ACE/angiotensin-converting enzyme

Rescinnamine as a first lead in inducing MSH2-dependent cell death [1]
It has previously been shown that reserpine and rescinnamine are capable of inducing cell death in an MSH2-dependent manner (Vasilyeva 2009, 2010). Further experiments demonstrated that this indole alkaloid is capable of overcoming resistance to cisplatin in ovarian cancer cells (Fig. 1).

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Effects of rescinnamine analogs on cell viability [1]
We next tested these new rescinnamine analogs in a well-defined cellular system with an endometrial cell line deficient (HEC59) and proficient (via chromosome transfer, HEC59 + chr.2) for MSH2. This cell system allows to determine whether our new analogs hit their target and, generally, induce cell death. The assays identified a few compounds that induced cell killing in the micromolar range (Fig. 6 and Table 1, compounds 1, 6, 7, 13, 15 and to a lesser extent 12).

These initial results suggested that computational modeling of docking to a specific target protein can identify lead compounds that have specific characteristics. In our case, rescinnamine was identified to be a good lead toward the development of an effective cancer drug that targets a very specific, pro-apoptotic mechanism that is more prevalent in cancer cells. When applied to a xenograft model, however, the hypotensive activity of the drug prevented administration of statistically effective doses, though a significant trend toward tumor inhibition is observed (Fig. 2). Since none of the commercially available reserpine analogs showed much promise in inducing MSH2-dependent cell death (Vasilyeva et al., 2010), we engaged computational modeling and chemical synthesis to generate novel rescinnamine analogs [2].

Cell Biology [1]
HEC59 cells (msh2 deficient) and the paired cell line HEC59 chr. 2 that restores the MSH2-deficiency via chromosome transfer have been extensively characterized (Umar et al., 1997). Cells were grown in standard growth media (DMEM-F12 + 10% FBS). Cells were plated in microtiter plates at an appropriate concentration in 100 µl media and incubated overnight. Media was replaced with media containing drug and allowed to incubate for 24 hours at indicated concentrations. Untreated cells received fresh media with vehicle only. One solution reagent (CellTiter 96)(r) Aqueous One Solution) was added to existing media (20 µl/well) and allowed to incubate 3–4 hrs. A plate reader was used to record the absorbance at 490 nm. Assays were performed at least in triplicates. Cell viability at each concentration was analyzed for IC50 values using GraphPad Prism 4™. Graphs represent mean values and standard deviations.

Xenograft [1]
SW416 or HEC59 cells in PBS mixed with Matrigel (1:1; BD Biosciences) were subcutaneously injected into the flank of nude donor mice. Tumors were grown for up to 3 weeks. Mice were euthanized, tumors excised, minced into 3 mm pieces, and surgically implanted into the right flank of acceptor mice (10 per group). Isoflurane anesthesia was provided during tumor inoculation. Injection of compounds was started 3 days following tumor implantation to allow recovery from surgery. Compounds were given intraperitoneally, in a volume of 0.5 ml/mouse with a ¼ inch, 23-gauge needle in a 6 ml plastic syringe. Mice were monitored based on survival and body weight. Tumor size, measured by caliper, and body weight were monitored daily for 57 days, and the prolongation of median survival time after ip treatment determined. Any animals showing signs of distress, unnatural movements, severe loss of appetite, severe signs of hypotension, tumor size of 1000g, or weight loss exceeding 10% before the end of the study were euthanized. Tumors were measured twice a week for each group. Tumor volume is calculated as length (mm) x width (mm)2. Initial measurements were performed when the tumor reached 150–200 mg. Tumor weight (in mg) is calculated as tumor weight (mg) - (length(mm) of tumor x width (mm) of tumor2)/2.

