TS ( Ki = 1.3 nM ); DHFR ( Ki = 7.2 nM ); GARFT ( Ki = 1.3 nM )

Statistically speaking, the mice in the PC61 plus Pemetrexed group survive longer than the other groups. The mice treated with PC61 plus Pemetrexed showed a significantly higher survival rate in a survival analysis when compared to the mice treated with PC61 alone, rat IgG plus Pemetrexed, or no treatment[2].

TS activity is measured by spectrophotometric means, which entails tracking the rise in absorbance at 340 nm brought about by the production of 7,8-dihydrofolate. The assay buffer has the following contents: 25 mM MgC1₂, 6.5 mM formaldehyde, 1 mM EDTA, 75 mM 2-mercaptoethanol, 50 mM N-tris[hydroxymethyljmethyl-2-aminoethanesulfonic acid]. The concentrations of hIS, 6R-MTHF, and deoxyuridylate monophosphate are 30 μM, 100 μM, and 30 nM (1.7 milliunits/mL), respectively. An uninhibited reaction and six inhibitor concentrations are tested at the 6R-MTHF concentration. K_{i app} values are obtained by applying nonlinear regression analysis, with the assistance of the ENZFITTER program, to fit the data to the Morrison equation. K_{i app}= K_i(1 + [S]/K_m is the formula used to calculate K_i values, where [S] is equivalent to 30 μM and K_m is equivalent to 3 μM. Using spectrophotometry, DHFR activity is measured by tracking the disappearance of 7,8-dihydrofolate and NADPH at 340 nm. In 0.5 mL of 50 mM potassium phosphate buffer, pH 7.5, 150 mM KC1, 10 nM 2-mercaptoethanol, and 14 nM (0.34 milliunitlmL) DHFR are present during the reaction, which occurs at 25°C. 10, μM is the concentration of NADPH, while 5, 10, or 15 μM is the concentration of 7,8-dihydrofolate. Seven inhibitor concentrations and an uninhibited reaction are measured at each 7,8-dihydrofolate concentration. K_{i app} values are obtained by fitting the data to the Morrison equation using nonlinear regression analysis and the ENZFITI'ER microcomputer program. K_{i app}= K_i(1 + [S]/K_m), where K_m of 7,8-dihydrofolate is equal to 0.15 μM and [S] is the concentration of 7,8-dihydrofolate used. GARFT activity is measured spectrophotometrically by tracking the rise in absorbance at 295 nm that is brought about by the formation of the product 5,8-dideazafolate. At 25°C and pH 7.5, the reaction solvent is composed of 50% a-thioglycerol, 20% glycerol, and 75 mM HEPES. Substrates and enzyme were used at the following concentrations: 10 μM α,β-glycinamide ribonucleotide, 0–10 μM 10-formyl–5,8–dideazafolic acid, and 10 nM (1.9 milliunits/mL) GARFT. Ki values are determined using the Beckman DU640 spectrophotometer's Enzyme Mechanism program, which fits data to the Michaelis-Menten equation for competitive inhibition using nonlinear regression analysis.

To ascertain the concentration necessary for 50% inhibition of growth, dose-response curves are produced (IC50). Initially, 4 mg/mL of pemetrexed disodium is dissolved in DMSO, and the concentration is then further diluted with cell culture medium. 24-well Cluster plates are filled to a total capacity of 2.0 mL with CCRF-CEM leukemia cells in complete medium. Duplicate wells are filled with varying concentrations of pemetrexed disodium until the total volume of DMSO is 0.5%. The plates are incubated for seventy-two hours at 37 °C in an air atmosphere containing 5% CO₂. Cell counts on a ZBI Coulter counter are measured at the conclusion of the incubation. AICA at 300 μM, thymidine at 5 μM, hypoxanthine at 100 μM, or a combination of 5 μM hymidine and 100 μM hypoxanthine are the conditions under which the IC50s for each compound are discovered for a number of investigations. The MTT colorimetric assay is modified to quantify the cytotoxicity of adherent tumor cells. 96-well tissue culture plates with a flat bottom are used to seed the human tumor cells at a density of 100 μL assay medium/well. In the assay medium, the only sources of folate are 2.3 μM or 2 nM folic acid, along with 10% FCS and folic acid-free RPMI 1640. It is left empty in Well 1A. Antifolate stock solutions (one milligram per milliliter) are prepared in Dulbecco's PBS, and then successive 2-fold dilutions are made in PBS. Triplicate wells are filled with ten-μL aliquots of each concentration. Plates are incubated at 37 °C for 72 hours in an atmosphere that is humidified with 5% CO₂ in the air. 10 μL of stock MTF solution is added to each well of an assay after MTT has been dissolved in PBS at a concentration of 5 mg/mL. The plates are then incubated for an additional two hours at 37 °C. 100 μL of DMSO is added to each well after incubation. The plates are read on a Dynatech MR600 reader using a test wavelength of 570 nm and a reference wavelength of 630 nm following complete formazan solubilization. The amount of medication needed to impede cell growth by 50% in comparison to untreated controls is known as the IC50.

