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To discover new variants associated with low ovarian reserve after gonadotoxic treatment among adult female childhood cancer survivors using a genome-wide association study approach. Genome-wide association study. Not applicable. A discovery cohort of adult female childhood cancer survivors from the pan-European PanCareLIFE cohort (n = 743; median age: 25.8 years), excluding those who received bilateral ovarian irradiation, bilateral oophorectomy, central nervous system or total body irradiation, or stem cell transplantation. Replication was attempted in the US-based St. Jude Lifetime Cohort (n = 391; median age: 31.3 years). Female childhood cancer survivors are at risk of therapy-related gonadal impairment. Alkylating agents are well-established risk factors, and the interindividual variability in gonadotoxicity may be explained by genetic polymorphisms. Data were collected in real-life conditions, and cyclophosphamide equivalent doses were used to quantify alkylation agent exposure. Anti-Müllerian hormone (AMH) levels served as a proxy for ovarian function, and the findings were combined in a meta-analysis. Three genome-wide significant (<5.0 × 10−8) and 16 genome-wide suggestive (<5.0 × 10−6) loci were associated with log-transformed AMH levels, adjusted for cyclophosphamide equivalent dose of alkylating agents, age at diagnosis, and age at study in the PanCareLIFE cohort. On the basis of the effect allele frequency (EAF) (>0.01 if not genome-wide significant), and biologic relevance, 15 single nucleotide polymorphisms were selected for replication. None of the single nucleotide polymorphisms were statistically significantly associated with AMH levels. A meta-analysis indicated that rs78861946 was associated with borderline genome-wide statistical significance (reference/effect allele: C/T; effect allele frequency: 0.04, beta (SE): −0.484 (0.091). This study found no genetic variants associated with a lower ovarian reserve after gonadotoxic treatment because the findings of this genome-wide association study were not statistically significant replicated in the replication cohort. Suggestive evidence for the potential importance of 1 variant is briefly discussed, but the lack of statistical significance calls for larger cohort sizes. Because the population of childhood cancer survivors is increasing, large-scale and systematic research is needed to identify genetic variants that could aid predictive risk models of gonadotoxicity as well as fertility preservation options for childhood cancer survivors. [ABSTRACT FROM AUTHOR]

The human alkylation B (AlkB) homologs, ALKBH2 and ALKBH3, respond to methylation damage to maintain genomic integrity and cellular viability. Both ALKBH2 and ALKBH3 are direct reversal repair enzymes that remove 1-methyladenine (1meA) and 3-methylcytosine (3meC) lesions commonly generated by alkylating chemotherapeutic agents. Thus, the existence of deficiencies in ALKBH proteins can be exploited in synergy with chemotherapy. In this study, we investigated possible interactions between ALKBH2 and ALKBH3 with other proteins that could alter damage response and discovered an interaction with the mismatch repair (MMR) system. To test whether the lack of active MMR impacts ALKBH2 and/or ALKBH3 response to methylating agents, we generated cells deficient in ALKBH2, ALKBH3, or both in addition to Mlh homolog 1 (MLH1), another MMR protein. We found that MLH1koALKBH3ko cells showed enhanced resistance toward SN1- and SN2-type methylating agents, whereas MLH1koALKBH2ko cells were only resistant to SN1-type methylating agents. Concomitant loss of ALKBH2 and ALKBH3 (ALKBH2ko3ko) rendered cells sensitive to SN1- and SN2-agents, but the additional loss of MLH1 enhanced resistance to both types of damage. We also showed that ALKBH2ko3ko cells have an ATR-dependent arrest at the G2/M checkpoint, increased apoptotic signaling, and replication fork stress in response to methylation. However, these responses were not observed with the loss of functional MLH1 in MLH1koALKBH2ko3ko cells. Finally, in MLH1koALKBH2ko3ko cells, we observed elevated mutant frequency in untreated and temozolomide treated cells. These results suggest that obtaining a more accurate prognosis of chemotherapeutic outcome requires information on the functionality of ALKBH2, ALKBH3, and MLH1. [ABSTRACT FROM AUTHOR]

Persulfides (RSSH/RSS−) participate in sulfur metabolism and are proposed to transduce hydrogen sulfide (H2S) signaling. Their biochemical properties are poorly understood. Herein, we studied the acidity and nucleophilicity of several low molecular weight persulfides using the alkylating agent, monobromobimane. The different persulfides presented similar pKa values (4.6–6.3) and pH-independent rate constants (3.2–9.0 × 10³ M−1 s−1), indicating that the substituents in persulfides affect properties to a lesser extent than in thiols because of the larger distance to the outer sulfur. The persulfides had higher reactivity with monobromobimane than analogous thiols and putative thiols with the same pKa, providing evidence for the alpha effect (enhanced nucleophilicity by the presence of a contiguous atom with high electron density). Additionally, we investigated two enzymes from the human mitochondrial H2S oxidation pathway that form catalytic persulfide intermediates, sulfide quinone oxidoreductase and thiosulfate sulfurtransferase (TST, rhodanese). The pH dependence of the activities of both enzymes was measured using sulfite and/or cyanide as sulfur acceptors. The TST half-reactions were also studied by stopped-flow fluorescence spectroscopy. Both persulfidated enzymes relied on protonated groups for reaction with the acceptors. Persulfidated sulfide quinone oxidoreductase appeared to have a pKa of 7.8 ± 0.2. Persulfidated TST presented a pKa of 9.38 ± 0.04, probably due to a critical active site residue rather than the persulfide itself. The TST thiol reacted in the anionic state with thiosulfate, with an apparent pKa of 6.5 ± 0.1. Overall, our study contributes to a fundamental understanding of persulfide properties and their modulation by protein environments. [ABSTRACT FROM AUTHOR]

To determine the incidence of all-cause and cancer mortality (CM) in association with immunosuppression. Retrospective cohort study at ocular inflammatory disease (OID) subspecialty centers. We harvested exposure and covariate data retrospectively from clinic inception (earliest in 1979) through 2010 inclusive. Then we ascertained overall and cancer-specific mortalities by National Death Index linkage. We constructed separate Cox models to evaluate overall and CM for each class of immunosuppressant and for each individual immunosuppressant compared with person-time unexposed to any immunosuppression. Patients with noninfectious OID, excluding those with human immunodeficiency infection or preexisting cancer. Tumor necrosis factor (TNF) inhibitors (mostly infliximab, adalimumab, and etanercept); antimetabolites (methotrexate, mycophenolate mofetil, azathioprine); calcineurin inhibitors (cyclosporine); and alkylating agents (cyclophosphamide) were given when clinically indicated in this noninterventional cohort study. Overall mortality and CM. Over 187 151 person-years (median follow-up 10.0 years), during which 15 938 patients were at risk for mortality, we observed 1970 deaths, 435 due to cancer. Both patients unexposed to immunosuppressants (standardized mortality ratio [SMR] = 0.95, 95% confidence interval [CI], 0.90–1.01) and those exposed to immunosuppressants but free of systemic inflammatory diseases (SIDs) (SMR = 1.04, 95% CI, 0.95–1.14) had similar mortality risk to the US population. Comparing patients exposed to TNF inhibitors, antimetabolites, calcineurin inhibitors, and alkylating agents with patients not exposed to any of these, we found that overall mortality (adjusted hazard ratio [aHR] = 0.88, 0.89, 0.90, 1.11) and CM (aHR = 1.25, 0.89, 0.86, 1.23) were not significantly increased. These results were stable in sensitivity analyses whether excluding or including patients with SID, across 0-, 3-, or 5-year lags and across quartiles of immunosuppressant dose and duration. Our results, in a cohort where the indication for treatment was proven unassociated with mortality risk, found that commonly used immunosuppressants—especially the antimetabolites methotrexate, mycophenolate mofetil, and azathioprine; the TNF inhibitors adalimumab and infliximab, and cyclosporine—were not associated with increased overall and CM over a median cohort follow-up of 10.0 years. These results suggest the safety of these agents with respect to overall and CM for patients treated with immunosuppression for a wide range of inflammatory diseases. Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article. [ABSTRACT FROM AUTHOR]

An important target in toxicology is the ion channel known as human transient receptor potential ankyrin 1 (hTRPA1). It is triggered by a variety of chemicals, including the alkylating chemical warfare agent sulfur mustard (SM). The activation potentials of structural analogs including O- and sesquimustard, nitrogen mustards (HN1, HN2, and HN3), and related chemotherapeutic drugs (bendamustine, cycylophosphamide, and ifosfamide) were examined in the current study. The aequorin assay was used to measure changes in intracellular calcium levels in human hTRPA1 overexpressing HEK293 cells. The XTT assay was used to determine cytotoxicity. The data presented here highlight that all investigated alkylating substances, with the exception of cyclophosphamide and ifosfamide, cause the activation of hTRPA1. Cytotoxicity and activation of hTRPA1 were found to be related. Compounds with high reactivity had higher cytotoxicity and vice versa. However, inhibiting hTRPA1 with the specific inhibitor AP18 could not reduce the cytotoxicity induced by alkylating agents. As a result, hTRPA1 does not play a significant role in the cytotoxicity of alkylating agents. • hTRPA1 is activated by SM-related alkylating compounds. • Cytotoxicity and hTRPA1 activation were shown to be related. • Substances that prominently activate hTRPA1 were shown to be more cytotoxic. • hTRPA1 inhibition does not generally reduce cytotoxicity. [ABSTRACT FROM AUTHOR]

