Iron chelator

Dexrazoxane binds iron to stop the production of superhydroxide radicals, thereby averting mitochondrial damage. It is hydrolyzed to its active form inside cells.

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Advances in cancer treatment utilizing multiple chemotherapies have dramatically increased cancer survivorship. Female cancer survivors treated with doxorubicin (DXR) chemotherapy often suffer from an acute impairment of ovarian function, which can persist as long-term, permanent ovarian insufficiency. Dexrazoxane (Dexra) pretreatment reduces DXR-induced insult in the heart, and protects in vitro cultured murine and non-human primate ovaries, demonstrating a drug-based shield to prevent DXR insult. The present study tested the ability of Dexra pretreatment to mitigate acute DXR chemotherapy ovarian toxicity in mice through the first 24 hours post-treatment, and improve subsequent long-term fertility throughout the reproductive lifespan. Adolescent CD-1 mice were treated with Dexra 1 hour prior to DXR treatment in a 1:1 mg or 10:1 mg Dexra:DXR ratio. During the acute injury period (2-24 hours post-injection), Dexra pretreatment at a 1:1 mg ratio decreased the extent of double strand DNA breaks, diminished γH2FAX activation, and reduced subsequent follicular cellular demise caused by DXR. In fertility and fecundity studies, dams pretreated with either Dexra:DXR dose ratio exhibited litter sizes larger than DXR-treated dams, and mice treated with a 1:1 mg Dexra:DXR ratio delivered pups with birth weights greater than DXR-treated females. While DXR significantly increased the "infertility index" (quantifying the percentage of dams failing to achieve pregnancy) through 6 gestations following treatment, Dexra pretreatment significantly reduced the infertility index following DXR treatment, improving fecundity. Low dose Dexra not only protected the ovaries, but also bestowed a considerable survival advantage following exposure to DXR chemotherapy. Mouse survivorship increased from 25% post-DXR treatment to over 80% with Dexra pretreatment. These data demonstrate that Dexra provides acute ovarian protection from DXR toxicity, improving reproductive health in a mouse model, suggesting this clinically available drug may provide ovarian protection for cancer patients[3].

Cells were treated with 0.1% DMSO (solvent control) or 200 μmol/L Dexrazoxane for 5 h followed by coincubation with doxorubicin for 1 day or VP-16 for 2 days. MTT (0.1 mg) was then added to each well and cells were incubated for an additional 4 h at 37°C. After removal of medium, DMSO was added and absorbance at 570 nm was measured using the Microplate Reader. Average IC50 values (mean ± SE) were determined in triplicate or quadruplicate.[1]

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Neutral comet assay. Primary MEFs were treated with DMSO or doxorubicin for 1.5 h in a CO2 incubator at 37°C followed by additional 30-min incubation in fresh medium to reverse Top2 cleavage complexes. H9C2 cells were treated with DMSO or Dexrazoxane (100 μmol/L) for 3 h, washed, and replenished with fresh medium. Cells were then treated with DMSO or doxorubicin for 1.5 h followed by additional 30-min incubation in fresh medium to reverse Top2 cleavage complexes. Cells were then washed and trypsinized using 0.005% trypsin and resuspended in DMEM supplemented with 10% FetalPlex animal serum complex (10,000/mL). Cell suspension (50 μL) was then mixed with 500 μL 0.5% low-melting point agarose at 37°C. Cell/agarose mixture (75 μL) was transferred onto glass slides. Slides were then immersed in prechilled lysis buffer [2.5 mol/L NaCl, 100 mmol/L EDTA, 10 mmol/L Tris (pH 10.0), 1% Triton X-100, 10% DMSO] for 1 h followed by equilibration in 1× Tris-borate EDTA (TBE) buffer for 30 min. Slides were electrophoresed in 1× TBE at 1.0 V/cm for 10 min and stained with Vistra Green. Images were visualized under a fluorescence microscope and captured with a charge-coupled device camera. The average comet tail moment was determined from measuring at least 100 cells for each treatment group as described previously. Statistical analysis of the mean comet tail moments was done using Student's t test.[1]

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Band depletion assay. H9C2 cells (1.2 × 105) were treated with 250 μmol/L VP-16 in the presence or absence of Dexrazoxane (150 μmol/L) for 15 min. Cells were either lysed immediately or incubated in drug-free medium for another 30 min at 37°C (to reverse Top2 cleavage complexes) before lysis. Cell lysates were analyzed by Western blotting using the anti-Top2α/Top2β and anti–α-tubulin antibody. The amount of Top2 cleavage complexes can be estimated from the difference between the amount of free Top2 after reversal and the amount of free Top2 without reversal[1].

