Midterm 2 Flashcards
What is the best safeguard for cancer?
Apoptosis
What does BAX do?
It induces apoptosis through the release of cytochrome C from the mitochondria.
Can apotosis be induced by a receptor?
Yes, ligands can bind to certain receptors that signal for caspase activation.
What is the paradox of current cancer therapies?
Cancer is caused by DNA damage and cancer therapies induce DNA damage to kill cancer cells (it also attacks healthy cells).
What are cancer therapies that target DNA/induce apoptosis?
- Topoisomerase II inhibitor
- DNA replication inhibitor
- Radiation therapy
What does tamoxifen do?
Estrogen receptor blocker.
What is a downside to tamoxifen?
Cancer cells tend to develop resistance to these drugs.
What does taxol do?
A natural compound that stabilizes tubulin.
What is a disadvantage of taxol?
Recently shown to be genotoxic; very toxic to immune cells.
What compound was extracted from Hawaiian Spider Lily?
Pancratistatin
What has pancratistatin been shown to do?
- induces apoptosis in many human cancer cell lines
- is non-toxic to normal cell lines
- does not target DNA
What category in health does pancratistatin fall under?
Natural products
What do most chemo regiments target?
Geno-toxic compounds or intermediary filaments.
How do animal and bacterial cells differ?
Cell wall vs cell membrane and different translation and transcriptional components.
What is the difference between normal cell and cancer cell?
Cancer cells are smaller because they are constantly dividing.
What are possible targets for selective elimination of cancer cells?
- Vulnerability of mitochondria
- Interfering with DNA replication and repair
- Tubulin stabilization
What are selective targeting strategies (new ones)?
- Mitochondrial vulnerability
- Metabolic vulnerability
- Oxidative vulnerability
How would selective targeting of cancer cells be performed?
- natural compounds and synthetic analogues
- natural extracts
- combination of chemo and natural extracts
How are mitochondria more vulnerable in cancer cells?
- Increased in oxidative stress
- More mtDNA mutations
- Hyperpolarization of organelle (more H+ outside; more acidic in cytosol)
What is the metabolic vulnerability of cancer cells?
Increased glycolysis in the cell resulting in 10x more sugar consumption.
How does oxidative vulnerability appear in cancer cells?
Increased.
How is oxidative vulnerability targeted in cancer cells?
Since they already have increased amounts, therapies increase it higher, which trip the balance to cell death. Normal cells are able to withstand the small increase.
What NP exists in tumeric for cancer therapy?
Curcumin
What natural health products can be used for cancer therapies?
Dandelion root, hibiscus, long pepper, white tea.
What is white tea used for cancer instead or black or green tea?
The leaves aren’t processed at all, just dried at room temp, so all compounds are present in the tea.
What are procedures for identifying chemotherapies and NHPs?
Identify mechanism of action > test efficacy in animals > test efficacy in cells
What is the treatment of in cell culture models (cancer)?
- normal cells can be obtained for healthy volunteers, especially blood
- normal and cancer cells are treated with anti-cancer natural; extracts or drugs
What are the in cell culture models monitored or assayed for?
- Cell growth or cell viability (cancer cell should decrease)
- Induction of apoptosis using apoptosis related markers (re-activated in cancer)
If apoptosis is happening in the cancer cell culture models, what are the possible mechanisms of action?
- Caspase activation
- Oxidative stress induction
- Mitochondrial potential decrease
What is an ex-vivo model?
When cancer cells from a patient are extracted and tested in a lab (done before chemotherapy; blood). Nucleated cells are obtained and put in culture plates; NHP or drugs applied to see effectiveness in death. Normal human blood tested for selectivity too.
What are in-vivo models?
Human cancer cells are transplanted under the skin (sub-cutaneous) of immunocompromised mice (nude; so they don’t reject transplants). Known as xenografts. Animals fed with natural extracts or injected with anticancer drugs, and the size of the tumor is monitored over 4-8 weeks to see if the growth has stopped.
Why are the weights of in-vivo models recorded regularly?
Related to the toxicity of drugs; any changes in weight (either gain or loss) can indicate toxicity.
