyzed. The present results showed that IC50 level of telomerase and inhibition of mitochondria respiration induced apoptosis but did not leave any significant effect on telomere length. The results also indicated that telomerase inhibition induced extrinsic-apoptosis in MDA-MB-231 and caused intrinsic- apoptosis in MDA-MB-468 cells. Furthermore, it was found that the expression of p53 decreased and was ineffective in cell apoptosis. The expressions of DNMT3a and TET2 increased in cells. In addition, combination treatment was better than BIBR1532 and tigecycline alone. The inhibition of telomerase and mitochondria respiration caused intrinsic- and extrinsic- apoptosis and increased DNMT3a and TET2 expression and it could be utilized in breast cancer treatment.

New 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinazoline and 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinoline Derivatives: Synthesis and Biological Evaluation as Novel Anticancer Agents by Targeting G-Quadruplex.

The syntheses of novel 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinazolines 12 and 2,4-bis[(substituted-aminomethyl)phenyl]phenylquinolines 13 are reported here in six steps starting from various halogeno-quinazoline-2,4-(1H,3H)-diones or substituted anilines. The antiproliferative activities of the products were determined in vitro against a panel of breast (MCF-7 and MDA-MB-231), human adherent cervical (HeLa and SiHa), and ovarian (A2780) cell lines. Disubstituted 6- and 7-phenyl-bis(3-dimethylaminopropyl)aminomethylphenyl-quinazolines 12b, 12f, and 12i displayed the most interesting antiproliferative activities against six human cancer cell lines. In the series of quinoline derivatives, 6-phenyl-bis(3-dimethylaminopropyl)aminomethylphenylquinoline 13a proved to be the most active. G-quadruplexes (G4) stacked non-canonical nucleic acid structures found in specific G-rich DNA, or RNA sequences in the human genome are considered as potential targets for the development of anticancer agents. Then, as small aza-organic heterocyclic derivatives are well known to target and stabilize G4 structures, their ability to bind G4 structures have been determined through FRET melting, circular dichroism, and native mass spectrometry assays. Finally, telomerase inhibition ability has been also assessed using the MCF-7 cell line.

Lead Optimization of BIBR1591 To Improve Its Telomerase Inhibitory Activity: Design and Synthesis of Novel Four Chemical Series with In Silico, In Vitro, and In Vivo Preclinical Assessments.

Herein, modifications to the previously reported BIBR1591 were conducted to obtain bioisosteric candidates with improved activities. The % inhibition of the newly afforded candidates against the telomerase target was investigated. Notably, 6f achieved superior telomerase inhibition (63.14%) compared to BIBR1532 and BIBR1591 (69.64 and 51.58%, respectively). In addition, 8a and 8b showed comparable promising telomerase inhibition with 58.65 and 55.57%, respectively, which were recorded to be frontier to that of BIBR1591. 6f, 8a, and 8b were tested against five cancer cell lines related to the lung and liver subtypes. Moreover, 6f was examined on both cell cycle progression and apoptosis induction in HuH7 cancer cells. Furthermore, the in vivo antitumor activity of 6f was further assessed in female mice with solid Ehrlich carcinoma. In addition, molecular docking and molecular dynamics simulations were carried out. Collectively, 6f, 8a, and 8b could be considered potential new telomerase inhibitors to be subjected to further investigation and/or optimization.

Telomerase inhibition in haematological neoplasms—are we ready for primetime?

Telomerase inhibition in haematological neoplasms—are we ready for primetime?

Imetelstat in patients with lower-risk myelodysplastic syndromes who have relapsed or are refractory to erythropoiesis-stimulating agents (IMerge): a multinational, randomised, double-blind, placebo-controlled, phase 3 trial

Unmet medical needs remain in patients with red blood cell transfusion-dependent (RBC-TD) lower-risk myelodysplastic syndromes (LR-MDS) who are not responding to or are ineligible for erythropoiesis-stimulating agents (ESAs). Imetelstat, a competitive telomerase inhibitor, showed promising results in a phase 2 trial. We aimed to compare the RBC transfusion independence (RBC-TI) rate with imetelstat versus placebo in patients with RBC-TD LR-MDS. In phase 3 of IMerge, a double-blind, placebo-controlled trial conducted in 118 sites including university hospitals, cancer centres, and outpatient clinics in 17 countries, patients (aged ≥18 years) with ESA-relapsed, ESA-refractory, or ESA-ineligible LR-MDS (low or intermediate-1 risk disease as per International Prognostic Scoring System [IPSS] criteria) were randomly assigned via a computer-generated schedule (2:1) to receive imetelstat 7·5 mg/kg or placebo, administered as a 2-h intravenous infusion, every 4 weeks until disease progression, unacceptable toxic effects, or withdrawal of consent. Randomisation was stratified by previous RBC transfusion burden and IPSS risk group. Patients, investigators, and those analysing the data were masked to group assignment. The primary endpoint was 8-week RBC-TI, defined as the proportion of patients without RBC transfusions for at least 8 consecutive weeks starting on the day of randomisation until subsequent anti-cancer therapy, if any. Primary efficacy analyses were performed in the intention-to-treat population, and safety analyses were conducted in patients who received at least one dose of trial medication or placebo. This trial is registered with ClinicalTrials.gov (NCT02598661; substudy active and recruiting). Between Sept 11, 2019, and Oct 13, 2021, 178 patients were enrolled and randomly assigned (118 to imetelstat and 60 to placebo). 111 (62%) were male and 67 (38%) were female. 91 (77%) of 118 patients had discontinued treatment by data cutoff in the imetelstat group versus 45 (75%) in the placebo group; a further one patient in the placebo group did not receive treatment. Median follow-up was 19·5 months (IQR 12·0-23·4) in the imetelstat group and 17·5 months (12·1-22·7) in the placebo group. In the imetelstat group, 47 (40% [95% CI 30·9-49·3]) patients had an RBC-TI of at least 8 weeks versus nine (15% [7·1-26·6]) in the placebo group (rate difference 25% [9·9 to 36·9]; p=0·0008). Overall, 107 (91%) of 118 patients receiving imetelstat and 28 (47%) of 59 patients receiving placebo had grade 3-4 treatment-emergent adverse events. The most common treatment-emergent grade 3-4 adverse events in patients taking imetelstat were neutropenia (80 [68%] patients who received imetelstat vs two [3%] who received placebo) and thrombocytopenia (73 [62%] vs five [8%]). No treatment-related deaths were reported. Imetelstat offers a novel mechanism of action with durable transfusion independence (approximately 1 year) and disease-modifying activity for heavily transfused patients with LR-MDS who are not responding to or are ineligible for ESAs. Janssen Research & Development before April 18, 2019, and Geron Corporation thereafter.

