Pentostatin 10mg powder for solution for injection vials

Pentostatin (PNT), a nucleoside antibiotic with a 1,3-diazo ring structure, is distributed in several actinomycetes and fungi species. Its special structure makes PNT possess a wide spectrum of biological and pharmacological properties, such as antibacterial, antitrypanosomal, anticancer, antiviral, herbicidal, insecticidal, and immunomodulatory effects. Because of the promising adenosine deaminase inhibitory activity of PNT, its extensive application in the clinical treatment of malignant tumors has been extensively studied. However, the fermentation level of microbial-derived PNT is low and cannot meet medical needs. Because the biosynthesis pathway of PNT is obscure, only high-yield mutant screening and optimization of medium components and fermentation processes have been conducted for enhancing its production. Recently, the biosynthesis pathways of PNT in actinomycetes and fungi hosts have been revealed successively, and the large-scale production of PNT by systematic metabolic engineering will become an inevitable trend. Therefore, this review covers all aspects of PNT research, in which major advances in understanding the resource microorganisms, mechanism of action, and biosynthesis pathway of PNT were achieved and diverse clinical applications of PNT were emphasized, and it will lay the foundation for commercial transformation and industrial technology of PNT based on systematic metabolic engineering.

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Integrating immunotherapy with natural compounds holds promise in enhancing the immune system's ability to eliminate cancer cells. Cordyceps militaris, a traditional Chinese medicine, emerges as a promising candidate in this regard. This study investigates the effects of cordycepin and C. militaris ethanolic extract (Cm-EE) on sensitizing cancer cells and regulating immune responses against breast cancer (BC) and hepatocellular carcinoma (HCC) cells. Cordycepin, pentostatin and adenosine were identified in Cm-EE. Cordycepin treatment decreased HLA-ABC-positive cells in pre-treated cancer cells, while Cm-EE increased NKG2D ligand and death receptor expression. Additionally, cordycepin enhanced NKG2D receptor and death ligand expression on CD3-negative effector immune cells, particularly on natural killer (NK) cells, while Cm-EE pre-treatment stimulated IL-2, IL-6, and IL-10 production. Co-culturing cancer cells with effector immune cells during cordycepin or Cm-EE incubation resulted in elevated cancer cell death. These findings highlight the potential of cordycepin and Cm-EE in improving the efficacy of cancer immunotherapy for BC and HCC.

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Nonmyeloablative (NMA) conditioning is being used increasingly with success in matched related donor (MRD) and alternative donor allogeneic hematopoietic cell transplantation (allo-HCT) in individuals with sickle cell disease (SCD). Advantages include decrease toxicity and applicability in patients otherwise unable to tolerate conditioning regimens due to end-organ damage or age. We aimed to add to published data outcomes of two similar NMA conditioning protocols, termed Protocol 1 (ClinicalTrials.gov ID NCT00061568) and Protocol 2 (ClinicalTrials.gov ID: NCT02105766)) in mainly adult patients with SCD to evaluate the safety, toxicity, and success of these regimens in individuals at high-risk for poor transplantation-related outcomes. We also evaluated the tolerability and outcomes of Protocol 2, which included preconditioning immunodepletion, in patients at even higher risk of T cell-mediated rejection or plasma/B cell-mediated anti-donor erythrocyte antibody production-the latter due to ABO incompatibility or recipient RBC alloimmunization to a donor antigen. Finally, we evaluated the incidence and trajectory of mixed donor myeloid chimerism over time following allo-HCT. In this retrospective analysis of the 2 prospective phase 2 NMA transplant protocols, 91 individuals with SCD or transfusion-dependent β-thalassemia underwent MRD allo-HCT at the National Heart, Lung, and Blood Institute; regimens contained alemtuzumab, low-dose radiation, and sirolimus for graft-versus-host disease (GVHD) prophylaxis with or without preconditioning immunodepletion with pentostatin and oral cyclophosphamide (Protocol 2). In the total cohort of 91 transplantation recipients, outcomes were favorable with timely neutrophil and platelet engraftment (median, 21 days [range, 7 to 67 days] and 21 days [range, 10 to 112 days], respectively), minimal high-grade acute GVHD and no chronic GVHD, overall survival of 90%, sickle-free survival of 85%, and mixed donor myeloid chimerism in 43% at a median follow up of 7.3 years (range, 0.8 to 20 years). Most patients with mixed myeloid chimerism at 2-years post-HCT remained stable in their values. In analyzing each protocol separately, outcomes were comparable except for higher cytomegalovirus reactivation necessitating treatment in Protocol 2 without an associated increase in graft failure. In the combined cohort, graft failure occurred in 11 patients, and hematologic malignancy or abnormal cytogenetics on bone marrow evaluation developed in 7 patients. In a subanalysis of factors that may implicate transplantation outcomes, the number of RBC units transfused post-HCT was significantly higher in recipients with pre-HCT history of alloimmunization to donor RBC antigens. There was no difference in the number of RBC units transfused, duration of transfusion, or red cell engraftment in those with major ABO incompatibility; preconditioning immunodepletion and pretreatment with rituximab likely were helpful. Both NMA allo-HCT protocols were successful in achieving adequate engraftment and sickle-free survival with minimal toxicity, including in individuals with mixed donor myeloid chimerism. The addition of preconditioning immunodepletion was well-tolerated and reduced the rate of graft failure in high-risk recipients.