Glioblastoma (GB) is the worst and most common primary brain tumor. Temozolomide (TMZ), an alkylating agent, is widely used for treating primary and recurrent high-grade gliomas. However, at least 50% of TMZ treated patients do not respond to TMZ and the development of chemoresistance is a major problem. Here, we designed a lipid nanoemulsion containing a thermoresponsive polymer (poloxamer 407) aiming to improve TMZ release into the brain via nasal delivery. Increasing amounts of poloxamer 407 were added to preformed nanoemulsions (250 nm-range) obtained by spontaneous emulsification. The influence of the polymer concentration (from 2.5% to 12.5%) and temperature on viscosity was clearly evidenced. Such effect was also noticed on the mucoadhesiveness of formulations, as well as TMZ release rate and retention/permeation through nasal porcine mucosa using Franz-type diffusion cells. From these results, a formulation containing 10% of poloxamer (NTMZ-P10) was selected for further experiments by nasal route. A significantly higher TMZ amount was observed in the brain of rats from NTMZ-P10 in comparison with controls. Finally, our results show that formulation reduced significantly tumor growth by three-fold: 103.88 ± 43.67 mm3 (for NTMZ-P10) and 303.28 ± 95.27 mm3 (control). Overall, these results suggest the potential of the thermoresponsive formulation, administered by the non-invasive nasal route, as a future effective glioblastoma treatment. [Display omitted] [ABSTRACT FROM AUTHOR]

Busulfan is an alkylating agent commonly used in cancer chemotherapy. It is also an ideal agent for preparing transplant recipients of spermatogonial stem cells because of its high efficiency in destroying endogenous germ cells in the testis. However, its toxicity mechanism remains unclear, affecting its clinical use and applications. Based on reports of busulfan causing orchitis and a previous study by our team, this article summarizes the relationship between busulfan and orchitis, cytokines, the blood–testis barrier, and the cytoskeleton, unravels the regulatory pathways and mechanism behind busulfan-induced orchitis, and reveals the molecular mechanism underlying impaired spermatogenic function in orchitis, providing new ideas for the clinical application of busulfan while reducing its testicular toxicity. • This article reviewed the relationship between busulfan, orchitis, and cytokines. • Analysed the effects of orchitis on the cytoskeleton and BTB. • Infer the possible regulatory pathways by Busulfan. • This article explored the regulatory mechanisms of busulfan-induced orchitis. [ABSTRACT FROM AUTHOR]

Conditioning regimens with high-dose chemotherapy and autologous stem cell transplantation (ASCT) are the mainstays of treatment in lymphoma patients. Although the most frequently used conditioning regimen is the BEAM regimen (Carmustine, Etoposide, Cytarabine, and Melphalan), and alternatives are also used in certain circumstances. The TEAM regimen (carmustine is substituted by the alkylating agent thiotepa) is one of these alternatives; however, data regarding the comparisons of efficacy and safety profiles of these 2 regimens is scarce. This study compared the outcomes of patients who received conditioning regimens with BEAM and TEAM and underwent an ASCT. This study was conducted as a retrospective assessment of 294 patient outcomes in terms of efficacy and safety. Adult patients with lymphoma diagnosis who received BEAM or TEAM conditioning regimens and underwent an ASCT between January 1, 2016 and December 31, 2019 were included in the analyses. A total of 294 patients (median age at ASCT: 50 years, males: 60.5%, diffuse large B-cell lymphoma: 35%) were included. Eighty patients (27.2%) received the TEAM regimen, and 214 (72.8%) received the BEAM regimen. Regarding safety profiles, the thrombocyte engraftment time was significantly higher in the TEAM group (P =.003) and fever of unknown etiology was significantly higher in the BEAM group (P =.042). Also, nausea was more in the TEAM group (P =.031). The complete remission rate was 57.5% and 70.3% in the TEAM and BEAM regimens, respectively. The overall mortality rate was 37.3% and not significantly different between the groups (43% and 35% in the TEAM and BEAM groups, P =.22) over a similar median follow-up of 1667 days (P =.28). The 3-year survival rate was 66% and 67% and the 5-year survival rate was 52% and 58% in the TEAM and BEAM regimens, respectively, without significant difference. To the best of our knowledge, this is one of the few studies in the literature that compared the TEAM and BEAM as conditioning regimens for ASCT in lymphoma patients. The 2 regimens may provide similar overall survival outcomes and have a comparable safety profile. Although the BEAM regimen may be associated with longer progression-free survival times, the difference may be covered by the similar survival after ASCT. [ABSTRACT FROM AUTHOR]

A soluble, painless, photothermally enhanced detoxification MNA coated with a ZnInS/UIO catalyst (named ZnInS/UIO-MNA) for the treatment of CEES-poisoned wounds under SSL irradiation has been proposed. [Display omitted] • ZnInS/UIO can rapidly degradate CEES under SSL. • Microneedles array can effectively transport ZnInS/UIO to the subcutaneous. • ZnInS/UIO-MNA reduces inflammation and promotes healing of poisoned wounds. Sulfur mustard (2,2′-dichloroethylsulfide; SM) is a bifunctional alkylating agent that can easily penetrate skin and cause persistent pain and damage. Effective biological dressings are required to treat wounds caused or poisoned by SM. Though the use of SM is regulated under the Chemical Weapons Convention, it is still a threat during wars and terrorist attacks. Herein, we present a photothermal-enhanced detoxification microneedles array (MNA) encapsulated with ZnIn 2 S 4 @UiO-66-NH 2 (ZnInS/UIO) catalysts for the treatment of 2-chloroethyl ethyl sulfide (CEES, SM analog)-poisoned wounds under simulated sunlight (SSL) irradiation. Due to the excellent photothermal detoxification capability possessed by ZnInS/UIO, the conversion rate of CEES can be significantly increased under SSL exposure. When encased in a polyvinyl alcohol (PVA) MNA and piercing into the skin, ZnInS/UIO catalysts can be released quickly from MNA for detoxification. After applying the resultant ZnInS/UIO-MNA to the CEES-poisoned wound bed, acceleration of the wound healing process and a reduced inflammatory response can be confirmed. In conclusion, ZnInS/UIO-MNA has encouraging potential as a first-aid dressing for CEES-poisoned wound healing in battlefields and injuries related to acts of terrorism. [ABSTRACT FROM AUTHOR]

To determine whether there is an interplay effect between radiation dose and alkylating agent dose for the endpoint of local control in patients with Group III RMS. The study population consisted of patients with Group III RMS enrolled on Children's Oncology Group (COG) studies D9602, D9803, ARST0331 or ARST0531. Data collected included patient, gender, histology, tumor site, TNM stage, radiation (RT) dose and fractionation, and local, regional or distant failure. RT BED using an α/β = 10 was calculated for each patient, and cyclophosphamide equivalent dose (CED) was assigned to patients based on the study treatment arm to which they were assigned. Local failure (LF) was defined as progression or relapse at the primary tumor site and was calculated from time of study entry. BED was segmented into 5 "bins" (< 42, 42-46, 47-52, 53-58, and > 58) and CED was segmented into 4 bins (< = 4.8, 8.4, 16.8, and > 20). Chi square or Fisher exact tests were used to compare patient characteristics. Logistic regression analysis was used to assess the impact of a variety of BED, CED, and clinicopathologic factors on LF. There were 792 patients who met inclusion criteria. Multivariable analysis of tumor/demographic factors showed tumor size > 5 cm to be predictive of local failure as seen in prior reports (P < 0.01). No trend for local control was observed for combinations of BED and CED. This analysis failed to show a relationship between local control and various combinations of BED and CED for patients with Group III RMS. Tumor size > 5 cm was again shown to be predictive of local failure. Additional analysis including results of the recently completed ARST1431 study will be performed to further test the hypothesis that BED and CED both contribute to local control in patients with Group III RMS. [ABSTRACT FROM AUTHOR]

[Display omitted] • The yield of benzene alkylation can achieve 99% under the optimal process conditions. • The ionic liquid catalyst achieve continuous flow with slug flow pattern. • Industrial mixed olefins cause faster deactivation of ionic liquids compared to pure olefins. Alkylbenzene (LAB) is an important benzene derivative that can be used as a raw material for high-performance anionic surfactants such as alkylbenzene sulfonates (LAS) to meet the new requirements for surfactants in many fields. LAB can be synthesized through Friedel-Crafts reaction in the presence of acidic catalysts as alkylating agents, which is commonly used in the production of fine chemicals. In this work, a novel and efficient continuous flow micro reaction system was introduced to enhance the production of long-chain alkylbenzene catalyzed by [(CH 3) 3 NH][Al 2 Cl 7 ] ionic liquid. The optimal conditions were investigated for the synthesis of long-chain alkylbenzenes with the 1-tetradecene and mixing α-alkenes of C 14 -C 16 from Fischer-Tropsch process. The results showed that the yield of benzene alkylation can achieve 99 % under the optimal process conditions which the reaction tube length was 2000 mm, the reaction temperature was 30 ℃, the catalyst dosage was 0.30 mL/min and the molar ratio of aluminum chloride to trimethylamine hydrochloride was 2:1. In addition, α-alkenes with C 14 -C 16 lead to deactivation of the ionic liquids catalyst easier because of transfer of AlCl 3 from the ionic liquid to the organic phase compared to the pure 1-tetradecene as an alkylating agent. Continuous synthesis of the long-chain alkylbenzenes catalyzed by ionic liquids can achieve in the microchannel reaction system by controlling the flow patterns. [ABSTRACT FROM AUTHOR]