Early Generation Telomerase Deficient Mice to Investigate Clonal Hematopoiesis in Aging

Clonal hematopoiesis is an age-associated condition characterized by the expansion of hematopoietic stem cell (HSC) pools with somatic mutations, observed in 10-30% of individuals older than 70 years. This phenomenon is primarily driven by mutations in transcriptional regulators such as DNMT3A, TET2, and ASXL1. While the clonal expansion of mutated HSCs is ubiquitous with age, it appears largely benign in the absence of strong selective pressures. These benign clones might compensate for the loss of regenerative HSC capacity during aging, and skew differentiation towards the myeloid lineage. Telomere erosion plays a significant role in this process. Telomeres normally prevent the activation of the DNA damage response (DDR) machinery. However, with each cell division, telomeres shorten until they reach a critical length, eventually leading to apoptosis or senescence. Previous studies have suggested a potential involvement of telomere attrition in the development of clonal hematopoiesis (Hinds et al. Blood 2016; Zink et al. Blood 2017). Late generation telomerase deficient (Terc-/- G5) mice develop premature bone marrow failure and mortality, however earlier generation Terc-/- G3 (3 homozygous crosses) have shortened telomeres but appear relatively normal. Importantly, inhibitors of telomerase have recently entered clinical trials for haematological malignancies and therefore understanding the long-term effects of telomerase inhibition is important. By studying the interplay between telomere attrition, HSC aging, and the development of CHIP, we aim to gain insights into the mechanisms underlying age-associated hematopoietic changes and their potential clinical implications. We examined young (age 8-10 week) and aged (10-12 month) Terc-/- G3 mice. Telomere lengths were significantly reduced in bone marrow cells from young Terc G3 (15 kb) when compared to wild-type mice (30 kb) with mild reduction in BM cellularity. Absolute long-term HSC numbers were reduced, as were megakaryocyte erythroid progenitors, but there was preservation of granulocyte macrophage progenitors and common myeloid progenitors. Spleen progenitor numbers were not affected. In comparison, aged (10-12 months old) Terc G3 mice showed enhanced differentiation skewing in the peripheral blood with significantly increased myeloid (n = 8 wt; n = 12 Terc ko; p = 0.0004) and concomitantly reduced B cell (p = 0.0002) and T cell (p = 0.0320) populations. There was marked mobilization of bone marrow progenitors to the spleen, most notably GMP (p = 0.0004) but also MEP and CMP in aged Terc G3. Furthermore, aged Terc G3 ko now exhibited marked reduction in bone marrow CMP (p < 0.0001) and MEP (p < 0.0001), but skewing towards GMP (p<0.001) resulting in normal total GMP numbers, but markedly reduced CMP and MEP numbers (p < 0.0001). Compared to young Terc G3 mice, the aged Terc G3 mice had relative expansion of the BM LT-HSC population, and therefore normal numbers of BM LT-HSC compared to aged matched controls. Aged Terc G3 BM LTHSC have been isolated for gene expression and clonality analysis. Together, these findings are consistent with accelerated aging in G3 Terc-/- mice, providing a model for examining clonal hematopoiesis. In order to assess the cell intrinsic function of Terc G3 LTHSC, we next performed non-competitive bone marrow transplantation experiments from Terc ko or wild-type donors into lethally irradiated young wild-type recipients (n = 9 per group), and analysed these at 4 months post-transplant. There was reduced, but stable reconstitution by Terc G3 cells, consistent with previous competitive bone marrow transplants in Terc G3 (Bruedigam et al. Cell Stem Cell 2014). We observed a similar reduction in bone marrow cellularity in Terc ko transplanted recipients (p = 0.003). Lineage skewing was less pronounced but CMP percentages were significantly reduced in Terc ko compared to wild-type conditions (p = 0.03). In contrast to primary mice, mobilization of progenitors to the spleen was not observed, suggesting that this finding was ameliorated by normal bone marrow and splenic supporting cells. The aged G3 Terc ko model faithfully recapitulates the haematopoietic phenotype of aging in humans and we propose to utilise this model to further our understanding of the mechanisms and physiological consequences of clonal hematopoiesis, to eventually help identify strategies to mitigate its clinical consequences.

