Uncovering the expression of circPVT1 in the extracellular vesicles of acute myeloid leukemia patients.

Extracellular vesicles (EVs) act as molecular mediators in the tumor microenvironment, by shuttling information contained within malignant cells and functioning as regulators of the immune system. Circular (circ)RNAs are characterized by a closed loop-like structure that makes them more stable in the extracellular milieu and suitable to be packaged inside EVs. circPVT1 (hsa_circ_0001821) showed an oncogenic role in several cancer types and immunosuppressive properties in myeloid and lymphoid cell subsets. In this study, we characterized EVs from acute myeloid leukemia (AML) patients in terms of size, concentrations, surface markers and circPVT1 cargo. We showed that circPVT1 is overexpressed by primary blast cells from newly-diagnosed AML patients compared with hematopoietic stem-progenitor cells and is released as cell-free RNA in the plasma. We isolated EVs from the plasma of AML patients and healthy subjects by size exclusion chromatography and characterized them by nanoparticle tracking analysis. EVs from patients' plasma are larger compared with those from healthy subjects and their surface profile is characterized by higher levels of the leukemic cell markers CD133, CD105, CD49e and other immune-related epitopes, with differences according to AML molecular profile. Moreover, digital PCR analysis revealed that circPVT1 is more abundant inside EVs from the plasma of AML patients compared with healthy subjects. Our findings provide new insights on the features and content of AML EVs and suggest a role of circPVT1 in the crosstalk between AML cells and the tumor microenvironment.

Venetoclax durable response in adult relapsed/refractory Philadelphia-negative acute lymphoblastic leukemia with JAK/STAT pathway alterations.

High-risk relapsed/refractory adult Philadelphia-negative (Ph-) B-cell acute lymphoblastic leukemia (B-ALL) is a great challenge due to limited possibilities to achieve and maintain a complete response. This also applies to cases with extramedullary (EM) involvement that have poor outcomes and no accepted standard therapeutic approaches. The incidence of EM localization in relapsed/refractory B-ALL is poorly investigated: data on patients treated with blinatumomab reported a 40% rate. Some responses were reported in EM patients with relapsed/refractory B-ALL treated with inotuzumab ozogamicin or CAR-T. However, molecular mechanisms of response or refractoriness are usually investigated neither at the medullary nor at EM sites. In the complex scenario of pluri-relapsed/refractory B-ALL patients, new target therapies are needed. Our analysis started with the case of an adult pluri-relapsed Ph- B-ALL patient, poorly sensitive to inotuzumab ozogamicin, donor lymphocyte infusions, and blinatumomab in EM disease, who achieved a durable/complete response after treatment with the BCL2-inhibitor venetoclax. The molecular characterization of medullary and EM samples revealed a tyrosine kinase domain JAK1 mutation in the bone marrow and EM samples at relapse. By comparing the expression level of BCL2- and JAK/STAT pathway-related genes between the patient samples, 136 adult JAK1 wt B-ALL, and 15 healthy controls, we identified differentially expressed genes, including LIFR, MTOR, SOCS1/2, and BCL2/BCL2L1, that are variably modulated at diverse time points and might explain the prolonged response to venetoclax (particularly in the EM site, which was only partially affected by previous therapies). Our results suggest that the deep molecular characterization of both medullary and EM samples is fundamental to identifying effective and personalized targeted therapies.

Targeting Proliferation Signals and the Cell Cycle Machinery in Acute Leukemias: Novel Molecules on the Horizon.

