-β-cyclodextrin Chemical Structure.jpg)
(2-Hydroxypropyl)-β-cyclodextrin is a cyclodextrin oligosaccharide that has been widely used drug delivery vehicle to improve the aqueous solubility, stability and bioavailability of chemical compounds.
Physicochemical Properties
| Molecular Formula | C63H12O42 |
| Molecular Weight | 1541.54 |
| Exact Mass | 1540.662 |
| CAS # | 128446-35-5 |
| Related CAS # | 128446-35-5 ;107745-73-3; |
| PubChem CID | 4363642 |
| Appearance | Typically exists as White to off-white solids at room temperature |
| Density | 1.4±0.1 g/cm3 |
| Boiling Point | 1521.9±60.0 °C at 760 mmHg |
| Melting Point | 278ºC (dec.) |
| Flash Point | 874.2±32.9 °C |
| Vapour Pressure | 0.0±0.6 mmHg at 25°C |
| Index of Refraction | 1.545 |
| LogP | -6.23 |
| Hydrogen Bond Donor Count | 21 |
| Hydrogen Bond Acceptor Count | 42 |
| Rotatable Bond Count | 28 |
| Heavy Atom Count | 105 |
| Complexity | 2010 |
| Defined Atom Stereocenter Count | 0 |
| SMILES | CO[C@@H]1[C@H](O[R])[C@@H](O)[C@H](C)[C@@H](CO[R])O1.[C;D2]CC(O)C.[R].[7].[R=H or] |
| InChi Key | ODLHGICHYURWBS-FOSILIAISA-N |
| InChi Code | InChI=1S/C63H112O42/c1-22(64)8-85-15-29-50-36(71)43(78)57(92-29)100-51-30(16-86-9-23(2)65)94-59(45(80)38(51)73)102-53-32(18-88-11-25(4)67)96-61(47(82)40(53)75)104-55-34(20-90-13-27(6)69)98-63(49(84)42(55)77)105-56-35(21-91-14-28(7)70)97-62(48(83)41(56)76)103-54-33(19-89-12-26(5)68)95-60(46(81)39(54)74)101-52-31(17-87-10-24(3)66)93-58(99-50)44(79)37(52)72/h22-84H,8-21H2,1-7H3/t22?,23?,24?,25?,26?,27?,28?,29-,30-,31-,32-,33-,34-,35-,36-,37-,38-,39-,40-,41-,42-,43-,44-,45-,46-,47-,48-,49-,50-,51-,52-,53-,54-,55-,56-,57-,58-,59-,60-,61-,62-,63-/m0/s1 |
| Chemical Name | (1R,3S,5S,6R,8S,10S,11R,13S,15S,16R,18S,20S,21R,23S,25S,26R,28S,30S,31R,33S,35S,36S,37S,38S,39S,40S,41S,42S,43S,44S,45S,46S,47S,48S,49S)-5,10,15,20,25,30,35-heptakis(2-hydroxypropoxymethyl)-2,4,7,9,12,14,17,19,22,24,27,29,32,34-tetradecaoxaoctacyclo[31.2.2.23,6.28,11.213,16.218,21.223,26.228,31]nonatetracontane-36,37,38,39,40,41,42,43,44,45,46,47,48,49-tetradecol |
| Synonyms | Hydroxypropyl betadex (2-Hydroxypropyl)-β-cyclodextrinHydroxypropyl-β-cyclodextrin; HP-β-CD; (2-Hydroxypropyl)-; A-cyclodextrin; MFCD00069372; HP-??cyclodextrin; ODLHGICHYURWBS-FOSILIAISA-N; HMS3885J22; s4760; |
| HS Tariff Code | 2934.99.9001 |
| Storage |
Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month |
| Shipping Condition | Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs) |
Biological Activity
| Targets |
Vehicle for drug delivery Transcription Factor EB (TFEB) activator, cholesterol depletion agent[1] Cholesterol homeostasis modulator[2] |
| ln Vitro |
Treatment of cells with HP-β-CD activates the transcription factor EB, a fundamental regulator of lysosomal activity and autophagy, and promotes autophagic clearance [1]. HP-β-CD therapy lowered intracellular cholesterol and effectively suppressed leukemic cell proliferation through G2/M cell cycle arrest and death. After 72 hours of exposure, the IC50 values of HP-β-CD were in the range of 3.86-10.09 mM. HP-β-CD also displayed anticancer effects on CML cells bearing the T315I BCR-ABL mutation (conferring resistance to most ABL tyrosine kinase inhibitors) and hypoxia-adapted CML cells with leukemic stem cell features. In addition, the colony-forming ability of human primary AML and CML cells is reduced by HP-β-CD [2]. Treatment with HPβCD (0.1-10 mM) induced nuclear translocation of TFEB in HeLa cells stably expressing TFEB-3×FLAG, indicating TFEB activation. The extent of nuclear translocation was concentration-dependent, with maximum activation observed at 1-10 mM[1] HPβCD (1 mM) treatment upregulated mRNA expression of TFEB target genes involved in lysosomal function (GBA: 5.1-fold; HEXA: 2.7-fold; LAMP1: 3.5-fold) and autophagy (MAPLC3B: 4.8-fold; SQSTM1: 2.1-fold; BECN1: 3.0-fold) in HeLa/TFEB cells after 24 hours[1] HPβCD (1 mM) treatment induced autophagy activation, as shown by increased formation of GFP-LC3 puncta, increased LC3-II/LC3-I ratio in Western blot, and enhanced autophagic flux in the presence of bafilomycin A1 (100 nM)[1] In LINCL patient-derived fibroblasts, HPβCD (0.1-10 mM) treatment reduced the accumulation of autofluorescent