[Display omitted] • Deposition of FLa on a MALDI target is a facile and low-cost procedure. • Stable coating of the target surface with FLa is achieved. • Human globin adducts with chlorinated compounds are enriched in on-target workflow. • FLa shows excellent selectivity towards chlorine-containing human globin adducts. "Lab-on-a-plate" methodology encapsulates a number of different approaches aimed to shorten extensive sample processing for matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) analysis. Lately, we have proposed two approaches to the enrichment of chlorine-containing adducts of human globin (HHb) in a "lab-on-a-plate" format based on different techniques for the sorbent application to MALDI target spots: (i) droplet-free electrospraying of metal oxide nanoparticles and (ii) deposition of lanthanum stearate thin films (FLa). The latter approach has turned out to be more time- and cost-effective and, therefore, seems to be more promising. In this work, we assessed the quantitative characteristics (sensitivity, selectivity) of the "thin-film" procedure in order to estimate its analytical potential. The FLa-modified MALDI target was used for the on-spot enrichment of HHb adducts from the tryptic digests of the protein incubated with two model mono- and dichlorine-containing alkylating agents. The sensitivity of the technique is in the range of hundreds of fmol. Furthermore, the FLa-based procedure shows high selectivity (maximum molar ratio of modified/unmodified form of LLGNVLVC 112 VLAHHFGK is 1:500). As a case study, on-plate extraction of HHb adducts, formed due to the interaction of the protein with in vitro generated diclofenac metabolites, was performed on FLa-functionalized spots. This resulted in specific extraction of the HHb adducts with two diclofenac oxidation products from the tryptic digest of modified HHb. The developed approach appears to be effective for in situ enrichment of chlorine-containing adducts of HHb. [ABSTRACT FROM AUTHOR]

[Display omitted] • Studying the alkylation processes of nucleotides using a spectroscopic method. • Observing severe changes in spectra due to incubation of nucleotides with Melphalan. • The SERS method could produce enhanced Raman signals of drug compared to its powder sample. • The dramatic intensity of Raman spectra of nucleotides with AgNPs in solid state. • High intensity of SERS signals of AgNPs in solid state compared to liquid form. SERS (Surface Enhanced Raman Spectroscopy) is a new Raman spectroscopy which relies on Surface Plasmon Resonance (SPR) of metal nanoparticles. We have applied colloidal silver and gold nanoparticles as amplifier agents to enhance nucleotide Raman signals. It is observed that without these enhancing agents, it is impossible to investigate nucleotide spectrum due to weak Raman signals. Interaction mechanism of Melphalan, an anticancer drug with four nucleotides (Adenine, Cytosine, Guanine, Thymine) was investigated using SERS to detect and identify changes due to alkylating process in Raman spectra. After incubating Melphalan drug with nucleotides for 24 h at 37 °C, some changes occurred in SERS spectrum and interpretation of SERS spectra revealed the influence of the alkyl substitution on peaks and Raman shifts. After incubation of Melphalan with each nucleotide, intensity of relevant SERS signals assigned to Amid III group of Cytosine and Amid I of Thymine decreased significantly, confirming alkylating taking place. In this study, we also investigated the effect of nanoparticles type on nucleotide spectrum. We could not obtain useful information in the cases of guanine nucleotide. The SERS spectrum of Cytosine as an example of nucleotides in aqueous solution compared to solid state and results demonstrated that in solid state better signals were obtained than in liquid state. [ABSTRACT FROM AUTHOR]

The human DNA repair enzyme AlkB homologue-2 (ALKBH2) repairs methyl adducts from genomic DNA and is overexpressed in several cancers. However, there are no known inhibitors available for this crucial DNA repair enzyme. The aim of this study was to examine whether the first-generation HIV protease inhibitors having strong anti-cancer activity can be repurposed as inhibitors of ALKBH2. We selected four such inhibitors and performed in vitro binding analysis against ALKBH2 based on alterations of its intrinsic tryptophan fluorescence and differential scanning fluorimetry. The effect of these HIV protease inhibitors on the DNA repair activity of ALKBH2 was also evaluated. Interestingly, we observed that one of the inhibitors, ritonavir, could inhibit ALKBH2-mediated DNA repair significantly via competitive inhibition and sensitized cancer cells to alkylating agent methylmethane sulfonate (MMS). This work may provide new insights into the possibilities of utilizing HIV protease inhibitor ritonavir as a DNA repair antagonist. • Four HIV drugs with anti-cancer properties were evaluated as ALKBH2 antagonist. • Intrinsic Trp fluorescence quenching showed that ritonavir binds to ALKBH2. • Thermal unfolding showed ritonavir binding requires ALKBH2 R110 residue. • Enzyme kinetics results suggested that ritonavir competitively inhibits ALKBH2. • Ritonavir exposure reduced cancer cell survival following MMS treatment. [ABSTRACT FROM AUTHOR]

Temozolomide (TMZ), an imidazotetrazine, is a second-generation DNA alkylating agent used as a first-line treatment of glioblastoma multiforme (GBM). It was approved by FDA in 2005 and declared a blockbuster drug in 2008. Although TMZ has shown 100% oral bioavailability and crosses the blood-brain barrier effectively, however it suffers from limitations such as a short half-life (∼1.8 h), rapid metabolism, and lesser accumulation in the brain (∼10–20%). Additionally, development of chemoresistance has been associated with its use. Since it is a potential chemotherapeutic agent with an unmet medical need, advanced delivery strategies have been explored to overcome the associated limitations of TMZ. Nanocarriers including liposomes, solid lipid nanoparticles (SLNs), nanostructure lipid carriers (NLCs), and polymeric nanoparticles have demonstrated their ability to improve its circulation time, stability, tissue-specific accumulation, sustained release, and cellular uptake. Because of the appreciable water solubility of TMZ (∼5 mg/mL), the physical loading of TMZ in these nanocarriers is always challenging. Alternatively, the conjugation approach, wherein TMZ has been conjugated to polymers or small molecules, has been explored with improved outcomes in vitro and in vivo. This review emphasized the practical evidence of the conjugation strategy to improve the therapeutic potential of TMZ in the treatment of glioblastoma multiforme. [Display omitted] [ABSTRACT FROM AUTHOR]

The busulfan, an alkylating agent, suppresses endogenous spermatogenesis in recipient testes. However, considering a wide variation in the effects of busulfan among animal species, its dosage and route of infusion need optimization to prepare effective and safe recipients. Thus, the current study aimed to create a suitable recipient goat model for germ cell (Gc) transplantation through a single intra-testicular (i.t.) busulfan infusion under ultrasonographic (USG) guidance. As observed through the infusion of trypan blue under USG guidance into mediastinum testis (MT) of pre-pubertal Barbari bucks, 3–5 mL of trypan blue solution could fill almost 80% of seminiferous tubules. Thereafter, in Experiment-1, the effect of different busulfan doses (mg/kg) i.e. 0 [negative control, Group (Gr) 1; 0 mg/kg-MT], 1 (Gr 2; 1 mg/kg-MT), 2 (Gr 3; 2 mg/kg-MT), and 3 (Gr 4; 3 mg/kg-MT) were studied. Further, in Experiment-2, sterilizing effects of busulfan infusion through two different routes [MT or cavum vaginale (CV)] were compared. Following i.t. busulfan treatment, no adverse physiological effects or body weight loss were detected. The histological analyses demonstrate a dose-dependent depletion of Gc with almost complete loss of Gc and spermatogenic activities in Gr 3 and 4, and extensive fibrosis in Gr 4. A considerable suppression of spermatogenesis marked with devoid of endogenous spermatogonial population and absence of significant (P > 0.05) effect on key hematological variables were observed in 2 mg/kg-MT Gr. These findings coupled with the results of significant (P < 0.05) down-regulation of marker genes of undifferentiated spermatogonia (THY-1 and PLZF), Gc pluripotency (UCHL-1 , OCT-4, and DDX-4), and adhesion (E-cadherin and β -integrin); up-regulation of apoptotic genes (ID - 4 and BCL-6), and unchanged expression of Sertoli cell marker (vimentin), confirmed the safe and efficient depletion of endogenous Gc in 2 mg/kg-MT Gr. Furthermore, the effect of busulfan infusion on scrotal-testicular biometry, endocrine variables (plasma cortisol and testosterone), and Gc removal was more evident when busulfan was infused into MT than into CV. Overall, the results demonstrated that 2.0 mg/kg is an optimal single dose of busulfan when infused into the MT under USG guidance for the preparation of pre-pubertal recipient bucks. Overall, this study provides a basis to prepare suitable recipients through providing an available niche for efficient Gc transplantation in goats. • Intra-testicular busulfan infusion resulted into a dose-dependent depletion of germ-cells. • Busulfan infusion triggered down-regulation of pluripotency and adhesion marker genes in the testis. • Scrotal-testicular biometry variables affected more when busulfan is infused into MT. • Hematological variables mostly remain unaffected after intra-testicular busulfan infusion. • 2 mg/kg is the most optimal busulfan dose when infused into the MT with USG guidance. [ABSTRACT FROM AUTHOR]

Metallodrugs are important for anticancer treatments. They bind mainly to human serum albumin (HSA) in blood circulation, greatly modulating their pharmacokinetics and anticancer efficacy. Fatty acid (FA) is one of the most important endogenous ligands of HSA with tight binding to HSA and affecting its conformation. However, the effect of fatty acids on metallodrugs interaction with HSA is unknown. Here we identify the binding sites of a widely used metallodrug, cisplatin, in HSA in the presence or absence of a representative fatty acid, myristate, by X-ray crystallography. Our crystal structures indicate that the sidechain of residue Met548 becomes more exposed to solvent in the presence of fatty acid, and is the main Pt binding site together with Met329 in HSA:Myr:cisplatin ternary structure. An undoubted new Pt binding site is detected at His338 in the presence of fatty acid, and additional two sites are also identified at His146 and His440 + K436. In addition, we revealed the mechanism of cisplatin-induced HSA aggregation, which is due to the crosslinking between Met298 and His510 of two HSA molecules. [ABSTRACT FROM AUTHOR]

