Preclinical Pharmacology

Preclinical Pharmacology

  • Rajendra Kumar Singh added an answer:
    7
    Which is better to use sub-acute or sub-chronic toxicity to investigate anti-diabetic activity of medicinal plants?

    i would like to use albino rats for this experiment and also try to induce diabetes by using alloxan , but am confused about using which one of sub-acute or sub-chronic toxicity

    Rajendra Kumar Singh

    Sub acute or sub chronic word used according to duration of study. As per the design of study you follow the protocol. Acute, suub acute or sub chronic and chronic study mention treatment duration. 

  • Prabhakaran Selvam added an answer:
    6
    Is curcumin a good candidate for wound healing besides its cytotoxic effect on Keratinocytes and fibroblasts?

    Many studies showed high dose of curcumin induces ROS formation and caspase medicated apoptosis in both fibroblasts and keratinocytes which should in theory make curcumin unfit for the treatment of cutaneous wound healing. The curcumin I have always known was cytoprotective, antioxidant, anti-inflammatory and has potent wound healing ability but these study results prove otherwise.I know Curcumin interacts with multiple pathways and has multiple pharmacological properties, so some how It might pleiotropically promote wound healing besides its cytotoxic effect on these cells.but I am not sure how inhibitory effect of curcumin on fibroblast and keratinocytes proliferation could help with cutaneous wound healing.

    • Source
      [Show abstract] [Hide abstract]
      ABSTRACT: Wound healing is a complex process that involves the well-coordinated interactions of different cell types. Topical application of high doses of curcumin, a plant-derived polyphenol, enhances both normal and diabetic cutaneous wound healing in rodents. For optimal tissue repair interactions between epidermal keratinocytes and dermal fibroblasts are essential. We previously demonstrated that curcumin increased reactive oxygen species (ROS) formation and apoptosis in dermal fibroblasts, which could be prevented by pre-induction of the cytoprotective enzyme heme oxygenase (HO)-1. To better understand the effects of curcumin on wound repair, we now assessed the effects of high doses of curcumin on the survival of HaCaT keratinocytes and the role of the HO system. We exposed HaCaT keratinocytes to curcumin in the presence or absence of the HO-1 inducers heme (FePP) and cobalt protoporphyrin (CoPP). We then assessed cell survival, ROS formation, and caspase activation. Curcumin induced caspase-dependent apoptosis in HaCaT keratinocytes via a ROS-dependent mechanism. Both FePP and CoPP induced HO-1 expression, but only FePP protected against curcumin-induced ROS formation and caspase-mediated apoptosis. In the presence of curcumin, FePP but not CoPP induced the expression of the iron scavenger ferritin. Together, our data show that the induction of ferritin, but not HO, protects HaCaT keratinocytes against cytotoxic doses of curcumin. The differential response of fibroblasts and keratinocytes to high curcumin doses may provide the basis for improving curcumin-based wound healing therapies. Copyright © 2015. Published by Elsevier Inc.
      Full-text · Article · Jun 2015 · Experimental Cell Research
    Prabhakaran Selvam

    Thank you so much Dr.Daniel Lerda.

  • Obinna Uchenna chukwuemeka Adumanya added an answer:
    4
    How can I determine a clinically relevant concentration?

    Hi!

    I am testing a new compound on colorectal cancer cell lines and I want to compare it's efficacy with the standard drug utilized in the treatment, which is the 5-FU. What concentrations should I use? I am using plasmatic concentrations found in blood samples collected from patients in treatment 2-3 hours after the infusion, are those concentrations considered clinically relevant? 

    Thanks in advance.

    Obinna Uchenna chukwuemeka Adumanya

    IC50 determination will be very useful pls.

  • Anne Goubier added an answer:
    2
    Does anyone have an idea for syngeneic tumour models for testing human tumour targeting Ab (e.g. Cetuximab) in immune competent mice ?

    Hi,

    I am looking for syngenic tumor models for evaluating human-specific tumour targeting Ab in immune competent mice. More specifically, I am looking for models expressing either huHER2, huEGFR or huCD20 and which have already shown some therapeutic response to Trastuzumab, Cetuximab or Rituximab, respectively. I have identified potential models (TUBO-EGFR, BL3750), but they are not commercially available and I cannot get access to them. Do you have any suggestion? That would greatly help our research program!