Transgenic model of cancer
When the genotype of mice are genetically engineered to mimic cancer. They can understand how cancer develops. Knock out a certain gene to see if cancer develops (automatically develop cancer).
Patient-derived xenograft model
Same as the in-vivo models; however, the actual cancer cells from patients are taken and xenografted in immunocompromised mice.
What is the purpose of patient-derived xenograft models?
Provide efficacy of treatment on actual tumor of the patients and can indicate appropriate treatment for a given patient (personal medicine).
What is the final model of cancer for treatment?
Clinical trials:
- therapeutic
- supplementary clinical trial with anti-cancer NHP
What was the first case of targeting cancer cells selectively?
In 2003, liver cancer cells treated with PST were shown to selectively target mitochondria.
What were the cells with PST treated with?
Spider lily compound
How did the cells with PST die?
Committed suicide via apoptosis.
What did the dandelion tea exhibit in the leukemia patient?
50 decrease in the cancer cells.
What kingdom do dandelions belong to?
Taraxacum spp.
What did the taraxacum leaf initially treat?
A potent diuretic.
What did the taraxacum root initially treat?
Treat inflammation and congestion of the liver and gall bladder.
What did Pandey do with the dandelion roots?
They collected them from the ground, and perform a water extraction to pull out the cells.
What were the findings of the hot-water extract of the dandelion root?
Cancer (leukemia) cells: began to die through apoptosis shown by condensed nuclei.
Non-cancer cells: nothing happened, which exhibited the extreme selectivity for cancer cells.
What did the dandelion root extract show in human pancreatic cancer cells?
Cancer cell viability decreased significantly. They responded in a dose dependent manner.
Why did Pandey not use the dandelion root from the ground for future experiments?
Soils could exhibit and influence different activities of compounds in the root. They needed reproducible results.
What does the water-soluble solution of dandelion root consist of?
Sugar and phospholipids. Make up a great weight in the concentration.
What happened when you used a water extract of dandelion root?
When you extract the root with water, you pull out loads of salt and sugar. 95% of the powder was nothing but sugar and salt.
What did dandelion root extract show in human chronic myelomonocytic leukemia cells?
DRE induces apoptosis and reduces the viability of various myelomonocytic leukemia cells in a dose and time dependent manner:
- increased apoptosis with dosage
- decreased viability with time
What stages are p53 positive cancers?
Not very advanced colon cancer as p53 is still active.
What stages are p53 negative cancers?
Very aggressive stage 4 colon cancer. HT-29.
What did DRE show in colon cancer cells?
It induced apoptosis and the viability of colon cancer cells in a dose and time dependent manner.
When can you collect blood samples from leukemia patients?
Cannot collect the sample if already undergoing chemotherapy. The sample is collected before they start so no medications are found in their blood samples. Only 5 mL is taken.
What did DRE show in patient-derived leukemia samples?
Compared to healthy controls, DRE induced apoptosis in cancer cells in a dose dependent manner.
What was the importance of the result of DRE in patient-derived leukemia samples?
DRE could induce apoptosis in both commercial cell lines and ex vivo samples of leukemia.
What must you look at to determine whether or not DRE affects normal cells?
Look at the cell division and proliferation.
Why does apoptosis occur in a healthy person?
Healthy samples have 5-10% of apoptotic cells because blood cells have a limited lifetime.
Does the DRE extract affect the viability of healthy cells?
All doses did not affect growth.
Does the DRE hot water extract affect apoptosis of healthy cells?
No, it did not even as the dose increased. This meant that DRE selectively induces apoptosis in cancer cells only.
What molecule is exhibited in late stage apoptosis?
Caspase 3
How may DRE activate extrinsic apoptosis?
By activating caspase enzymes involved in apoptosis, specifically in cancer cells.
What did DRE induce in blood cells?
In leukaemia, DRE induced caspase 8 immediately (15 minutes), and this activity kept growing.
Do we know what compound in DRE induces apoptosis?
No, this is because we are using an extract not an isolated compound.
What does the fas-associated death domain do in cancer cells?
The receptor is there, but the adaptor is defective. This means there is no receptor-mediated cell death, so it inhibits extrinsic apoptosis in cancer cells.
