Chlorambucil (CLB) belongs to the class of nitrogen mustards (NMs), which are highly reactive bifunctional alkylating agents and were the first chemotherapeutic agents developed. They form DNA interstrand crosslinks (ICLs), which cause a blockage of DNA strand separation, inhibiting essential processes in DNA metabolism like replication and transcription. In fast replicating cells, e.g., tumor cells, this can induce cell death. The upregulation of ICL repair is thought to be a key factor for the resistance of tumor cells to ICL-inducing cytostatic agents including NMs. To monitor induction and repair of CLB-induced ICLs, we adjusted the automated reversed fluorometric analysis of alkaline DNA unwinding assay (rFADU) for the detection of ICLs in adherent cells. For the detection of monoalkylated DNA bases we established an LC-MS/MS method. We performed a comparative analysis of adduct formation and removal in five human cell lines and in peripheral blood mononuclear cells (PBMCs) after treatment with CLB. Dose-dependent increases in adduct formation were observed, and suitable treatment concentrations were identified for each cell line, which were then used for monitoring the kinetics of adduct formation. We observed significant differences in the repair kinetics of the cell lines tested. For example, in A2780 cells, hTERT immortalized VH10 cells, and in PBMCs a time-dependent repair of the two main monoalkylated DNA-adducts was confirmed. Regarding ICLs, repair was observed in all cell systems except for PBMCs. In conclusion, LC-MS/MS analyses combined with the rFADU technique are powerful tools to study the molecular mechanisms of NM-induced DNA damage and repair. By applying these methods to a spectrum of human cell systems of different origin and transformation status, we obtained insight into the cell-type specific repair of different CLB-induced DNA lesions, which may help identify novel resistance mechanisms of tumors and define molecular targets for therapeutic interventions.

USP1 (Ubiquitin-specific protease 1) is closely related to the prognosis of patients with liver cancer and plays an important role in DNA damage repair. C527 is a selective USP1 inhibitor (USP1i), which can regulate the protein ubiquitination to effectively inhibit the proliferation of cancer cells. However, its clinical application is hindered due to the poor water solubility and lack of tumor targeting. Moreover, the efficacy of single use of USP1i is still limited. Herein, a glutathione (GSH) sensitive amphiphilic polymer (poly (2-HD-co-HPMDA)-mPEG, PHHM) with disulfide bonds in the main chain was designed to encapsulate the USP1i as well as platinum (IV) prodrug (Pt (IV)-C12), resulting in the formation of composite nanoparticles, i.e., NP-Pt-USP1i. NP-Pt-USP1i can inhibit the DNA damage repair by targeting USP1 by the encapsulated USP1i, which ultimately increases the sensitivity of tumor cells to cisplatin and enhances the anti-cancer efficacy of cisplatin. Finally, an intraperitoneal tumor mice model and a patient-derived xenograft (PDX) of liver cancer mice model were established to prove that NP-Pt-USP1i could effectively inhibit the tumor growth. This work further validated the possibility of therapeutically target USP1 by USP1i in combination with DNA damaging alkylating agents, which could become a promising cancer treatment modality in the future.

