Topo I (DU-145 Luc cells) ( IC50 = 2 nM ); Topo I (MCF-7 Luc cells) ( IC50 = 13 nM )

In a dose- and time-dependent way, topotecan hydrochloride hydrate strongly suppresses the growth of human glioma cells and glioma stem cells (GSC) [1]. Topotecan hydrochloride hydrate (0–40 μM) drastically reduced cell viability in a dose-dependent manner when compared to the control group [1]. Topotecan hydrochloride hydrate exhibited anti-proliferative activity with IC50 values of 2.73±0.25, 2.95±0.23, 5.46±0.41 and 5.95±0.24 μM, respectively, on U251, U87, GSCs-U251 and GSCs-U87 cells [1].

Topotecan hydrochloride hydrate (0.5, 1.0, and 1.5 mg/kg; oral, daily) led in larger decreases in microvessel density in ovarian cancer models, whereas patients treated with topotecan 1.5 mg/kg orally daily Rats showed lower food intake and poor antitumor effects [2].

Topotecan [(S)-9-dimethylaminomethyl-10-hydroxycamptothecin hydrochloride; SK&F 104864-A, NSC 609699], a water soluble semisynthetic analogue of the alkaloid camptothecin, is a potent topoisomerase I inhibitor. Here we show that topotecan stabilizes topoisomerase I/DNA cleavable complexes in radiation-resistant human B-lineage acute lymphoblastic leukemia (ALL) cells, causes rapid apoptotic cell death despite high-level expression of bcl-2 protein, and inhibits ALL cell in vitro clonogenic growth in a dose-dependent fashion. Furthermore, topotecan elicited potent antileukemic activity in three different severe combined immunodeficiency (SCID) mouse models of human poor prognosis ALL and markedly improved event-free survival of SCID mice challenged with otherwise fatal doses of human leukemia cells at systemic drug exposure levels that can be easily achieved in children with leukemia. [Blood. 1995 May 15;85(10):2817-28]

Gliomas, the most malignant form of brain tumors, contain a small subpopulation of glioma stem cells (GSCs) that are implicated in therapeutic resistance and tumor recurrence. Topoisomerase I inhibitors, shikonin and topotecan, play a crucial role in anti-cancer therapies. After isolated and identified the GSCs from glioma cells successfully, U251, U87, GSCs-U251 and GSCs-U87 cells were administrated with various concentrations of shikonin or topotecan at different time points to seek for the optimal administration concentration and time point. The cell viability, cell cycle and apoptosis were detected using cell counting kit-8 and flow cytometer to observe the inhibitory effects on glioma cells and GSCs. We demonstrated that shikonin and topotecan obviously inhibited proliferation of not only human glioma cells but also GSCs in a dose- and time-dependent manner. According to the IC50 values at 24 h, 2 μmol/L of shikonin and 3 μmol/L of topotecan were selected as the optimal administration concentration. In addition, shikonin and topotecan induced cell cycle arrest in G0/G1 and S phases and promoted apoptosis. The down-regulation of Bcl-2 expression with the activation of caspase 9/3-dependent pathway was involved in the apoptosis process. Therefore, the above results showed that topoisomerase I inhibitors, shikonin and topotecan, inhibited growth and induced apoptosis of GSCs as well as glioma cells, which suggested that they might be the potential anticancer agents targeting gliomas to provide a novel therapeutic strategy[1].

In vivo antitumor efficacies of the LDM topotecan and pazopanib as single agents and in combination were tested on 4 subcutaneous xenograft models and on 2 neuroblastoma metastatic models. Circulating angiogenic factors such as circulating endothelial cells (CEC), circulating endothelial pro genitor cells (CEP), and microvessel densities were used as surrogate biomarker markers of antiangiogenic activity.[2]