ceroid lipopigment in a concentration- and time-dependent manner, with maximal clearance at 1-10 mM after 3 days[1] HPβCD (1 mM) treatment in LINCL fibroblasts also induced TFEB nuclear translocation and upregulated lysosomal (GBA: 2.7-fold; HEXA: 2.8-fold; LAMP1: 1.6-fold) and autophagy (MAPLC3B: 3.2-fold; SQSTM1: 2.5-fold; BECN1: 2.5-fold) gene expression after 3 days[1] TFEB siRNA silencing attenuated HPβCD-mediated clearance of ceroid lipopigment and upregulation of lysosomal/autophagy genes, confirming TFEB's role in the process[1] ATG7 siRNA silencing also reduced HPβCD-mediated clearance of ceroid lipopigment, indicating the clearance is autophagy-dependent[1] HPβCD treatment (0.1-10 mM, 3 days) did not induce early or late apoptosis in LINCL fibroblasts, as measured by annexin V and propidium iodide staining[1] HP-β-CyD inhibited the growth of various leukemic cell lines (acute myeloid leukemia, acute lymphoblastic leukemia, chronic myeloid leukemia) in a dose- and time-dependent manner. IC₅₀ values after 72 hours exposure ranged from 3.86 mM to 10.09 mM across 13 cell lines[2] HP-β-CyD (5-15 mM) induced apoptosis in BV173 and K562 cells, as measured by Annexin V/7-AAD staining after 24 hours[2] HP-β-CyD (5-15 mM) induced G₂/M cell-cycle arrest in BV173, K562, and other leukemic cell lines after 12 hours, as determined by propidium iodide staining and flow cytometry[2] HP-β-CyD (5-10 mM) treatment for 1-3 hours promoted cholesterol efflux from Ba/F3 BCR-ABLWT and BV173 cells in a time- and dose-dependent manner, and decreased intracellular cholesterol content[2] HP-β-CyD (10 mM) treatment modulated signaling pathways in leukemic cells: it inhibited phosphorylation of Stat5, Akt, and Lyn in BV173 cells, and reduced p-Lyn levels in K562 cells (with recovery after 8-24 hours), while increasing p-ERK1/2 levels in both cell lines[2] HP-β-CyD inhibited the growth of Ba/F3 cells expressing the T315I BCR-ABL mutation (conferring resistance to imatinib and dasatinib) with an IC₅₀ of 6.87 ± 0.76 mM, comparable to its effect on wild-type cells[2] HP-β-CyD inhibited the growth of hypoxia-adapted (stem cell-like) CML cell lines K562/HA and KCL22/HA, with IC₅₀ values of 3.86 mM and 5.61 mM, respectively[2] HP-β-CyD (5-15 mM) inhibited the colony-forming ability of primary mononuclear cells from acute myeloid leukemia (AML) and accelerated phase chronic myeloid leukemia (CML-AP) patients in methylcellulose assays[2] HP-β-CyD (up to 15 mM) did not significantly inhibit the growth of normal human primary hepatocytes (IC₅₀: 18.65 mM)[2] In a colony formation assay using normal mouse bone marrow mononuclear cells, HP-β-CyD at 5 mM and 15 mM reduced colony numbers to 93% and 84% of control, respectively, while 25 mM reduced it to 52.4%[2] |
| ln Vivo |
Because cells generated from patients with lysosomal storage illnesses have reduced activity of the lysosomal autophagy system, HP-β-CD treatment increases transcription factor EB-mediated clearance of proteolipid aggregates and accumulates [1]. Leukemia mice models can have a much higher survival rate when HP-β-CD is injected intraperitoneally. Mice given HP-β-CD systemically did not exhibit any overt negative effects [2]. Intraperitoneal administration of HP-β-CyD (50 mM or 150 mM in saline, 200 μL, twice daily for 20 days) significantly prolonged survival in nude mice transplanted with Ba/F3 BCR-ABLWT leukemia cells compared to vehicle-treated controls[2] Intraperitoneal administration of HP-β-CyD (50 mM or 150 mM in saline, 200 μL, for 5 consecutive days per week for 13 weeks) significantly prolonged survival in NOD/SCID mice xenotransplanted with human BV173 leukemia cells compared to vehicle-treated controls[2] |
| Enzyme Assay |
Tripeptidyl peptidase 1 (TPP1) enzymatic activity assay was performed to confirm the null activity in LINCL patient-derived fibroblasts compared to healthy individual fibroblasts. Cells were lysed, and the supernatant was collected for the assay using a specific substrate to measure TPP1 activity[1] |
| Cell Assay |