    Anne Goubier

    Dear Beatrice, Thanks a lot, that is very helpful!

    Anne

  • Uldis Berzins asked a question:
    Open
    Preclinical studies: what risks are assessed with lengthening telomeres of autologous stem cells (by 20% or more from baseline)?

    1) Why lengthening telomeres stem cells begin expressing MHC-1 ?

    2) Why lengthening telomeres stem cells stop expressing CXCR4 ?

  • Rafik Karaman added an answer:
    2
    Any advice about long-acting PTH analogue or PTHR modulator?

    Does anyone have any information about new drugs from these classes?

    Rafik Karaman

    Dear Olivera,

    Please read the following text:

    Long-Acting Parathyroid Hormone Analog for the Treatment of Hypoparathyroidism
    Hypoparathyroidism is a rare hormone-deficiency syndrome in which the body lacks parathyroid hormone (PTH). Due to PTH’s central role in maintaining the balance of calcium and phosphate in the blood, symptoms of hypoparathyroidism include muscle cramping, convulsions, intellectual disabilities, cataracts and abnormal heart rhythm. Hypoparathyroidism can occur after an injury to the parathyroid glands or following surgery or radiation treatment for thyroid cancer. It also may occur as a consequence of other rare genetic disorders or toxic exposures.

    Hypoparathyroidism represents one of the few remaining hormone deficiencies for which an approved replacement therapy does not exist. Attempts to replace PTH have shown limited usefulness. Due to a persistent lack of calcium, patients must receive high-dose calcium supplements, which can have negative effects on the kidneys. The goal of this project is to develop a PTH replacement that will demonstrate a more normal, stable level of PTH activity and lessen the need for chronic high-dose calcium supplements.

    Scientific Synopsis
    The primary symptoms of hypoparathyroidism are due to low serum calcium (hypocalcemia). Replacement of PTH has been explored to remedy the calcium deficiency, but maintaining an optimal calcium level has proven problematic because hypercalcemia can occur as a result of excess PTH. Multiple efforts are under way targeting either full-length PTH (PTH 1–84) or the active amino-terminal domain (PTH 1–34), but these molecules have undesirable pharmacokinetic properties for chronic daily management of calcium levels in patients with hypoparathyroidism.

    Eli Lilly scientists have identified a PTH receptor modulator (PTH-RM) that can normalize serum calcium. At fairly low doses, the PTH-RM was shown to normalize calcium levels in parathyroidectomized rats, with a profile similar to basal insulin use in patients with diabetes. The investigators are using TRND support to develop this PTH-RM toward a Phase II proof-of-concept study for hypoparathyroidism by leveraging the existing data package.

    Lead Collaborator
    Lilly Research Laboratories, Eli Lilly & Company(link is external), Indianapolis

    Henry U. Bryant, Ph.D.

    Public Health Impact
    There is no approved hormone replacement therapy for hypoparathyroidism. Insufficient PTH causes a range of symptoms, which must be managed in part through lifelong, high-dose supplements of calcium and vitamin D. Hospital and emergency room visits are common, and the high-dose supplements can cause kidney damage.

    Outcomes
    TRND, the Eunice Kennedy Shriver National Institute of Child Health and Human Development, and Eli Lilly teams are performing a gap analysis to determine the pre-clinical and clinical development plan. Bridging studies are anticipated to support a Phase II proof-of-concept clinical trial. TRND experts will support the preparation and filing of the Investigational New Drug application with the Food and Drug Administration to conduct the proof-of-concept trial in hypoparathyroidism patients.

    https://ncats.nih.gov/trnd/projects/active/parathyroid-hormone-hypoparathyroidism

    Hoping this will be helpful,

    Rafik

  • Mallappa Shalavadi added an answer:
    3
    How can I select oral doses [3 doses--lower, moderate and high doses] of extracts from LD50?

    How can I select oral doses [3 doses--lower, moderate and high doses] of extracts from LD50?

    Mallappa Shalavadi

    Thank you sir

  • Nissar Reshi added an answer:
    5
    Can Silymarin tablet (which I purchased) be used for an in vivo hepatoprotective activity study?

    how to make standard drug? I bought SILYMARIN tablet for invivo hepaptoprotective activity. Can Silymarin tablet be used for this study?