Absorption, Distribution and Excretion
Renal clearance is a crucial determinant of topotecan elimination. In a mass balance/excretion study of four patients with solid tumors, the mean recovery of total topotecan and its N-demethyl metabolites in urine and feces over 9 days was 73.4 ± 2.3% of the intravenously administered dose. The fecal excretion rate of total topotecan was 9 ± 3.6%, while that of N-demethyltopotecan was 1.7 ± 0.6%. The pharmacokinetics of topotecan have been extensively studied in patients with normal renal function, and one study was conducted in patients with mild to moderate renal impairment. However, the effect of hemodialysis on the distribution of topotecan in the body has not been reported. This study aimed to describe the distribution of topotecan in patients with severe renal impairment undergoing hemodialysis. The distribution of topotecan lactone in a patient undergoing hemodialysis and a patient not undergoing hemodialysis was characterized. The clearance rates of topotecanolactone, measured when used alone and in combination with hemodialysis, were 5.3 L/hr/m² and 20.1 L/hr/m², respectively. Thirty minutes after the end of hemodialysis, the plasma concentration of topotecan was higher than that at the end of dialysis (i.e., 8.0 ng/mL vs. 4.9 ng/mL), suggesting a rebound effect. The terminal half-life of topotecan was 13.6 hours when not undergoing hemodialysis, while the apparent half-life measured during hemodialysis was 3.0 hours. These results indicate that the plasma clearance of topotecan increased approximately fourfold during hemodialysis. Hemodialysis may be an effective method for systemic clearance of topotecan and should be considered in specific clinical situations (e.g., accidental overdose, severe renal impairment). In lactating rats receiving intravenous topotecan (at a dose of 4.72 mg/m²), drug concentrations were high (i.e., 48 times higher than plasma concentrations) and distributed into breast milk. It is currently unclear whether topotecan is distributed into human breast milk. Following oral administration, approximately 57% of topotecan (once daily for 5 consecutive days) is excreted in the urine as unchanged drug (20%) and N-desmethyl metabolite (2%). Approximately 33% of the oral topotecan is excreted in the feces as total topotecan, and approximately 2% as N-desmethyltopotecan. Following intravenous administration, approximately 74% of the topotecan dose is excreted within 9 days, primarily as unchanged drug in the urine (51%) and feces (18%); approximately 3% of N-desmethyltopotecan is excreted in the urine and approximately 2% in the feces. Following oral and intravenous administration (intravenous dose less than 2% of the administered dose), topotecan and the O-glucuronide metabolite of N-desmethyltopotecan were also detected in the urine.
No significant sex-based pharmacokinetic differences were reported in patients receiving oral topotecan. The mean plasma clearance of topotecan via intravenous administration was 24% higher in men than in women, primarily due to body size differences.
For more complete data on absorption, distribution, and excretion of topotecan (6 items in total), please visit the HSDB record page.

---

Metabolism/Metabolites
The lactone moiety of topotecan undergoes reversible pH-dependent hydrolysis; the lactone form of topotecan is pharmacologically active.
The lactone moiety of topotecan undergoes reversible pH-dependent hydrolysis; the lactone form is the pharmacologically active form. At pH=4, the lactone form is predominant, while under physiological pH conditions, the open-ring hydroxy acid form is predominant. In vitro human liver microsomal studies indicate that topotecan is metabolized to an N-demethylated metabolite. After intravenous administration, the mean AUC ratio of total topotecan and topotecan lactone metabolites to the parent drug is approximately 3%.

---

Biological Half-Life
2-3 hours
The pharmacokinetics of topotecan have been evaluated in cancer patients at doses of 0.5 to 1.5 mg/m², administered via intravenous infusion over 30 minutes. Topotecan exhibits multi-exponential decay pharmacokinetics with a terminal half-life of 2 to 3 hours. The terminal half-life of oral topotecan is 3 to 6 hours, while that after intravenous administration is 2 to 3 hours. …This study aimed to describe the distribution of topotecan in patients with severe renal impairment undergoing hemodialysis. …After dialysis, the terminal half-life of topotecan was 13.6 hours, compared to an apparent half-life of 3.0 hours measured during dialysis. …

Effects During Pregnancy and Lactation
◉ Overview of Lactation Use
Most data suggest that breastfeeding is not advisable while the mother is receiving high-dose anti-tumor drug treatment. The manufacturer recommends that women not breastfeed during topotecan treatment and for one week after the last dose. Chemotherapy may adversely affect the normal microbiota and chemical composition of breast milk. Women receiving chemotherapy during pregnancy are more likely to experience breastfeeding difficulties. ◉ Effects on Breastfed Infants
No published information found as of the revision date. ◉ Effects on Lactation and Breast Milk
No published information found as of the revision date.

---

Protein Binding Rate
35%

[1]. PLoS One. 2013 Nov 26;8(11):e81815.Topoisomerase I inhibitors, Shikonin and Topotecan, inhibit growth and induce apoptosis of glioma cells andglioma stem cells.

[2]. Metronomic oral topotecan with pazopanib is an active antiangiogenic regimen in mouse models of aggressive pediatric solid tumor. Clin Cancer Res. 2011 Sep 1;17(17):5656-67.