Background: Mosunetuzumab is a CD20 × CD3 T-cell-engaging bispecific monoclonal antibody that redirects T cells to eliminate malignant B cells. In a phase 1 study, mosunetuzumab was well tolerated and active in patients with relapsed or refractory B-cell lymphoma. We, therefore, aimed to evaluate the safety and anti-tumour activity of fixed-duration mosunetuzumab in patients with relapsed or refractory follicular lymphoma who had received two or more previous therapies.Methods: We conducted a single-arm, multicentre, phase 2 study at 49 centres in seven countries (Australia, Canada, Germany, South Korea, Spain, UK, and USA). All patients were aged 18 years or older with histologically confirmed follicular lymphoma (grade 1-3a) and an Eastern Cooperative Oncology Group performance status of 0-1. Patients had disease that was relapsed or refractory to two or more previous lines of treatment, including an anti-CD20 therapy and an alkylating agent. Intravenous mosunetuzumab was administered in 21-day cycles with cycle 1 step-up dosing: 1 mg on cycle 1 day 1, 2 mg on cycle 1 day 8, 60 mg on cycle 1 day 15 and cycle 2 day 1, and 30 mg on day 1 of cycle 3 and onwards. Patients with a complete response by investigator assessment using the International Harmonisation Project criteria completed treatment after cycle 8, whereas patients with a partial response or stable disease continued treatment for up to 17 cycles. The primary endpoint was independent review committee-assessed complete response rate (as best response) in all enrolled patients; the primary efficacy analysis compared the observed IRC-assessed complete response rate with a 14% historical control complete response rate in a similar patient population receiving the pan class I PI3K inhibitor copanlisib. Safety was assessed in all enrolled patients. This study is registered with ClinicalTrials.gov, number NCT02500407, and is ongoing.Findings: Between May 2, 2019, and Sept 25, 2020, we enrolled 90 patients. As of the data cutoff date (Aug 27, 2021), the median follow-up was 18·3 months (IQR 13·8-23·3). According to independent review committee assessment, a complete response was recorded in 54 patients (60·0% [95% CI 49·1-70·2]). The observed complete response rate was significantly higher than the historical control complete response rate with copanlisib of 14% (p<0·0001), thereby meeting the primary study endpoint. Cytokine release syndrome was the most common adverse event (40 [44%] of 90 patients) and was predominantly grade 1 (23 [26%] of 90) and grade 2 (15 [17%]), and primarily confined to cycle 1. The most common grade 3-4 adverse events were neutropenia or neutrophil count decreased (24 [27%] of 90 patients), hypophosphataemia (15 [17%]), hyperglycaemia (seven [8%]), and anaemia (seven [8%]). Serious adverse events occurred in 42 (47%) of 90 patients. No treatment-related grade 5 (ie, fatal) adverse event occurred.Interpretation: Fixed-duration mosunetuzumab has a favourable safety profile and induces high rates of complete remissions, allowing potential administration as an outpatient regimen, in patients with relapsed or refractory follicular lymphoma and two or more previous therapies.Funding: F Hoffmann-La Roche and Genentech. [ABSTRACT FROM AUTHOR]

[Display omitted] • The adsorption of CMT and NU drugs on B 36 was both physical and exothermic. • The behavior of NU∥ B 36 was superior due to its strong agreement with all tryouts. • CMT/NU adsorbed on the B 36 do not damage the protein as they interact with amino acids. • Drugs could be released from the B 36 surface without significantly altering its structure. • An alternative and improved approach for B 36 -based drug delivery vehicles. Various techniques have been devised so far to enhance cancer treatment efficacy and address drug conflicts. We explored Borophene (B 36) as a nanocarrier for the anti-cancer drugs carmustine (CMT) and nitrosourea (NU) in vacuum and liquid phases using density functional theory. Both drugs were placed at perpendicular and parallel positions of B 36 and the adsorption energies of – 2.83, and −3.28 eV for CMT and NU respectively. The parallel site was found good due to higher adsorption energy than the perpendicular site. We examine the structural stability, electronic properties, adsorption properties work function (φ), recovery time (τ), and universal signifiers of the CMT and NU drugs for both configurations. Hirshfeld and DOS analysis shows that the drug acts as a donor of electrons to the B 36 sheet in the configurations studied. Computed results show that, compared to perpendicular configurations, the parallel configurations of CMT and NU exhibit better performance, with shorter recovery times at body temperature. We have confirmed the water solubility of both drugs and their distribution. We have verified that the drugs interact effectively with the amino acids without causing any damage to the protein. Additionally, our calculations indicate that in the slightly acidic environment of tumor tissue, CMT and NU drugs could begin to release from the B 36 surface without causing significant changes to the structural properties. The findings of this study indicated that, between the two drugs, CMT is likely to be used as a nanocarrier for B 36 , while NU/ B 36 is an extremely hopeful biological carrier for drug delivery purposes. [ABSTRACT FROM AUTHOR]

Cyclophosphamide and ifosfamide are major alkylating agents but their therapeutics uses are limiting by the toxicity due to several toxicities. Indeed conventional chemotherapies are generally used with the maximum tolerated dose. In contrast, metronomic schedule aims to get a minimum dose for efficacy with a good safety. Depending on the dose, their mechanisms of action are different and offer a dual activity: at high dose, cyclophosphamide is mainly used in graft conditioning for its immunosuppressive properties, while at metronomic dose it is used as an immunoactive agent. Currently, at metronomic dose, cyclophosphamide is studied in clinic against various types of cancer, alone or in combination with others anticancer drugs (anti-angiogenic, immune-modulating agents, immune checkpoints blockers, vaccines, radiotherapy, others conventional anticancer agents), as a nth-line or first-line treatment. More than three quarters of clinical studies show promising results, mostly in breast, ovarian and prostate cancers. Taking advantage of the immune system, use dual antitumor action's chemotherapy is clearly a therapeutic strategy that deserves to be confirmed in order to improve the efficacy/toxicity balance of anticancer treatments, and to use CPM or analogues as a standard of care. [ABSTRACT FROM AUTHOR]

[Display omitted] • BPL is a common chemical inactivating reagent used in vaccines. • Via BPL carcinogenicity, its residual should be hydrolyzed completely in vaccines to avoid cytotoxicity. • BPL residual determination in vaccines is challenging due to its low concentration, presence in a complex mixture of vaccine and lack of suitable chromophore. • BPL derivatization with RBH was used to enhance analytical sensitivity and stabilize the BPL molecule for measurement with HPLC-UV. β-propiolactone (BPL) is an alkylating agent used for inactivation of biological samples such as vaccines. Due to its known carcinogenic properties, complete hydrolysis of BPL is essential, and the detection of trace amounts is crucial. In this study a novel High-Performance Liquid Chromatography-Ultraviolet (HPLC-UV) method was developed. Rhodamine B hydrazide (RBH) was synthesized and utilized as a derivatizing reagent to react with BPL. The reaction was optimized in a weak acidic solution, resulting in a high yield. The separation of the RBH-derivatized BPL was achieved on a C8 column and detected by a UV detector at a wavelength of 560 nm. The method's validation demonstrated a high linearity (r2 > 0.99) over a concentration range of 0.5–50 µg/mL, with detection and quantification limits of 0.17 µg/mL and 0.5 µg/mL, respectively. The average recovery of samples was 85.20 % with a relative standard deviation (RSD) of 1.75 %. This method was successfully applied for BPL residue analysis in inactivated COVID-19 vaccines. This novel derivatization method offers a promising solution for monitoring BPL residues in the vaccine production process for quality control purposes and compliance with regulatory standards. [ABSTRACT FROM AUTHOR]

Mutations in isocitrate dehydrogenase isoform 1 (IDH1) are primarily found in secondary glioblastoma (GBM) and low-grade glioma but are rare in primary GBM. The standard treatment for GBM includes radiation combined with temozolomide, an alkylating agent. Fortunately, IDH1 mutant gliomas are sensitive to this treatment, resulting in a more favorable prognosis. However, it's estimated that up to 75 % of IDH1 mutant gliomas will progress to WHO grade IV over time and develop resistance to alkylating agents. Therefore, understanding the mechanism(s) by which IDH1 mutant gliomas confer sensitivity to alkylating agents is crucial for developing targeted chemotherapeutic approaches. The base excision repair (BER) pathway is responsible for repairing most base damage induced by alkylating agents. Defects in this pathway can lead to hypersensitivity to these agents due to unresolved DNA damage. The coordinated assembly and disassembly of BER protein complexes are essential for cell survival and for maintaining genomic integrity following alkylating agent exposure. These complexes rely on poly-ADP-ribose formation, an NAD+-dependent post-translational modification synthesized by PARP1 and PARP2 during the BER process. At the lesion site, poly-ADP-ribose facilitates the recruitment of XRCC1. This scaffold protein helps assemble BER proteins like DNA polymerase beta (Polβ), a bifunctional DNA polymerase containing both DNA synthesis and 5′-deoxyribose-phosphate lyase (5'dRP lyase) activity. Here, we confirm that IDH1 mutant glioma cells have defective NAD+ metabolism, but still produce sufficient nuclear NAD+ for robust PARP1 activation and BER complex formation in response to DNA damage. However, the overproduction of 2-hydroxyglutarate, an oncometabolite produced by the IDH1 R132H mutant protein, suppresses BER capacity by reducing Polβ protein levels. This defines a novel mechanism by which the IDH1 mutation in gliomas confers cellular sensitivity to alkylating agents and to inhibitors of the poly-ADP-ribose glycohydrolase, PARG. • D-2-Hydroxglutarate suppresses Polβ protein levels in IDH1 mutant glioma cells. • IDH1 mutant glioma cells are selectively sensitive to alkylating agents and PARGi • NAD+ can be enhanced in IDH1 mutant cells to improve PARGi efficacy • NADP(H) phosphatases MESH1 and NOCT are downregulated in IDH1 mutant cells • Polβ levels regulate the response to alkylating agents and PARGi [ABSTRACT FROM AUTHOR]

