Camptothecin is a quinoline alkaloid isolated from the plant Camptotheca acuminata of the Davidiaceae family. It is a natural topoisomerase I inhibitor that forms a complex with topoisomerase I-DNA, thereby preventing DNA replication and RNA synthesis to produce anti-tumor effects. Camptothecin and its derivatives are a class of alkaloids with broad-spectrum anti-tumor activity, which have significant inhibitory effects on malignant tumors. Clinical drugs such as irinotecan and topotecan have been developed. The main defects of camptothecin anti-tumor drugs are low water solubility, poor structural stability, and strong toxic side effects. Therefore, the corresponding structural modification strategies are gradually iterated with the progress of drug development.

Camptothecin Analogs

Camptothecin (CPT) is an alkaloid extracted from Camptotheca acuminata, with a unique five-ring structure of pyrroloquinoline ring, conjugated pyridine ring and six-membered hydroxy lactone ring. Because of its extensive anti-tumor activity in both in vitro and in vivo experiments and its significant inhibitory effect on DNA topoisomerase I, camptothecin compounds have entered the pharmaceutical field as small molecule chemotherapy drugs. In 1958, American chemists Monroe E. Wall and Mansukh C. Wani first extracted a crude extract of camptothecin from Camptotheca acuminata. In the same year, the National Cancer Chemotherapy Service Center (CCNSC) of the United States found in the CA-755 experiment that the crude extract of camptothecin had high anti-tumor activity. Therefore, scholars believed that camptothecin had the possibility of clinical application and started related research on it. With the advancement of separation technology, the chemical structure of camptothecin was discovered by single crystal X-ray analysis in 1966 and reported in the Journal of the American Chemical Society. The pure form of camptothecin was discovered by countercurrent chromatography in 1963. In clinical studies for malignant tumors like stomach cancer and colon cancer, the sodium salt of camptothecin was utilized because of the instability and poor solubility of the lactone ring of the drug. It was discovered during the study phase that it would result in severe adverse effects such as bone marrow suppression, diarrhea, and vomiting. Research on anti-tumor medications containing camptothecin has since reached a low point. It was not until 1985 that Yaw-Huei Hsiang discovered that camptothecin inhibits DNA topoisomerase I. At that point, camptothecin was once again brought back into the scientific realm and DNA topoisomerase I was confirmed to be an important therapeutic target for cancer chemotherapy. Medical professionals then created a range of derivatives of camptothecin for use in clinical trials.

Camptothecin Structure

Camptothecin molecules are composed of five rings, A, B, C, D, and E. Rings A and B are quinoline rings, ring C is a pyrrole ring, ring D is a pyridone, and ring E is an α-hydroxy lactone with an S-type chiral carbon. The molecular structure is highly unsaturated, and there is a continuous conjugated system between the five rings, which makes camptothecin have strong natural fluorescence. Camptothecin is sensitive to light and heat. When light is irradiated on camptothecin solution, its absorbance decreases. The stronger the light, the more the absorbance decreases. It is very stable when it is protected from light, and heating can decompose camptothecin.

Fig. 1. Structure and source of camptothecin.

Camptothecin Solubility

Camptothecin is a special, neutral alkaloid that is insoluble in acid and difficult to dissolve in general organic solvents. It is not easy to form salts with acid and is insoluble in water. It is soluble in a few solvents such as pyridine, chloroform, methanol, and dimethyl sulfoxide. It dissolves in concentrated sulfuric acid and is yellow-green. It is easily opened to form a water-soluble carboxylate salt after being treated with dilute alkali at room temperature, and it is re-lactonized to form a ring when acidified. In the buffer medium, there is a dynamic equilibrium between the lactone ring and the open-ring carboxylate. When pH>7.4, the open-ring carboxylate is the main form, and when pH<4.5, the lactone form is the main form.

Camptothecin Apoptosis

Camptothecins exert their anticancer effects by binding to and stabilizing the cleavable complex formed between topoisomerase I and DNA, leading to DNA strand breaks and ultimately triggering apoptosis in cancer cells. This mechanism of action makes camptothecins particularly effective against a variety of solid tumors, including colorectal cancer, ovarian cancer, and small cell lung cancer. In addition to their direct cytotoxic effects, camptothecins have shown promise in combination therapy with other anticancer drugs, increasing their overall effectiveness against drug-resistant tumors. BOC Sciences provides high-quality camptothecins for research purposes, ensuring that researchers have access to reliable compounds for their studies.