Impact of Mutational Status on Clinical Response to Imetelstat in Patients with Lower-Risk Myelodysplastic Syndromes in the IMerge Phase 3 Study

CONCLUSIONS Imetelstat, a first-in-class telomerase inhibitor, showed higher rates of red blood cell (RBC) transfusion independence (TI) for ≥8 weeks, ≥24 weeks, and ≥1 year (39.8%, 28.0%, and 17.8%) than placebo (15.0%, 3.3%, and 1.7%) in the IMerge (NCT02598661) phase 3 study of patients with RBC transfusion-dependent non-del(5q) lower-risk myelodysplastic syndromes (LR-MDS) relapsed/refractory to erythropoiesis-stimulating agents (Platzbecker et al. EHA 2023. Abstr S165). To evaluate the impact of MDS-associated mutations on clinical efficacy of imetelstat, we performed next-generation sequencing of a panel of 36 genes, recurrently mutated in MDS, using DNA samples from peripheral blood collected at study entry. Further analysis of TI responses to imetelstat was performed across different mutation subgroups, defined based on genes involved in different biological functions, including the splicing process, epigenetic modifiers, transcription regulation, and receptors/kinases. Baseline mutation data were available in 165 of 178 patients (imetelstat, n = 110; placebo, n = 55; 93.2% and 91.7% of total in each group, respectively). Of patients with mutation data, 161 (97.6%) had ≥1 mutation detected, among whom, 75 (70.1%), 33 (30.8%), and 9 (8.4%) patients in the imetelstat group and 38 (70.4%), 15 (27.8%), and 7 (13%) patients in the placebo group had >1, >2, and >3 mutations, respectively. The ≥8-week TI rates in the imetelstat vs placebo groups were 42.7% vs 15.8% ( P = .006) for patients with >1 mutation, 45.5% vs 6.7% ( P = .012) for patients with >2 mutations, and 55.6% vs 14.3% ( P = .089) for patients with >3 mutations, respectively. The ≥24-week TI rates were 26.7% vs 2.6% ( P = .003), 33.3% vs 0% ( P = .014), and 33.3% vs 0% ( P = .117), respectively. In patients with mutations associated with poor prognosis ( TP53, ETV6, RUNX1, ASXL1 or EZH2), 31.8% and 9.1% of patients in the imetelstat group achieved ≥8-week and ≥24-week TI vs 0 of those in the placebo group. TP53 mutations were detected in 2 patients in each group; both patients in the imetelstat group and none in the placebo group had ≥8-week TI. Among patients with ASXL1 mutations, 5 of 18 patients (27.8%) in the imetelstat and 0 of 6 (0) in placebo group had ≥8-week TI. Among patients with ETV6 mutations, 1 of 2 (50%) in the imetelstat group and 0 of 1 patients in the placebo group had ≥8-week TI. Two patients in each group had RUNX1 mutations; none achieved TI. Imetelstat ≥8-week TI rates were 43.8%, 37.7%, 40.0%, and 80.0% for patients harboring mutated genes in the RNA spliceosome, epigenetic modifiers, transcription regulation, and receptors/kinases, respectively; ≥24-week TI was 30.2%, 27.5%, 20.0%, and 80.0%, respectively. The most frequently mutated gene was SF3B1 (125 of 165; 75.8%), the ≥8-week and ≥24-week TI rates were 48.8% vs 16.3% ( P = .001) and 35.4% vs 2.3% ( P < .001) with imetelstat vs placebo. In the imetelstat group, ≥8-week TI was achieved in patients with different spectrum of SF3B1 hot-spot mutations: 2 of 8 patients (25.0%) with E622D, 4 of 7 (57.1%) with R625C/L/G, 7 of 12 (58.3%) with H662Q/N/D/Y, 2 of 2 (100%) with T663P, 2 of 6 (33.3%) with K666R/T/Q/N, 18 of 41 (43.9%) with K700E, 2 of 2 (100%) with A744P, and 1 of 2 patients (50.0%) with E783K. Durable ≥24-week TI was also observed in patients with these hot-spot mutations. Other genes with mutation frequency >10% were TET2 (32.7%), DNMT3A (17.0%), ASXL1 (14.5%), and CUX1 (12.7%). The ≥8-week TI rates in the imetelstat group vs placebo group were 50% vs 21.4% for TET2 mutations; 31.6% vs 22.2% for DNMT3A mutations; 27.8% vs 0 for ASXL1 mutations; 35.7% vs 14.3% for CUX1 mutations (Figure). The ≥24-week TI rates were 40% vs 0, 26.3% vs 11.1%, 11.1% vs 0, and 14.3% vs 0, respectively, for those mutations (Figure). Higher RBC-TI rates were observed in patients with various baseline mutational profiles treated with imetelstat compared with placebo in IMerge. While the sample size for specific mutations was small, consistent with the observation that patients with LR-MDS have a low number of specific mutations, TI responses in patients receiving imetelstat occurred regardless of the presence of mutations associated with poor prognosis or the number of mutations. Imetelstat showed comparable TI rates across different molecularly defined subgroups, suggesting that clinical benefit of imetelstat in patients with LR-MDS is independent of the underlying molecular pattern.