BACKGROUND: Chemotherapy-induced neutropenia increases the risk of febrile neutropenia (FN) and infection with resultant hospitalizations, with substantial health care resource utilization (HCRU) and costs. Granulocyte-colony stimulating factor (GCSF) is recommended as primary prophylaxis for chemotherapy regimens having more than a 20% risk of FN. Yet, for intermediate-risk (10%-20%) regimens, it should be considered only for patients with 1 or more clinical risk factors (RFs) for FN. It is unclear whether FN prophylaxis for intermediate-risk patients is being optimally implemented. OBJECTIVE: To examine RFs, prophylaxis use, HCRU, and costs associated with incident FN during chemotherapy. METHODS: This retrospective study used administrative claims data for commercial and Medicare Advantage enrollees with nonmyeloid cancer treated with intermediate-risk chemotherapy regimens during January 1, 2009, to March 31, 2020. Clinical RFs, GCSF prophylaxis, incident FN, HCRU, and costs were analyzed descriptively by receipt of primary GCSF, secondary GCSF, or no GCSF prophylaxis. Multivariable Cox regression analysis was used to examine the association between number of RFs and cumulative FN risk. RESULTS: The sample comprised 13,937 patients (mean age 67 years, 55% female). Patients had a mean of 2.3 RFs, the most common being recent surgery, were aged 65 years or greater, and had baseline liver or renal dysfunction; 98% had 1 or more RFs. However, only 35% of patients received primary prophylaxis; 12% received secondary prophylaxis. The hazard ratio of incident FN was higher with increasing number of RFs during the first line of therapy, yet more than 54% of patients received no prophylaxis, regardless of RFs. Use of GCSF prophylaxis varied more by chemotherapeutic regimen than by number of RFs. Among patients treated with rituximab, cyclophosphamide, hydroxydaunorubicin hydrochloride (doxorubicin hydrochloride), vincristine, and prednisone, 76% received primary prophylaxis, whereas only 22% of patients treated with carboplatin/paclitaxel received primary prophylaxis. Among patients with a first line of therapy FN event, 78% had an inpatient stay and 42% had an emergency visit. During cycle 1, mean FN-related coordination of benefits-adjusted medical costs per patient per month ($13,886 for patients with primary prophylaxis and $18,233 for those with none) were driven by inpatient hospitalizations, at 91% and 97%, respectively. CONCLUSIONS: Incident FN occurred more often with increasing numbers of RFs, but GCSF prophylaxis use did not rise correspondingly. Variation in prophylaxis use was greater based on regimen than RF number. Lower health care costs were observed among patients with primary prophylaxis use. Improved individual risk identification for intermediate-risk regimens and appropriate prophylaxis may decrease FN events toward the goal of better clinical and health care cost outcomes. DISCLOSURES: This work was funded by Sandoz Inc., which participated in the design of the study, interpretation of the data, writing and revision of the manuscript, and the decision to submit the manuscript for publication. The study was performed by Optum under contract with Sandoz Inc. The author(s) meet criteria for authorship as recommended by the International Committee of Medical Journal Editors. The authors received no direct compensation related to the development of the manuscript. Dr Li is an employee of Sandoz Inc. Drs Bell and Lal and Mr Peterson-Brandt were employees of Optum at the time of the study. Ms Anderson and Dr Aslam are employees of Optum. Dr Lyman has been primary investigator on a research grant from Amgen to their institution and has consulted for Sandoz, G1 Therapeutics, Partners Healthcare, BeyondSpring, ER Squibb, Merck, Jazz Pharm, Kallyope, Teva; Fresenius Kabi, Seattle Genetics, and Samsung.

Methotrexate (MTX) is used to treat rheumatoid arthritis, acute leukemia, and psoriasis. MTX can cause certain side effects, such as myelosuppression, while the exact mechanism of myelosuppression caused by MTX is unknown. Notch signaling pathway has been considered to be essential to regulate hematopoietic stem cell (HSC) regeneration and homeostasis, thus contributing to bone marrow hematopoiesis. However, whether MTX affects Notch signaling remains unexplored. Here, our study provides evidence that MTX strongly suppresses the Notch signaling pathway. We found that MTX inhibited the interaction between Nedd4 with Numb, thus restricting K48-linked polyubiquitination of Numb and stabilizing Numb proteins. This in turn inhibited the Notch signaling pathway by reducing Notch1 protein levels. Interestingly, we found that a monomeric drug, Triptolide, is capable of alleviating the inhibitory effect of MTX on Notch signaling pathway. This study promotes our understanding of MTX-mediated regulation of Notch signaling and could provide ideas to alleviate MTX-induced myelosuppression.