Mice [3]
\nAll surgery was performed under Ketamine and isofluorane anesthesia. Female CD-1 mice were allowed to acclimate to the laboratory environment for one week prior to the start of an experiment under the supervision and care of the animal facility staff. At 4 weeks of age, the adolescent mice were injected with Dexrazoxane/Dexra or vehicle control (0.0167 M lactate in saline) via intraperitoneal injection using ≤ 200 μL/injection 1 hour prior to DXR injection. DXR or vehicle (saline) was subsequently administered via intraperitoneal injection.\n

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\nAcute treatment [3]
\nAt 4 weeks of age, mice were treated with 1) Vehicle for Dexrazoxane/Dexra + Vehicle for DXR, 2) Vehicle for Dexra + 20 mg/kg DXR, 3) 20 mg/kg Dexra + Vehicle for DXR, or 4) 20 mg/kg Dexra + 20 mg/kg DXR; doses were calculated based on the average weight of a 4-week-old CD-1 mouse. The 20 mg/kg DXR dose represents twice the maximum human equivalent DXR dose and was chosen in order to engage ample acute DXR toxicity. The 20 mg/kg Dexra dose represents a 1:1 Dexra/DXR mg ratio, providing a significant dose reduction from that used in cardioprotection to limit potential side effects of Dexra. The chosen Dexra dose was based on our previous in vitro study demonstrating a 2 μM Dexra dose, 100-folds lower than that used in in vitro cardiac protection studies, preserved granulosa cell viability against DXR. Animals were euthanized with CO2 followed by cervical dislocation and ovaries removed surgically 0, 2, 4, 10, 12 or 24 h after the second injection. Experiments were carried out in 4 biological replicates in which 3 mice were treated per drug group and harvested for each time point per biological replicate; in sum, n = 12 animals per treatment were totaled across all replicates. Ovaries were placed in 2 mL phosphate buffered saline, pH 7.4, and cleared of fat and attached bursa. For each ovarian pair, one was fixed in 10% formalin and processed for TUNEL assay, and the second was processed for a neutral comet assay. Separate mice were treated to provide ovaries utilized for protein extraction followed by Western blot analysis as previously described.\n

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\nBreeding trial [3]
\nFemale CD-1 mice were housed in Innovive system cages from 3 weeks until 8 months of age. At 4 weeks of age, mice were treated with: 1) Vehicle for Dexrazoxane/Dexra + Vehicle for DXR, 2) Vehicle for Dexra + 10mg/kg DXR, 3) 10mg/kg Dexra (1:1 mg ratio) + 10mg/kg DXR, 4) 100 mg/kg Dexra (10:1 mg ratio) + 10mg/kg DXR, 5) 10mg/kg Dexra (1:1 mg ratio) + Vehicle for DXR, or 6) 100mg/kg Dexra (10:1 mg ratio) + Vehicle for DXR. DXR was administered at 10 mg/kg body weight (a human equivalent dose of 30mg/m2) to minimize long-term cardiotoxicity. Dexra dose is expressed as a ratio to DXR dose throughout the manuscript. Dexra was administered at either a 1:1 mg ratio (labeled as Dexra1:DXR1, groups 3 above) or 10:1 mg ratio (labeled as Dexra10:DXR1, group 4 above, currently used in cardioprotective protocols) to DXR as indicated. Dexra control-treated animals (groups 5 and 6, above) are labeled as DexraC (DexraC1 and DexraC10 respectively) throughout the manuscript. At 6 weeks of age and prior to breeding, animals were treated for two weeks with drinking water medicated with enrofloxacin (22.7 mg/ml) at a calculated dose of 5 mg/kg (0.5 mL/300 mL ddH2O bottle) as a prophylactic to mitigate the side effects of a compromised immune system brought on by DXR treatment. At 8 weeks of age, females were moved to breeder cages where two females were paired with one male. Females were continuously mated from 8 weeks of age to 8 months of age or until 6 litters were achieved. Males were rotated following each breeding round to minimize any potential male-specific infertility effect. Animals within the breeder cage were fed a maintenance chow diet with protein: 24%; Fat: 4%; Fiber: 4.5% as well as irradiated sunflower seeds. Bi-weekly assessment of animal health was conducted, and additional nutritive support via DietGel® and sunflower seeds was given to females having difficulty maintaining body condition. Females remained within the breeder cage until they showed visual or palpable signs of pregnancy, at which point they were separated and maintained on a breeder irradiated diet (Protein: 19%; Fat: 9%; Fiber: 5%) until parturition. The health of the breeding mice was monitored at least three times daily when the mice were near parturition.\n [3]
br>\n\nFollowing delivery, pups were separated and the females were returned to the breeder cage within 24 h post-partum. The pups were counted, weighed, and euthanized on post-natal day 1 (PND1). At 8 months of age, the now non-pregnant dams were weighed, anesthetized with isoflurane (confirmed with limb pinch) and sacrificed via terminal blood draw followed by cervical dislocation. A terminal blood draw was carried out for future studies. Ovaries were removed from each female and weighed. Mice that did not survive to breeding age or that displayed signs of deteriorating health were removed from the breeding trial to minimize any suffering. The breeding trial was carried out in 4 replicates, with 3–6 mice per group per replicate, where the total number of female mice in each group at the start of breeding was 16 control, 16 DXR, 21 Dexrazoxane/Dexra1:DXR1, 16 Dexra10:DXR1, 12 DexraC1, and 12 DexraC10 across all 4 replicates. Data for survival analysis, pup weights, and litter sizes were included for analysis at the intervals for which the dam was present in the trial. Infertility index was conducted on mice that gave birth at each mating round and ovarian weight analysis was conducted at 8 months.