Durable Continuous Transfusion Independence (TI) with Imetelstat in IMerge Phase 3 for Patients with Heavily Transfused Non-Del(5q) Lower-Risk Myelodysplastic Syndromes (LR-MDS) Relapsed/Refractory (R/R) to or Ineligible for Erythropoiesis-Stimulating Agents (ESAs)

CONCLUSIONS MDS are serious life-threatening hematologic malignancies in which a heterogeneous group of clonal disorders results in ineffective hematopoiesis. An unmet need remains for new therapies for patients (pts) with LR-MDS who are red blood cell transfusion dependent (RBC-TD) and R/R to or ineligible for ESAs. Imetelstat, an oligonucleotide, is a first-in-class telomerase inhibitor that targets cells with high telomerase activity by direct binding to the RNA template of telomerase. In the IMerge phase 3 trial, imetelstat produced higher rates of TI for ≥8 weeks, ≥24 weeks, and ≥1 year (39.8%, 28.0%, and 17.8%) than placebo (15.0%, 3.3%, and 1.7%) in pts with non-del(5q) LR-MDS that was RBC-TD, R/R to/ineligible for ESAs, and naïve to lenalidomide or hypomethylating agents (HMAs; Platzbecker et al. EHA 2023. Abstr S165). We report characteristics and clinical benefit for pts with sustained TI for ≥1 year from this trial. IMerge (MDS3001, NCT02598661) is a global, double-blind, randomized, placebo-controlled, phase 3 trial of imetelstat in RBC-TD, ESA-R/R, non-del 5(q) lenalidomide/HMA-naïve LR-MDS. The primary end point was 8-week TI rate; secondary end points included safety, 24-week TI, duration of response, hematologic improvement, and MDS response. Exploratory end points included assessment of cytogenetic response and mutational status with clinical response. The proportion of pts with>1-year TI and other binary end points, were summarized with percentage and 95% 2-sided exact Clopper-Pearson CI. The Kaplan-Meier method was used to estimate the distribution of TI. Of 118 pts receiving imetelstat, 21 (17.8%; 95% CI, 11.4-25.9) achieved ≥1-year sustained TI, representing 45% of ≥8-week TI (21 of 47 pts) and 64% of ≥24-week TI (21 of 33 pts); of 60 pts receiving placebo plus supportive care, 1 (1.7%; 95% CI, 0-8.9) achieved ≥1-year TI. Of the ≥1-year TI imetelstat responders, 15/21 (71.4%) had ring sideroblasts, as did the 1 placebo pt. The median prior RBC transfusion burden was 6 U over 8 weeks (range, 4-9 U) for the imetelstat group and 5 U for the placebo pt. Additional baseline characteristics are in the table. Pts received imetelstat for a median of 101.1 weeks (range, 75.1-163.9 weeks) and a median of 24 cycles (range, 18-41 cycles). The median duration of TI for imetelstat ≥1-year TI responders was 123 weeks (95% CI, 80.4 to not evaluable); the median increase in hemoglobin during the longest TI interval was 5.18 g/dL (range, 2.67-13.76 g/dL) for the imetelstat group vs 1.67 g/dL for the placebo pt. After a median follow-up of 125 weeks, none of the patients with ≥1 year TI on either arm progressed to acute myeloid leukemia (AML). Of the pts receiving imetelstat, 7 had an abnormal karyotype at baseline, of which 6 had reduction in the cytogenetic abnormal clones (4 with cytogenetic complete response and 2 with cytogenetic partial response by independent review committee). Mutation data were available for 18 pts receiving imetelstat, all with SF3B1 mutations present at baseline, and multiple of these pts concurrently had TET2, DNMT3A, ASXL1, or JAK2 mutations. The maximal reduction ranged from −6% to −100% in SF3B1 variant allele frequency (VAF) in these pts, and 13 of 18 (72.2%) achieved ≥50% VAF reduction, including 7 with complete elimination of the VAF. Reduction in other concurrent mutations was also observed in these pts. Safety was consistent with that previously reported; most frequent adverse events were reversible grade 3 or 4 thrombocytopenia and neutropenia. At the time of data cutoff (May 10, 2023), 13 pts receiving imetelstat and the pt receiving placebo were ongoing (Figure); of the 8 who discontinued treatment, 7 had loss of response, and 1 was due to adverse event. Analyses for progression-free and overall survival were not evaluable as of this cutoff date (insufficient follow-up). Treatment with imetelstat resulted in ≥1-year sustained, continuous TI in 17.8% of pts in the IMerge phase 3 trial. In this ESA-R/R/ineligible population with a high prior transfusion burden, a reduction to 0 RBC transfusions for ≥1 year represents an opportunity to achieve relief from iron overload and other transfusion associated complications, and decreased demand on already limited blood product supply. Furthermore, durable TI and meaningful reductions in mutational burden suggest imetelstat may have disease-modifying activity.

