Ibandronate inhibits the mevalonate pathway, specifically targeting the farnesyl-IPP-synthase (farnesyl diphosphate synthase). This inhibition prevents protein prenylation, particularly farnesylation, which is crucial for the function of signaling proteins like Ras. [2]
Ibandronate is a nitrogen-containing bisphosphonate that exerts its therapeutic effect by reducing osteoclast-mediated bone resorption. It inhibits bone turnover and increases bone mineral density (BMD). [3]

Ibandronate, administered in a metronomic fashion (daily 1-hour exposure for 8 days), significantly reduced the growth of human umbilical vein endothelial cells (HUVECs). A concentration of 1.25 µM caused a 42% reduction compared to 1 µM. Concentrations of 1.5, 1.75, and 2 µM caused reductions of 46%, 43%, and 50%, respectively, compared to 1 µM. Concentrations of 2 µM and higher (10 µM, 100 µM) caused significant reductions in cell growth compared to untreated controls, with the effect plateauing above 2 µM. [1]
Ibandronate (2, 10, 100 µM, metronomic administration) significantly inhibited capillary-like tube formation of HUVECs on Matrigel compared to untreated controls. [1]
Ibandronate (2 and 10 µM, metronomic administration) significantly increased apoptosis in HUVECs compared to untreated controls, as measured by an ELISA detecting mono- and oligonucleosomes. There was no significant increase in apoptosis at 1 µM or 100 µM. [1]
Western blot analysis showed that Ibandronate exposure increased the expression of vascular endothelial growth factor (VEGF) in HUVECs stimulated with basic fibroblast growth factor (bFGF). Compared to controls, 2 µM ibandronate resulted in a 1.4-fold increase, 10 µM in a 2-fold increase, and 100 µM in a 2.5-fold increase in VEGF expression. [1]
Ibandronate inhibited the growth of prostate cancer cell lines LNCaP (androgen-sensitive) and PC-3 (androgen-independent) in a dose-dependent manner after 5 days of treatment. The IC50 value for LNCaP cells was 3.7 µM, and for PC-3 cells it was 33.69 µM. [2]
The combination of Ibandronate (10 µM) and Docetaxel (4 nM) exhibited synergistic growth inhibitory effects on both LNCaP and PC-3 cells. The combination index (CI) was 0.659 for LNCaP and 0.76 for PC-3, indicating synergism (CI < 1). [2]
The growth inhibitory effect of Ibandronate on both LNCaP and PC-3 cells was completely reversed by the addition of farnesol (FOH, 50-100 µM) or farnesyl pyrophosphate (FPP, 1-10 µM). [2]
Surprisingly, farnesol (FOH, 75 µM) alone exhibited strong growth inhibitory effects on LNCaP cells, reducing viable cell number to 8.8% of the control. When combined with Ibandronate (10 µM) and Docetaxel (4 nM), FOH (75 µM) did not reverse the inhibition but instead further enhanced the growth inhibitory effect. [2]

In preclinical studies conducted in rats, dogs, and monkeys, intermittent treatment with ibandronate produced equivalent benefits to daily regimens in reversing bone loss, increasing bone strength, and restoring or preventing loss of normal bone architecture. [3]

For the cell growth assay, HUVECs were seeded in culture flasks and allowed to adhere overnight. Cells were then treated with ibandronate at specified concentrations for 1 hour daily for 8 consecutive days. After each exposure, the drug was washed off with PBS, and fresh medium was added. On day 9, cells were trypsinized and counted using an automated cell counter. An untreated control group underwent the same washing and incubation procedure without the drug. [1]
For the tube formation assay, 24-well plates were pre-coated with growth factor-reduced Matrigel. HUVECs from the growth assay (including untreated controls) were transferred onto the Matrigel-coated wells. After incubation for 10 hours, the formation of capillary-like tubular networks was observed under a phase-contrast microscope. Tube length was measured and calculated as mean length per area. [1]
Apoptosis was determined using a specific ELISA kit that detects histone-associated DNA fragments (mono- and oligonucleosomes) in the cytoplasmic fraction of cell lysates. HUVECs were treated with ibandronate following the same metronomic protocol as the growth assay. On day 9, a specific number of cells from each treatment group were lysed, and the cytoplasmic fraction was analyzed according to the kit's instructions. Absorbance was measured at 405 nm, which is proportional to the amount of nucleosomes. [1]
For Western blot analysis, total protein was extracted from HUVECs after the metronomic treatment protocol. Equal amounts of protein were separated by SDS-PAGE, transferred to a nitrocellulose membrane, and probed with a primary antibody against VEGF. After washing, the membrane was incubated with a horseradish peroxidase-conjugated secondary antibody. Protein bands were visualized using a chemiluminescence detection system and quantified by densitometry. GAPDH was used as a loading control. [1]
Prostate cancer cells (LNCaP and PC-3) were seeded in 96-well plates and allowed to adhere overnight. The cells were then treated with increasing concentrations of the test compounds (Ibandronate, Docetaxel, Farnesol, FPP) alone or in combination. Control cultures received an equivalent volume of the respective diluent (PBS or DMSO). After 5 days of incubation, cell viability was assessed. [2]
Cell viability was measured using both direct cell counting with a hemocytometer and the MTT assay. For the MTT assay, the culture medium was removed, cells were washed, and MTT reagent was added to each well. After a 4-hour incubation at 37°C to allow formazan crystal formation, the MTT solution was removed, and the crystals were dissolved using DMSO. The absorbance of the resulting solution was measured at 570 nm. Results were expressed as a percentage of viable cells compared to untreated controls. [2]
The combined effects of Ibandronate and Docetaxel were quantitatively analyzed for synergism using the median effect method and calculation of the combination index (CI) as described by Chou. A CI < 1 indicates synergism. [2]