Uncontrolled proliferative signals and cell cycle dysregulation due to genomic or functional alterations are important drivers of the expansion of undifferentiated blast cells in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) cells. Therefore, they are largely studied as potential therapeutic targets in the field. We here present the most recent advancements in the evaluation of novel compounds targeting cell cycle proteins or oncogenic mechanisms, including those showing an antiproliferative effect in acute leukemia, independently of the identification of a specific target. Several new kinase inhibitors have been synthesized that showed effectiveness in a nanomolar to micromolar concentration range as inhibitors of FLT3 and its mutant forms, a highly attractive therapeutic target due to its driver role in a significant fraction of AML cases. Moreover, we introduce novel molecules functioning as microtubule-depolymerizing or P53-restoring agents, G-quadruplex-stabilizing molecules and CDK2, CHK1, PI3Kδ, STAT5, BRD4 and BRPF1 inhibitors. We here discuss their mechanisms of action, including the downstream intracellular changes induced by in vitro treatment, hematopoietic toxicity, in vivo bio-availability and efficacy in murine xenograft models. The promising activity profile demonstrated by some of these candidates deserves further development towards clinical investigation.

Exploring the ATR-CHK1 pathway in the response of doxorubicin-induced DNA damages in acute lymphoblastic leukemia cells.

Doxorubicin (Dox) is one of the most commonly used anthracyclines for the treatment of solid and hematological tumors such as B-/T cell acute lymphoblastic leukemia (ALL). Dox compromises topoisomerase II enzyme functionality, thus inducing structural damages during DNA replication and causes direct damages intercalating into DNA double helix. Eukaryotic cells respond to DNA damages by activating the ATM-CHK2 and/or ATR-CHK1 pathway, whose function is to regulate cell cycle progression, to promote damage repair, and to control apoptosis. We evaluated the efficacy of a new drug schedule combining Dox and specific ATR (VE-821) or CHK1 (prexasertib, PX) inhibitors in the treatment of human B-/T cell precursor ALL cell lines and primary ALL leukemic cells. We found that ALL cell lines respond to Dox activating the G2/M cell cycle checkpoint. Exposure of Dox-pretreated ALL cell lines to VE-821 or PX enhanced Dox cytotoxic effect. This phenomenon was associated with the abrogation of the G2/M cell cycle checkpoint with changes in the expression pCDK1 and cyclin B1, and cell entry in mitosis, followed by the induction of apoptosis. Indeed, the inhibition of the G2/M checkpoint led to a significant increment of normal and aberrant mitotic cells, including those showing tripolar spindles, metaphases with lagging chromosomes, and massive chromosomes fragmentation. In conclusion, we found that the ATR-CHK1 pathway is involved in the response to Dox-induced DNA damages and we demonstrated that our new in vitro drug schedule that combines Dox followed by ATR/CHK1 inhibitors can increase Dox cytotoxicity against ALL cells, while using lower drug doses. • Doxorubicin activates the G2/M cell cycle checkpoint in acute lymphoblastic leukemia (ALL) cells. • ALL cells respond to doxorubicin-induced DNA damages by activating the ATR-CHK1 pathway. • The inhibition of the ATR-CHK1 pathway synergizes with doxorubicin in the induction of cytotoxicity in ALL cells. • The inhibition of ATR-CHK1 pathway induces aberrant chromosome segregation and mitotic spindle defects in doxorubicin-pretreated ALL cells.

ETV6::ABL1-Positive Myeloid Neoplasm: A Case of a Durable Response to Imatinib Mesylate without Additional or Previous Treatment.

ETV6::ABL1 rearranged neoplasms are rare hematological diseases. To date, about 80 cases have been reported, including myeloid and lymphoid leukemias. The ETV6 gene codes for an ETS family transcription factor and several fusion partners have been described. When translocated, ETV6 causes the constitutive activation of the partner genes. Here, we report the case of a 54-year-old woman with a cryptic insertion of the 3' region of ABL1 in the ETV6 gene. The patient was first diagnosed with idiopathic hypereosinophilic syndrome, according to the clinical history, conventional cytogenetics, standard molecular analyses and pathologist description. Next generation sequencing of diagnosis samples unexpectedly detected both ETV6::ABL1 type A and B fusion transcripts, which were then confirmed by FISH. The diagnosis was Myeloid/Lymphoid neoplasm with ETV6::ABL1 fusion, and the patient received imatinib mesylate treatment. In a follow-up after more than one year, the patient still maintained the molecular and complete hematological responses. This case highlights the importance of timely and proper diagnostics and prompt tyrosine kinase inhibitor treatment.