What does DRE require for apoptosis of cancer cells?
The fas-associated death domain.
Is p53 mutated in colon cancer HCT-116?
No the anti-oncogene is not mutated.
Is p53 mutated in colon cancer HT-29?
Yes it’s negative in this aggressive colon cancer.
How did HCT-116 and HT-29 respond to DRE?
Dose dependent and time-dependent, thus killing more cancer cells as time and dose increased.
Why might a combination of chemotherapy drugs for advanced colon cancer be bad?
Advanced colon cancer becomes almost resistant to these treatments and they are highly toxic.
What are the chemotherapy drugs used for advanced colon cancer?
Folinic acid, oxliplatin, and fluorouracil.
How did the DRE affect the viability of NCM-460 cells?
They did not affect the viability, which is good because these are normal colon mucosal cells.
What did the chemotherapy drugs do to the NCM-460 cells?
Decreased the viability of them, which isn’t good because they are killing normal colon mucosal cells.
Do many people die from colon cancer? What instead?
No they do not, instead they die from the side effects of the chemotherapy drugs which compromise their immune system.
What did the wound-healing assay of DRE on NCM-460 cells show?
No effect, these means the cells grew and healed this gap.
What did the wound-healing assay of DRE on colon cancer cells?
The gap kept increasing in size, which showed that DRE induced apoptosis in cancer cells and was able to stop their migration and division.
Describe the mice DRE/water experiment.
Healthy mice were either given water (control) or DRE in water (treatment). Their weight gain was monitored over time.
What was the result of the mice DRE/water experiment? Meaning?
No difference in weight gain from either drinking water or DRE/water. This meant that DRE was non-toxic and well tolerated as they gained weight the same way.
What did urinalysis of the mice DRE/water experiment show?
Those with the dandelion root extract had less albumin proteins in their urine, which means non-toxic.
In mice with HT-29 and HCT116 tumors, what did DRE with food do to the tumor size (colon and melanoma)?
Tumor size did not increase and lower doses of DRE were even effective.
How did the DRE compound reach the tumor in the mice?
The active compounds were absorbed by intestine, circulated in blood, went to tumor site, then excreted anti-cancer effect on the xenographed tumour by inhibiting apoptosis (proof of principle).
How does JC-1 look in healthy mitochondria?
If the mitochondria is healthy and maintains proper potential, the fluorescent molecules get into mitochondria, they oligomerize and start giving red fluorescence as they accumulate.
How does JC-1 look in unhealthy mitochondria?
If anything has caused mitochondrial opening like apoptosis, the fluorescent molecules escape from the mitochondria and you do not get the fluorescent.
What happens to the mitochondria fluorescent when DRE given to cancerous colon cells?
Mitochondria collapse due to little to none fluorescent, this meant that there was a compound in DRE that would selectively target the mitochondria of cancer cells.
What happens to the mitochondria fluorescent when DRE given to normal colon cells?
No effect on mitochondrial potential as the fluorescence is still red.
What is the mechanism of action for DRE and mitochondria?
A simple crude DRE has compounds that selectively depolarize mitochondria in cancer cells.
What happened to the gene expression of anti-apoptotic genes of colorectal cancer cells, and healthy colon epithelial cells treated with DRE?
Cancer = down regulation, so cell death can occur
Healthy = increase in expression
ex. Bcl-2
What happened to the gene expression of pro-apoptotic genes of colorectal cancer cells, and healthy colon epithelial cells treated with DRE?
Cancer = elevated, trigger cell death
Healthy = decrease in expression
ex. caspase 1, TNF
What compounds are found in DRE?
alpha-amyrin, beta-amyrin, lupeol, and taraxasterol.
How much of the following compounds (alpha-amyrin, beta-amyrin, lupeol, and taraxasterol) are found in DRE?
Concentrations of all these compounds in the crude extract were in nanometer amounts (micromolar of individual).
Why is a low concentration of the compounds in DRE more efficient that the purified compounds?
Indicate a syngeristic effect.
How to prevent toxicity in a crude extract?
Make its antidote in the same extract.