Absorption, Distribution and Excretion
RAUWOLFIA ALKALOIDS ARE ABSORBED READILY FROM GI TRACT & FROM PARENTERAL SITES OF INJECTION.
SEE RAUWOLFIA ALKALOIDS. ...PASS PLACENTAL BARRIER & MAY AFFECT NEWBORN. /RAUWOLFIA ALKALOIDS/

human TDLo oral 4 ug/kg/D SENSE ORGANS AND SPECIAL SENSES: OTHER CHANGES: OLFACTION; BEHAVIORAL: SOMNOLENCE (GENERAL DEPRESSED ACTIVITY); BEHAVIORAL: ANTIPSYCHOTIC Toxicology of Drugs and Chemicals, Deichmann, W.B., New York, Academic Press, Inc., 1969, -(517), 1969
rat LD50 oral 1 gm/kg Drugs in Japan, 6(898), 1982
rat LD50 intraperitoneal 250 mg/kg Drugs in Japan, 6(898), 1982
rat LD50 subcutaneous 540 mg/kg Drugs in Japan, 6(898), 1982
mouse LD50 oral 1420 mg/kg Psychotropic Drugs and Related Compounds, 2nd ed., Usdin, E., and D.H. Efron, Washington, DC, 1972, -(109), 1972

[1]. Computational and synthetic studies towards improving rescinnamine as an inducer of MSH2-dependent apoptosis in cancer treatment. Mol Cancer Biol. 2013;1(1):44.

[2]. Pharmacological studies with rescinnamine, a new alkaloid isolated from Rauwolfia serpentina. Proc Soc Exp Biol Med. 1954 May;86(1):120-4.

Rescinnamine is an odorless white to cream colored crystalline powder. (NTP, 1992)
Rescinnamine is a methyl ester, an organic heteropentacyclic compound and an indole alkaloid. It has a role as an antihypertensive agent. It derives from a hydride of a yohimban.
Rescinnamine is an angiotensin-converting enzyme inhibitor used as an antihypertensive drug. It is an alkaloid obtained from Rauwolfia serpentina and other species of Rauwolfia.
Rescinnamine has been reported in Rauvolfia, Rauvolfia serpentina, and other organisms with data available.

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Drug Indication
For the treatment of hypertension.

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Mechanism of Action
Rescinnamine Binds to and inhibits the angiotensin converting enzyme. Rescinnamine competes with angiotensin I for binding at the angiotensin-converting enzyme, blocking the conversion of angiotensin I to angiotensin II. Inhibition of ACE results in decreased plasma angiotensin II. As angiotensin II is a vasoconstrictor and a negative-feedback mediator for renin activity, lower concentrations result in a decrease in blood pressure and stimulation of baroreceptor reflex mechanisms, which leads to decreased vasopressor activity and to decreased aldosterone secretion.
SEE RAUWOLFIA ALKALOIDS. ...PRODUCE VARIABLE LOWERING OF MEAN ARTERIAL BLOOD PRESSURE...BY BLOCKING AFFERENT IMPULSES WHICH...STIMULATE SYMPATHETIC VASOPRESSOR REFLEXES NEAR LEVEL OF HYPOTHALAMUS &/OR BY DECR REACTIVITY OF PERIPHERAL SYMPATHETIC...SYSTEM THROUGH DEPLETION OF NOREPINEPHRINE @ NERVE ENDINGS. /RAUWOLFIA.../
SEE RAUWOLFIA ALKALOIDS. ALSO EXHIBIT A NON-HYPNOTIC SEDATIVE EFFECT WHICH MAY BE ASSOCIATED WITH THEIR ABILITY TO MODIFY CONCN OF SEROTONIN &/OR NOREPINEPHRINE IN BRAIN STEM. /RAUWOLFIA ALKALOIDS/

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Therapeutic Uses
ORAL ADMIN...OF LITTLE VALUE IN MGMNT OF SEVERE HYPERTENSION & THAT ASSOCIATED WITH TOXEMIA OF PREGNANCY EXCEPT IN CONJUNCTION WITH THIAZIDE COMPD OR MORE POTENT ANTIHYPERTENSIVE DRUGS... /RAUWOLFIA ALKALOIDS/
ANTIHYPERTENSIVE /SRP: LARGELY REPLACED BY MORE EFFECTIVE THERAPY/
TRANQUILIZER /SRP: LARGELY REPLACED BY MORE EFFECTIVE THERAPY/