CDC20 in and out of mitosis: a prognostic factor and therapeutic target in hematological malignancies.

Cell division cycle 20 homologue (CDC20) is a well-known regulator of cell cycle, as it controls the correct segregation of chromosomes during mitosis. Many studies have focused on the biological role of CDC20 in cancer development, as alterations of its functionality have been linked to genomic instability and evidence demonstrated that high CDC20 expression levels are associated with poor overall survival in solid cancers. More recently, novel CDC20 functions have been demonstrated or suggested, including the regulation of apoptosis and stemness properties and a correlation with immune cell infiltration. Here, we here summarize and discuss the role of CDC20 inside and outside mitosis, starting from its network of interacting proteins. In the last years, CDC20 has also attracted more interest in the blood cancer field, being overexpressed and showing an association with prognosis both in myeloid and lymphoid malignancies. Preclinical findings showed that selective CDC20 and APC/CCDC20/APC/CCDH1 inhibitors, namely Apcin and proTAME, are effective against lymphoma and multiple myeloma cells, resulting in mitotic arrest and apoptosis and synergizing with clinically-relevant drugs. The evidence and hypothesis presented in this review provide the input for further biological and chemical studies aiming to dissect novel potential CDC20 roles and targeting strategies in hematological malignancies.

Targeting PARP proteins in acute leukemia: DNA damage response inhibition and therapeutic strategies.

The members of the Poly(ADP-ribose) polymerase (PARP) superfamily are involved in several biological processes and, in particular, in the DNA damage response (DDR). The most studied members, PARP1, PARP2 and PARP3, act as sensors of DNA damages, in order to activate different intracellular repair pathways, including single-strand repair, homologous recombination, conventional and alternative non-homologous end joining. This review recapitulates the functional role of PARPs in the DDR pathways, also in relationship with the cell cycle phases, which drives our knowledge of the mechanisms of action of PARP inhibitors (PARPi), encompassing inhibition of single-strand breaks and base excision repair, PARP trapping and sensitization to antileukemia immune responses. Several studies have demonstrated a preclinical activity of the current available PARPi, olaparib, rucaparib, niraparib, veliparib and talazoparib, as single agent and/or in combination with cytotoxic, hypomethylating or targeted drugs in acute leukemia, thus encouraging the development of clinical trials. We here summarize the most recent preclinical and clinical findings and discuss the synthetic lethal interactions of PARPi in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL). Despite the low frequency of genomic alterations of PARP and other DDR-related genes in acute leukemia, selective vulnerabilities have been reported in several disease subgroups, along with a "BRCAness phenotype." AML carrying the RUNX1-RUNX1T1 or PML-RARA fusion genes or mutations in signaling genes (FLT3-ITD in combination with TET2 or TET2 and DNMT3A deficiency), cohesin complex members (STAG2), TP53 and BCOR as co-occurring lesions, IDH1/2 and ALL cases expressing the TCF3-HLF chimera or TET1 was highly sensitive to PARPi in preclinical studies. These data, along with the warning coming from the observation of cases of therapy-related myeloid malignancies among patients receiving PARPi for solid tumors treatment, indicate that PARPi represents a promising strategy in a personalized medicine setting. The characterization of the clonal and subclonal genetic background and of the DDR functionality is crucial to select acute leukemia patients that will likely benefit of PARPi-based therapeutic regimens.

Therapeutic Targeting of Acute Myeloid Leukemia by Gemtuzumab Ozogamicin.

Acute myeloid leukemia (AML) is a complex hematological malignancy characterized by genetic and clinical heterogeneity and high mortality. Despite the recent introduction of novel pharmaceutical agents in hemato-oncology, few advancements have been made in AML for decades. In the last years, the therapeutic options have rapidly changed, with the approval of innovative compounds that provide new opportunities, together with new challenges for clinicians: among them, on 1 September, 2017 the Food and Drug Administration granted approval for Gemtuzumab Ozogamicin (GO) in combination with daunorubicin and cytarabine for the treatment of adult patients affected by newly diagnosed CD33+ AML. Benefits of GO-based regimens were also reported in the pre- and post-transplantation settings. Moreover, several biomarkers of GO response have been suggested, including expression of CD33 and multidrug resistance genes, cytogenetic and molecular profiles, minimal residual disease and stemness signatures. Among them, elevated CD33 expression on blast cells and non-adverse cytogenetic or molecular risk represent largely validated predictors of good response.

