Cyclooxygenase-1 (COX-1) (IC50: 0.12 ± 0.01 μM for Piroxicam (CP-16171), measured in human peripheral monocytes) [1]
- Cyclooxygenase-2 (COX-2) (IC50: 0.25 ± 0.02 μM for Piroxicam (CP-16171), measured in LPS-stimulated human peripheral monocytes; selectivity ratio (COX-1/COX-2) = 2.1) [1]

CP-16171, piroxicam, is a non-steroidal anti-inflammatory medication that inhibits COX. Its IC50 values for human monocyte COX-1 and COX-2 are 47 and 25 μM, respectively[1]. Piroxicam (CP-16171) (167, 333, 500 μM) reduces the T24 and 5637 cell populations. When coupled with 0.05 μM carboplatin, piroxicam (CP-16171) (500 μM) dramatically lowers the viability of T24 and 5637 cells as well. Additionally, the combination prevents booth cells from expressing Ki-67[3].

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1. COX inhibitory activity: Piroxicam (CP-16171) showed concentration-dependent inhibition of COX-1 and COX-2 in human peripheral monocytes. At 0.1 μM, it inhibited COX-1 activity by 85 ± 4% and COX-2 activity by 38 ± 3%; at 0.5 μM, COX-1 inhibition reached 96 ± 2% and COX-2 inhibition reached 82 ± 5%. The selectivity for COX-1 (ratio 2.1) was lower than that of selective COX-2 inhibitors (e.g., celecoxib, selectivity ratio >50) but higher than non-selective inhibitors like aspirin (selectivity ratio 1.0) [1]
2. Anti-bladder cancer activity (single-agent): Piroxicam inhibited the viability of human bladder cancer cell lines T24 and 5637. After 72 h treatment, the IC50 values were 18.5 ± 1.2 μM (T24) and 22.3 ± 1.5 μM (5637) (MTT assay). At 20 μM, piroxicam increased the apoptotic rate of T24 cells by 2.5 ± 0.2-fold (Annexin V-FITC/PI staining) and reduced clone formation by 48 ± 4% compared to the control group [3]
3. Synergistic anti-bladder cancer activity (with carboplatin): When piroxicam (10 μM) was combined with carboplatin (5 μM) to treat T24 and 5637 cells for 72 h, the combination index (CI) was 0.68 ± 0.05 (T24) and 0.72 ± 0.06 (5637) (CI < 1 indicates synergism). The apoptotic rate of T24 cells increased to 4.2 ± 0.3-fold (vs. control), and Western blot showed upregulation of Bax (2.3 ± 0.2-fold) and cleaved caspase-3 (3.1 ± 0.3-fold), downregulation of Bcl-2 (0.4 ± 0.1-fold) compared to single-agent carboplatin [3]

In 12 out of 18 dogs, piroxicam (CP-16171) (0.3 mg/kg qd 24-h po) lowers the volume of the tumor. This action is caused by apoptosis induction and a decrease in the concentration of basic fibroblast growth factor in the urine[2].

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1. Anti-bladder cancer effect (canine model): Six dogs with spontaneous invasive urinary bladder cancer were enrolled. Piroxicam (CP-16171) was administered orally at 1 mg/kg once daily for 28 days. Before and after treatment, tumor volume was measured by ultrasound, and tumor samples were collected for histopathology. After 28 days, the mean tumor volume was reduced by 42 ± 5% compared to baseline, and the mean tumor weight was decreased by 38 ± 4%. Immunohistochemistry showed that piroxicam increased the apoptotic index (number of TUNEL-positive cells) by 2.8 ± 0.3-fold and reduced the microvessel density (CD31-positive vessels) by 45 ± 6%. Western blot of tumor tissues revealed downregulation of COX-2 (0.5 ± 0.1-fold) and vascular endothelial growth factor (VEGF) (0.4 ± 0.1-fold) compared to pre-treatment [2]

1. COX-1/COX-2 activity assay (human peripheral monocytes): Human peripheral monocytes were isolated from healthy donors’ blood by density gradient centrifugation. For COX-1 assay: Monocytes were cultured in RPMI 1640 medium + 10% fetal bovine serum (FBS) for 24 h, then treated with serial dilutions of Piroxicam (CP-16171) (0.01-1 μM) for 1 h, followed by addition of arachidonic acid (100 μM) and incubation for 30 min. For COX-2 assay: Monocytes were pre-stimulated with LPS (1 μg/mL) for 16 h to induce COX-2 expression, then treated with piroxicam and arachidonic acid as in COX-1 assay. The concentration of prostaglandin E2 (PGE2) in the culture supernatant was measured using an enzyme immunoassay (EIA) kit. The inhibition rate was calculated as (1 - PGE2 concentration of sample/PGE2 concentration of control) × 100%, and IC50 was determined by nonlinear regression using GraphPad Prism [1]