Effects of HP-β-CyD on in vitro cell growth[2] \nCell viability was assessed using a trypan blue dye exclusion method and cell proliferation was evaluated using a modified methyl-thiazol-diphenyl- tetrazolium (MTT) assay with SF reagent as described previously. Cells, including human primary hepatocytes were seeded in flat-bottomed 96-well plates at a density of 1×104 cells in 100 μL medium per well, and incubated with HP-β-CyD at various concentrations for 72 hours. The mean of three replicates was calculated for each concentration. \n\nWestern blot analysis[2] \nWhole cell lysates of leukemic cells treated with or without HP-β-CyD were prepared from cells using lysis buffer, as reported previously, with minor modification. Protein was separated using a 10% NuPAGE electrophoresis system, transferred to a nitrocellulose membrane, blocked with 5% bovine serum albumin at room temperature for 1 hour, and incubated with primary antibodies at 4°C overnight. Antibodies against Akt, phosphorylated-Akt (Thr308 or Ser473), phospholyrated-Erk1/2 (Thr202/Thr204), phospholyrated-Stat5, Lyn, Stat5, Erk1/2, Actin, and phospholyrated-Lyn were used as primary antibodies. Horseradish peroxidase-coupled immunoglobulin IgG was used as the secondary antibody. An enhanced chemiluminescence kit was used for detection. The results are representative of at least two independent experiments. Intensity of the immunoblot signals after background subtraction was quantified using ImageJ software. \n\nCell-cycle analysis[2] \nCell-cycle analyses of human leukemic cell lines were performed as described previously. In brief, 1×106 cells were treated with the indicated concentration of HP-β-CyD. Twelve or twenty-four hours after HP-β-CyD treatment, cells were collected and fixed with 70% ethanol. Cells were then incubated with 0.1% Triton X-100 and 0.5% RNase A at room temparature for 30 minutes and stained with 50 μg/mL propidium iodide. Cellular DNA content was analyzed by flow cytometry, and cell-cycle profiles were determined using a FACS Caliber flow cytometer with CellQuest software. Data are the mean ± SD of three independent experiments. \n\nApoptosis assays[2] \nApoptosis assay was performed by staining cells with 7-amino-actinomycin D (7-AAD) and annexin V, according to the manufacturer’s instructions. Cells were cultured in 6-well plate at a density of 4×105 cells, and incubated with various concentrations of HP-β-CyD for 12 or 24 hours. Then, cells were stained with 7-amino-actinomycin D (7-AAD) and Annexin V-APC, and analyzed using a FACSAriaII system with Diva software. \n\nHematopoietic colony-forming assays[2] \nHP-β-CyD toxicity in normal hematopoietic progenitors was investigated using a standard methylcellulose culture assay as described previously. A total of 2×104 mononuclear cells from the BM of 10-week-old C57BL/6N mice were exposed to 0, 5, 15, or 25 mM HP-β-CyD in 1 mL MethoCult M3434 . After 8 days of culture, the number of colonies was counted using an inverted microscope. Data represent the mean number of colonies ± SD (n = 3). Clinical samples were obtained with informed consent. Mononuclear cells from leukemia patients were cultured in semi-solid medium containing recombinant cytokines. \n\nCholesterol assays[2] \nLeukemic cells (3×106) were incubated with 5 or 10 mM HP-β-CyD in HBSS (pH 7.4) at 37°C for 1, 2, or 3 hours. Cell culture supernatants were recovered by centrifugation (3,000 rpm, 5 min). The concentration of total cholesterol in the supernatants was determined using a Cholesterol E-test Wako. Data are the mean ± SD of three experiments. Cellular lipids were extracted with methanol:chloroform (1:2), and total cholesterol and free cholesterol were determined enzymatically. The amount of esterified cholesterol was calculated by subtracting free cholesterol from total cholesterol. Cellular protein concentration was determined by BCA assay. Data are the mean ± SD of three experiments. For filipin staining, cells were incubated with β-CyDs (10 mM) for 1 hour. Thereafter, cellular cholesterol was detected using a Cholesterol Cell-Based Detection Assay Kit. For immunofluorescence assays, cells were seeded on coverslips and treated with HPβCD, then fixed and permeabilized. Primary antibodies against targets such as FLAG, LC3, LAMP2, or TFEB were applied, followed by fluorescent secondary antibodies. Images were