Integrated genomic-metabolic classification of acute myeloid leukemia defines a subgroup with NPM1 and cohesin/DNA damage mutations.

Although targeting of cell metabolism is a promising therapeutic strategy in acute myeloid leukemia (AML), metabolic dependencies are largely unexplored. We aimed to classify AML patients based on their metabolic landscape and map connections between metabolic and genomic profiles. Combined serum and urine metabolomics improved AML characterization compared with individual biofluid analysis. At intracellular level, AML displayed dysregulated amino acid, nucleotide, lipid, and bioenergetic metabolism. The integration of intracellular and biofluid metabolomics provided a map of alterations in the metabolism of polyamine, purine, keton bodies and polyunsaturated fatty acids and tricarboxylic acid cycle. The intracellular metabolome distinguished three AML clusters, correlating with distinct genomic profiles: NPM1-mutated(mut), chromatin/spliceosome-mut and TP53-mut/aneuploid AML that were confirmed by biofluid analysis. Interestingly, integrated genomic-metabolic profiles defined two subgroups of NPM1-mut AML. One was enriched for mutations in cohesin/DNA damage-related genes (NPM1/cohesin-mut AML) and showed increased serum choline + trimethylamine-N-oxide and leucine, higher mutation load, transcriptomic signatures of reduced inflammatory status and better ex-vivo response to EGFR and MET inhibition. The transcriptional differences of enzyme-encoding genes between NPM1/cohesin-mut and NPM1-mut allowed in silico modeling of intracellular metabolic perturbations. This approach predicted alterations in NAD and purine metabolism in NPM1/cohesin-mut AML that suggest potential vulnerabilities, worthy of being therapeutically explored.

CART-Cell Therapy: Recent Advances and New Evidence in Multiple Myeloma.

Despite the improvement in survival outcomes, multiple myeloma (MM) remains an incurable disease. Chimeric antigen receptor (CAR) T-cell therapy targeting B-cell maturation antigen (BCMA) represents a new strategy for the treatment of relapsed/refractory MM (R/R). In this paper, we describe several recent advances in the field of anti-BCMA CAR T-cell therapy and MM. Currently, available data on anti-BCMA CART-cell therapy has demonstrated efficacy and manageable toxicity in heavily pretreated R/R MM patients. Despite this, the main issues remain to be addressed. First of all, a significant proportion of patients eventually relapse. The potential strategy to prevent relapse includes sequential or combined infusion with CAR T-cells against targets other than BCMA, CAR T-cells with novel dual-targeting vector design, and BCMA expression upregulation. Another dark side of CART therapy is safety. Cytokine release syndrome (CRS) andneurologic toxicity are well-described adverse effects. In the MM trials, most CRS events tended to be grade 1 or 2, with fewer patients experiencing grade 3 or higher. Another critical point is the extended timeline of the manufacturing process. Allo-CARs offers the potential for scalable manufacturing for on-demand treatment with shorter waiting days. Another issue is undoubtedly going to be access to this therapy. Currently, only a few academic centers can perform these procedures. Recognizing these issues, the excellent response with BCMA-targeted CAR T-cell therapy makes it a treatment strategy of great promise.

Pharmacological Inhibition of WIP1 Sensitizes Acute Myeloid Leukemia Cells to the MDM2 Inhibitor Nutlin-3a.