    Nissar Reshi

    Of course you can use as +ve control with your plant extract or synthetic compound as test drug in in vivo as well as in vitro experimant

  • Syed Muhammad Sharib added an answer:
    7
    Computational Drug screening methods must be complemented with in-vivo models to assess the degree of hepatotoxicity. Do you agree?

    In vivo drug screening assays are far superior than computational models that are employed to predict drug-induced toxicity since the former often miss subclinical hepatotoxicity that can be ascertained only from in vivo studies. Hepatotoxicity still accounts for 50% of acute liver failure owing to drug toxicity, and to rely exclusively on QSAR-based techniques or related systems preclude us from determining the actual toxicity of potential hepatotoxins/drugs.

    Cell culture techniques, in vivo studies in rat, mice, and assays should be regularly complemented with computational toxicity studies to determine the actual levels of subclinical toxicity related to Type A and Type B ADRs. Conclusive evidence should be drawn only after bioassays, and not based on computational predictions. This would likely minimize the incidence of hepatotoxicity during drug therapy, reduce the cost associated with hospitalization, and the incidence of overall hospitalization due to ADRs.

    Syed Muhammad Sharib

    Yes there are limitations, i am not denying about that. You have made exact points regarding technique, But my point is we can go with both modeling together. That will lead to cover all the ethical issues. We can first be precised by using In-Silico models. Yes PBPK/PD is valuable in drug discovery. Field researchers and scientists emerging the tool in variety of aspects now a days. Bayer Technology Services have came upon with a great revolution. 

  • Monica Cavali added an answer:
    4
    How much time do we need to give metformin to rats to get diabetic in preclinical trial ?

    how much time we need to give metformin for rats after it get diabetic in clinical trial ?

    Monica Cavali

    Sorry, but I didn't understand your question. Could you explain your study?

  • Denise Priolli added an answer:
    10
    Does anybody has any experience with 786-O(RCC) induced xenograft nude mice models?
    Any suggestions about the optimum cell no. for inoculation as I had inoculated 786-O (5 *10 ^ 6 cells/animal, Subcutaneously in flank region) in CD1 nude mice. After one week I observed tumorous swellings up to 100 mm3 at the site of inoculation and then it disappeared after the 2nd week. What could be the reason?
    Denise Priolli

    The routine for renal adenocarcinoma/786 is the fast growth followed by disappearance of tumor due to NK response in nude mice. In a pilot that I conducted with three animals  I had successonly in one animal, but after re-inoculation of 4X10E6 cells. The literature reports that the xenograft works in BALB/c nu beige or SCID but I have no experience with these strains.

  • Sujit Kashyap added an answer:
    6
    Can or should, a monoclonal antibody for western blot, IHC etc application be used for an animal study?

    I wish to study the effect of the monoclonal antibody against a certain protein. Can i purchase this monoclonal antibody for animal study from the widely known companies that are generally used for western blot, IHC etc? If not then why?

    Sujit Kashyap

    Dear Bret Lum, i was talking about the use of rat monoclonal antibody in the rat model of disease. I guess this will not develop antidrug antibody if so then please explain how.

  • Mohammad Sadegh Amiri added an answer:
    19
    Can a plant with antioxidant effect also increase lipid peroxidation?

    I am investigating the hepatoprotective effect of some medicinal plant on chronic carbon tetrachloride induced liver injury. I observed that although some of the medicinal plants increased the antioxidant enzyme (SOD, CAT and GSH) level in rats co-exposed to CCl4 and the extracts while they also increased lipid peroxidation and markers of hepatic injury such as ALT and AST.Has anyone experienced something similar? If yes, what are the likely reasons why this happened?

    Mohammad Sadegh Amiri

    Dear Ifeoluwa T Oyeyemi

    I am attaching literature on the anticancer properties. I hope they will be useful to you.
    I recommend the fruit of Rhus coriaria L. (Anacardiaceae) for your research.

    Good luck.

  • Alessandra Tiziana Peana added an answer:
    6
    Do you know how to administer bromocriptine s.c. or i.p. without using alcohol?