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The special structure of pentostatin causes it to possess a wide spectrum of biological and pharmacological properties, and it has been extensively employed to treat malignant tumors and is the first-line treatment for hairy cell leukemia. Pentostatin is mainly distributed in several actinomycetes and fungi species. However, its low titer in microbes is not able to meet medical needs. Here, we report a strain improvement strategy based on combined atmospheric and room-temperature plasma (ARTP) mutagenesis and ribosome engineering screening, as well as fermentation optimization, for enhanced pentostatin production. The original strain, Actinomadura sp. ATCC 39365, was treated with ARTP and screened by ribosome engineering to obtain one stable pentostatin high-yield mutant Actinomadura sp. S-15, which produced 86.35 mg/L pentostatin, representing a 33.79% increase compared to Actinomadura sp. ATCC 39365. qRT-PCR analysis revealed that pentostatin biosynthesis-related gene expression was significantly upregulated in Actinomadura sp. S-15. Then, to further enhance pentostatin production, the fermentation medium was optimized in flask culture and the pentostatin production of Actinomadura sp. S-15 reached 152.06 mg/L, which is the highest pentostatin production reported so far. These results demonstrate the effectiveness of combined ARTP mutation, ribosome engineering screening, and medium optimization for the enhancement of pentostatin production, and provide a methodology enabling the sustainable production of pentostatin on an industrial scale.

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Hairy cell leukemia (HCL) is caused by the kinase-activating BRAF-V600E mutation in > 95% of patients, making this disease sensitive to oral BRAF inhibitors [1-3]. Rare HCL cases lacking BRAF-V600E have been recently described that respond suboptimally to standard chemotherapy with purine analogs and harbor alternative, potentially targetable, kinase-activating short deletions of BRAF or missense mutations of MAP2K1/MEK1, the kinase phosphorylated by BRAF within the RAF → MEK → ERK signaling pathway [4, 5]. Here, we describe a 60-year-old male with HCL who had received multiple treatments over the previous 23 years, that is splenectomy and six subsequent lines of chemo-immunotherapy, with refractoriness to the last one (pentostatin plus rituximab) resulting in persistent cytopenias (hemoglobin 5.7 g/dL, platelets 27 000/mm3; neutrophils 1190/mm3). This case had clinico-pathological features typical of classic HCL, including “fried-egg” histological pattern (Figure 1A) with marked reticulin fibrosis (not shown), as well as circulating leukemic cells with circumferential hairy projections and inconspicuous nucleoli (Figure 1B). However, leukemic cells lacked BRAF-V600E (Figure 1C), yet showed prominent ERK phosphorylation (Figure 1D) that was due to a clonal activating Q56P mutation of MAP2K1/MEK1 identified upon next-generation sequencing. Indeed, in vitro treatment of patients' leukemic cells with the MEK inhibitor cobimetinib strongly dephosphorylated ERK and elicited marked apoptosis (not shown). Because the patient had exhausted all standard therapeutic options, before the genetic and functional characterization of the patient's leukemia was complete, he was empirically started on the BRAF inhibitor vemurafenib (960 mg twice daily for 8 days) plus rituximab 375 mg/sqm (for 1 dose), but was then quickly switched to oral cobimetinib (60 mg daily) for 4 cycles (each comprising 21 days of dosing followed by 7 days of rest), with resolution of cytopenias and achievement of a complete remission immunohistochemically confirmed through a bone marrow biopsy. Since recurrent grade-2 cutaneous rash due to cobimetinib precluded long-term continuous treatment with the drug, this response was consolidated with an additional two cycles of cobimetinib (mostly at a reduced dose of 40 mg daily) combined with rituximab (375 mg/sqm intravenously) for eight doses (3 during cobimetinib therapy and 5 thereafter). A complete remission was re-confirmed at the end of such treatment, including clearing of the MAP2K1/MEK1 mutation, and progression-free survival was 15 months from the start of treatment. MAP2K1 mutations were first identified in HCL-variant [6], a distinct disease that has a worse prognosis than classic HCL [7], and in a patient with relapsed HCL-variant harboring mutant MAP2K1 (K57N) treatment with the MEK inhibitor trametinib was only modestly active (stable disease after 6 cycles) [8]. The current case highlights the importance of correctly diagnosing classic HCL even in the absence of its hallmark BRAF-V600E mutation and of searching for alternative, targetable kinase mutations to optimally guide patient management. Supported by grants from Associazione Italiana per la Ricerca sul Cancro (“Metastasis 5-per-mille” grant number, 21198, to Dr. Enrico Tiacci) and the Hairy Cell Leukemia Foundation–Leukemia and Lymphoma Society (HCL2025 Initiative–Translational Research grant number, HCL8029-22, to Dr. Enrico Tiacci). Open access publishing facilitated by Universita degli Studi di Perugia, as part of the Wiley - CRUI-CARE agreement. This report was prepared in accordance with the Declaration of Helsinki. Written informed consent was obtained from the patient. All identifying details have been omitted or anonymized to protect patient confidentiality. The authors declare no conflicts of interest.

