Chemistry, Cyclophosphamide, Cancer Chemotherapy, and Serendipity: Sixty Years On

Cambridge Dictionary: serendipity | noun | the phenomenon of finding interesting or valuable things by chance. The year 2019 marked the 60th anniversary of the approval of cyclophosphamide (CP) as an anticancer by the U.S. Food & Drug Administration in 1959 for the treatment of lymphoma. Between 1959 and 2019 there were ~50,000 publications listed in PubMed that have CP in the title and/or abstract, with these annual numbers showing a continual increase, and over 1,800 such articles in 2019 alone. The discovery of CP is a prime example of serendipity in science, which also applies to key elements of the metabolism and pharmacological basis for the specificity of the cytotoxicity of CP toward cancer cells. Phosphoramide mustard (PM), HO(H2N)P(O)N(CH2CH2Cl)2, the principal metabolite of CP with DNA alkylating activity, was synthesized and reported by Friedman and Seligman in 1954 prior to the discovery of CP. Interestingly, the original drug design premise for synthesizing PM, which was based on elevated phosphamidase enzyme activity in cancer cells proved to be incorrect. While this wrong premise also led to the synthesis of CP, as a six-membered ring cyclic phosphamidase-activated precursor of PM, the actual metabolic conversion of CP to PM was subsequently found to involve a surprisingly complex array of metabolites and metabolic pathways, all completely unrelated to phosphamidase. Although the molecular structure of CP has an asymmetrically substituted, i.e. chiral phosphorus center, the racemic mixture of the Rp and Sp enantiomers of CP was used throughout its initial investigations and subsequent clinical trials despite the involvement of an initial enzyme-mediated metabolic activation step, which could, in principle, be stereoselective for only one of the enantiomers of CP. Stereochemical investigations along those lines were eventually carried out, but the results did not warrant replacement of racemic CP with either enantiomer in the clinic. Amazingly, there are now ~4,000 structural congeners of PM listed Chemical Abstracts, but none have led to an anticancer drug superior to CP. This account provides a synopsis of the key chemistry and stereochemistry investigations that comprise this story of CP, as a remarkable instance of serendipity in science, and my chance involvement in the unfolding of this fascinating story.

Evolution of Nitrogen-Based Alkylating Anticancer Agents

Despite the significant progress in anticancer drug development over recent years, there is a vital need for newer agents with unique, but still effective, mechanisms of action in order to treat the disease, particularly the highly aggressive and drug-resistant types. Alkylating agents, in particular nitrogen-based alkylators, are commonly used to treat hematological and solid malignancies; they exert their antineoplastic effects at all phases of the cell cycle and prevent reproduction of tumor cells. Certain alkylating agents have been designed to be more lipophilic, enabling the compound to penetrate the cell and enhance its alkylating activity against tumors. This review details the evolution of currently available alkylating agents and their profiles, with a focus on nitrogen-based alkylating agents, as important anticancer therapy strategies.

Phase I pharmacokinetic (PK) and pharmacodynamic (PD) study of intravenous dimethane sulfonate (DMS612, NSC 281612) in advanced malignancies.

2553 Background: DMS612 is a dimethane sulfonate compound that was identified as preferentially cytotoxic to renal cell carcinoma (RCC) cell lines in a chemical screen of the NCI-60 panel. DMS612 has bifunctional alkylating activity in vitro. Objectives of this first-in-human phase I study included determining the dose-limiting toxicity (DLT), maximum tolerated dose (MTD), recommended phase 2 dose (RP2D), PK and PD of DMS612 administered by 10 minute intravenous infusion on day 1, 8 and 15 of a 28 day cycle. Methods: Eligibility criteria included adults with advanced solid malignancies or lymphoma with ECOG performance status 0-2, life expectancy > 3 months and adequate organ and marrow function. Patients were enrolled using a standard “3+3” dose escalation scheme. Plasma PK of DMS612 and metabolites was assessed by LC-MS/MS. DNA damage PD was assessed by γ-H2AX immunofluorescence. Results: 35 subjects were enrolled (22 male, 13 female) with median age 59 years (41-75). Tumor types included colorectal (8), RCC (4), cervix (2), and urothelial (2). Doses administered were 1.5, 3, 5, 7, 9 and 12 mg/m2. The MTD was determined to be 9 mg/m2, with only one DLT of grade 4 thrombocytopenia in 12 subjects enrolled. The maximum administered dose of 12 mg/m2 was considered to be intolerable after 1 of 3 subjects had grade 4 neutropenia and 1 had prolonged grade 3 thrombocytopenia. Prolonged thrombocytopenia in later cycles was observed in other subjects, including one patient naïve to prior cytotoxic chemotherapy. One subject with RCC had a confirmed partial response at 7 mg/m2. DMS612 was rapidly converted into carboxy, chloroethyl and hydroxyethyl analogues and their glucuronides, some of which retained alkylating activity in vitro. Dose-dependent pharmacodynamic evidence of DNA damage induced by DMS612 in vivo was observed by γ-H2AX immunofluorescance in both peripheral blood lymphocytes and plucked scalp hairs. Conclusions: The MTD of DMS12 administered by intravenous infusion on day 1, 8 and 15 of a 28-day cycle was 9 mg/m2. Pre-clinical and clinical observations suggest that further study of DMS612 in RCC is warranted.