In acute myeloid leukemia (AML), the restoration of p53 activity through MDM2 inhibition proved efficacy in combinatorial therapies. WIP1, encoded from PPM1D, is a negative regulator of p53. We evaluated PPM1D expression and explored the therapeutic efficacy of WIP1 inhibitor (WIP1i) GSK2830371, in association with the MDM2 inhibitor Nutlin-3a (Nut-3a) in AML cell lines and primary samples. PPM1D transcript levels were higher in young patients compared with older ones and in core-binding-factor AML compared with other cytogenetic subgroups. In contrast, its expression was reduced in NPM1-mutated (mut, irrespective of FLT3-ITD status) or TP53-mut cases compared with wild-type (wt) ones. Either Nut-3a, and moderately WIP1i, as single agent decreased cell viability of TP53-wt cells (MV-4-11, MOLM-13, OCI-AML3) in a time/dosage-dependent manner, but not of TP53-mut cells (HEL, KASUMI-1, NOMO-1). The drug combination synergistically reduced viability and induced apoptosis in TP53-wt AML cell line and primary cells, but not in TP53-mut cells. Gene expression and immunoblotting analyses showed increased p53, MDM2 and p21 levels in treated TP53-wt cells and highlighted the enrichment of MYC, PI3K-AKT-mTOR and inflammation-related signatures upon WIP1i, Nut-3a and their combination, respectively, in the MV-4-11 TP53-wt model. This study demonstrated that WIP1 is a promising therapeutic target to enhance Nut-3a efficacy in TP53-wt AML.

Synthesis of Novel Tryptamine Derivatives and Their Biological Activity as Antitumor Agents.

We synthesized five novel tryptamine derivatives characterized by the presence of an azelayl chain or of a 1,1,1-trichloroethyl group, in turn connected to another heterocyclic scaffold. The combination of tryptamin-, 1,1,1-trichloroethyl- and 2-aminopyrimidinyl- moieties produced compound 9 identified as the most active compound in hematological cancer cell lines (IC50 = 0.57-65.32 µM). Moreover, keeping constant the presence of the tryptaminic scaffold and binding it to the azelayl moiety, the compounds maintain biological activity. Compound 13 is still active against hematological cancer cell lines and shows a selective effect only on HT29 cells (IC50 = 0.006 µM) among solid tumor models. Compound 14 loses activity on all leukemic lines, while showing a high level of toxicity on all solid tumor lines tested (IC50 0.0015-0.469 µM).

Therapeutic afucosylated monoclonal antibody and bispecific T-cell engagers for T-cell acute lymphoblastic leukemia.

BACKGROUND: T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive disease with a poor cure rate for relapsed/resistant patients. Due to the lack of T-cell restricted targetable antigens, effective immune-therapeutics are not presently available and the treatment of chemo-refractory T-ALL is still an unmet clinical need. To develop novel immune-therapy for T-ALL, we generated an afucosylated monoclonal antibody (mAb) (ahuUMG1) and two different bispecific T-cell engagers (BTCEs) against UMG1, a unique CD43-epitope highly and selectively expressed by T-ALL cells from pediatric and adult patients. METHODS: UMG1 expression was assessed by immunohistochemistry (IHC) on a wide panel of normal tissue microarrays (TMAs), and by flow cytometry on healthy peripheral blood/bone marrow-derived cells, on 10 different T-ALL cell lines, and on 110 T-ALL primary patient-derived cells. CD43-UMG1 binding site was defined through a peptide microarray scanning. ahuUMG1 was generated by Genetic Glyco-Engineering technology from a novel humanized mAb directed against UMG1 (huUMG1). BTCEs were generated as IgG1-(scFv)2 constructs with bivalent (2+2) or monovalent (2+1) CD3ε arms. Antibody dependent cellular cytotoxicity (ADCC), antibody dependent cellular phagocytosis (ADCP) and redirected T-cell cytotoxicity assays were analysed by flow cytometry. In vivo antitumor activity of ahUMG1 and UMG1-BTCEs was investigated in NSG mice against subcutaneous and orthotopic xenografts of human T-ALL. RESULTS: Among 110 T-ALL patient-derived samples, 53 (48.1%) stained positive (24% of TI/TII, 82% of TIII and 42.8% of TIV). Importantly, no expression of UMG1-epitope was found in normal tissues/cells, excluding cortical thymocytes and a minority (<5%) of peripheral blood T lymphocytes. ahUMG1 induced strong ADCC and ADCP on T-ALL cells in vitro, which translated in antitumor activity in vivo and significantly extended survival of treated mice. Both UMG1-BTCEs demonstrated highly effective killing activity against T-ALL cells in vitro. We demonstrated that this effect was specifically exerted by engaged activated T cells. Moreover, UMG1-BTCEs effectively antagonized tumor growth at concentrations >2 log lower as compared with ahuUMG1, with significant mice survival advantage in different T-ALL models in vivo. CONCLUSION: Altogether our findings, including the safe UMG1-epitope expression profile, provide a framework for the clinical development of these innovative immune-therapeutics for this still orphan disease.