acquired by confocal microscopy and analyzed for co-localization and intensity[1] Quantitative RT-PCR was used to measure mRNA expression. Total RNA was extracted from treated cells, cDNA was synthesized, and qPCR was performed using gene-specific primers. Expression levels were normalized to GAPDH and compared to untreated controls[1] Western blot analysis was performed to detect protein levels. Cells were lysed, proteins were separated by SDS-PAGE, transferred to membranes, and probed with primary antibodies against LC3 and GAPDH, followed by HRP-conjugated secondary antibodies and chemiluminescent detection[1] Apoptosis assay was conducted using an annexin V-FITC/propidium iodide kit. Treated cells were stained and analyzed by flow cytometry to quantify early and late apoptotic populations[1] For siRNA transfection, cells were seeded in plates and transfected with TFEB-specific or control siRNA using a transfection reagent. After 2 days, cells were treated with HPβCD for an additional 3 days before analysis[1] Cell viability and proliferation were assessed using trypan blue dye exclusion and a modified MTT assay. Cells were seeded in 96-well plates and treated with HP-β-CyD for 72 hours. IC₅₀ values were determined using nonlinear regression analysis[2] For apoptosis assay, cells were cultured with HP-β-CyD for 12 or 24 hours, then stained with Annexin V-APC and 7-AAD, and analyzed by flow cytometry[2] For cell-cycle analysis, cells treated with HP-β-CyD for 12 or 24 hours were fixed, permeabilized, treated with RNase A, stained with propidium iodide, and analyzed by flow cytometry to determine DNA content[2] For cholesterol efflux assay, leukemic cells were incubated with HP-β-CyD in buffer, supernatants were collected by centrifugation, and total cholesterol concentration was measured using a commercial enzymatic kit[2] For intracellular cholesterol measurement, lipids were extracted from treated cells using methanol:chloroform, and total and free cholesterol were determined enzymatically. Esterified cholesterol was calculated by subtraction[2] For filipin staining, cells treated with HP-β-CyD were stained with filipin solution and visualized by fluorescence microscopy to assess cellular cholesterol levels[2] For Western blot analysis, whole cell lysates from treated cells were prepared, proteins were separated by SDS-PAGE, transferred to membranes, and probed with specific primary and secondary antibodies for detection[2] For hematopoietic colony-forming assays, bone marrow mononuclear cells or primary leukemic cells were plated in methylcellulose medium containing HP-β-CyD and cytokines. Colonies were counted after 8-13 days of culture[2] |
| Animal Protocol |
Murine leukemia model[2] Two different experimental settings were used. The protocol was approved by the Committee on the Ethics of Animal Experiments of the Saga University (Permit number: 25-028-0). First, nude mice were intravenously transplanted with 1×106 EGFP+ Ba/F3 BCR-ABLWT cells. These mice were intraperitoneally injected with 200 μL vehicle (saline) or HP-β-CyD (50 or 150 mM) for 20 consecutive days 3 days after transplantation, and survival was monitored daily. Leukemic cell engraftment was confirmed by detection of GFP-positive cells in the recipient’s BM using flow cytometry.[2] The second experimental setting involved a human leukemia xenograft model. BV173 cells (1×106) were intravenously injected into sublethally irradiated (2 Gy) NOD/SCID mice. After 72 hours, xenotransplanted mice were intraperitoneally injected with 200 μL vehicle or HP-β-CyD (50 or 150 mM) for 5 consecutive days every week for 13 weeks, and survival was monitored daily. The percentage of human leukemic cells in BM was determined by flow cytometry after double staining with FITC-conjugated anti-human CD19 and PE/Cy7-conjugated anti-mouse CD45 antibodies. All surgery was performed under sodium pentobarbital anesthesia, and all efforts were made to minimize suffering. Mice were euthanized with ether when they became moribund or unable to obtain food or water, as recommended by the institutional guidelines of Saga University. Survival data were analyzed by a log-rank nonparametric test and shown as Kaplan-Meier survival curves (n = 10 for each group).