[Display omitted] • GEM-loaded PLGA NPs were produced using a double emulsion method. • Factorial design was employed to optimize the physicochemical properties of NPs. • Coating with CH enhanced the in vitro mucoadhesive properties of the NPs. • GEM encapsulation improved drug cytotoxicity in human GBM cells. • CH-GEM-NPs sensitize tumor cells to the effect of GBM gold-standard treatment. Glioblastoma (GBM) is a highly deadly brain tumor that does not respond satisfactorily to conventional treatment. The non-alkylating agent gemcitabine (GEM) has been proposed for treating GBM. It can overcome MGMT protein-mediated resistance, a major limitation of conventional therapy with the alkylating agent temozolomide (TMZ). However, GEM's high systemic toxicity and poor permeability across the blood–brain barrier (BBB) pose significant challenges for its delivery to the brain. Thus, mucoadhesive poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) coated with chitosan (CH), suitable for intranasal GEM delivery, were proposed in this work. A central composite design (CCD) was implemented for NPs optimization, and NPs with appropriate characteristics for intranasal administration were obtained. in vitro studies revealed that the NPs possess excellent mucoadhesive properties and the ability to selectively release GEM in the simulated tumor tissue environment. in vitro studies using two human GBM cell lines (U215 and T98G) revealed the NPs' ability to promote GEM's antiproliferative activity to sensitize cells to the effect of TMZ. The findings of this work demonstrate that the developed CH-GEM-NPs are suitable delivery systems for GEM, both as a single therapy and as a chemosensitizer to the GBM gold standard therapy. [ABSTRACT FROM AUTHOR]

Apoptosis has traditionally been regarded as the desired cell death pathway activated by chemotherapeutic drugs due to its controlled and non-inflammatory nature. However, recent discoveries of alternative cell death pathways have paved the way for immune-stimulatory treatment approaches in cancer. Ferroptosis (dependent on iron) and cuproptosis (dependent on copper) hold promise for selective cancer cell targeting and overcoming drug resistance. Copper ionophores and iron-bearing nano-drugs show potential for clinical therapy as single agents and as adjuvant treatments. Here we review up-to-date evidence for the involvement of metal ion-dependent cell death pathways in the cytotoxicity of classical chemotherapeutic agents (alkylating agents, topoisomerase inhibitors, antimetabolites, and mitotic spindle inhibitors) and their combinations with cuproptosis and ferroptosis inducers, indicating the prospects, advantages, and obstacles of their use. [ABSTRACT FROM AUTHOR]

Renal cell carcinoma (RCC) is rare in the pediatric population, comprising about 5% of renal neoplasms in children.1 Out of all childhood cases of RCC, translocation RCCs (tRCC) is the most common.2 It is well described in the literature that exposure to alkylating agents such as cyclophosphamide and/or topoisomerase II inhibitors such as doxorubicin and etoposide, is a risk factor for the development of Xp11 (or TFE3) tRCC.3 Herein is a case of tRCC development in a patient with history of exposure to topoisomerase II inhibitors and alkylating agents to treat a common childhood malignancy. [ABSTRACT FROM AUTHOR]

Cancer is one of the leading causes of death in the modern world. Nowadays, most often treatment methods used in clinical oncology are drug therapies applied as monotherapy or combined therapy. Additionally, recent studies focus on developing approaches with the use of a drug in combination with other factors, not only chemical, to improve the probability and magnitude of therapeutic responses and reduce the possibility of chemoresistance. Such a promising factor seems to be an electromagnetic field (EMF) application. Here, we tested the effect of continuous or pulsed EMF on human cancer cells of different origin treated or not with 3-bromopyruvate, a small and powerful alkylating agent with a broad spectrum of anticancer activities. We provide strong evidence suggesting that ELF-EMF potentiates the anti-cancer activity of 3BP in human cancer cells through inhibition of TNFα secretion leading to irreversible p21/p27-dependent G2/M cell cycle arrest and finally cancer cell death. Our findings suggest a novel approach combining pharmacotherapy with ELF-EMF. In conclusion, electromagnetic field seems to be a potential modulator of anti-cancer efficacy of 3BP while combined therapy offers off-target activity. These features contribute to the development of innovative therapeutic strategies for cancer treatment. [Display omitted] • EMF potentiates the 3BP activity inhibition of TNFα secretion. • EMF supports anti-cancer 3BP action via p21/p27-dependent G2/M cell cycle arrest. • 3BP-mediated cancer cell death is dependent on EMF off-target activity. • Enhancement of 3BP action by EMF may represent a novel approach for cancer treatment. [ABSTRACT FROM AUTHOR]

Chemotherapy constitutes the backbone of cancer treatment. Several predictive assays assist personalized administration of cytotoxic drugs and are recommended for use in a clinical setting. The deficiency of DNA repair by homologous recombination (HRD), which is caused by inactivation of BRCA1/2 genes or other genetic events, is associated with high tumor responsiveness to platinum compounds, bifunctional alkylating agents and topoisomerase II poisons. Low activity of MGMT predicts the efficacy of nitrosoureas and tetrazines. Some clinically established pharmacogenetic tests allow for the adjustment of drug dosage, for example, the analysis of DPYD allelic variants for administration of fluoropyrimidines and UGT1A1 genotyping for the use of irinotecan. While there are promising molecular predictors of tumor sensitivity to pemetrexed, gemcitabine and taxanes, they remain in the investigational stage and require additional validation. Comprehensive molecular analysis of tumors obtained from drug responders and non-responders is likely to reveal new clinically useful predictive markers for cytotoxic therapy. [ABSTRACT FROM AUTHOR]

Forisomes are giant polyprotein complexes that undergo reversible conformational rearrangements from a spindle-like to a plug-like state in response to Ca2+ or changes in pH. They act as valves in the plant vasculature, and reproduce this function in vitro to regulate flow in microfluidic capillaries controlled by electro-titration. Heterologous expression in yeast or plants allows the large-scale production of tailor-made artificial forisomes for technical applications. Here we investigated the unexpected disintegration of artificial forisomes in response to Ca2+ following the deletion of the M1 motif in the MtSEO-F1 protein or the replacement of all four conserved cysteine residues therein. This phenomenon could be mimicked in wild-type forisomes under reducing conditions by adding a thiol alkylating agent. We propose a model in which reversible changes in forisome structure depend on cysteine residues with ambiguous redox states, allowing the formation of intermolecular disulfide bridges (confirmed by mass spectrometry) as well as noncovalent thiol interactions to connect forisome substructures in the dispersed state. This is facilitated by the projection of the M1 motif from the MtSEO-F1 protein as part of an extended loop. Our findings support the rational engineering of disintegrating forisomes to control the release of peptides or enzymes in microfluidic systems. • Deletion of conserved M1 motif leads to calcium-triggered forisome disintegration. • Conserved cysteines in the M1 motif preserve forisome functionality. • Disulfides and reduced cysteines are equally important for forisome integrity. [ABSTRACT FROM AUTHOR]

Proposed hypothesis in this study. Brain tumours overexpress SPARC protein which is responsible for the uptake of albumin nanoparticles (HSA NPs). Temozolomide (TMZ), a DNA alkylating agent, is the first line drug for treatment of glioma. Thus the delivery of Temozolomide acid (TMZA), a derivative of TMZ into HSA NPs (TMZA-HSA NPs) is hypothesized to ensure delivery of TMZA into glioma cells, overcoming the BBB and ensuring TMZ delivery in sufficient amounts for improved glioma therapy. [Display omitted] Glioblastoma is one of the most difficult to treat cancers with poor prognosis and survival of around one year from diagnosis. Effective treatments are desperately needed. This work aims to prepare temozolomide acid (TMZA) loaded albumin nanoparticles, for the first time, to target glioblastoma (GL261) and brain cancer stem cells (BL6). TMZA was loaded into human serum albumin nanoparticles (HSA NPs) using the desolvation method. A response surface 3-level factorial design was used to study the effect of different formulation parameters on the drug loading and particle size of NPs. The optimum conditions were found to be: 4 mg TMZA with 0.05% sodium cholate. This yielded NPs with particle size and drug loading of 111.7 nm and 5.5% respectively. The selected formula was found to have good shelf life and serum stability but with a relatively fast drug release pattern. The optimized NPs showed excellent cellular uptake with ∼ 50 and 100% of cells were positive for NP uptake after 24 h incubation with both GL261 and BL6 glioblastoma cell lines, respectively. The selected formula showed high cytotoxicity with ̴ 20% cell viability at 1 mM TMZA after 72 h incubation time. Finally, the fluorescently labelled NPs showed co-localization with the bioluminescent syngeneic BL6 intra-cranial tumour mouse model after intravenous administration. [ABSTRACT FROM AUTHOR]

RAD51 paralog gene mutations are observed in both hereditary breast and ovarian cancers. Classically, defects in RAD51 paralog function are associated with homologous recombination (HR) deficiency and increased genomic instability. Several recent investigative advances have enabled characterization of non-canonical RAD51 paralog function during DNA replication. Here we discuss the role of the RAD51 paralogs and their associated complexes in integrating a robust response to DNA replication stress. We highlight recent discoveries suggesting that the RAD51 paralogs complexes mediate lesion-specific tolerance of replicative stress following exposure to alkylating agents and the requirement for the Shu complex in fork restart upon fork stalling by dNTP depletion. In addition, we describe the role of the BCDX2 complex in restraining and promoting fork remodeling in response to fluctuating dNTP pools. Finally, we highlight recent work demonstrating a requirement for RAD51C in recognizing and tolerating methyl-adducts. In each scenario, RAD51 paralog complexes play a central role in lesion recognition and bypass in a replicative context. Future studies will determine how these critical functions for RAD51 paralog complexes contribute to tumorigenesis. [ABSTRACT FROM AUTHOR]