What did the case study of the 70 year old man with acute myeloid luekemia and DRE show?
The man was only given 2 months to live, and felt better within 48 hours. His leukemia count went down by 10% in two weeks and be became leukemia free. A sustained remission of over 4 years.
Why boil DRE for 10 minutes?
- Extraction of compound better
- After 10 minutes of boiling, none of the bacteria/viruses will be alive (good for immunocompromised people)
What is the purpose of pharmaco 4 library?
It has derivatives of all compounds, which is a huge resource for people to screen these compounds against cancer treatment, enzymes, etc.
What was the cancer lines tested in pharmaco 4 library?
Transfected with Ras to become cancer.
What did they do with the healthy and cancerous cell lines on pharmaco 4 library?
They tested each compound to determine which one could selectively induce cancer cell death but NOT by DNA or tubulin.
What did they want the compounds to exhibit on pharmaco 4 library?
Selective induction via oxidative stress in cancer cells only.
What was the best compound for oxidative stress and apoptosis in cancer cells?
Piper longum, which is long pepper.
What is the problem with using long pepper in a clinical trial?
- is a pure compound, so very dangerous
- you can buy it yourself, no patent ability
What extract was effective for long pepper?
Only effective in 100% ethanolic extract.
What did long pepper extract exhibit in aggressive human cancers?
It showed selective induction of apoptosis in ovarian cancer, melanoma, and in normal colon mucosa.
How does long pepper affect cancer cells?
In dose and time dependent ratios since cell viability continuously decreases (in ovarian).
Do long pepper induced apoptosis affect the caspase?
No, as inhibition of caspases via Z-VAD-FMK still showed that apoptosis occurred. Cells still died in cancerous lines (colon).
What happened to the mitochondrial fluorescence in NCM460 and HEK293 (healthy cells) with long pepper extract?
Very fluorescent, indicating that long pepper is selective for cancer cells.
What happened to the mitochondrial fluorescence in cancerous cells with long pepper extract?
Little to none, indicating apoptosis.
How does long pepper target cancer?
Via mitochondria and causing oxidative stress.
What is released when long pepper targets the mitochondria?
Studies showed that endonuclease G (a pro-apoptotic factor) and apoptosis inducing factor (AIF) were increased in cancerous models.
What did long pepper do to tumor growth in xenotransplanted mice of colon cancer?
It reduced tumor growth in the the ones that were given long pepper extract. There was no increase in size.
How did long pepper extract effect mice weight gain?
Weight stayed around the same between mice that had water and those that were forced to be injected orally with long pepper extract (turbid water). This meant the extract was well-tolerated (not the ethanol one).
Why do a supplemental trial?
Not risky as they are still able to take drugs and chemotherapy, but they ALSO take DRE for test.
EXTRA: no current negative interaction of DRE with chemotherapy
What is neccesary before supplemental trial?
NHP-drug interaction studies.
How did post-docs choose herbs to treat colon cancer in TCM?
Looked at herbs that were used to treat gastric/digestive issues and asked people what they used to treat certain symptoms.
What did the post-docs call the final formulation of herbs for colon cancer in TCM? How many?
A combination of 4 herbs was called PHY906.
What does PHY906 do?
Used in combination with chemotherapy to help reduce gastrointestinal toxicity. It promotes the regeneration of stem cells to restore the intestinal lining.
What four herbs were used in PHY906?
Licorice root, white peony, red date, and baikal scullcap.
What chemotherapies did they use with PHY906?
CPT-11, camptosar, 5-fluorouracil, and leucovorin.
Describe the oral experiment of PHY906 with mice.
Xenografted mice with tumors. Control group was given nothing, and then one group had CPT-11 chemotherapy. In the other groups CPT-11 was given with increasing dosage of PHY906.
What did the increase of the PHY906 extract do in the oral experiment of mice?
It had a positive interaction with the chemotherapy, increasing its efficacy by reducing the tumor size. Increase the anti-tumor activity of CPT-11.
What happened when the mice began to take CPT-11?
The mice started to lose weight right away, which is consistent to what happens when people take chemotherapy.
What did PHY 906 to do the mice’s weight in the oral experiment?