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Adenosine deaminase (ADA) is one of the most important enzymes in nucleoside metabolism, regulating the levels of adenosine and deoxyadenosine triphosphate (ADT/dATP) on either side of the cell membrane. This small protein (weighing approximately 40 kDa) exhibits deamination properties towards other pharmaceuticals built on adenine as the leading structure, which requires co-administration of ADA inhibitors. 3′-deoxyadenosine (Cordycepin, Cord) is an active compound isolated from the fungus Cordyceps, which has been used in traditional Chinese medicine for over 2000 years. Its anticancer activity is likely related to the inhibition of primer elongation of lagging strands during genetic information replication. Unfortunately, Cord is rapidly deaminated by ADA into inactive 3′-deoxyinosine, necessitating its co-administration with ADA inhibitors. Here, for the first time, the synthesis and discussion of the oxidised form of Cord are presented. The 7,8-dihydro-8-oxo-3′-deoxyadenosine (CordOXO) exhibits high resistance to ADA because of its syn conformation, as shown experimentally by UV spectroscopy and RP-HPLC monitoring. Theoretical Density Functional based Tight Binding (DFTB) studies of the Michaelis complex ADA-CordOXO have revealed significant distance increases between the “active” H2O molecule and C6 of the 8-oxo-adenine moiety of CordOXO, i.e., 4 Å as opposed to 2.7 Å in the cases of ADA-dAdo and Cord. In conclusion, it can be postulated that the conversion of Cord to CordOXO enhances its therapeutic potential; however, this needs to be verified in vitro and in vivo. It should be emphasised that the therapeutic effect, if any, can be achieved theoretically without ADA inhibitors, e.g., pentostatin, thus reducing adverse effects. These promising preliminary results, presented here, warrant further investigations.

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ABSTRACT: The primary objective in multiply relapsed hairy cell leukemia and variant (HCL/HCLv) was to determine whether pentostatin-rituximab (DCFR) and bendamustine-rituximab (BR) each achieve an overall response rate (ORR) exceeding that historically achieved by rituximab alone (∼40%) in favor of 65%. Prospective data were unreported for either regimen. Fifty-six patients received 6 28-day cycles of rituximab (375 mg/m2, days 1 and 15) with either bendamustine (90 mg/m2, days 1 and 2) or pentostatin (4 mg/m2, days 1 and 15). Eligibility required ≥2 purine analogs, or 1 purine analog plus rituximab for response of <1 year to the initial purine analog. Although patients were assigned to either regimen through randomization to increase homogeneity of the 2 treatment groups, the DCFR arm had fewer previous purine analogs (P = .021) and lower baseline marrow HCL/HCLv infiltration (P = .013). ORRs for DCFR and BR were 93% (95% confidence intervals [CI], 83-102) and 86%, (95% CI, 73-99), respectively, exceeding 40% (P< .0001) for each group. Rates for complete remission (CR) and minimal residual disease-free CR and median progression-free survival (141 vs 50 months; HR, 0.63; 95% CI, 0.32-1.25) numerically favored DCFR, but that arm was significantly enriched with less previous purine analogs and marrow infiltration, each of which was associated post hoc with better response. Post hoc subgroup analysis, particularly for 41 patients with classic HCL, suggested any superiority of DCFR vs BR might apply to patients with more favorable disease. DCFR and BR were highly effective in multiply relapsed HCL/HCLv. Possible DCFR superiority was hypothesis-generating, given uneven baseline risks and trial design. This trial was registered at www.clinicaltrials.gov as #NCT01059786.

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