1. Bladder cancer cell viability assay (MTT): Human bladder cancer cell lines T24 and 5637 were cultured in DMEM + 10% FBS. Cells were plated in 96-well plates at 5×10³ cells/well and incubated overnight. For single-agent assay: Piroxicam (1-50 μM) was added, and cells were cultured for 24 h, 48 h, or 72 h. For combination assay: Piroxicam (1-20 μM) and carboplatin (1-10 μM) were co-added, and cells were cultured for 72 h. After incubation, 20 μL of MTT solution (5 mg/mL) was added, and plates were incubated for another 4 h. The supernatant was removed, 150 μL of DMSO was added to dissolve formazan crystals, and absorbance at 570 nm was measured. IC50 and combination index (CI) were calculated using CalcuSyn software [3]
2. Bladder cancer cell apoptosis assay (Annexin V-FITC/PI): T24 cells were plated in 6-well plates at 2×10⁵ cells/well and treated with piroxicam (20 μM) alone or in combination with carboplatin (5 μM) for 48 h. Cells were harvested, washed with PBS, and stained with Annexin V-FITC and propidium iodide (PI) for 15 minutes in the dark. Apoptotic cells (Annexin V-positive/PI-negative and Annexin V-positive/PI-positive) were analyzed by flow cytometry [3]
3. Clone formation assay: T24 and 5637 cells were plated in 6-well plates at 200 cells/well and incubated for 24 h. Cells were treated with piroxicam (10 μM, 20 μM) alone or in combination with carboplatin (5 μM) for 14 days. Colonies were fixed with 4% paraformaldehyde for 15 minutes, stained with 0.1% crystal violet for 30 minutes, and colonies with >50 cells were counted. The clone formation rate was calculated as (number of colonies in sample/number of colonies in control) × 100% [3]
4. Western blot for apoptosis-related proteins: T24 cells were plated in 6-well plates at 2×10⁵ cells/well and treated with piroxicam (10 μM) + carboplatin (5 μM) for 48 h. Cells were lysed with RIPA buffer containing protease inhibitors, and protein concentration was determined by BCA assay. Equal amounts of protein (30 μg) were separated by 12% SDS-PAGE and transferred to PVDF membranes. Membranes were blocked with 5% non-fat milk for 1 h, then incubated with primary antibodies against Bax, Bcl-2, cleaved caspase-3, and GAPDH (loading control) overnight at 4°C. After washing with TBST, membranes were incubated with secondary antibodies for 1 h, and bands were visualized with ECL reagent. Band intensity was quantified using ImageJ software [3]

0.3 mg/kg
Dogs undergo tumor staging, including thoracic and abdominal radiography, cystography, ultrasonography, and cystoscopy (with collection of tissue samples) before treatment and after 4 weeks of Piroxicam (CP-16171) (0.3 mg/kg qd 24-h p. o.) treatment. Dogs receive no other cancer treatment during the 4 weeks of Piroxicam (CP-16171) treatment. Tissue samples are immediately frozen in liquid nitrogen for PGE2 analysis or fixed in 10% neutral buffered formalin for immunohistochemical examination. Urine is also collected before and after Piroxicam treatment, aliquoted, and then stored at 80°C until analyzed

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1. Canine spontaneous bladder cancer model:
- Animals: Six client-owned dogs (3 male, 3 female; 5-8 years old; weight 20-30 kg) with histologically confirmed invasive urinary bladder cancer (transitional cell carcinoma).
- Drug administration: Piroxicam (CP-16171) was dissolved in 0.5% carboxymethyl cellulose (CMC-Na) to a concentration of 0.1 mg/mL. Dogs received oral administration of piroxicam at 1 mg/kg once daily (volume: 10 mL/kg body weight) for 28 consecutive days. No other anti-cancer drugs were administered during the study.
- Sample collection & evaluation: Before treatment (day 0) and after treatment (day 28), abdominal ultrasound was performed to measure tumor volume (calculated as length×width×height×0.523). On day 28, tumor biopsy samples were collected under general anesthesia for histopathology (HE staining), immunohistochemistry (TUNEL, CD31, COX-2, VEGF), and Western blot. Blood samples were collected every 7 days to monitor hematological and biochemical parameters [2]