DNA-protein cross-links are formed when proteins become covalently trapped with DNA in the presence of exogenous or endogenous alkylating agents. If left unrepaired, they inhibit transcription as well as DNA unwinding during replication and may result in genome instability or even cell death. The DNA repair protein O6-alkylguanine DNA-alkyltransferase (AGT) is known to form DNA cross-links in the presence of the carcinogen 1,2-dibromoethane, resulting in G:C to T:A trans-versions and other mutations in both bacterial and mammalian cells. We hypothesized that AGT-DNA cross-links would be processed by nuclear proteases to yield peptides small enough to be bypassed by translesion (TLS) polymerases. Here, a 15-mer and a 36-mer peptide from the active site of AGT were cross-linked to the N2 position of guanine via conjugate addition of a thiol containing a peptide dehydroalanine moiety. Bypass studies with DNA polymerases (pols) η and κ indicated that both can accurately bypass the cross-linked DNA peptides. The specificity constant (kcat/Km) for steady-state incorporation of the correct nucleotide dCTP increased by 6-fold with human (h) pol κ and 3-fold with hpol η, with hpol η preferentially inserting nucleotides in the order dC > dG > dA > dT. LC-MS/MS analysis of the extension product also revealed error-free bypass of the cross-linked 15-mer peptide by hpol η. We conclude that a bulky 15-mer AGT peptide cross-linked to the N2 position of guanine can retard polymerization, but that overall fidelity is not compromised because only correct bases are inserted and extended. [ABSTRACT FROM AUTHOR]

Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults. Despite the gold standard treatment combining surgical resection, radiation and adjuvant plus concomitant chemotherapy with the alkylating agent temozolomide (TMZ), the prognosis remains poor (5-year survival rate < 10%). Over the last three decades, a vast array of drug delivery systems (DDS) have been developed for the local treatment of GBM, with the majority of the characterization being undertaken in pre-clinical models. We aimed to gain an overview of the potential efficacy of such local delivery systems in comparison to the systemic drug administration. In this paper, a systematic search of Pubmed, Web of Science, and Scopus was performed using pre-determined search terms. Studies were assessed for eligibility based on specific inclusion and exclusion criteria. A total of fifteen publications were included for analysis of local vs systemic group median survival, tumor volume and adverse events, with five brought forward for a meta-analysis. The majority of studies showed local delivery to be more efficacious than systemic administration, regardless of the drug, animal model, type of DDS used, or duration of the study. The meta-analysis also showed that the mean difference between median survival ratios was statistically significantly in favor of local delivery. Preclinical evidence shows that there is a firm rationale for further developing DDS for local therapeutic delivery to GBM and other brain cancers. [Display omitted] • Gliadel wafers for glioblastoma have achieved limited success in the clinic. • New drug delivery systems are being designed to overcome current drawbacks. • Preclinical data from local systems is compared against systemic administration. • Local delivery gives a higher median survival time than systemic administration. • This data provides a firm rationale for development of local drug delivery devices. [ABSTRACT FROM AUTHOR]

Objective: To normalize age-dependent effects on standardized measures of spermatogonial quantity such as the number of spermatogonia per tubular cross-section (S/T) or fertility index.Design: Published quantitative histologic data on human spermatogonial numbers were used to create Z-scores for reference means and tested on archived testicular tissue samples.Setting: Retrospective cohort study.Patient(s): The sample cohort comprised testicular samples from 24 boys with cancer diagnosis and 10 with Klinefelter syndrome, as part of the fertility preservation programs NORDFERTIL and Androprotect, as well as archived histologic samples from 35 prepubertal boys with acute lymphoblastic leukemia and 20 testicular biobank samples.Intervention(s): None.Main Outcome Measure(s): Z-score values for S/T and fertility index on the basis of morphology and germ cell-specific markers (MAGEA4 and/or DDX4) were calculated, and the impact of cancer therapy exposure and genetic disorders on Z-score values was evaluated.Result(s): The Z-scores for S/T values in the nontreated samples (-2.08 ± 2.20, n = 28) and samples treated with nonalkylating agents (-1.90 ± 2.60, n = 25) were comparable within ±3 standard deviations of the reference mean value but differed significantly from samples exposed to alkylating agents (-12.14 ± 9.20, n = 22) and from patients with Klinefelter syndrome (-11.56 ± 4.89, n = 8). The Z-scores for S/T were correlated with increasing cumulative exposure to alkylating agents (r = -0.7020).Conclusion(s): The Z-score values for S/T allow for the quantification of genetic and cancer treatment-related effects on testicular tissue stored for fertility preservation, facilitating their use for patient counseling. [ABSTRACT FROM AUTHOR]

Glioblastoma multiforme (GBM) is abnormal cell proliferation of glial cells. GBM is the grade IV glioma brain cancer which is life-threatening to many individuals affected by this cancer. The DNA alkylating agent Temozolomide (TMZ) has the distinctiveness of being FDA approved anticancer drug for the first line treatment for GBM. However, treatment of GBM still remains a challenge. This is attributed to TMZ's toxic nature, severe side effects, and fast degradation in vivo. In addition, the lack of targeting ability increases the chances of systemic toxicities. A nano enabled targeted delivery system not only improves the efficiency of TMZ by making it cross the blood brain barrier, have specificity to target, but also reduces toxicity to healthy tissues. Over the last decade the significant advances in the area of nanotechnology applied to medicine have developed many multifunctional therapeutics. In this context, the present review article comprehends the significant progress in the field of TMZ loaded nanocarriers showing promise for futuristic nanomedicine therapies in treating GBM. [Display omitted] • The challenges presented by BBB are overcome by nanocarrier system strategies. • Nano-enabled delivery systems improve the pharmacokinetics of Temozolomide. • Targeting of nanocarriers with a peptide or a ligand are beneficial. • Nanocarriers improve the efficiency of TMZ delivered to the target site (GBM cells). • TMZ loaded nanocarriers show promise for futuristic nanomedicine therapies in GBM [ABSTRACT FROM AUTHOR]

• Mustard gas is a highly toxic alkylating agent that rapidly penetrates into ocular tissues. • Sulfur mustard gas exposure to eye causes multiple ophthalmic manifestations including acute and chronic vision loss. • MGK involves unique acute, chronic, and delayed ocular manifestations that can persist for months and years post-exposure. Sulfur mustard (SM) is a chemical warfare agent that has been used throughout recent history and remains a threat today. Exposed soldiers and civilians experience a variety of symptoms primarily in the respiratory system, skin, and eyes. The ocular tissues are highly sensitive to damage by SM and undergo unique manifestations of acute, chronic, and delayed complications that can persist for months and years after exposure. The mechanisms of this unique mustard gas keratopathy are still not fully understood and animal models for the study of this disease are discussed. Recent advances in mechanisms of injury are included in this review. Ophthalmic manifestations of SM injury including persistent epithelial defects, limbal stem cell deficiency, corneal neovascularization, dry eye, and corneal opacification have been reported. A wide variety of medical and surgical therapies have been studied and are reviewed here along with potential future therapies. [ABSTRACT FROM AUTHOR]

One of the key strategies in chemotherapy involves crosslinking the DNA strands of cancer cells to impede their replication, with platinum (Pt) coordination compounds being a prominent class and cisplatin being its major representative. Steroidal ligands tethered to DNA interactive Pt core act as drug carriers for targeted therapy. While crosslinking of nuclear or mitochondrial DNA strands using coordination complexes has been studied for years, there remains a lack of comprehensive reviews addressing the advancements made in steroidal-Pt derivatives. This review specifically focuses on advancements made in steroid-tethered structural derivatives of Pt(II) or prodrug Pt(IV) for targeted chemotherapy, synthesized between 2000 and 2023. This period was deliberately chosen due to the widespread use of computational techniques for more accurate structure-based drug-design in last two decades. This review discusses the strategy behind tethering steroidal ligands such as testosterone, estrogen, bile acids, and cholesterol to the central DNA interactive Pt core through specific linker groups. The steroidal ligands function as drug delivery vehicles of DNA interactive Pt core and bind with their respective target receptors or proteins that are often overexpressed in cancer cells, thus enabling targeted delivery of Pt moiety to interact with DNA. We discussed structural features such as the location of the linker group on the steroid, the mono, bi, and tridentate configuration of the chelating arm in coordination with Pt, and the rigidity and flexibility of the linker group. The comparative in vitro, in vivo activities, and relative binding affinities of the designed compounds against standard Pt drugs are also discussed. We also provided a critique of observed trends and shortcomings. Our review will provide insights into future molecular designing of targeted DNA crosslinkers and their structural optimization to achieve desired drug properties. From this analysis, we proposed further research directions leading to the future of targeted chemotherapy. [Display omitted] • Review focuses on advancements in steroidal-Pt chemotherapy derivatives (2000–2023). • Focus on tethering steroidal ligands to DNA-interactive Pt cores for targeted therapy. • Details of strategy behind molecular structural design of steroid-linker-Pt. • Details of in vitro, in vivo activities, binding affinity and computational studies. • Identified gaps in current research and need for in-depth molecular understanding. [ABSTRACT FROM AUTHOR]

Ifosfamide is an alkylating antineoplastic drug used in chemotherapy, but it is also detected in wastewater. Here, the objectives were to (1) determine teratogenic, cardiotoxic, and mitochondrial toxicity potential of ifosfamide exposure; (2) elucidate mechanisms of toxicity; (3) characterize exposure effects on larval behavior. Survival rate, hatch rate, and morphological deformity incidence were not different amongst treatments following exposure levels up to 1000 µg/L ifosfamide over 7 days. RNA-seq reveled 231 and 93 differentially expressed transcripts in larvae exposed to 1 µg/L and 100 µg/L ifosfamide, respectively. Several gene networks related to vascular resistance, cardiovascular response, and heart rate were affected, consistent with tachycardia observed in exposed embryonic fish. Hyperactivity in larval zebrafish was observed with ifosfamide exposure, potentially associated with dopamine-related gene networks. This study improves ecological risk assessment of antineoplastics by elucidating molecular mechanisms related to ifosfamide toxicity, and to alkylating agents in general. • Heart rate was increased with ifosfamide exposure in embryos. • Ifosfamide altered cardiovascular and immunological gene networks. • Hyperactivity was observed with 0.1 µg/L and 1 µg/L ifosfamide exposure. • Ifosfamide exposure was not related to anxiolytic/anti-anxiolytic behavior. [ABSTRACT FROM AUTHOR]