Decreased the animal weight loss caused by CPT-11.
What did the CPT-11 do to the mucosal cells in the oral mice experiment?
Almost kills the mucosa cells, attacking both healthy and cancerous cells.
What happened to the mucosal cells when PHY906 was combined with CPT-11?
The mucosal structure was much better showing clear scientific proof that PHY906 directly affected the toxicity of CPT-11 on mucosal cells (decrease). There was no difference in the healthy mucosa (outer layer of intestine) after given the extract.
What does TUNEL test?
It looks for DNA fragementation during apoptosis by endonuclease. The greater the amount of DNA fragmentation, the more cells there are that are dying.
How does TUNEL work?
The assay uses an enzyme called TdT. TdT is able to add labeled nucleotides to the 3’ ends of DNA breaks.
These labeled nucleotides can be detected using fluorescence or other methods (probe with antibodies), allowing researchers to visualize and quantify DNA fragmentation. Each cell will broken DNA will light up.
What cells are TUNEL used for?
Intestinal cells
What does CPT-11 do to intestinal cells in terms of apoptosis?
Lots of cells were dying by apoptosis, including healthy cells.
What happened when PHY906 was added to CPT-11 via intestinal cells and apoptosis?
Less apoptosis were dying, and it helped with the prevention of cell death in healthy cells. It recovered the mucosal layer.
What molecules indicate inflammation?
TNF alpha a and macrophages.
What does CPT-11 do for inflammation?
Induces it
What did PHY906 added to CPT-11 do for inflammation?
Inhibited inflammation in intestineal cells. Interleukins (IL-6) and other inflammatory factors were decreased with the extract. Normal cells were protected with an anti-inflammatory effect, but increased inflammation in cancer cells to help kill them.
When was PHY906 given to pancreatic cancer patients?
Progression-free survival 12.3/28 weeks. This is the time in which the disease stops to progress.
What was the result of PHY906 in pancreatic cancer patients?
There was an increase in survival with the extract, even if the disease may be progressing, they are not dying quicker. There was an overall quality of life improvement with the extract.
What did a mushroom extract (LEM) in combination with chemotherapy show in gastrointestinal patients?
Exerted inhibitory effects on the incidence of adverse effects caused by chemotherapy, like nausea, and also improved immune system.
Why are postoperative breast cancer patients still given chemotherapy?
Even though the surgery and tumor are removed, a dose of chemotherapy is given to ensure all cancer cells are killed and prevent a relapse.
What were the results of postoperative breast cancer patients who received anthracycline-based chemotherapy (drugs)?
Decreased quality of life and immune function.
What were the results of postoperative breast cancer patients who received anthracycline-based chemotherapy with the addition of LEM (mushroom extract)?
Better quality of life and increased immune function.
How to ensure the results of postoperative breast cancer patients who received anthracycline-based chemotherapy with the addition of LEM wasn’t the same as the placebo?
They may feel “better” with the placebo, but if the quality of life remains for an extended period of time, it’s more than just a psychosomatic effect.
How much dosage of LEM were the postoperative breast cancer patients given?
1.8 g per day for 6 weeks of 2 rounds of chemotherapy.
Did quality of life when using LEM with postoperative breast cancer patients decrease or increase?
It did decrease a bit but not as much compared to only chemotherapy being given on its own.
When was there a significant result in the quality of life with LEM in postoperative breast cancer patients?
In the 2nd cycle of chemotherapy.
How to measure quality of life?
FACT-BRM (functional assessment of cancer therapy, which is a biological response modifier).
When would Health Canada approve a supplementary clinical trial for DRE?
If only scientists can show that DRE does not have negative interaction with chemotherapy.
What did lemongrass extract and DRE show in human prostate cancer cells?
Induction of apoptosis in both cell lines compared to taxol alone.
What did DRE and lemongrass extract do on normal human fibroblasts when combined with taxol (chemo)?
There was a decrease in the the apoptosis of healthy cell lines, meaning the extracts provided some type of protection.
What does taxol affect in healthy cells?
Targets the tublin in them, killing them.
What did DRE do in combination with taxol/mitoxantrone to human prostate cells?