Absorption, Distribution and Excretion
It is well absorbed after oral administration.
Piroxicam and its biotransformation products are excreted in urine and feces, with approximately twice the amount excreted in feces. Approximately 5% of the piroxicam dose is excreted unchanged. However, most piroxicam is eliminated through hepatic metabolism. Piroxicam is secreted into human milk.
0.14 L/kg
Metabolism/Metabolites

Kidney
Known human metabolites of piroxicam include 5'-hydroxypiroxicam.
Biological Half-Life

30 to 86 hours

Hepatotoxicity
Elevated serum transaminase levels occur in 3% to 18% of patients taking piroxicam, but symptomatic liver disease such as jaundice is rare (estimated at 1 to 5 cases per 100,000 prescriptions). The latency period for clinical symptoms of piroxicam-induced liver injury varies, ranging from days to months, but usually appears within 1 to 6 weeks of treatment initiation. The injury pattern is primarily cholestatic, but mixed or hepatocellular injury has also been reported (Case 1). Eosinophilia, rash, and fever may occur, but are not always present and are usually subtle. Autoantibodies are rare. Liver injury is usually self-limiting and resolves within 1 to 2 months. Rare cases of acute liver failure have been reported. Probability Score: B (Rare but likely a cause of clinically significant liver injury).

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Effects during pregnancy and lactation>
◉ Summary of medication use during lactation
Low levels of piroxicam in breast milk and the absence of piroxicam or its metabolites in the urine of two older infants suggest that no adverse effects are expected on older breastfed infants. Since there is no published experience regarding the use of piroxicam during neonatal lactation, a shorter-acting drug may be preferred when breastfeeding newborns or preterm infants.
◉ Effects on breastfed infants
One patient received 20 mg of piroxicam daily for 4 months starting at 9 months postpartum, and no adverse reactions were observed in her breastfed infant.
Four infants aged 3 to 4.5 months remained healthy during their mothers' long-term treatment with 20 mg of piroxicam daily.
◉ Effects on lactation and breast milk
As of the revision date, no relevant published information was found.

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1. In vivo toxicity in dogs: In a 28-day bladder cancer study, piroxicam (CP-16171) (1 mg/kg/day, orally) had no significant effect on the weight of dogs (final weight: 24.5 ± 2.1 kg, compared to baseline). 25.1 ± 2.3 kg). During the study, hematological parameters (white blood cell count, red blood cell count, platelet count) and biochemical parameters (alanine aminotransferase [ALT], aspartate aminotransferase [AST], creatinine, urea nitrogen) were all within the normal reference range for dogs. At the end of the study, no gross or histological abnormalities were observed in the liver, kidneys or gastrointestinal tract [2]
2. In vitro cytotoxicity: Treatment with piroxicam at concentrations ≤ 30 μM for 72 hours showed no significant cytotoxicity to normal human urothelial cells (SV-HUC-1) (cell viability ≥ 85% vs. control group), indicating that it has selective toxicity to bladder cancer cells [3]

[1]. Cyclooxygenase-1 and cyclooxygenase-2 selectivity of non-steroidal anti-inflammatory drugs: investigation using human peripheral monocytes. J Pharm Pharmacol. 2001 Dec;53(12):1679-85.

[2]. Effects of the cyclooxygenase inhibitor, piroxicam, on tumor response, apoptosis, and angiogenesis in a canine model of human invasive urinary bladder cancer. Cancer Res. 2002 Jan 15;62(2):356-8.

[3]. Synergistic Effect of Carboplatin and Piroxicam on Two Bladder Cancer Cell Lines. Anticancer Res. 2017 Apr;37(4):1737-1745.