Effects During Pregnancy and Lactation
◉ Summary of Use during Lactation
Most sources consider breastfeeding to be contraindicated during maternal high-dose antineoplastic drug therapy. The manufacturer recommends that mothers should not to breastfeed during treatment with pemetrexed and for one week after the last dose. Chemotherapy may adversely affect the normal microbiome and chemical makeup of breastmilk.[1] Women who receive chemotherapy during pregnancy are more likely to have difficulty nursing their infant.[2]
◉ Effects in Breastfed Infants
Relevant published information was not found as of the revision date.
◉ Effects on Lactation and Breastmilk
Relevant published information was not found as of the revision date.
28170295

[1]. Cancer Res . 1997 Mar 15;57(6):1116-23.

[2]. Int J Cancer . 2013 Jan 15;132(2):459-71.

[3]. Clin Cancer Res . 2000 Mar;6(3):1016-23.

Pemetrexed disodium is an organic sodium salt that is the disodium salt of N-{4-[2-(2-amino-4-oxo-4,7-dihydro-1H-pyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl}-L-glutamic acid. Inhibits thymidylate synthase (TS), 421 dihydrofolate reductase (DHFR), and glycinamide ribonucleotide formyltransferase (GARFT). It has a role as an antineoplastic agent, an antimetabolite, an EC 1.5.1.3 (dihydrofolate reductase) inhibitor, an EC 2.1.2.2 (phosphoribosylglycinamide formyltransferase) inhibitor and an EC 2.1.1.45 (thymidylate synthase) inhibitor. It contains a pemetrexed(2-).
Pemetrexed Disodium is the disodium salt of a synthetic pyrimidine-based antifolate. Pemetrexed binds to and inhibits the enzyme thymidylate synthase (TS) which catalyses the methylation of 2'-deoxyuridine-5'-monophosphate (dUMP) to 2'-deoxythymidine-5'-monophosphate (dTMP), an essential precursor in DNA synthesis.
A guanine-derived ANTINEOPLASTIC AGENT that functions as a NUCLEIC ACID SYNTHESIS INHIBITOR through its binding to, and inhibition of, THYMIDYLATE SYNTHASE.
See also: Pemetrexed (broader).

---

Drug Indication
Malignant pleural mesotheliomaPemetrexed Krka in combination with cisplatin is indicated for the treatment of chemotherapy naïve patients with unresectable malignant pleural mesothelioma. Non-small cell lung cancer Pemetrexed Krka in combination with cisplatin is indicated for the first-line treatment of patients with locally advanced or metastatic non-small cell lung cancer other than predominantly squamous cell histology. Pemetrexed Krka is indicated as monotherapy for the maintenance treatment of locally advanced or metastatic non-small cell lung cancer other than predominantly squamous cell histology in patients whose disease has not progressed immediately following platinum-based chemotherapy. Pemetrexed Krka is indicated as monotherapy for the second-line treatment of patients with locally advanced or metastatic non-small cell lung cancer other than predominantly squamous cell histology.
Malignant pleural mesotheliomaAlimta in combination with cisplatin is indicated for the treatment of chemotherapy-naïve patients with unresectable malignant pleural mesothelioma. Non-small-cell lung cancer Alimta in combination with cisplatin is indicated for the first-line treatment of patients with locally advanced or metastatic non-small-cell lung cancer other than predominantly squamous cell histology. Alimta is indicated as monotherapy for the maintenance treatment of locally advanced or metastatic non-small-cell lung cancer other than predominantly squamous cell histology in patients whose disease has not progressed immediately following platinum-based chemotherapy. Alimta is indicated as monotherapy for the second line treatment of patients with locally advanced or metastatic non-small-cell lung cancer other than predominantly squamous cell histology.
Carcinoma of the head and neck (Covered by class waiver: oropharyngeal epithelial carcinoma, excluding nasopharyngeal carcinoma), Malignant pleural mesothelioma

C20H19N5NA2O6

471.37

471.113

150399-23-8

135413520

Solid powder

160°C 20mm

36-38°C

160°C/20mm

4

7

7

33

737

1

C1=CC(=CC=C1CCC2=CNC3=C2C(=O)NC(=N3)N)C(=O)N[C@@H](CCC(=O)[O-])C(=O)[O-].[Na+].[Na+]

NYDXNILOWQXUOF-GXKRWWSZSA-L

InChI=1S/C20H21N5O6.2Na/c21-20-24-16-15(18(29)25-20)12(9-22-16)6-3-10-1-4-11(5-2-10)17(28)23-13(19(30)31)7-8-14(26)27;;/h1-2,4-5,9,13H,3,6-8H2,(H,23,28)(H,26,27)(H,30,31)(H4,21,22,24,25,29);;/q;2*+1/p-2/t13-;;/m0../s1

disodium;(2S)-2-[[4-[2-(2-amino-4-oxo-3,7-dihydropyrrolo[2,3-d]pyrimidin-5-yl)ethyl]benzoyl]amino]pentanedioate

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Note: Please store this product in a sealed and protected environment, avoid exposure to moisture.

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

Solubility in Formulation 1: 50 mg/mL (106.07 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with sonication.

---

Solubility in Formulation 2: 100 mg/mL (212.15 mM) in Saline (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

---

View More

Solubility in Formulation 3: Saline: 30 mg/mL

---

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