This clinical study refers to prior preclinical studies but does not describe specific animal experiment protocols (e.g., formulation, dosing regimen) within its own methods. [3]
The clinical trial itself was a human study. Postmenopausal osteoporotic women received oral ibandronate either daily (2.5 mg) or intermittently (20 mg every other day for 12 doses every 3 months). Tablets were taken immediately after rising and 1 hour before food, non-study medications, and fluids other than plain water. All participants received daily calcium (500 mg) and vitamin D (400 IU) supplementation. [3]

Absorption, Distribution and Excretion
The bioavailability of oral ibandronate sodium is 0.63%. In a study of healthy men, the time to peak concentration (Tmax) of a 10 mg oral dose was 1.1 ± 0.6 hours, and the peak plasma concentration (Cmax) was 4.1 ± 2.6 ng/mL. The time to peak concentration was approximately 1 hour, while the peak plasma concentration varied with the dose. The AUC of ibandronate sodium after intravenous injection was 316 nGh/mL for 2 mg, 581 nGh/mL for 4 mg, and 908 nGh/mL for 6 mg. Ibandronate sodium is primarily excreted in the urine, with unabsorbed drug excreted unchanged in the feces. The apparent terminal volume of distribution of ibandronate sodium was 90–368 L in healthy subjects and 103 L in postmenopausal women with osteopenia.
The total clearance of ibandronate sodium is 84-160 mL/min.
Metabolism/Metabolites
Ibandronate sodium is not metabolized in the human body.
There is no evidence that ibandronate sodium is metabolized in the human body.
Excretion route: Ibandronate sodium is excreted via the kidneys. Unabsorbed ibandronate sodium is excreted unchanged in the feces.
Half-life: 10-60 hours

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Biological half-life The half-life of ibandronate sodium in postmenopausal women is 37-157 hours.

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The half-life of bisphosphonates (including ibandronate sodium) in circulation is 0.5 to 2 hours. [1]
Oral bisphosphonates are poorly absorbed in the gastrointestinal tract and vary greatly from person to person, with bioavailability of approximately 1-10%. [1]
After oral administration of ibandronate sodium
, plasma concentrations increase linearly, nonsaturatedly, and in a dose-dependent manner. Daily oral administration leads to plasma drug accumulation, reaching steady state after 8 days. After 12 months of daily oral administration, plasma drug concentrations can accumulate to 1.5 to 2 times. [1]
After intravenous administration of 6 mg ibandronate sodium, peak plasma concentrations are 1–2 µM, and rapidly decrease to 10% of the maximum concentration within 3–8 hours after administration. [1]
The bioavailability and plasma concentrations of daily oral administration of 50 mg ibandronate sodium are comparable to those of monthly intravenous administration of 6 mg ibandronate sodium. [1]

Toxicity Summary
Ibandronate sodium's effects on bone tissue are partly based on its affinity for hydroxyapatite, a component of the bone mineral matrix. Nitrogenous bisphosphonates (such as pamidronate sodium, alendronate sodium, risedronate sodium, ibandronate sodium, and zoledronic acid sodium) appear to act as analogs of isoprene diphosphate lipids, thereby inhibiting farnesyl pyrophosphate (FPP) synthase, an enzyme in the mevalonate pathway. Inhibition of this enzyme in osteoclasts prevents the biosynthesis of isoprene lipids (FPP and GGPP), which are crucial for the post-translational farnesylation and geranylation of small GTPase signaling proteins. This activity inhibits osteoclast activity and reduces bone resorption and turnover. In postmenopausal women, it reduces elevated bone turnover, resulting in a net increase in bone mass on average.

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Effects during pregnancy and lactation>
◉ Overview of use during lactation
Ibandronate sodium has a very low oral absorption rate (average absorption rate of 6% in adults on an empty stomach, negligible when taken with food), therefore breastfed infants are unlikely to absorb ibandronate sodium. However, since there is currently no information on the use of ibandronate sodium during lactation, other medications may be preferred, especially for breastfed newborns or premature infants.
◉ Effects on breastfed infants
As of the revision date, no relevant published information was found.
◉ Effects on lactation and breast milk
As of the revision date, no relevant published information was found.