Camptothecin Mechanism of Action

Camptothecin is a potent anticancer agent that works by interfering with DNA replication and transcription. The primary target of camptothecin is topoisomerase I, an enzyme that plays a crucial role in DNA replication and transcription. The role of topoisomerase I is to relieve torsional stress that accumulates during DNA unwinding and prevent the formation of supercoils in the DNA double helix. In normal cells, topoisomerase I cuts a DNA strand, allowing it to rotate around the intact strand, then reseals the break and releases the torsional strain. However, camptothecin disrupts this process by binding to the topoisomerase I-DNA complex and stabilizing it in a way that prevents the DNA strand from resealing. This results in the formation of a stable ternary complex of camptothecin, topoisomerase I, and DNA, which ultimately leads to the accumulation of DNA strand breaks. If these breaks are not repaired, they may trigger cell cycle arrest and ultimately lead to apoptosis in cancer cells.

In addition, camptothecin-induced DNA damage can also interfere with RNA transcription by hindering the progression of RNA polymerase along the DNA template. This disruption of transcription further exacerbates the cytotoxic effects of camptothecin on cancer cells. The cellular response to camptothecin-induced DNA damage involves activation of DNA damage response pathways, which include signaling cascades that regulate cell cycle checkpoints and apoptosis. A key regulator in this pathway is the ataxia telangiectasia mutated (ATM) kinase, which is activated in the DNA damage response and coordinates the cellular response to genotoxic stress.

In addition to inhibiting topoisomerase I, camptothecins exhibit other mechanisms of action. For example, camptothecin derivatives such as irinotecan and topotecan can be converted into active metabolites in vivo, which also contribute to their antitumor effects. These metabolites can further inhibit topoisomerase I, leading to enhanced DNA damage and cytotoxicity. In addition, camptothecins have been reported to induce oxidative stress in cancer cells, resulting in the generation of reactive oxygen species (ROS), which lead to DNA damage and cell death. The interaction between camptothecin-induced ROS and DNA can exacerbate the cytotoxic effects of this drug and provide an additional mechanism for its anticancer activity.

Camptothecin ADC

ADCs are antibody-drug conjugates that combine the potency of cytotoxic drugs with the selectivity of monoclonal antibodies, minimizing damage to healthy cells and reducing systemic toxicity. The structure of ADCs consists of three main components: monoclonal antibodies, (cytotoxic drug) payloads, and linkers. Clonal antibodies are engineered to specifically bind to target antigens overexpressed on cancer cells. This enables ADCs to selectively target cancer cells while sparing normal cells. The cytotoxic drug payload is a potent chemotherapy drug that is highly effective in killing cancer cells. The linker molecule attaches the cytotoxic drug to the antibody, and its stability is critical for controlling the release of the drug within the target cell. Therefore, ADCs are an effective and promising class of cancer treatments. The FDA's first approval of Mylotarg (gemtuzumab ozogamicin) for adult acute myeloid leukemia marked the beginning of the ADC era for targeted cancer treatment. To date, 15 ADC drugs have been launched and more than 170 ADCs are currently in clinical development, and such therapies are reshaping cancer treatment across tumor types.

In recent years, camptothecin-based topoisomerase I inhibitors have emerged as one of the most promising classes of ADC payloads. ADCs address many of the major challenges associated with camptothecin small molecules. The antibody component enhances solubility and improves half-life and exposure of the conjugated payload. In addition, trafficking of ADCs into endosomal and lysosomal compartments exposes the camptothecin payload to a low pH environment (∼4.5-6), shifting the equilibrium from the less active carboxylate form to the active lactone form. This is not the case with camptothecin small molecules, which rapidly establish an equilibrium favoring the less active form after injection. Therefore, ADC cellular uptake and trafficking provides a unique mechanism for converting the inactive camptothecin form into the active form, highlighting how the lactone equilibrium can be tuned depending on the delivery method.

To date, according to incomplete statistics from the UmabsDB global database, there are nearly 80 ADC drugs using camptothecin derivatives as small molecule toxins being explored in the clinical stage, and these ADCs use more than 15 different payloads and more than 21 different linkers in 24 different targets. The major (>60%) payloads of ADCs currently in clinical development are exatecan or exatecan-derived compounds closely related to DXd (e.g., SHR9265, Ed-04, DDDXd, MH30010008). To date, three camptothecin-derived molecules have been reported as warheads for ADCs. One is the active ingredient of the clinical drug irinotecan, called SN-38. The second is a metabolite of DX-8951f, called DXd, which is a product of metabolic hydrolysis of the mAb construct DS-8201a. The third is exatecan, which was initially approved as a standalone drug in South Korea. The novel camptothecin payloads differ from DXd, SN-38, and exatecan not only in their association with novel linkers, but also in their intrinsic properties, such as in vitro cytotoxicity, hydrophilicity, permeability, metabolic stability, pharmacokinetics, and sensitivity to multidrug resistance efflux pumps.