Characterization and Management of Cytopenias after Imetelstat Treatment in the IMerge Phase 3 Trial of Patients with Lower-Risk Myelodysplastic Syndromes (LR-MDS)

CONCLUSIONS Imetelstat, an oligonucleotide and a first-in-class telomerase inhibitor, selectively targets malignant hematopoietic stem and progenitor cells with high telomerase activity by direct binding to the RNA template. In patients with MDS the on-target effects of imetelstat are associated with the development of hematologic treatment-emergent adverse events (TEAEs). In the IMerge phase 3 trial, among patients treated with imetelstat, 68% experienced grade 3-4 neutropenia, and 62% had grade 3-4 thrombocytopenia; however, clinical consequences of grade 3-4 infection or bleeding were similar in patients treated with imetelstat and placebo (Platzbecker et al. EHA 2023. Abstr S165). Here, we report additional data on the occurrence and management of cytopenias after treatment with imetelstat. IMerge phase 3 trial (NCT02598661) is a multicenter study involving 178 patients with a median age of 72 years with red blood cell (RBC) transfusion dependency and LR-MDS relapsed/refractory to or ineligible for erythropoiesis-stimulating agents. Patients were randomized 2:1 to receive either imetelstat or placebo and stratified by prior RBC transfusion burden (4-6 or >6 U) and by International Prognostic Scoring System risk group. The safety population included all patients who received ≥1 dose of study drug and comprised 118 patients treated with imetelstat and 59 with placebo. TEAEs of neutropenia and thrombocytopenia with imetelstat treatment were more prevalent in cycles 1-3 (68.6% and 62.7% respectively), and their frequency decreased over time: 42.7% and 48.5% in cycles 4-6, 36.8% and 44.7% in cycles 7-12, and 31.3% and 37.5% in cycle 13 and beyond. Based on laboratory assessment, neutrophil and platelet counts decreased from baseline levels in patients treated with imetelstat versus those treated with placebo (91.5% and 95.8% vs 47.5% and 33.9%, respectively). Neutropenia decreased to a maximum of grade 4 and 3 in 55 and 29 of imetelstat-treated patients, respectively, and >80% of cytopenia events stabilized to grade ≤2 within 4 weeks (Figure). The median time to grade 4 neutropenia was 5.0 weeks (range,1.1-90.3 weeks) for the imetelstat group vs 23.0 weeks (range, 23.0-23.0 weeks) for the placebo group. Similarly, median time to grade 4 thrombocytopenia was 5.2 weeks (range, 2.0-29.9 weeks) with imetelstat vs 35.5 weeks (range, 28.0-43.0 weeks) with placebo. It is noteworthy that among 47 patients who achieved the primary end point of 8-week RBC transfusion independence with imetelstat, 34 (72.3%) and 28 patients (59.6%) had grade 3-4 neutropenia and grade 3-4 thrombocytopenia, respectively. In the imetelstat group, 3 patients had grade 3-4 neutropenia concurrent with grade 3-4 infections, and no patients experienced grade 3-4 thrombocytopenia associated with grade 3-4 bleeding events. There were no grade 5 cytopenias for either imetelstat or placebo groups. In the imetelstat group, cytopenias were managed with protocol-specified treatment delays and dose adjustments. Dose reductions due to neutropenia and thrombocytopenia occurred in 39 patients (33.1%) and 27 patients (22.9%), respectively. Of imetelstat-treated patients, 6 (5.1%) discontinued treatment due to neutropenia (1 case, grade 4; all others, grade 3), and 4 (3.4%) discontinued due to thrombocytopenia (1 case, grade 2; 3 cases, grade 3). In addition to dose reductions and discontinuation of imetelstat, thrombocytopenia and neutropenia were managed by cycle delays in 46.6% and 50.8% of patients, by platelet transfusions in 17.8% of patients, and by concomitant therapy with growth factor support (mostly during cycles 2-4) in 34.7% of patients. Similar patterns of imetelstat dose modifications were seen in a pooled phase 2 and phase 3 safety analysis of IMerge. In the IMerge phase 3 trial, thrombocytopenia and neutropenia were the most common and most frequently reported grade 3-4 AEs during treatment cycles 1-3. However, these AEs were generally transient, reversible, and manageable through treatment delays and dose adjustments, allowing for patients to remain on treatment and continue to experience clinical benefit. Moreover, occurrence of grade 3-4 cytopenias in responders suggests that these TEAEs do not affect the efficacy of imetelstat.

BIBR1532 inhibits proliferation and enhances apoptosis in multiple myeloma cells by reducing telomerase activity

Background Multiple myeloma (MM) is a rare haematological disorder with few therapeutic options. BIBR1532, a telomerase inhibitor, is widely used in cancer treatment and has promising outcomes. In this study, we investigated the efficacy and mechanism of action of BIBR1532 in MM. Methods K562 and MEG-01 cells were cultured with BIBR1532 at different concentrations. After 24 and 48 h, cell survival was analyzed. Next, these cells were cultured with 25 and 50 µM BIBR1532 for 48 h, then, cell proliferation, apoptosis, and the expression of the telomerase activity related markers were tested by 5-Ethynyl-2′-deoxyuridine (EdU) staining, flow cytometric analysis, western blot and quantitative real-time PCR (qRT-PCR), respectively. Expression of Bcl-xL, Bad, Survivin, phosphorylation of PI3K, AKT, mTOR, ERK1/2, and MAPK were tested via western blotting. Further experiments were conducted to evaluate the synergistic effects of BIBR1532 and doxorubicin (Dox) or bortezomib (Bor). Results BIBR1532 inhibited K562 and MEG-01 cell survival in a dose- and time-dependent manner. In addition, BIBR1532 hindered cell proliferation while promoting apoptosis, and this effect was enhanced by increasing the BIBR1532 concentration. Moreover, BIBR1532 inhibited TERT and c-MYC expression, PI3K, AKT, mTOR phosphorylation, and facilitated ERK1/2 and MAPK phosphorylation. Additionally, BIBR1532 combined with Dox or Bor showed synergistic effects in MM treatment. Conclusion BIBR1532 inhibits proliferation and promotes apoptosis in MM cells by inhibiting telomerase activity. Additionally, BIBR1532 combined with Dox or Bor exhibited synergistic effects, indicating that BIBR1532 may be a novel medicine for the treatment of MM.