Alkylating agents are potent anticancer compounds that exert their anticancer properties through the inhibition of cell replication and transcription leading to cell death. Despite the numerous benefits, these agents also have serious drawbacks such as their high toxicity and low specificity towards cancer cells. As previously reported by our group, conjugation of alkylating agents with azasteroids can reduce their systemic toxicity and enhance their anticancer activity. In this work, novel steroidal alkylating agents bearing POPAM-OH were synthesized and their anticancer efficacy was evaluated in vitro and in vivo. All the novel hybrids demonstrated high antiproliferative effects against 5 different cancer cell lines in the low micromolar range. Treatment of SCID mice bearing SKOV-3 or PC-3 tumor xenografts with the most potent hybrid 19 led to significant reduction of tumor size (tumor inhibition TI = 95% in SKOV3 models and TI = 85.2% in PC3 models). Importantly, the acute toxicity of hybrid 19 (LD10 = 36 μΜ, LD50 = 62 μΜ) in CB17 SCID mice exhibited three-fold decrease compared to the acute toxicity of previously reported hybrids of POPAM-NH2. This is an important finding since systemic cytotoxicity is a critical limitation of alkylating agents. Collectively, the steroidal conjugates of POPAM-OH displayed significant anticancer efficacy and reduced toxicity in vitro and in vivo rendering them as good candidates for cancer therapy.

Busulfan (Bu) is an alkylating agent commonly used at high doses in the preparative regimens of hematopoietic stem cell transplantation (HSCT). It has been shown that such high doses of Bu are associated with generalized seizures which are usually managed by prophylactic antiepileptic drugs (AEDs) such as valproic acid (VPA). Being a strong enzyme inhibitor, VPA may inhibit Bu metabolism and thus increase its potential toxicity. Despite its clinical relevance, the potential interaction between Bu and VPA has not yet been evaluated. The aim of the present study was to assess and evaluate the potential drug-drug interaction (DDI) between Bu and VPA. This study was carried out by incubating Bu in laboratory-prepared rat liver-subcellular fractions including S9, microsomes, and cytosol, alone or in combination with VPA. The liver fractions were prepared by differential centrifugation of the liver homogenate. Analysis of Bu was employed using a fully validated LC-MS/MS method. The validation parameters were within the proposed limits of the international standards guidelines. Bu metabolic stability was assessed by incubating Bu at a concentration of 8 μg/ml in liver fractions at 37°C. There were significant reductions in Bu levels in S9 and cytosolic fractions, whereas these levels were not significantly (P ˃ 0.05) changed in microsomes. However, in presence of VPA, Bu levels in S9 fraction remained unchanged. These results indicated, for the first time, the potential metabolic interaction of Bu and VPA being in S9 only. This could be explained by inhibiting Bu cytosolic metabolism by the interaction with VPA either by sharing the same metabolic enzyme or the required co-factor. In conclusion, the present findings suggest, for the first time, a potential DDI between Bu and VPA in vitro using rat liver fractions. Further investigations are warranted in human-derived liver fractions to confirm such an interaction.

Since 2006, ophthalmic artery chemosurgery (OAC) has been used for ocular-sparing treatment of retinoblastoma. Systemic exposure to melphalan is known to cause ovarian dysfunction, but the effect of melphalan-based OAC has not yet been determined. Here, we assess biochemical and symptomatic measures of ovarian function in a cohort of pubertal female survivors of retinoblastoma treated with melphalan-based OAC. These 13 patients all had normal gonadotropins at a median age of 11.1 years, 9.6 years from the completion of therapy. None had symptoms of ovarian dysfunction. This study provides initial evidence that ovarian function remains intact after melphalan-based OAC.

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.