Gliomas are among the most malignant brain tumors, containing a small subset of glioma stem cells (GSCs), which are closely associated with treatment resistance and tumor recurrence. The topoisomerase I inhibitors shikonin and topotecan play crucial roles in anticancer therapy. We successfully isolated and identified GSCs from glioma cells, and then treated U251, U87, GSCs-U251, and GSCs-U87 cells with different concentrations of shikonin or topotecan at different time points to identify the optimal dosage concentrations and time points. We used a cell counting kit-8 and flow cytometry to detect cell viability, cell cycle, and apoptosis to observe the inhibitory effects of shikonin and topotecan on glioma cells and GSCs. The results showed that shikonin and topotecan significantly inhibited not only the proliferation of human glioma cells in a dose- and time-dependent manner but also the proliferation of GSCs. Based on the 24-hour IC50 value, 2 μmol/L shikonin and 3 μmol/L topotecan were selected as the optimal dosage concentrations. In addition, shikonin and topotecan can induce cell cycle arrest in the G0/G1 phase and S phase and promote apoptosis. Downregulation of Bcl-2 expression and activation of caspase 9/3-dependent pathway are involved in the apoptosis process. Therefore, the above results indicate that the topoisomerase I inhibitors shikonin and topotecan can inhibit the growth of glioma stem cells (GSCs) and glioma cells and induce their apoptosis, suggesting that they may be potential anticancer drugs targeting gliomas, providing a new strategy for glioma treatment. [1] Objective: Low-dose rhythmic chemotherapy (LDM) combined with VEGF signaling pathway inhibitors is an effective strategy for synergistically inhibiting angiogenesis and tumor growth in many adult preclinical cancer models. We tested the efficacy of daily oral low-dose topotecan (LDM) monotherapy and its combination with vascular endothelial growth factor (VEGF) receptor inhibitor pazopanib in three pediatric extracranial solid tumor mouse models. Experimental design: We conducted an in vitro dose-response study of topotecan and pazopanib in several neuroblastoma, osteosarcoma and rhabdomyosarcoma cell lines. In four subcutaneous xenograft tumor models and two neuroblastoma metastatic tumor models, we tested the in vivo antitumor efficacy of low-dose topotecan and pazopanib monotherapy and combination therapy. Circulating angiogenic factors, such as circulating endothelial cells (CEC), circulating endothelial progenitor cells (CEP), and microvessel density, were used as surrogate biomarkers for anti-angiogenic activity. Results: In vitro studies showed that topotecan dose-dependently reduced the viability of all cell lines, while pazopanib did not. In vivo studies showed that topotecan combined with pazopanib (TP+PZ) exhibited significant antitumor activity in all models and significantly prolonged survival compared to monotherapy with either drug. The reduction in extracellular fragment (CEP) and/or circulating endothelial cell (CEC) levels and tumor microvessel density was associated with tumor response, thus confirming the anti-angiogenic activity of this regimen. No drug interactions were found in the pharmacokinetic studies of the two drugs. Conclusion: In a mouse model of pediatric tumors, the rhythmic administration of the TP+PZ regimen showed statistically significant antitumor activity compared with the respective monotherapy, and can be used as an effective option for maintenance therapy of aggressive pediatric solid tumors. [2]

475.922045230865

475.151

1044663-62-8

Topotecan hydrochloride;119413-54-6; 123948-87-8; 1044663-62-8 (Topotecan HCl hydrate)

73440824

Typically exists as solid at room temperature

4

8

3

33

867

1

CC[C@@]1(C2=C(COC1=O)C(=O)N3CC4=CC5=C(C=CC(=C5CN(C)C)O)N=C4C3=C2)O.O.Cl

XVYBIGDRQOMVJX-IFUPQEAVSA-N

InChI=1S/C23H23N3O5.ClH.H2O/c1-4-23(30)16-8-18-20-12(9-26(18)21(28)15(16)11-31-22(23)29)7-13-14(10-25(2)3)19(27)6-5-17(13)24-20;;/h5-8,27,30H,4,9-11H2,1-3H3;1H;1H2/t23-;;/m0../s1

(19S)-8-[(dimethylamino)methyl]-19-ethyl-7,19-dihydroxy-17-oxa-3,13-diazapentacyclo[11.8.0.02,11.04,9.015,20]henicosa-1(21),2,4(9),5,7,10,15(20)-heptaene-14,18-dione;hydrate;hydrochloride

Topotecan hydrochloride hydrate; Topotecan (hydrochloride hydrate); 1044663-62-8; SCHEMBL13731135; Topotecan hydrochloride hydrate; Topotecan (hydrochloride hydrate); 1044663-62-8; SCHEMBL13731135; Tox21_500905; NCGC00261590-01; Tox21_500905; CCG-222209; NCGC00261590-01;

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.

---

Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)
*Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution.
Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)
Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil)
Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals).

View More

Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)]
*Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.
Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline)
Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline)
Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline)
Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline
Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil)
Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline)

---

Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium)
Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose
Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals).

View More

Oral Formulation 3: Dissolved in PEG400
Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose
Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose
Oral Formulation 6: Mixing with food powders

---

Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.

---

 (Please use freshly prepared in vivo formulations for optimal results.)