Efficacy of Imetelstat in Achieving Red Blood Cell Transfusion Independence (RBC-TI) across Different Risk Subgroups in Patients with Lower-Risk Myelodysplastic Syndromes (LR-MDS) Relapsed/Refractory (R/R) to Erythropoiesis-Stimulating Agents (ESAs) in IMerge Phase 3 Study

CONCLUSIONS IMerge (NCT02598661) is a phase 2/3 global study of imetelstat, a first-in-class telomerase inhibitor, for patients with RBC transfusion-dependent non-del(5q) LR-MDS R/R to or ineligible for ESAs, a population with unmet needs. Phase 3 results showed that the rate of ≥8-week, ≥24-week and 1-year RBC-TI was higher with imetelstat than placebo (Zeidan et al. ASCO 2023. Abstr 7004; Platzbecker et al. EHA 2023. Abstr S165). Toevaluate clinical efficacy of imetelstat across different International Prognostic Scoring System (IPSS), revised IPSS (IPSS-R), IPSS-R cytogenetic, or IPSS-molecular (IPSS-M) risk categories,cytogenetic analysis was performed centrally on bone marrow aspirates by karyotyping, and mutation profile was analyzed using peripheral blood samples by next-generation sequencing on a panel of 36 genes commonly mutated in MDS. Baseline IPSS, IPSS-R, IPSS-R cytogenetic, or IPSS-M risk categories were derived for patients in the phase 3 portion of IMerge based on specific classification criteria, and rates of RBC-TI ≥8-week, ≥24-week, ≥1-year were assessed for each risk subgroup. In 105 IPSS-low patients, 95 (90.5%) remained IPSS-M very low/low/moderate low, and 10 patients (9.5%) upstaged to IPSS-M moderate high and high. In 50 IPSS-intermediate-1 patients, 39 (78%) remained IPSS-M very low/low/moderate low, and 11 patients (22%) upstaged to IPSS-M moderate high/high. Among 129 IPSS-R low/very low patients, 118 (91.5%) remained IPSS-M very low/low/moderate low, and 11 (8.5%) patients upstaged to IPSS-moderate high/high. In 25 IPSS-R-intermediate patients, 16 (64%) remained IPSS-M low/moderate low, and 9 patients (36%) upstaged to IPSS-M moderate high/high/very high. One IPSS-R high-risk patient remained IPSS-M high. Overall, significantly higher rates of ≥8-week, ≥24-week, and ≥1-year RBC-TI were achieved in patients receiving imetelstat (39.8%, 28%, and 13.6%) vs those receiving placebo (15%, 3.3%, and 1.7%). Further subgroup analysis demonstrated that imetelstat consistently had higher TI response rates than placebo across different risk subgroups, irrespective of classification system. For IPSS risk groups, the TI rates of ≥8-week, ≥24-week, and ≥1-year with imetelstat vs placebo were 40.0% vs 20.5% ( P = 0.034), 28.8% vs 5.1% ( P = 0.003), and 12.5% vs 2.6% ( P = 0.082), respectively, in patients with low risk and 39.5% vs 4.8% ( P = 0.004), 26.3% vs 0% ( P = 0.009), and 15.8% vs 0% ( P = 0.048), respectively, in patients with intermediate-1 risk. For IPSS-R risk groups, the TI rates of ≥8-week, ≥24-week, and ≥1-year with imetelstat vs placebo were 42.5% vs 19.6%, 29.9% vs 4.3%, and 11.5% vs 2.2%, respectively, in the low-risk subgroup. The TI rates of ≥8-week, ≥24-week, ≥1-year TI with imetelstat were 35%, 25%, and 20%, respectively, in the intermediate-risk subgroup, whereas placebo treatment had no response. The number of patients was too low in both arms of the very low or high IPSS-R subgroups to assess difference in TI response. For IPSS-R cytogenetic risk groups, the TI rates of ≥8-week, ≥24-week, and ≥1-year with imetelstat vs placebo were 37.1% vs 17.0%, 24.7% vs 4.3%, and 10.1% vs 2.1%, respectively, in very good/good risk groups and 54.5% vs 11.1%, 40.9% vs 0%, and 22.7% vs 0%, respectively, in the intermediate-risk group. For IPSS-M risk groups, the TI rates of ≥8-week, ≥24-week, and ≥1-year with imetelstat vs placebo were 47.8% vs 21.2%, 34.8% vs 3%, and 14.5% vs 0%, respectively, in patients with very low/low risk and 20.7% vs 6.3%, 10.3% vs 0%, and 6.9% vs 0%, respectively, in patients with moderate low/moderate high risk. In patients with high/very high risk, ≥8-week TI rates were 40% vs 0% with imetelstat vs placebo, with no ≥24-week or ≥1-year TI observed in either arm. All results are summarized in the Table. Improvement in RBC-TI rates was observed in patients treated with imetelstat vs placebo across different risk subgroups as defined by IPSS, IPSS-R, IPSS-R cytogenetic, or IPSS-M risk profiles. Notably, placebo had not achieved durable (≥24-week and ≥1-year) TI response in the higher-risk groups irrespective of the risk classification assessment model used, while TI response rates with imetelstat in higher-risk subgroups with poor prognosis were similar to TI response rates in lower-risk subgroups of heavily transfused R/R ESA LR-MDS, indicating that clinical efficacy of imetelstat is independent of risk categories.