In this study an efficient method to facilitate the sustainable synthesis of O -methyl dehydrozingerone dimer, and its monomer was developed. This one-pot two-step process involves O -methylation of phenolic compounds, using dimethyl carbonate as an alkylating green agent, followed by a Claisen Schmidt condensation in the presence of acetone under microwave irradiation without changing the nature of the base used for the entire synthetic process. The starting material vanillin dimer [6,6′-dihydroxy-5,5′-dimethoxy-[1,1′-biphenyl]-3,3′-dicarbaldehyde], was prepared by an innovative high-speed oxidative coupling reaction. [Display omitted] [ABSTRACT FROM AUTHOR]

Poly (ADP-ribose) polymerase inhibitors (PARPi) are used for patients with BRCA1/2 mutations, but patients with other mutations may benefit from PARPi treatment. Another mutation that is present in more cancers than BRCA1/2 is mutation to the TP53 gene. In 2D breast cancer cell lines, mutant p53 (mtp53) proteins tightly associate with replicating DNA and Poly (ADP-ribose) polymerase (PARP) protein. Combination drug treatment with the alkylating agent temozolomide and the PARPi talazoparib kills mtp53 expressing 2D grown breast cancer cell lines. We evaluated the sensitivity to the combination of temozolomide plus PARPi talazoparib treatment to breast and lung cancer patient-derived tumor organoids (PDTOs). The combination of the two drugs was synergistic for a cytotoxic response in PDTOs with mtp53 but not for PDTOs with wtp53. The combination of talazoparib and temozolomide induced more DNA double-strand breaks in mtp53 expressing organoids than in wild-type p53 expressing organoids as shown by increased γ-H2AX protein expression. Moreover, breast cancer tissue microarrays (TMAs) showed a positive correlation between stable p53 and high PARP1 expression in sub-groups of breast cancers, which may indicate sub-classes of breast cancers sensitive to PARPi therapy. These results suggest that mtp53 could be a biomarker to predict response to the combination of PARPi talazoparib-temozolomide treatment. • We demonstrated that breast and lung patient derived tumor organoids (PDTOs) carrying mutant p53 (mtp53) present synergistic cytotoxicity to PARP inhibitor (PARPi) talazoparib and DNA damaging agent temozolomide. • Combination drug synergism was achieved when treating mtp53 containing, but not wild-type p53 expressing, PDTO. • Synergy resulted most likely from double-strand breaks in mtp53 expressing PDTOs that were not able to be repaired and subsequently led to cell death. • Breast cancer tissue microarrays (TMAs) showed a positive correlation between stable p53 and high PARP1 expression in sub-groups of breast cancers, which may indicate sub-classes of breast cancers sensitive to PARPi therapy. • We found that the combination drug synergism occurred in both breast and lung PDTO carrying wild type BRCA1/2 which demonstrated high strength for mtp53 as a significant biomarker. • This work suggests that in addition to BRCA1 mutation, TP53 mutation may be an additional predictor for cancer cell death following combination PARPi treatment. [ABSTRACT FROM AUTHOR]

Temozolomide (TMZ) is the most preferred and approved chemotherapeutic drug for either first- or second-line chemotherapy for glioma patients across the globe. In glioma patients, resistance to treatment with alkylating drugs like TMZ is known to be conferred by exalted levels of MGMT gene expression. On the contrary, epigenetic silencing through MGMT gene promoter methylation leading to subsequent reduction in MGMT transcription and protein expression, is predicted to have a response favoring TMZ treatment. Thus, MGMT protein level in cancer cells is a crucial determining factor in indicating and predicting the choice of alkylating agents in chemotherapy or choosing glioma patients directly for a second line of treatment. Thus, in-depth research is necessary to achieve insights into MGMT gene regulation that has recently enticed a fascinating interest in epigenetic, transcriptional, post-transcriptional, and post-translational levels. Furthermore, MGMT promoter methylation, stability of MGMT protein, and related subsequent adaptive responses are also important contributors to strategic developments in glioma therapy. With applications to its identification as a prognostic biomarker, thus predicting response to advanced glioma therapy, this review aims to concentrate on the mechanistic role and regulation of MGMT gene expression at epigenetic, transcriptional, post-transcriptional, and post-translational levels functioning under the control of multiple signaling dynamics. [Display omitted] • MGMT is a major contributor of TMZ-based chemo resistance in Glioma patients. • MGMT has been identified as a prognostic biomarker of Glioma due to its involvement at epigenetic, transcriptional, post-transcriptional and post-translational levels. • Several approaches have been explored to combat MGMT based manipulation in Glioma therapy. [ABSTRACT FROM AUTHOR]

Glioblastoma multiforme remains a therapeutic challenge. We offer a historical review of the outcomes of patients with glioblastoma from the earliest report of surgery for this lesion through the introduction of modern chemotherapeutics and aggressive approaches to tumor resection. We reviewed all major surgical series of patients with glioblastoma from the introduction of craniotomy for glioma (1884) to 2020. The earliest reported craniotomy for glioblastoma resulted in the patient's death less than a month after surgery. Improved intracranial pressure management resulted in improved outcomes, reducing early postoperative mortality from 50% to 6% in Harvey Cushing's series. In the first major surgical series (1912), the mean survival was 10.1 months. This figure did not improve until the introduction of radiotherapy in the 1950s, which doubled survival relative to those who had surgery alone. The most recent significant advance, chemotherapy with the alkylating agent temozolomide, extended survival by 2.5 months compared with surgery and radiotherapy alone (14.6 and 12.1 months, respectively). This protocol remains the standard regimen for newly diagnosed glioblastoma. The innovative treatments being investigated have yet to show a survival benefit. With advancements in localization, imaging, anesthesia, surgical technique, control of cerebral edema, and adjuvant therapies, outcomes in glioblastoma improved incrementally from Cushing's time until the introduction of magnetic resonance imaging enabled better degrees of resection in the 1990s. Modest improvements came with the advent of biomarker-driven targeted chemotherapy in the first decade of the current century. [ABSTRACT FROM AUTHOR]

Objective: To investigate mechanisms of primordial follicle (PMF) loss in vivo in human ovaries shortly after alkylating agent (AA) chemotherapy.Design: Cohort study.Setting: Tertiary university medical center.Patient(s): Ninety-six women aged 15-39 years who underwent ovarian tissue cryopreservation for fertility preservation.Intervention(s): Fresh ovarian tissue samples were harvested from women treated with AA (n = 24) or non-AA (n = 24) chemotherapy <6 months after treatment and age-matched untreated women (n = 48).Main Outcome Measure(s): Differential follicle counts, time from chemotherapy exposure, immunostaining for apoptosis (cleaved caspase-3) and FOXO3A on tissue harvested within ultrashort time intervals (4-12 days), collagen (Sirius red) and neovascularization (CD34).Result(s): AA-treated ovaries had significant loss of PMFs, and significant increase in absolute numbers of growing follicles compared with untreated control ovaries. The number of growing follicles was inversely correlated with time from chemotherapy. Representative staining for FOXO3A observed decreased nuclear localization in PMF oocytes in AA-treated ovaries removed within the ultrashort time interval compared with untreated ovaries. Neither significant loss of PMFs, increase in growing follicles, nor decrease in nuclear FOXO3A were observed in non-AA-treated ovaries. No increased expression of cleaved caspase-3 was seen in PMFs within the ultrashort time interval after AA or non-AA chemotherapy. Significant stromal fibrosis and neovascularization were observed in AA-treated ovaries only after follicle loss had already occurred (4-6 months).Conclusion(s): Follicle activation occurs in vivo in ovaries of patients treated with AA, indicating a pathologic mechanism which may contribute to chemotherapy-induced follicle loss. [ABSTRACT FROM AUTHOR]

• Toxic effects of 300−1,500 mg min/m3 sulfur mustard (SM) vapor were characterized in rabbit corneas. • Recurrent edematous lesions (RELs) emerged in a stereotyped fashion over a 20 wk period. • Likelihood of REL emergence was strongly correlated to SM vapor dose. • However, REL pathophysiologies were similar regardless of SM vapor dose. • These data illustrate optimal SM vapor doses for future therapeutic and mechanistic studies. Sulfur mustard (SM) is a lipid soluble alkylating agent that causes genotoxic injury. The eye is highly sensitive to SM toxicity and exposures exceeding 400 mg min/m3 can elicit irreversible corneal pathophysiologies. Development of medical countermeasures for ocular SM exposure has been hindered by a limited understanding of dose-dependent effects of SM on corneal injury. Here, clinical, histological and ultrastructural analyses were used to characterize the effects of SM dose on corneal injury progression. Corneas were evaluated for up to 20 wk following exposure to saturated SM vapor for 30−150 s, which corresponds to 300−1,500 mg min/m3. In acute studies, a ceiling effect on corneal edema developed at doses associated with full-thickness corneal lesions, implicating endothelial toxicity in corneal swelling. Recurrent edematous lesions (RELs) transiently emerged after 2 wk in a dose-dependent fashion, followed by the development of secondary corneal pathophysiologies such as neovascularization, stromal scarring and endothelial abnormalities. RELs appeared in 96 % of corneas exposed for ≥ 90 s, 52 % of corneas exposed for 60 s and 0 % of corneas exposed for 30 s. While REL latency was variable in corneas exposed for 60 s, REL emergence was synchronized at exposures ≥ 90 s. Corneas did not exhibit more than one REL, suggesting RELs are part of a programmed pathophysiological response to severe alkylating lesions. In post-mortem studies at 12 wk, corneal edema was positively correlated to severity of endothelial pathologies, consistent with previous findings that endothelial toxicity influences long-term outcomes. These results provide novel insight into long-term corneal pathophysiological responses to acute toxicity and identify exposure conditions suitable for therapeutic testing. [ABSTRACT FROM AUTHOR]