Absorption, Distribution and Excretion
Intravenous administration achieves complete bioavailability. Urinary excretion plays a crucial role in the elimination of dexrazoxane. 42% of a 500 mg/m² dose of dexrazoxane is excreted in the urine.
9 to 22.6 L/m²
7.88 L/h/m² [50 mg/m² doxorubicin and 500 mg/m² dexrazoxane]
6.25 L/h/m² [60 mg/m² doxorubicin and 600 mg/m² dexrazoxane]
Following intravenous administration, the drug rapidly distributes into tissue fluids, with the highest concentrations of the parent drug and its hydrolysates in the liver and kidneys.
At the end of a 15-minute infusion of 500 mg/m² doxorubicin, the mean peak plasma concentration of dexrazoxane was 36.5 mcg/mL. After a rapid distribution phase, dexrazoxane reaches post-distribution equilibrium within 2 to 4 hours.
The estimated steady-state volume of distribution of dexrazoxane indicates that it is primarily distributed in systemic water (25 L/m²).
In vitro studies have shown that dexrazoxane does not bind to plasma proteins.
For more complete data on the absorption, distribution, and excretion of dexrazoxane (9 items), please visit the HSDB record page.

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Metabolites/Metabolites
Dexrazoxane is hydrolyzed in the liver and kidneys by dihydropyrimidine amide hydrolases to active metabolites that bind to metal ions.
Metabolites include the active drug, diacid-diacid cleavage products, and two monoacid-monoamide cyclic products of unknown concentration.
In vitro studies have shown that dexrazoxane is hydrolyzed in the liver and kidneys by dihydropyridinase (DHPase), but not in the heart extract. This study aimed to determine the metabolism of dexrazoxane (ICRF-187) into monocyclic open-ring hydrolysates and bicyclic open-ring metal chelates (ADR-925) in brain metastases treated with high-dose etoposide. In this phase I/II clinical trial, dexrazoxane was used as a rescue agent to reduce the extracranial toxicity of etoposide. High-performance liquid chromatography (HPLC) was used to determine dexrazoxane and its monocyclic open-ring hydrolysates, and fluorescence flow injection assay was used to determine ADR-925. Following dexrazoxane infusion, both monocyclic open-ring hydrolysates appeared in plasma at low concentrations, which then rapidly decreased, with half-lives of 0.6 hours and 2.5 hours, respectively. A plasma ADR-925 concentration of 10 μM was detected after the intravenous infusion of dexrazoxane, indicating rapid metabolism of dexrazoxane in vivo. The plasma concentration of ADR-925 plateaued at 30 μM for 4 hours before slowly decreasing. The pharmacokinetics of dexrazoxane are similar to other reported low-dose data under different conditions. The rapid appearance of ADR-925 in plasma may enable its uptake by cardiac tissue and binding of free iron. These results indicate that the dexrazoxane intermediate is enzymatically metabolized to ADR-925, providing a pharmacodynamic basis for the antioxidant cardioprotective activity of dexrazoxane. Dexrazoxane is hydrolyzed in the liver and kidneys by dihydropyrimidine amide hydrolases to an active metabolite capable of binding to metal ions. Elimination pathway: Urinary excretion plays a significant role in the elimination of dexrazoxane. 42% of a 500 mg/m² dose of dexrazoxane is excreted in the urine. Half-life: 2.5 hours. The distribution half-life is approximately 12 to 60 minutes… Elimination - 2.5 hours.