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Drug Warnings
PT WITH ALLERGIC DISORDERS MAY SUFFER EXACERBATION OF SYMPTOMS. /RAUWOLFIA ALKALOIDS/
HYPOTENSIVE & SEDATIVE EFFECTS ARE ACCOMPANIED BY BRADYCARDIA, BUT CARDIAC OUTPUT & RENAL PLASMA FLOW ARE NOT MARKEDLY AFFECTED. /RAUWOLFIA ALKALOIDS/
EXCEPT THAT SEDATION & BRADYCARDIA OCCUR LESS FREQUENTLY & IN MILDER FORM WITH RESCINNAMINE, INCIDENCE OF OTHER SIDE EFFECTS SUCH AS WEAKNESS & FATIGUE, NASAL CONGESTION, DIZZINESS, CONFUSION, INCR APPETITE, & WT GAIN IS ABOUT THE SAME AS WITH RESERPINE.
UNLESS DOSAGE IS CAREFULLY ADJUSTED, DRUG MAY INDUCE A PARADOXICAL FORM OF ANXIETY & ADVERSE REACTIVE DEPRESSION. ... SUICIDAL DEPRESSION IS MOST SERIOUS UNTOWARD EFFECT. ...SHOULD BE USED CAUTIOUSLY IN PT WITH HISTORY OF PEPTIC ULCER. ... CONTRAINDICATED IN ECLAMPSIA & IN NURSING MOTHERS... /RESERPINE/
For more Drug Warnings (Complete) data for RESCINNAMINE (11 total), please visit the HSDB record page.

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Pharmacodynamics
Used to treat hypertension. Rescinnamine inhibits angiotensin-converting enzyme. ACE is a peptidyl dipeptidase that catalyzes the conversion of angiotensin I to the vasoconstrictor substance, angiotensin II. Angiotensin II also stimulates aldosterone secretion by the adrenal cortex and general vasoconstriction, both of which lead to increases vascular resistance. By inhibiting angiotensin II, aldosterone reabsorption is decreased as well as vasoconstriction. This combined effect serves to decrease blood pressure.

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We, and others, have previously shown that mismatch repair proteins, in addition to their repair function, contribute to cell death initiation. In response to some drugs, this cell death activity is independent of the repair function of the proteins. Rescinnamine, a derivative of the indole alkaloid reserpine, a drug used to treat hypertension several decades ago, was shown to target the cell death-initiating activity of mismatch repair proteins. When used in animals, the hypotensive action of this drug prevents applying appropriate concentrations for statistically significant tumor reduction. Using a combination of computational modeling, chemical synthesis and cell assays, we determine how rescinnamine can be structurally modified and what effect these modifications have on cell survival. These results inform further computational modeling to suggest new synthetic lead molecules to move toward further biological testing.[1]

C35H42N2O9

634.726

634.289

C, 66.23; H, 6.67; N, 4.41; O, 22.69

24815-24-5

5280954

Fine needles from benzene
WHITE, OR PALE BUFF TO CREAM-COLORED CRYSTALLINE POWDER

1.31g/cm3

756.8ºC at 760 mmHg

238ºC

411.5ºC

8.5E-23mmHg at 25°C

1.621

4.508

1

10

11

46

1080

6

COC1=CC2=C(C=C1)C3=C([C@H]4C[C@H]5[C@H](C[C@H]([C@@H]([C@H]5C(=O)OC)OC)OC(=O)/C=C/C6=CC(=C(C(=C6)OC)OC)OC)CN4CC3)N2

SZLZWPPUNLXJEA-QEGASFHISA-N

InChI=1S/C35H42N2O9/c1-40-21-8-9-22-23-11-12-37-18-20-15-29(46-30(38)10-7-19-13-27(41-2)33(43-4)28(14-19)42-3)34(44-5)31(35(39)45-6)24(20)17-26(37)32(23)36-25(22)16-21/h7-10,13-14,16,20,24,26,29,31,34,36H,11-12,15,17-18H2,1-6H3/b10-7+/t20-,24+,26-,29-,31+,34+/m1/s1

methyl (1R,15S,17R,18R,19S,20S)-6,18-dimethoxy-17-[(E)-3-(3,4,5-trimethoxyphenyl)prop-2-enoyl]oxy-1,3,11,12,14,15,16,17,18,19,20,21-dodecahydroyohimban-19-carboxylate

Anaprel; Apoterin; Apoterin S; Rescinnamine; CCRIS-4711; Cartric; CCRIS 4711; CCRIS4711; Cinnaloid; Cinnasil; Moderil; Rescisan; Resealoid; Reserpinine; Scinnamina

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.)