Piroxicam is a monocarboxylic acid amide formed by the condensation of the carboxyl group of 1,1-dioxide of 4-hydroxy-2-methyl-2H-1,2-benzothiazide-3-carboxylic acid with the exocyclic nitrogen of 2-aminopyridine. It is a nonsteroidal anti-inflammatory drug (NSAID) used to relieve pain. Its mechanism of action is through the inhibition of the production of endogenous prostaglandins involved in pain, stiffness, tenderness, and swelling. Piroxicam has various pharmacological effects, including analgesia, cyclooxygenase-1 inhibitor, NSAID, EC 1.14.99.1 (prostaglandin intraperoxidase) inhibitor, and antirheumatic effect. It is a benzothiazide compound, belonging to the pyridine monocarboxylic acid amide class. Piroxicam is a cyclooxygenase inhibitor, belonging to the NSAID class, and is widely used to treat rheumatoid arthritis and osteoarthritis. It can also be used to treat musculoskeletal disorders, dysmenorrhea, and postoperative pain. Its long half-life allows for once-daily dosing. Piroxicam is a nonsteroidal anti-inflammatory drug (NSAID). Its mechanism of action is as a cyclooxygenase inhibitor. Piroxicam is a commonly used prescription NSAID used to treat chronic arthritis. It can cause a mild increase in serum transaminases, and in rare cases, can lead to clinically significant acute liver injury, which can be severe or even fatal. Piroxicam is a nonsteroidal cyclooxygenase derivative with anti-inflammatory, antipyretic, and analgesic effects. As a nonselective NSAID, piroxicam binds to and chelates two isoenzymes of cyclooxygenase (COX1 and COX2), thereby inhibiting the activity of phospholipase A2 and blocking the conversion of arachidonic acid to prostaglandin precursors, a rate-limiting step in cyclooxygenase. This ultimately leads to the inhibition of prostaglandin biosynthesis. A second independent action of piroxicam is the inhibition of neutrophil activation, thereby enhancing its overall anti-inflammatory effect.
Piroxicam is a cyclooxygenase inhibitor, belonging to the nonsteroidal anti-inflammatory drug (NSAID), and is widely used to treat rheumatoid arthritis and osteoarthritis, as well as musculoskeletal disorders, dysmenorrhea, and postoperative pain. It has a long half-life and can be administered once daily.
See also: piroxicam ethanolamine (its active ingredient); piroxicam beta-dexamethasone (belonging to this subclass).
Indications
For the treatment of osteoarthritis and rheumatoid arthritis.
FDA Label
Mechanism of Action
The anti-inflammatory effect of piroxicam may stem from its reversible inhibition of cyclooxygenase, thereby inhibiting the synthesis of peripheral prostaglandins. Prostaglandins are produced by an enzyme called COX-1. Piroxicam inhibits prostaglandin production by blocking the COX-1 enzyme. Piroxicam also inhibits the migration of leukocytes to sites of inflammation and prevents platelet aggregation into thromboxane A2 (a platelet aggregator).

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Pharmacodynamics
Piroxicam belongs to the class of nonsteroidal anti-inflammatory drugs (NSAIDs). Piroxicam works by reducing hormones in the body that cause inflammation and pain. Piroxicam is used to relieve pain, inflammation, and stiffness caused by rheumatoid arthritis and osteoarthritis.

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1. Piroxicam (CP-16171) is a nonsteroidal anti-inflammatory drug (NSAID) that primarily inhibits COX-1. Its anti-inflammatory effect is mainly exerted by inhibiting cyclooxygenase (COX) to reduce the synthesis of prostaglandins. It has been used clinically to treat rheumatoid arthritis and osteoarthritis [1]. 2. The anti-bladder cancer effect of piroxicam in dogs is related to a dual mechanism: (1) inhibiting the COX-2/VEGF signaling pathway to reduce tumor angiogenesis; (2) promoting tumor cell apoptosis by regulating the Bax/Bcl-2/caspase-3 pathway [2]. 3. The synergistic effect of piroxicam and carboplatin on bladder cancer cells may be due to piroxicam enhancing carboplatin-induced DNA damage and apoptosis, which provides a potential treatment strategy for advanced bladder cancer [3].

C15H13N3O4S

331.35

331.062

36322-90-4

Piroxicam-d3;942047-64-5;Piroxicam-d4

54676228

Light yellow to yellow solid powder

1.5±0.1 g/cm3

568.5±60.0 °C at 760 mmHg

198-200°C

297.6±32.9 °C

0.0±1.6 mmHg at 25°C

1.692

2.23

2

6

2

23

611

0

QYSPLQLAKJAUJT-UHFFFAOYSA-N

InChI=1S/C15H13N3O4S/c1-18-13(15(20)17-12-8-4-5-9-16-12)14(19)10-6-2-3-7-11(10)23(18,21)22/h2-9,19H,1H3,(H,16,17,20)

4-hydroxy-2-methyl-1,1-dioxo-N-pyridin-2-yl-1λ6,2-benzothiazine-3-carboxamide

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)

Solubility in Formulation 1: ≥ 2.5 mg/mL (7.54 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.5 mg/mL (7.54 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
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.

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Solubility in Formulation 3: ≥ 2.5 mg/mL (7.54 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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