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Protein binding>
The protein binding rate of ibandronate sodium in serum is 85.7-99.5% in the concentration range of 0.5-10 ng/mL, but is generally 86% in the concentration range of 20-2000 ng/mL.

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Toxicity Data
LD₅₀ = 811 mg/kg (rat, oral)

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In this 3-year clinical trial, oral ibandronate sodium (once daily) and intermittent ibandronate sodium were well tolerated, with an overall incidence of adverse events similar to that in the placebo group. No statistically significant differences were found in overall adverse events, drug-related adverse events, or drug-related adverse events leading to discontinuation of treatment. [3]
Although approximately 30% of patients had a history of gastrointestinal disease, the incidence of upper gastrointestinal adverse events (e.g., dyspepsia, nausea, gastritis) was comparable in the placebo group, the daily ibandronate sodium group, and the intermittent ibandronate sodium group. The specific incidence of upper gastrointestinal events is shown in Table 3 of this publication. [3]
No changes in clinically significant laboratory parameters (renal function, liver function, serum electrolytes, blood cell count) were observed, and the distribution of significant laboratory abnormalities was balanced among the treatment groups. [3]
The number of serious adverse events, withdrawals due to adverse events, and deaths remained balanced in both the placebo group and the ibandronate sodium treatment group. [3]

[1]. Morgan, C., S. Jeremiah, and J. Wagstaff, Metronomic administration of ibandronate and its anti-angiogenic effects in vitro. Microvasc Res, 2009. 78(3): p. 453-8.

[2]. Differential effects of ibandronate, docetaxel and farnesol treatment alone and in combination on the growth of prostate cancer cell lines. Acta Oncol, 2011. 50(1): p. 127-33.

[3]. Effects of oral ibandronate administered daily or intermittently on fracture risk in postmenopausal osteoporosis. J Bone Miner Res, 2004. 19(8): p. 1241-9.

[4]. Effects of treatment with ibandronate on bone mass, architecture, biomechanical properties, and bone concentration of ibandronate in ovariectomized aged rats. J Rheumatol, 2002. 29(10): p. 2200-8.

Pharmacodynamics
Ibandronate sodium is a nitrogen-containing bisphosphonate used to treat and prevent osteoporosis in postmenopausal women. It has a wide therapeutic index, low overdose toxicity, and a long duration of action with a half-life of up to 157 hours. Patients should be informed of the risks of upper gastrointestinal adverse reactions, hypocalcemia, musculoskeletal pain, osteonecrosis of the jaw, atypical femoral fractures, and severe renal impairment. Ibandronate sodium is a third-generation bisphosphonate. [1] This study simulated a rhythmic dosing regimen of oral ibandronate sodium (i.e., daily low-dose administration) to investigate its in vitro anti-angiogenic effects. [1] The anti-angiogenic effect of rhythmic injection of ibandronate sodium is thought to be achieved at least in part by inhibiting the stimulatory effect of basic fibroblast growth factor (bFGF) on endothelial cells. However, this inhibition leads to a compensatory increase in vascular endothelial growth factor (VEGF) expression. [1] The study concluded that, due to the interaction between bFGF and VEGF, the inhibition of bFGF by ibandronate sodium alone is unlikely to completely suppress angiogenesis, suggesting that combination therapy with anti-VEGF drugs may be necessary. [1] Ibandronate sodium is a second-generation bisphosphonate. [2] The study showed that the main mechanism by which ibandronate sodium inhibits the growth of prostate cancer cells is by inhibiting the mevalonate pathway, thereby preventing protein farnesylation. This is different from zoledronic acid, which mainly works by inhibiting geranyl geranylation. [2] The observed synergistic effect between ibandronate sodium and docetaxel may have clinical significance for the treatment of metastatic prostate cancer. [2] Farnesol, while able to antagonize the effects of ibandronate sodium by supplementing the farnesyl pool, also has intrinsic properties that inhibit the growth of prostate cancer cells, and its mechanism may be unrelated to the mevalonate pathway. [2]

C9H23NO7P2

319.22902

319.094

114084-78-5

Ibandronate Sodium Monohydrate;138926-19-9;Ibandronic Acid-d3 sodium;1329834-28-7;Ibandronic acid-d3;1130899-41-0;Ibandronate Sodium;138844-81-2

60852

Typically exists as solid at room temperature

1.5±0.1 g/cm3

587.8±60.0 °C at 760 mmHg

113-115ºC

309.3±32.9 °C

0.0±3.7 mmHg at 25°C

1.538

-0.65

5

8

9

19

342

0

OC(P(O)(O)=O)(P(O)(O)=O)CCN(C)CCCCC

MPBVHIBUJCELCL-UHFFFAOYSA-N

InChI=1S/C9H23NO7P2/c1-3-4-5-7-10(2)8-6-9(11,18(12,13)14)19(15,16)17/h11H,3-8H2,1-2H3,(H2,12,13,14)(H2,15,16,17)

[1-hydroxy-3-[methyl(pentyl)amino]-1-phosphonopropyl]phosphonic acid

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.

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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).

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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)

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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).

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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

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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.

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