CPX-351 daunorubicin-cytarabine liposome: a novel formulation to treat patients with newly diagnosed secondary acute myeloid leukemia.

Over the last few years, we assisted to an increasing knowledge about acute myeloid leukemia (AML) pathobiology. However, outcomes remain unsatisfactory particularly for adult patients over 60 years old. Not surprisingly several cases of therapy-related AML (tAML) and secondary AML, both characterized by poorer prognosis, are more common in older population. For several decades initial therapy for AML remained unchanged and typically treatment consisted of an anthracycline combined with continuous infusion of cytarabine for 7 days, the so-called "7+3" standard regimen. The efforts made by the researchers to improve this standard schedule, have led to only modest improvement in the response rate (RR) but no change in overall survival (OS), until the recent evolution seen with new target specific mutation therapies. In 2017, a new liposomal-encapsulated formulation with daunorubicin and cytarabine (CPX-351) was approved by the US Food and Drug Administration for the treatment of newly diagnosed tAML or AML with myelodysplasia-related changes (AML-MRCs). Based on the findings that ratiometric delivery may be more effective than administration of either drug at their maximum tolerated dose (MTD), CPX-351 was designed to deliver a fixed 5:1 molar ratio of the two molecules historically used in the standard "7+3" regimen, cytarabine and daunorubicin respectively. CPX-351 did show improvements of overall survival compared to traditional "7+3" in newly diagnosed secondary and therapy-related AML in adult patients. However, questions remain regarding how to select across AML patient subgroups to maximize the clinical benefit. Possible future directions include evaluating CPX-351 dose intensification, combining this liposomal formulation with targeted therapies and not least important a better understanding about the mechanism of improved responses in tAML and AML-MRC, two entities recognized to be less chemo-sensitive than other hematologic malignancies. In summary, CPX-351 offers finally something new in the landscape of AML therapy. Herein we will review the rationale behind this new drug product development, the main pharmacological characteristics, and discuss the results of clinical trials that led to its FDA approval at first and by EMA in 2018.

Tagraxofusp and anti-CD123 in blastic plasmacytoid dendritic cell neoplasm: a new hope.

Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is a rare hematological malignancy, characterized by poor prognosis if treated with conventional therapy. Allogenic hematologic stem cell transplant can improve survival and can be curative, but it is available in a small percentage of patients given that the median age at diagnosis is 70 years. In this scenario it is assumed that only the development of precision medicine-driven therapy will change BPDCN patient prognosis. CD123 (the α-subunit of interleukin (IL)-3 receptor) is over-expressed on BPDCN cells surface and seems to be the ideal marker to develop antibody-based therapies. Tagraxofusp (Elzonris®), a recombinant immunotoxin consisting of human interleukin-3 fused to a truncated diphtheria toxin, has been approved by FDA in December 2018 for the treatment of BPDCN in adult and pediatric patients. tagraxofusp has shown promising clinical activity, with a high overall response rate and quite manageable safety profile even in elderly patients. It seems to improve overall survival too, but comparative trials are necessary to confirm this. Adverse events are commonly reported and the most important are transaminitis, thrombocytopenia and capillary leak syndrome (CLS). Therefore, to prevent the onset of severe CLS is recommended to reserve tagraxofusp for patients with preserved hepatic and cardiac functions, and to strictly observe serum albumin level. Further studies are required to resolve many several unanswered questions about tagraxofusp. In this review, we will resume and discuss pharmacological characteristic of tagraxofusp, results of clinical trials leading to its approval by FDA in 2018 and future perspectives about its use in BPDCN and other hematological malignancies.