[2] Lung histology[2] HP-β-CyD was administered to NOD/SCID mice for 13 weeks as described in the above section entitled “Murine leukemia model”. Age-matched mice were used as a control. Lungs were perfused with 10% buffered formalin and excised. Tissues were fixed in 10% buffered formalin and embedded in paraffin. These blocks were then sectioned and stained with hematoxylin and eosin (H&E). For the murine Ba/F3 BCR-ABLWT leukemia model, nude mice were intravenously transplanted with 1×10⁶ EGFP⁺ Ba/F3 BCR-ABLWT cells. Three days later, mice received intraperitoneal injections of 200 μL vehicle (saline), 50 mM HP-β-CyD (695.5 mg/kg), or 150 mM HP-β-CyD (2086.5 mg/kg) twice daily for 20 consecutive days. Survival was monitored daily[2] For the human leukemia xenograft model, NOD/SCID mice were sublethally irradiated (2 Gy) and intravenously injected with 1×10⁶ BV173 cells. Three days later, mice received intraperitoneal injections of 200 μL vehicle (saline), 50 mM HP-β-CyD (695.5 mg/kg), or 150 mM HP-β-CyD (2086.5 mg/kg) for 5 consecutive days every week for 13 weeks. Survival was monitored daily[2] |
| Toxicity/Toxicokinetics |
In the described mouse studies, systemic administration of HP-β-CyD (up to 150 mM, 2086.5 mg/kg, i.p.) did not cause gross lesions, hemolysis, or anemia[2] Histological examination of lungs from NOD/SCID mice treated with HP-β-CyD for 13 weeks showed no obvious changes compared to age-matched controls[2] In contrast, all mice injected with 150 mM methyl-β-cyclodextrin (M-β-CyD) died of diffuse alveolar hemorrhage within 24 hours[2] |
| References |
[1]. 2-Hydroxypropyl-β-cyclodextrin promotes transcription factor EB-mediated activation of autophagy: implications for therapy. J Biol Chem. 2014 Apr 4;289(14):10211-22. [2]. 2-Hydroxypropyl-β-Cyclodextrin Acts as a Novel Anticancer Agent. PLoS One. 2015 Nov 4;10(11):e0141946. |
| Additional Infomation |
Derivative of beta-cyclodextrin that is used as an excipient for steroid drugs and as a lipid chelator. HPβCD is an FDA-approved excipient used to improve drug stability and bioavailability[1] It can extract cholesterol from biological membranes by trapping it in its hydrophobic core[1] The study proposes a model where cellular uptake of HPβCD leads to TFEB activation, which in turn upregulates lysosomal and autophagy genes, enhancing clearance of storage material like ceroid lipopigment in lysosomal storage disorder models[1] HPβCD-mediated autophagy activation under the studied conditions was not associated with apoptosis induction, suggesting a pro-survival response[1] HP-β-CyD is an FDA-approved pharmaceutical excipient used to improve drug solubility and bioavailability, and is clinically used for Niemann-Pick Type C disease[2] The study proposes that HP-β-CyD exerts anticancer effects by disrupting cholesterol homeostasis in cancer cells, leading to apoptosis and cell-cycle arrest[2] HP-β-CyD showed efficacy against TKI-resistant leukemia cells (including T315I mutant) and hypoxia-adapted stem cell-like leukemic cells[2] The concentration of HP-β-CyD required for in vivo efficacy (approximately 2 g/kg) is comparable to doses used in clinical trials for Niemann-Pick Type C disease[2] |
Solubility Data
| Solubility (In Vitro) |
DMSO : ~50 mg/mL H2O : ~50 mg/mL |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.08 mg/mL (Infinity 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 20.8 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. Solubility in Formulation 2: ≥ 2.08 mg/mL (Infinity 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 20.8 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. Solubility in Formulation 3: ≥ 2.08 mg/mL (Infinity 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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 4: 100 mg/mL (Infinity mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication (<60°C). Solubility in Formulation 5: 200 mg/mL (Infinity 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. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 0.6487 mL | 3.2435 mL | 6.4870 mL | |
| 5 mM | 0.1297 mL | 0.6487 mL | 1.2974 mL | |
| 10 mM | 0.0649 mL | 0.3244 mL | 0.6487 mL |