The telomerase inhibitor imetelstat differentially targets JAK2V617F versus CALR mutant myeloproliferative neoplasm cells and inhibits JAK-STAT signaling.

Imetelstat shows activity in patients with myeloproliferative neoplasms, including primary myelofibrosis (PMF) and essential thrombocythemia. Here, we describe a case of prolonged disease stabilization by imetelstat treatment of a high-risk PMF patient enrolled into the clinical study MYF2001. We confirmed continuous shortening of telomere length (TL) by imetelstat treatment but observed emergence and expansion of a KRAST58I mutated clone during the patient's clinical course. In order to investigate the molecular mechanisms involved in the imetelstat treatment response, we generated induced pluripotent stem cells (iPSC) from this patient. TL of iPSC-derived hematopoietic stem and progenitor cells, which was increased after reprogramming, was reduced upon imetelstat treatment for 14 days. However, while imetelstat reduced clonogenic growth of the patient's primary CD34+ cells, clonogenic growth of iPSC-derived CD34+ cells was not affected, suggesting that TL was not critically short in these cells. Also, the propensity of iPSC differentiation toward megakaryocytes and granulocytes was not altered. Using human TF-1MPL and murine 32DMPL cell lines stably expressing JAK2V617F or CALRdel52, imetelstat-induced reduction of viability was significantly more pronounced in CALRdel52 than in JAK2V617F cells. This was associated with an immediate downregulation of JAK2 phosphorylation and downstream signaling as well as a reduction of hTERT and STAT3 mRNA expression. Hence, our data demonstrate that imetelstat reduces TL and targets JAK/STAT signaling, particularly in CALR-mutated cells. Although the exact patient subpopulation who will benefit most from imetelstat needs to be defined, our data propose that CALR-mutated clones are highly vulnerable.

POT1 involved in telomeric DNA damage repair and genomic stability of cervical cancer cells in response to radiation.

Though telomeres play a crucial role in maintaining genomic stability in cancer cells and have emerged as attractive therapeutic targets in anticancer therapy, the relationship between telomere dysfunction and genomic instability induced by irradiation is still unclear. In this study, we identified that protection of telomeres 1 (POT1), a single-stranded DNA (ssDNA)-binding protein, was upregulated in γ-irradiated HeLa cells and in cancer patients who exhibit radiation tolerance. Knockdown of POT1 delayed the repair of radiation-induced telomeric DNA damage which was associated with enhanced H3K9 trimethylation and enhanced the radiosensitivity of HeLa cells. The depletion of POT1 also resulted in significant genomic instability, by showing a significant increase in end-to-end chromosomal fusions, and the formation of anaphase bridges and micronuclei. Furthermore, knockdown of POT1 disturbed telomerase recruitment to telomere, and POT1 could interact with phosphorylated ATM (p-ATM) and POT1 depletion decreased the levels of p-ATM induced by irradiation, suggesting that POT1 could regulate the telomerase recruitment to telomeres to repair irradiation-induced telomeric DNA damage of HeLa cells through interactions with p-ATM. The enhancement of radiosensitivity in cancer cells can be achieved through the combination of POT1 and telomerase inhibitors, presenting a potential approach for radiotherapy in cancer treatment.

Recent advances in therapies for primary myelofibrosis.

Primary myelofibrosis (PMF), polycythemia vera (PV) and essential thrombocythemia (ET) form the classical BCR-ABL1-negative myeloproliferative neoplasms (MPNs) that are driven by a constitutive activation of JAK2 signaling. PMF as well as secondary MF (post-ET and post-PV MF) are the most aggressive MPNs. Presently, there is no curative treatment, except allogenic hematopoietic stem cell transplantation. JAK inhibitors, essentially ruxolitinib, are the therapy of reference for intermediate and high-risk MF. However, presently the current JAK inhibitors behave mainly as anti-inflammatory drugs, improving general symptoms and spleen size without major impact on disease progression. A better understanding of the genetics of MF, the biology of its leukemic stem cells (LSCs), the mechanisms of fibrosis and of cytopenia and the role of inflammatory cytokines has led to new approaches with the development of numerous therapeutic agents that target epigenetic regulation, telomerase, apoptosis, cell cycle, cytokines and signaling. Furthermore, the use of a new less toxic form of interferon-α has been revived, as it is presently one of the only molecules that targets the mutated clone. These new approaches have different aims: (a) to provide alternative therapy to JAK inhibition; (b) to correct cytopenia; and (c) to inhibit fibrosis development. However, the main important goal is to find new disease modifier treatments, which will profoundly modify the progression of the disease without major toxicity. Presently the most promising approaches consist of the inhibition of telomerase and the combination of JAK2 inhibitors (ruxolitinib) with either a BCL2/BCL-xL or BET inhibitor. Yet, the most straightforward future approaches can be considered to be the development of and/or selective inhibition of JAK2V617F and the targeting MPL and calreticulin mutants by immunotherapy. It can be expected that the therapy of MF will be significantly improved in the coming years.