Female infertility is a significant health issue worldwide with a rising incidence. Anti-cancer therapy is one of the most important reasons for increasing infertility. Although anti-cancer treatment increases the rate of survival, it decreases the quality of life through its side effects. The most substantial side effects are sexual dysfunction and infertility. Breast cancer is the most common cancer. The first-line treatment of breast cancer is chemotherapy by alkylating agents like cyclophosphamide, which leads to infertility. For instance, persistent chemotherapy-induced amenorrhea among breast cancer patients could affect almost half of the patients that undergo such therapy. However, some agents or therapeutic methods can ameliorate these intoxicating effects. Chemotherapy plus gonadotropin-releasing hormone agonist, in breast cancer patients, can not only improve overall survival but also reduce ovarian toxicity. Age plays an essential role in chemotherapy-induced amenorrhea. Chemotherapy at a younger age can reduce the risk of infertility. Gynecological cancers including uterine and ovarian cancer, which have high mortality rates, are the most related cancers to infertility. Surgery is the primary treatment of gynecological cancers. Studies demonstrated that fertility-sparing surgery is a better option than radical surgery. In addition, neoadjuvant chemotherapy is mostly a better option than primary cytoreductive surgery in terms of survival and fertility. Immune checkpoint inhibitors (ICIs) have recently played a major role in treating various cancer types. However, ICIs are associated with hypophysitis, which affects ovaries and can lead to infertility. There are some options for ovarian preservation such as embryo cryopreservation, oocyte cryopreservation, ovarian transposition, ovarian tissue cryopreservation, and ovarian suppression by GnRH agonists. Anti-müllerian hormone level can be utilized to monitor the ovarian reserve. Moreover, to avoid fertility loss, approaches such as using transplantation of human placenta mesenchymal stem cells, administrating anti-inflammatory agents and hormone therapy are under investigation.

For missions beyond low Earth orbit to the moon or Mars, space explorers will encounter a complex radiation field composed of various ion species with a broad range of energies. Such missions pose significant radiation protection challenges that need to be solved in order to minimize exposures and associated health risks. An innovative galactic cosmic ray simulator (GCRsim) was recently developed at the NASA Space Radiation Laboratory (NSRL) at Brookhaven National Laboratory (BNL). The GCRsim technology is intended to represent major components of the space radiation environment in a ground analog laboratory setting where it can be used to improve understanding of biological risks and serve as a testbed for countermeasure development and validation. The current GCRsim consists of 33 energetic ion beams that collectively simulate the primary and secondary GCR field encountered by humans in space over the broad range of particle types, energies, and linear energy transfer (LET) of interest to health effects. A virtual workshop was held in December 2020 to assess the status of the NASA baseline GCRsim. Workshop attendees examined various aspects of simulator design, with a particular emphasis on beam selection strategies. Experimental results, modeling approaches, areas of consensus, and questions of concern were also discussed in detail. This report includes a summary of the GCRsim workshop and a description of the current status of the GCRsim. This information is important for future advancements and applications in space radiobiology.

Alkylating agents are genotoxic chemicals that can induce and treat various types of cancer. This occurs through covalent bonding with cellular macromolecules, in particular DNA, leading to the loss of functional integrity under the persistence of modifications upon replication. O6-alkylguanine (O6-AlkylG) adducts are proposed to be the most potent DNA lesions induced by alkylating agents. If not repaired correctly, these adducts can result, at the molecular level, in DNA point mutations, chromosome aberrations, recombination, crosslinking, and single- and double-strand breaks (SSB/DSBs). At the cellular level, these lesions can result in malignant transformation, senescence, or cell death. O6-methylguanine-DNA methyltransferase (MGMT) is a DNA repair protein capable of removing the alkyl groups from O6-AlkylG adducts in a damage reversal process that can prevent the adverse biological effects of DNA damage caused by guanine O6-alkylation. MGMT can thereby defend normal cells against tumor initiation, however it can also protect tumor cells against the beneficial effects of chemotherapy. Hence, MGMT can play an important role in both the prevention and treatment of cancer; thus, it can be considered as a double-edged sword. From a clinical perspective, MGMT is a therapeutic target, and it is important to explore the rational development of its clinical exploitation.