Toxicity Summary
The mechanism by which dexrazoxane exerts its cardioprotective effect is not fully understood. Dexrazoxane is a cyclic derivative of EDTA that readily penetrates cell membranes. Laboratory studies have shown that dexrazoxane (a prodrug) is converted intracellularly into an open-ring bidentate chelator that chelates free iron and interferes with iron-mediated free radical generation, which is considered to be partly responsible for anthracycline-induced cardiomyopathy. Notably, dexrazoxane may also exert its protective effect through its inhibition of topoisomerase II. Pregnancy and Lactation Effects ◉ Overview of Use During Lactation There is currently no information regarding the use of dexrazoxane during lactation. The manufacturer recommends that women refrain from breastfeeding during treatment and for two weeks after the last dose of dexrazoxane. However, due to the combined use of dexrazoxane and doxorubicin, the withdrawal period may be longer depending on the doxorubicin dose.
◉ 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.

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Protein binding
Very low (< 2%)

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Toxicity data
Humans (intravenous): TDLo: 383 mg/kg
Mice (intraperitoneal): LDLo 800 mg/kg
Dogs (intravenous): LDLo: 2 g/kg
Mice intraperitoneal LD₁₀ = 500 mg/kg. Intravenous, dogs LD₁₀ = 2 g/kg.

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Interaction
In a crossover study of cancer patients, the pharmacokinetics of doxorubicin (50 mg/m²) and its major metabolite doxorubicinol were not significantly altered in the presence of dexorubicin (500 mg/m²).

[1]. Topoisomerase IIbeta mediated DNA double-strand breaks: implications in doxorubicin cardiotoxicity and prevention by dexrazoxane. Cancer Res. 2007 Sep 15;67(18):8839-46.

[2]. Daunorubicin-induced apoptosis in rat cardiac myocytes is inhibited by dexrazoxane. Circ Res. 1999 Feb 19;84(3):257-65.

[3]. Dexrazoxane Diminishes Doxorubicin-Induced Acute Ovarian Damage and Preserves Ovarian Function and Fecundity in Mice. PLoS One . 2015 Nov 6;10(11):e0142588.

Dexrazoxane hydrochloride is the hydrochloride salt of dioxanepiperazine with iron-chelating, chemoprotective, cardioprotective, and antitumor activities. Upon hydrolysis, dexrazoxane is converted to an active form similar to ethylenediaminetetraacetic acid (EDTA), chelating iron and thus limiting the formation of anthracycline-iron complexes that generate free radicals. This may minimize anthracycline-iron complex-mediated oxidative damage to the heart and soft tissues. The drug also inhibits the catalytic activity of topoisomerase II, thereby inhibiting tumor cell growth.
(+)-enantiomers of dexrazoxane.
See also: Dexrazoxane (with active moiety).
Drug Indications