    Dear all,

    I have to administer bromocriptine (BC) to mice to inhibit prolactin release to analyse the role of this hormone in behaviour. BC has a very low solubility in water. People usually do a stock (saturated) solution of BC in pure ethanol and then disolve it in PBS or water and inject it s.c. Since the final concentration of alcohol might be nearly 10%, BC injections are probably very painful, and this may invalidate the results of the ensuing behavioural analysis.

    I've thought of using DMSO, in which it is much more soluble. Has anybody experience of drug administration in solutions containing relatively high concentrations of DMSO (about 1%)? Could I use i.p. injections, in that case?

    Many thanks

    Fernando

  • Hafiz Iftikhar Hussain asked a question:
    Open
    Which breed of cat is mostly used for Pharmaco-kinetic studies?

    if more than one breeds used please inform.

    Thank you.

  • Marcus Vinicius P.S. Nascimento added an answer:
    7
    Can I use the OECD guideline #423 to test toxicity of a synthetic compound via intraperitoneal route in mice?

    I know this is a guideline for acute oral toxicity, but I can't find any current guideline to test synthetic compounds via the intraperitoneal route in vivo (mice). OECD 423 seems to be the protocol that uses less animals. Does anyone know if I can use it as described in the protocol changing only the administration route (Oral to Intraperitoneal)?

    guideline 423: http://ntp.niehs.nih.gov/iccvam/suppdocs/feddocs/oecd/oecd_gl423.pdf

    Marcus Vinicius P.S. Nascimento

    So OECD 401 can be used with intraperitoneal route but 423 can't? I thought OECD 423 would be better accepted because it uses less animals than 401. Don't you think that this might influence later publications (the use of a unnecessary number of animals)?

  • Christopher Jobdevairakkam added an answer:
    3
    Do you think that infusion pumps in preclinical studies can add variability to your results?

    There is a recent study by John Osborn that demonstrate that if you use an alzet pump for delivering angiotensin you get a lot variability in your blood pressure readouts as compared to new technology pump such I-precio or tethered.

    What is your opinion on that?

    Christopher Jobdevairakkam

    Usually all the infusion pumps are periodically calibrated and the variation can be less than 2%. Having said that, for a given application  if the infusion time is less than, say 3 hrs, the variation can be +/- 3.6 min. that is not a significant variation. However if the infusion time is more than 12 hrs, hope you can imagine the large variation that could lead a significant difference in dosage administration.

  • Stephen Haber added an answer:
    2
    Is NMS-P626 the same as RXDX-101 and NMS-e628?

    The drug name is entrectinib. I am looking for pre-clinical studies and I can not figure out if they are the same drug.

    Stephen Haber

    They are the same.

    See https://clinicaltrials.gov/ct2/show/NCT02097810

           http://pubchem.ncbi.nlm.nih.gov/compound/25141092#section=Top

          and

          http://www.medkoo.com/Anticancer-trials/NMS-E628.htm

  • Bharath Srinivasan added an answer:
    9
    Which terminology is suitable for this situation below?- 'drug target' is suitable?

    We have found a gene of which expression are known to play a crucial role in human disease. in our experiments, the levels of mRNA and proteins of the gene are reduced in a drug treatment. However, we did not have know whether the drug interact with the protein directly. Seemingly, the drug might be involved only in the gene expressional regulation with no direct interaction with the protein product.

    In this case, can we term the gene as the drug target? or etc..?

    (ex. identification of 'gene name' as a drug target of 'drug name')

    What is the most suitable term for the description in this situation?

    Reference information is also very appreciated.

    Bharath Srinivasan

    Dear Dong-Myung Kim,

        Your question is interesting. However, caution may need to be exercised in calling the gene a "drug target" of the small molecule. It is mandatory to demonstrate that  other genes do not show any effect vis-a-vis their expression and protein levels for you to specifically claim that the small-molecule targets the gene of interest. Lot of small molecules have been shown to have a global effect on the downregulation or upregulation of several different genes by targeting RNA polymereases, repressors and activators and transcription factors. I hope the answer helps.

  • Sandeep Rajput added an answer:
    1
    What are the protocols needed to manufacture injectable VEGF-C without a hydrogel delivery vehicle?