Opp-Dibenzoporphyrin Pyridinium Derivatives as Potential G-Quadruplex DNA Ligands.

Since the occurrence of tumours is closely associated with the telomerase function and oncogene expression, the structure of such enzymes and genes are being recognized as targets for new anticancer drugs. The efficacy of several ligands in telomerase inhibition and in the regulation of genes expression, by an effective stabilisation of G-quadruplexes (G4) DNA structures, is being considered as a promising strategy in cancer therapies. When evaluating the potential of a ligand for telomerase inhibition, the selectivity towards quadruplex versus duplex DNA is a fundamental attribute due to the large amount of double-stranded DNA in the cellular nucleus. This study reports the evaluated efficacy of three tetracationic opp-dibenzoporphyrins, a free base, and the corresponding zinc(II) and nickel(II) complexes, to stabilise G4 structures, namely the telomeric DNA sequence (AG3(T2AG3)3). In order to evaluate the selectivity of these ligands towards G4 structures, their interaction towards DNA calf thymus, as a double-strand DNA sequence, were also studied. The data obtained by using different spectroscopic techniques, such as ultraviolet-visible, fluorescence, and circular dichroism, suggested good affinity of the free-base porphyrin and of its zinc(II) complex for the considered DNA structures, both showing a pattern of selectivity for the telomeric G4 structure. A pattern of aggregation in aqueous solution was detected for both Zn(II) and Ni(II) metallo dibenzoporphyrins and the ability of DNA sequences to induce ligand disaggregation was observed.

PinX1-Promoted Autophagy Inhibits Cell Proliferation and Induces Cell Apoptosis by Inhibiting the NF-κB/p65 Signaling Pathway in Nasopharyngeal Carcinoma.

The role of Pin2 telomeric repeat factor 1-interacting telomerase inhibitor 1 (PinX1) in tumorigenesis and development has been extensively studied. As we previously demonstrated, PinX1 plays an important role in modulating epithelial-mesenchymal transition (EMT), stemness, cell proliferation, and apoptosis in nasopharyngeal carcinoma (NPC). However, the relationship between PinX1, autophagy, and cell function in NPC remains unclear. This study aimed to investigate the mechanisms by which PinX1 regulates autophagy in NPC, and to explore its biological role and clinical significance in disease progression. The proliferative capacity of NPC cells was assessed by MTT and xenograft tumorigenicity assays. Autophagic flux was monitored using a tandem monomeric DAPI-FITC-LC3 reporter assay. The rates of apoptosis and the cell cycle in NPC cells were analyzed using flow cytometry. The activation of autophagy and the signaling status of the AKT/mTOR and NF-κB/p65 pathways were evaluated by Western blot analysis. In addition to promoting autophagy and apoptosis, PinX1 overexpression suppressed proliferation, migration, invasion, and decelerated cell-cycle progression in NPC cells. These effects were reversed by inhibiting autophagy with 3-methyladenine. Mechanistic investigations clarified that PinX1 overexpression significantly reduced the expression of p-AKT, p-mTOR, p65, and p-p65. Chloroquine treatment in PinX1-overexpressing cells did not significantly alter p-AKT and p-mTOR levels, whereas 3-MA treatment in PinX1-overexpressing cells resulted in increased p65 and p-p65 expression, relative to untreated PinX1-overexpressing cells. It appears that PinX1 promotes autophagy by inhibiting the AKT/mTOR signaling pathway, which then inhibits NF-κB/p65 pathways, and consequently inhibiting cell proliferation and causing cell apoptosis in NPC cells.

Synthesis, characterization, anti-cancer evaluation, and DNA-binding study of new bay-substituted perylene derivatives

Two new perylene derivatives 1,7-di(3,5-diamino-pyrimidoxyl) perylene-3,4,9,10- tetracarboxylic acid bisanhydride (4) and 1,7-di(2-[3-[(4-amino-2-methylpyrimidin-5-yl)methyl]-4-methyl-1,3-thiazol-3-ium-5-yl]) ethoxyperylene-3,4,9,10-tetracarboxylic bisanhydride (6) have been synthesized. We aimed to study their interactions with G-quadruplex (G4) structures as potent G4 ligands and telomerase inhibitors. We used a PCR-amplified guanine-rich region from the human beta-globin gene, oligonucleotide from human telomeres (a-coreTT), an oncogene (c-kit), and SK-HEP-1 adenocarcinoma cells to characterize those compounds’ binding and stabilizing abilities to G4 structures and anti-cancer potential. All results obtained through UV-visible and fluorescence spectroscopies, agarose gel electrophoresis, and MTT assay on SK-HEP-1 adenocarcinoma cells were in good agreement. Compounds 4 and 6 are promising DNA-binding and cytotoxic compounds with a relatively antiproliferative effect on the selected tumour. In all studies, the formal positive charge carrier, compound 6, showed higher activity in terms of anti-cancer effects. These results may help elucidate the feasibility of the perylene derivatives as future chemo-therapeutic agents.