Savene is indicated for the treatment of anthracycline extravasation.
Doxorubicin is one of the most effective and widely used anticancer drugs in clinical practice. However, cardiotoxicity is one of the life-threatening side effects of doxorubicin treatment. Dexrazoxane (Zinecard, also known as ICRF-187) has been used clinically as a cardioprotective agent to counteract doxorubicin cardiotoxicity. However, the molecular mechanisms of doxorubicin cardiotoxicity and the cardioprotective effects of dexazosin remain not fully elucidated. In this study, we found that dexazosin specifically eliminated the doxorubicin-induced DNA damage signal γ-H2AX in H9C2 cardiomyocytes, but had no effect on camptothecin or hydrogen peroxide. The proteasome inhibitors bortezomib and MG132 also specifically eliminated doxorubicin-induced DNA damage, and in top2β(-/-) mouse embryonic fibroblasts (MEFs), DNA damage was significantly reduced compared to TOP2β(+/+) MEFs, suggesting the involvement of the proteasome and DNA topoisomerase IIβ (Top2β). Furthermore, in addition to antagonizing the formation of the Top2 cleavage complex, dexazosin also induced rapid degradation of Top2β, consistent with the reduction in doxorubicin-induced DNA damage. Our results collectively suggest that dexazosin antagonizes doxorubicin-induced DNA damage by interfering with topoisomerase 2β (Top2β), which may indicate that Top2β is involved in the cardiotoxicity of doxorubicin. The specific involvement of the proteasome and Top2β in doxorubicin-induced DNA damage is consistent with the following model: the processing of the doxorubicin-induced Top2β-DNA covalent complex by the proteasome exposes the Top2β-hidden DNA double-strand breaks. [1] The clinical efficacy of anthracycline antitumor drugs is limited by their high incidence of severe and often irreversible cardiotoxicity, the reasons for which remain controversial. In primary cultures of neonatal and adult rat ventricular myocytes, we found that daunorubicin at concentrations ≤1 μmol/L induced programmed cell death in cardiomyocytes within 24 hours, which has been confirmed by a variety of complementary techniques. In contrast, daunorubicin at concentrations ≥10 μmol/L induced necrotic cell death within 24 hours without the characteristic changes of apoptosis observed. To determine whether reactive oxygen species play a role in daunorubicin-mediated apoptosis, we monitored hydrogen peroxide generation using dichlorofluorescein (DCF). However, daunorubicin (1 μmol/L) did not increase the intensity of DCF fluorescence, and the combination of the antioxidants N-acetylcysteine or α-tocopherol with ascorbic acid also failed to prevent apoptosis. Conversely, dexzosen (10 μmol/L), known to reduce the cardiotoxicity of anthracyclines, prevents daunorubicin-induced cardiomyocyte apoptosis but not necrosis induced by higher concentrations of anthracyclines (≥10 μmol/L). The superoxide dismutase mimic porphyrin manganese (II/III) tetrakis(1-methyl-4-pyridyl)porphyrin (50 μmol/L) mimics the anti-apoptotic effect of dexzosen. The understanding that anthracycline-induced cardiomyocyte apoptosis (possibly mediated by superoxide anion generation) occurs at concentrations far below those leading to cardiomyocyte necrosis may help in designing new therapeutic strategies to limit the toxicity of these drugs. [2]

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Dexzosen/Dexra alleviates acute DXR-induced ovarian toxicity and improves the fertility window, manifested by increased fertility, offspring weight, litter size, and number of births after DXR treatment. A 1:1 Dexra:DXR dose protects the ovaries. Easy-to-take dexmedetomidine (Dexra) offers a timely, economical, and safe method of pharmacological ovarian protection, particularly suitable for pre-pubertal and adolescent girls where oocyte and embryo freezing is not a viable fertility preservation option. [3]

C11H16N4O4.XHCL

N/A

304.094

C, 43.36; H, 5.62; Cl, 11.63; N, 18.39; O, 21.00

149003-01-0

1263283-43-7 (HCl); 24584-09-6; 149003-01-0 (HCl)

6918223

White to off-white solid powder

531.5ºC at 760 mmHg

193ºC

275.3ºC

2.22E-11mmHg at 25°C

3

6

3

20

404

1

Cl[H].O=C1C([H])([H])N(C([H])([H])C(N1[H])=O)[C@@]([H])(C([H])([H])[H])C([H])([H])N1C([H])([H])C(N([H])C(C1([H])[H])=O)=O

BIFMNMPSIYHKDN-FJXQXJEOSA-N

InChI=1S/C11H16N4O4.ClH/c1-7(15-5-10(18)13-11(19)6-15)2-14-3-8(16)12-9(17)4-14;/h7H,2-6H2,1H3,(H,12,16,17)(H,13,18,19);1H/t7-;/m0./s1

4-[(2S)-2-(3,5-dioxopiperazin-1-yl)propyl]piperazine-2,6-dione;hydrochloride

ICRF-187 (ADR-529) HCl; (+)-Razoxane hydrochloride; ADR-529 hydrochloride; Cardioxan; Dexrazoxane HCl, Dexrazoxane hydrochloride; 149003-01-0; Dexrazoxane HCl; Totect; Cardioxane; Cardioxan; Savene; Zinecard; Dexrazoxane hydrochloride; ICRF-187 hydrochloride; Savene; ADR529; ADR-529; ADR 529; ICRF-187; ICRF187; ICRF 187; NSC169780; NSC-169780; NSC 169780; Cardioxan; Cardioxane; US brand names: Totect; Zinecard. Foreign brand names: Cardioxane Savene.

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 (e.g. under nitrogen), avoid exposure to moisture and light.

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: ≥ 3 mg/mL (8.79 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 30.0 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: ≥ 3 mg/mL (8.79 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 30.0 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: ≥ 3 mg/mL (8.79 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 30.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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Solubility in Formulation 4: 130 mg/mL (381.02 mM) in PBS (add these co-solvents sequentially from left to right, and one by one), clear solution; with ultrasonication.

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