    I'm looking at injecting VEGF-C into a rat model, specifically into the medial right thigh tissue. I'm going to do this after removing a lymph node.

    From the literature i have seen so far, most researcher's have combined VEGF-C with a hydrogel (HAMC [Baker et al., 2010] or Gelatin [Hwang et al., 2011]) before injecting into the lymph node region.

    Can VEGF-C brought stragiht from a manufacture (eg R &D - Recombinant Human VEGF‑C) be injetcted into rat tissue without further modification? if modification is needed, can VEGF-C be dissolved in PBS prior to injection?

    NB: Dont wont want to use Adenovirus VEGF-C.

    I'm asking this question because time is short to prepeare a injectable delivery vehicle for VEGF-C. However i'am open to suggestions which are simple and can be manufactured in a hour.

    The volume of VEGF-C i want to deliver are similar to the values mentioned by [Baker et al., 2010] and [Hwang et al., 2011

    Thank you.

    Sandeep Rajput

    The only reason they used Hydrogel is to increase the solubility or to localized the VEGFC into tissue only. If VEGFC is soluble in PBS , u can inject it directly.

  • Muzammil Najmi added an answer:
    3
    How can I make a good use of leftover human specimens?

    Recently, I may have a chance to receive a large batch of leftover specimens. Besides using these leftover specimens for research, I am also thinking about the possibility of turning these specimens into some products which can generate funding for future academic research development of my lab. Did anyone have experience in similar scenario or know any institute which is doing something similar? Thank you.

    Muzammil Najmi

    The human specimens should not be used for purposes other than the ones for which permission/consent was obtained from the subjects at the time of collection. Particularly, use for commercial purposes (raising funds, as it has been mentioned) will be attended with huge ethical concerns. These may however, be used to further extend the research for betterment of humanity.

  • Adelaeda Barrera added an answer:
    7
    Has anybody worked on SKOV-3 cell lines induced xenograft mouse model using CD1 NUDE mice?
    Can anybody suggest about cell number/animal as well as time consumption to grow tumors up to palpable limit?
    Adelaeda Barrera

    Hello, by any chance does anyone know if SKOV-3 cell lines can produce ascites when injected to nu/nu mice?

  • Ifeoma Obidike Ezenyi added an answer:
    15
    How can I reduce mortality in STZ induced type II diabetes in wistar rats?

    Dose injected - STZ 60mg/kg I.P

    Strain: wistar albino female rats

    Bwt-120 to 150g

    have given 5%glucose 2hr after STZ induction for 24hours

                                                    observations

    after 3 days- diabetic rats - 19 out of 30

    after 7 days-

    mortality observed -12 out of 19 ie. 7 rats alive

    Ifeoma Obidike Ezenyi

    Also increase sucrose concentration to 10%w/v

  • Stanton de Riel added an answer:
    2
    How to lower the pH of a furosemide buffer?

    Hi All,

    I just prepare 50 mg/ml furosemide in NaOH, then try to adjust pH with HCL. We hope the pH for this buffer is around 7, well we also accept 8 or 9.  however, when pH drop a little bit lower than 10, many white furosemide was coming out and didn't dissove any more. and pH 10 is too high for our animal study.  Is there any buffer I can try for prepare furosemide and let it pH become lower?

    Thanks

    Yiyun

    Stanton de Riel

    Looks like the furosemide molecule has several sites that can protonate/deprotonate -- your problem is likely to be that at pH 7 it's (functionally) neutral, hence not very soluble (the carboxylate will be deprotonated, but the secondary amine will be protonated, hence net neutral). You might try chelating it with a cyclodextrin. You can't cheat nature on pH issues, really. Depending on the administration route, could you use a co-solvent to increase solubility, perhaps, or if slower release is acceptable, load it as a suspension (by gavage, or subcutaneous depot injection)? 

  • Lana Krestetska added an answer:
    3
    Is there any knowledge in the litterature that Salmonella Typhimurium might be responsible for dizziness?