Hormone-dependent cancers, such as certain types of breast cancer are characterized by over-expression of estrogen receptors (ERs). Anticancer drug conjugates combining ER ligands with other classes of anticancer agents may not only benefit from dual action at both anti-cancer targets but also from selective delivery of cytotoxic agents to ER-positive tumor cells resulting in less toxicity and adverse effects. Moreover, they could also take advantage of overcoming resistance typical for anti-hormonal monotherapy such as tamoxifen. In this review, we discuss the design, structures and pharmacological effects of numerous series of drug conjugates containing ER ligands such as selective ER modulators (tamoxifen, 4-hydroxytamoxifen, endoxifen), selective ER degraders (ICI-164384) and ER agonists (estradiol) linked to diverse anti-cancer agents including histone-deacetylase inhibitors, DNA-alkylating agents, antimitotic agents and epidermal growth factor receptor inhibitors.

Compared with traditional oral and injection administration, the transdermal administration of traditional Chinese medicine has distinctive characteristics and advantages, which can avoid the "first pass effect" of the liver and the destruction of the gastrointestinal tract, maintain a stable blood concentration, and prolong drug action time. However, the basic theory and technology research in transdermal drug delivery are relatively limited at present, especially regarding research on new carriers of transdermal drug delivery and pharmacokinetic studies of the skin, which has become a bottleneck of transdermal drug delivery development. Triptolide is one of the main active components of Tripterygium wilfordii, which displays activities against mouse models of polycystic kidney disease and pancreatic cancer but its physical properties and severe toxicity limit its therapeutic potential. Due to the previously mentioned advantages of transdermal administration, in this study, we performed a detail analysis of the pharmacokinetics of a new transdermal triptolide delivery system. Triptolide nanoemulsion gels were prepared and served as new delivery systems, and the ex vivo characteristics were described. The metabolic characteristics of the different triptolide transdermal drug delivery formulations were investigated via skin-blood synchronous microdialysis combined with LC/MS. A multiscale modeling framework, molecular dynamics and finite element modeling were adopted to simulate the transport process of triptolide in the skin and to explore the pharmacokinetics and mathematical patterns. This study shows that the three-layer model can be used for transdermal drug delivery system drug diffusion research. Therefore, it is profitable for transdermal drug delivery system design and the optimization of the dosage form. Based on the drug concentration of the in vivo microdialysis measurement technology, the diffusion coefficient of drugs in the skin can be more accurately measured, and the numerical results can be verified. Therefore, the microdialysis technique combined with mathematical modeling provides a very good platform for the further study of transdermal delivery systems. This research will provide a new technology and method for the study of the pharmacokinetics of traditional Chinese medicine transdermal drug delivery. It has important theoretical and practical significance in clarifying the metabolic transformation of percutaneous drug absorption and screening for appropriate drugs and dosage forms of transdermal drug delivery.

Non-Hodgkin's lymphoma (NHL) is the fifth most common hematologic disorder in the United States, and its prevalence has been rising in Western countries. Among the subtypes of NHL, diffuse large B-cell lymphoma (DLBCL) mostly involves the lymph nodes, stomach, and gastrointestinal tract, whereas hepatic involvement of DLBCL is rare. On serologic testing, elevated immunoglobulin G (IgG) levels can be observed in DLBCL; however, elevated IgG levels are mainly observed in autoimmune hepatitis. A targeted-lesion biopsy is required for the diagnosis of DLBCL. Based on a final diagnosis, the patient was treated with rituximab-based chemotherapy, including cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy (R-CHOP). Herein, we report a case of DLBCL mimicking antinuclear antibody-negative autoimmune hepatitis, which was finally diagnosed as DLBCL involving the liver, and was confirmed by liver biopsy.

The pharmacological treatment of systemic lupus erythematosus (SLE) aims to decrease disease activity, progression, systemic compromise, and mortality. Among the pharmacological alternatives, there are chemically synthesized drugs whose efficacy has been evaluated, but which have the potential to generate adverse events that may compromise adherence and response to treatment. Therapy selection and monitoring will depend on patient characteristics and the safety profile of each drug. The aim of this review is to provide a comprehensive understanding of the most important synthetic drugs used in the treatment of SLE, including the current treatment options (mycophenolate mofetil, azathioprine, and cyclophosphamide), review their mechanism of action, efficacy, safety, and, most importantly, provide monitoring parameters that should be considered while the patient is receiving the pharmacotherapy.