Objective: To develop and internally validate a clinical predictive tool to assess the likelihood that a young cancer patient will experience diminished ovarian reserve (DOR) after chemotherapy.Design: Prospective cohort study.Setting: University hospitals.Patient(s): Postpubertal adolescent and young adult women with a new diagnosis of cancer requiring chemotherapy.Intervention: None.Main Outcome Measure(s): Diminished ovarian reserve after completion of and recovery from chemotherapy, defined as serum antimüllerian hormone (AMH) <1 ng/mL at 8-24 months after completion of chemotherapy.Result(s): A multivariable logistic regression model which includes age, cancer type, exposure to an alkylating agent, and baseline AMH value accurately predicts the diagnosis of DOR after chemotherapy with an area under the receiver operating characteristic curve of 0.89.Conclusion(s): Pretreatment information on age, cancer type, use of an alkylating agent, and baseline AMH levels make up a clinically useful predictive tool to identify which women are most at risk for DOR caused by chemotherapy. [ABSTRACT FROM AUTHOR]

As the Watson-Crick faces of nucleobases are protected in dsDNA, it is commonly assumed that deleterious alkylation damage to the Watson-Crick faces of nucleobases predominantly occurs when DNA becomes single-stranded during replication and transcription. However, damage to the Watson-Crick faces of nucleobases has been reported in dsDNA in vitro through mechanisms that are not understood. In addition, the extent of protection from methylation damage conferred by dsDNA relative to ssDNA has not been quantified. Watson-Crick base pairs in dsDNA exist in dynamic equilibrium with Hoogsteen base pairs that expose the Watson-Crick faces of purine nucleobases to solvent. Whether this can influence the damage susceptibility of dsDNA remains unknown. Using dot-blot and primer extension assays, we measured the susceptibility of adenine-N1 to methylation by dimethyl sulfate (DMS) when in an A-T Watson-Crick versus Hoogsteen conformation. Relative to unpaired adenines in a bulge, Watson-Crick A-T base pairs in dsDNA only conferred ~130-fold protection against adenine-N1 methylation, and this protection was reduced to ~40-fold for A(syn)-T Hoogsteen base pairs embedded in a DNA-drug complex. Our results indicate that Watson-Crick faces of nucleobases are accessible to alkylating agents in canonical dsDNA and that Hoogsteen base pairs increase this accessibility. Given the higher abundance of dsDNA relative to ssDNA, these results suggest that dsDNA could be a substantial source of cytotoxic damage. The work establishes DMS probing as a method for characterizing A(syn)-T Hoogsteen base pairs in vitro and also lays the foundation for a sequencing approach to map A(syn)-T Hoogsteen and unpaired adenines genome-wide in vivo. [ABSTRACT FROM AUTHOR]

Pamiparib, an investigational Poly (ADP-ribose) polymerase (PARP) inhibitor in clinical development, demonstrates excellent selectivity for both PARP1 and PARP2, and superb anti-proliferation activities in tumor cell lines with BRCA1/2 mutations or HR pathway deficiency (HRD). Pamiparib has good bioavailability and is 16-fold more potent than olaparib in an efficacy study using BRCA1 mutated MDA-MB-436 breast cancer xenograft model. Pamiparib also shows strong anti-tumor synergy with temozolomide (TMZ), a DNA alkylating agent used to treat brain tumors. Compared to other PARP inhibitors, pamiparib demonstrated improved penetration across the blood brain barrier (BBB) in mice. Oral administration of pamiparib at a dose as low as 3 mg/kg is sufficient to abrogate PARylation in brain tumor tissues. In SCLC-derived, TMZ-resistant H209 intracranial xenograft model, combination of pamiparib with TMZ overcomes its resistance and shows significant tumor inhibitory effects and prolonged life span. Our data suggests that combination of pamiparib with TMZ has unique potential for treatment of brain tumors. Currently, the combination therapy of pamiparib with TMZ is evaluated in clinical trial [NCT03150862]. [ABSTRACT FROM AUTHOR]

• Temozolomide inhibits mtDNA replication and transcription in NSC. • Temozolomide reduces hippocampal neuron dendritic branching and spine density. • Temozolomide causes mitochondrial dysfunction, oxidative stress, and apoptosis. Primary brain tumor patients often experience neurological, cognitive, and depressive symptoms that profoundly affect quality of life. The DNA alkylating agent, temozolomide (TMZ), along with radiation therapy forms the standard of care for glioblastoma (GBM) – the most common and aggressive of all brain cancers. Numerous studies have reported that TMZ disrupts hippocampal neurogenesis and causes spatial learning deficits in rodents; however, the effect of TMZ on mature hippocampal neurons has not been addressed. In this study, we examined the mitochondrial-mediated mechanisms involving TMZ-induced neural damage in primary rat neural stem/progenitor cells (NSC) and hippocampal neurons. TMZ inhibited mt DNA replication and transcription of mitochondrial genes (ND1 and Cyt b) in NSC by 24 h, whereas the effect of TMZ on neuronal mt DNA transcription was less pronounced. Transmission electron microscopy imaging revealed mitochondrial degradation in TMZ-treated NSC. Acute TMZ exposure (4 h) caused a rapid reduction in dendritic branching and loss of postsynaptic density-95 (PSD95) puncta on dendrites. Longer TMZ exposure impaired mitochondrial respiratory activity, increased oxidative stress, and induced apoptosis in hippocampal neurons. The presented findings suggest that NSC may be more vulnerable to TMZ than hippocampal neurons upon acute exposure; however long-term TMZ exposure results in neuronal mitochondrial respiratory dysfunction and dendritic damage, which may be associated with delayed cognitive impairments. [ABSTRACT FROM AUTHOR]

A microfluidic platform based on the integration of denaturation and online immobilized enzyme reactor (IMER) digestion for protein pretreatment was first developed on a glass chip. The design of three inlet channels and two groups of snake channel in glass chip can allow the protein solution, the reducing reagent and the alkylating agent to be simultaneously injected into the chip channel and ensured the reaction solution on-line efficient mixing and sufficient reacting. By thiol-ene click chemistry, the capillary-based and glass chip-based trypsin IMER on the surface of poly(trimethylolpropane trimethacrylate) monolith were fabricated. The wide range of flow rate tolerance (0.8–5.0 μL/min), and the acceptable reproducibility (RSD% = 3.1%, n = 5) and stability (13.8% decrease of enzyme activity in 2 months) indicated the feasibility of using IMER for online digestion of proteins. Compared with the solution denaturation-offline IMER digestion, the integrated microfluidic platform of chip denaturation-chip IMER and chip denaturation-online IMER have comparable protein identification ability for mouse liver protein with a similar number of protein (798 or 826 vs. 843) and unique peptides (3923 or 4593 vs. 3916). More importantly, the easy and fast digestion of protein samples and possible combination with MS revealed that this microfluidic platform can be a potential method for rapid proteomics analysis. Image 1 • On-line denaturation and digestion was achieved by integrated microfluidic chip. • Chemical reduction and alkylation were completed by two groups of snake channel. • The on-chip trypsin IMER was prepared by thiol-ene click chemistry. • Fast identification for mouse liver protein was achieved by microchip platform. • The microchip can simplify manual processing and reduce sample consumption. [ABSTRACT FROM AUTHOR]

Higher expression of the human DNA repair enzyme MUTYH has previously been shown to be strongly associated with reduced survival in a panel of 24 human lymphoblastoid cell lines exposed to the alkylating agent N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). The molecular mechanism of MUTYH-enhanced MNNG cytotoxicity is unclear, because MUTYH has a well-established role in the repair of oxidative DNA lesions. Here, we show in mouse embryonic fibroblasts (MEFs) that this MNNG-dependent phenotype does not involve oxidative DNA damage and occurs independently of both O6-methyl guanine adduct cytotoxicity and MUTYH-dependent glycosylase activity. We found that blocking of abasic (AP) sites abolishes higher survival of Mutyh-deficient (Mutyh-/-) MEFs, but this blockade had no additive cytotoxicity in WT MEFs, suggesting the cytotoxicity is due to MUTYH interactions with MNNG-induced AP sites. We found that recombinant mouse MUTYH tightly binds AP sites opposite all four canonical undamaged bases and stimulated apurinic/apyrimidinic endonuclease 1 (APE1)-mediated DNA incision. Consistent with these observations, we found that stable expression of WT, but not catalytically-inactive MUTYH, enhances MNNG cytotoxicity in Mutyh-/- MEFs and that MUTYH expression enhances MNNG-induced genomic strand breaks. Taken together, these results suggest that MUTYH enhances the rapid accumulation of AP-site intermediates by interacting with APE1, implicating MUTYH as a factor that modulates the delicate process of base-excision repair independently of its glycosylase activity. [ABSTRACT FROM AUTHOR]

The role of 3D genome organization in the precise regulation of gene expression is well established. Accordingly, the mechanistic connections between 3D genome alterations and disease development are becoming increasingly apparent. This opinion article provides a snapshot of our current understanding of the 3D genome alterations associated with cancers. We discuss potential connections of the 3D genome and cancer transcriptional addiction phenomenon as well as molecular mechanisms of action of 3D genome-disrupting drugs. Finally, we highlight issues and perspectives raised by the discovery of the first pharmaceutical strongly affecting 3D genome organization. Changes in spatial genome organization (3D genome) of cancer cells mostly occur on the level of topologically associating domains (TADs) and chromatin loops. Many cancers are addicted to high-level transcription of oncogenes, which requires involvement of transcriptional coactivators, 3D genome-mediated enhancer-promotor communication (EPCs), and specific chromatin-associated factors. There are several chemotherapeutic drugs targeting transcriptional coactivators that selectively kill cancer cells. A new group of epigenetic drugs targets the 3D genome organization via DNA-intercalating small molecules that alter DNA topology. This leads to the inability of CCCTC-binding factor (CTCF) to bind efficiently to its cognate DNA sites, results in large-scale perturbations in the 3D genome, and leads to preferential downregulation of enhancer and super-enhancer-driven transcription. [ABSTRACT FROM AUTHOR]