    I'm working with Salmonella enterice serovar Typhimurium and when I orally infect mice, I observe 10% of mice having strong dizziness. This bacteria is known to infect systemic tissues such as mesenteric lymph nodes, peyer's patches, spleen and liver but what about the dizziness phenotype? Is Salmonella able to infect the internal ear? Or is there any other explanation for this?
    Many thanks in advance for your help!
    Best,
    Thibault

    Lana Krestetska

    exaggerated lean to one side of the body and longitudinal spinning and rotary motion? http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1925087/

  • Imtiyaz Rather added an answer:
    3
    Is anyone doing research on coumarin, or 7-hydroxycoumarin in the management of lymphedema?

    Earlier research on coumarin showed very promising results, but some of the in vivo studies using rodents showed toxicity, which was discovered to be a metabolic pathway that is not usual in humans. But the compound was taken off the market in a number of countries, which is a real shame since it is one of the few and only pharmacological approaches that has shown success in reducing protein accumulation. If you are doing research in this area, please contact me.

    Thanks, 

    Elaine Weil, NP

    Imtiyaz Rather

    Coumarin and 7-hydroxycoumarin acts as anti coagulant and can definitely be a choice in treating lymphedema.

  • Adam L Vanwert added an answer:
    11
    Any tips/alternatives for mouse tail vein AAV injections?

    I've been attempting some mouse tail vein injections and seem to have trouble with consistency. Ultimately I would like to deliver viral vectors systemically.

    Any tips for mouse restraint? (They of course jump from the stick and I lose my vessel.)

    Are there simpler alternative sites for IV injection or other modes of systemic delivery that might serve my purpose?

    Are there any syringe/needle sizes that seem to work better?

    Adam L Vanwert

    Please keep in mind that intraperitoneally injected substances will go to the liver via the portal vein prior to the systemic circulation.  This may significantly reduce the systemic availability.  I would continue trying the tail vein.  It's just a matter of practice until you get good or at least good enough.

    Ryan, I think IP injection for infecting enteric nerves makes some sense.  Remember, the enteric nervous system is covered by cell layers.  The myenteric plexus is covered by the longitudinal muscle, and the submucosal plexus is covered by the longitudinal and circular muscle.

    Ryan, you might want to consider that the capillaries in the enteric wall are buried under the muscle layers.  Therefore, it might be more efffective to get the virus in a proximal artery (right before the mesenteric vessels) so it goes into the intestinal wall in a high concentration.  It may then diffuse out of the capillaries.  I actually have no idea how large the virus particles are.  Perhaps they don't cross capillary pores at all.

  • Lasse Giil added an answer:
    2
    How can I get functional analyses of antibodies to GPCR for a good price?

    Dear all.

    We are studying the pathophysiology og autoantibodies to GPCR that we induce in animal models. In addition to looking at the histological endpoints, we would like to purify the IgG from the animals and get dose-response curves for individual animals and compare, and if possible by price, charcterize their functionality (and not just titers we get) further. I have contacted several drug-development companies. They will do it, but at a steep price. Are there any academic GPCR service lab. that have analytical services? I have not  been able to find one. We got nothing in-house and the time and sallary spendt developing will be much more then the drug-developing companies charge. Thanks for potential response.

    Lasse Giil

    Thank you Markus!

    I would like to look at activation of the second messenger, or any other version of confirmation of GPCR activation and ability to create a dose-response curve (some use arrestin, some use IC Calcium, (euroscreen/discoverX/eurofits). We want to look at anti-AT1R and anti-beta1.

    Would love to use the human version, but we only have biobank material (With longitudinal data), so we dont want to spend alot of the serum, as its used by many other researchers.

    We were going to use polyclonal antibodies, because we are looking at their ability to cause pathology and polyclonals more "Natural", even though raising them in Rabbits is far from ideal. The peptide immunization protocol is well known and has been used in several other studies. We want to get "dose-response" curves, as part of mapping their pathophysiological pontential.

    Purification of the human forms will be a next step, but because of time/funding, we cannot do that now.

  • Geert C. Mudde added an answer:
    2
    Does anybody have experience with the use of gemcitabine (Gemzar or a generic product) in non human primates? What would be the correct dose?

    We are planning a chemo+vaccination combination study in NHP to mimic potential use in pancreatic cancer patients. The human clinical dose of gemcitabin is 1000mg/m2.  

    Geert C. Mudde

    Txs a lot!

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