Glioblastoma multiforme (GBM) is the most lethal adult brain cancer. Temozolomide (TMZ), the standard chemotherapeutic drug used in GBM, has limited benefit and alternate therapies are needed to improve GBM treatment. Nerve growth factor (NGF) and its precursor proNGF are increasingly recognized as stimulators of human tumor progression. The expression and stimulatory effect of NGF on GBM cell growth has previously been reported, but the status of proNGF in GBM is unreported. In this study, we have investigated proNGF expression and biological activity in GBM. A clinical cohort of GBM (n = 72) and low-grade glioma (n = 20) was analyzed by immunohistochemistry for proNGF and digital quantification. ProNGF expression was significantly increased in GBM compared to low grade gliomas and proNGF was also detected in patient plasma samples. ProNGF was also detected in most GBM cell lines by Western blotting. Although anti-proNGF blocking antibodies inhibited cell growth in GBM cells with methylated MGMT gene promoter, targeting proNGF could not potentiate the efficacy of TMZ. In subcutaneous xenograft of human GBM cells, anti-proNGF antibodies slightly reduced tumor volume but had no impact on TMZ efficacy. In conclusion, this data reveals that proNGF is overexpressed in GBM and can stimulate cancer cell growth. The potential of proNGF as a clinical biomarker and therapeutic target warrants further investigations.

Electrospun systems are becoming promising devices usable for topical treatments. They are eligible to deliver different therapies, from anti-inflammatory to antitumoral. In the current research, polycaprolactone electrospun membranes loaded with synthetic and commercial antitumoral active substances were produced, underlining how the matrix-filler affinity is a crucial parameter for designing drug delivery devices. Nanofibrous membranes loaded with different percentages of Dacarbazine (the drug of choice for melanoma) and a synthetic derivative of Dacarbazine were produced and compared to membranes loaded with AuM1, a highly active Au-complex with low affinity to the matrix. AFM morphologies showed that the surface profile of nanofibers loaded with affine substances is similar to one of the unloaded systems, thanks to the nature of the matrix-filler interaction. FTIR analyses proved the efficacy of the interaction between the amidic group of the Dacarbazine and the polycaprolactone. In AuM1-loaded membranes, because of the weak matrix-filler interaction, the complex is mainly aggregated in nanometric domains on the nanofiber surface, which manifests a nanometric roughness. Consequently, the release profiles follow a Fickian behavior for the Dacarbazine-based systems, whereas a two-step with a highly prominent burst effect was observed for AuM1 systems. The performed antitumoral tests evidence the high-cytotoxic activity of the electrospun systems against melanoma cell lines, proving that the synthetic substances are more active than the commercial dacarbazine.

Ataxia-telangiectasia (AT) is a multisystemic neurodegenerative inborn error of immunity (IEI) characterized by DNA repair defect, chromosomal instability, and hypersensitivity to ionizing radiation. Impaired DNA double-strand break repair determines a high risk of developing hematological malignancies, especially lymphoproliferative diseases. Poor response to treatment, excessive chemotherapy toxicities, and the need for avoiding exposure to ionizing radiation make the successful clinical management of patients with AT challenging for oncologists. We describe the favorable outcome of the LBCL with IRF4 rearrangement at stage III in a 7-year-old female patient diagnosed with AT. The patient was treated according to the B-HR arm of the INTER-B-NHL-COP 2010 protocol, including the administration of rituximab, cyclophosphamide, methotrexate, prednisone, etc. She presented excessive treatment toxicities despite individually reduced doses of methotrexate and cyclophosphamide. However, in the MRI there was no significant reduction in pathologic lymph nodes after three immunochemotherapy courses. Therefore, a lymph node biopsy was taken. Its subsequent histopathological examination revealed tuberculosis-like changes, though tuberculosis suspicion was excluded. After two following immunochemotherapy courses, PET-CT confirmed complete remission. From March 2022 onwards, the patient has remained in remission under the care of the outpatient children's oncology clinic.