Gemtuzumab ozogamicin in acute myeloid leukemia: past, present and future.

After being in the therapeutic wilderness for several decades, acute myeloid leukemia has been recently thrust into the limelight with a series of drug approvals. Technical refinements in production, genetic manipulation and chemical modification of monoclonal antibodies led to growing interest in antibodies-based treatment strategies. Much of the focus of these efforts in acute myeloid leukemia has been on CD33 as a target. On September 2, 2017, the U.S. Food and Drug Administration approved gemtuzumab ozogamicin for treatment of relapsed or refractory CD33+ acute myeloid leukemia. This signals a new chapter in the long and unusual story of gemtuzumab ozogamicin, which was the first antibody-drug conjugate approved for human use by the Food and Drug Administration. In this review we have analyzed the history of this drug which, among several mishaps, is experiencing a second youth and still represents a field to be further explored.

A WEE1 family business: regulation of mitosis, cancer progression, and therapeutic target.

The inhibition of the DNA damage response (DDR) pathway in the treatment of cancer has recently gained interest, and different DDR inhibitors have been developed. Among them, the most promising ones target the WEE1 kinase family, which has a crucial role in cell cycle regulation and DNA damage identification and repair in both nonmalignant and cancer cells. This review recapitulates and discusses the most recent findings on the biological function of WEE1/PKMYT1 during the cell cycle and in the DNA damage repair, with a focus on their dual role as tumor suppressors in nonmalignant cells and pseudo-oncogenes in cancer cells. We here report the available data on the molecular and functional alterations of WEE1/PKMYT1 kinases in both hematological and solid tumors. Moreover, we summarize the preclinical information on 36 chemo/radiotherapy agents, and in particular their effect on cell cycle checkpoints and on the cellular WEE1/PKMYT1-dependent response. Finally, this review outlines the most important pre-clinical and clinical data available on the efficacy of WEE1/PKMYT1 inhibitors in monotherapy and in combination with chemo/radiotherapy agents or with other selective inhibitors currently used or under evaluation for the treatment of cancer patients.

Combined Oral Fentanyl Citrate and Midazolam as Premedication for Bone Marrow Aspiration and Biopsy in Patients with Hematological Malignancies: A Randomized, Controlled and Patient-Blinded Clinical Trial.

Bone marrow aspiration and biopsy (BMAB) is a painful procedure, and the routinely used local infiltration anesthesia (LIA) with lidocaine is unable to provide pain relief during the most uncomfortable phases. The primary endpoint of the present randomized, patient-blinded trial was to evaluate the efficacy of an opioid and benzodiazepine combination plus LIA (sedoanalgesia) in patients undergoing BMAB for hematological malignancies. The secondary endpoint was the safety of the procedure in an outpatient setting. Ancillary assessments were anticipatory anxiety related to pain recall in the event of re-biopsy, and adequacy of bone tissue harvested. Patients were randomly assigned to one of 2 arms to receive either sedoanalgesic placebo plus LIA (standard group) or oral fentanyl citrate 200 µg plus oral midazolam 5 mg plus LIA (combo group) during BMAB. Pre-procedural anxiety and procedural pain were assessed according to the Numerical Rating Scale (NRS: 0-10), dividing the time of the procedure into five intervals (T0, T1, T2a, T2b and T3) and evaluating the degree of discomfort at each time (T) in both groups. One hundred and sixteen patients were eligible for the study. At T2b (time of biopsy) and T3 (time after biopsy), a significantly lower perception of pain was registered in the combo group. Moreover, there were no significant sedoanalgesia-related side-effects. Finally, histological specimens were higher in quality in the combo group. Sedoanalgesia was highly effective in reducing pain during biopsy, diminished anticipatory anxiety in patients undergoing re-biopsy and led to fewer non-diagnostic specimens being harvested.