There was an increase in cell death in cancer cells, indicated it enhanced the anticancer effects of chemo-regiments and they had a positive interaction.
What did LGE do in combination with taxol/mitoxantrone to human prostate cells?
There was an increase in cell death in cancer cells, indicated it enhanced the anticancer effects of chemo-regiments and they had a positive interaction.
What extract had a better effect on cancer cells: LGE or DRE?
LGE had a greater decrease in human protaste cancer cells.
What was the mechanism of DRE in the human prostate cell experiment?
It induced cancer cell apoptosis through caspase activation of these cells. When there was a caspase inhibitor used in conjuction, cells level went up again.
What was the mechanism of LGE in the human prostate cell experiment?
It induced cancer cell apoptosis independent of caspase (different pathway for cell death). When there was a caspase inhibitor used in conjuction, cells levels were not affected.
Describe the experiment of tumor size for xenographed prostate cancer mice.
Orally fed with water (absorbed through intestine and goes to cancer site), control; fed with water and DRE or LGE.
What happened to tumor size of prostate cancer when mice were fed water with DRE/LGE?
Overall reduction in tumor size.
How was the tolerability of DRE and LGE in prostate cancer xenographed mice?
Well-tolerated, meaning they were still able to gain weight.
What is folfox made of?
A combination of 3 chemotherapy drugs.
What was LGE extracted with?
Ethanol
What did white tea and LGE do on human lymphoma cells in vivo (xenographed mice) in terms of tumor size?
Reduced size of tumor.
What did white tea and LGE do on human lymphoma cells in vitro?
Induced apoptosis of cancer cells, but selectively targeted them.
What was the overall interaction of LG extract and FOLFOX? Why?
Positive interaction. The activity of FOLFOX did not decrease when used together in terms of killing cancer cells.
How much cell death did LGE cause of colorectal cancer cells?
80 percent
How much cell death did FOLFOX cause of colorectal cancer cells?
50 percent
How did LGE and FOLFOX work in mitochondria of colorectal cancer?
A positive interaction, targeting a higher percentage of the mitochondria of cancer cells, resulting in apoptosis.
What was the in vivo result of colorectal cancer tumor size in mice fed with LGE and FOLFOX?
Showed the best result when used in combination, reducing the size of the tumor.
What lines were LGE and FOLFOX tested in vivo on?
p53+ and p53-
Did LGE help with the toxicity of FOLFOX? If so, how?
Yes, it reduced the toxic effects of chemo as the mice with colorectal cancer had weight gain similar to the control mice (no cancer). There was a significant loss in weight in those mice that had only chemo (stopped eating).
What gene can be knocked out in humans to cause colon cancer?
APC min
What gene can be knocked out in mice to cause intestinal cancer?
APC min
What does an APC min knockout result in humans? Mice?
Humans = colon cancer
Mice = intestinal cancer
Describe the study of gene knockout in mice with LGE and DRE.
Mice had APC gene knockout knockout, resulting in intestinal tumors and cancer (transgenic).
What happened to the tumor size of DRE and LGE on APC min transgenic mice?
There was a great reduction in tumor size of intestinal tumors in mice with intestinal cancer, meaning it can be preventative in the development of cancer.
How do natural extracts help with the prevention of cancer development?
By reducing tumor size.
What is usually the best treatment for cancer?
NHP and chemo combination.
What is white tea extracted with?
Water extract
What is the process level of white tea leaves?
Completely unprocessed, so all natural compounds are present.
For lymphoma xenografts in vivo, how did cancer cell numbers do with LGE? What extract worked?
LGE has decreased cancer cell viability ONLY and ONLY worked for the ethanolic extract.
For lymphoma xenografts in vivo, how did cancer cell numbers do with white tea extract? What extract worked?
White tea extract reduced cancer cell viability ONLY and the water extract worked the best for anti-cancer activity (ethanolic extract still worked, but not as good).
For lymphoma xenografts in vivo, how did tumor size do with white tea extract?
Reduced
For lymphoma xenografts in vivo, how did tumor size do with LGE?
Reduced
Why is a higher extract of white tea needed to achieve better results?
Mostly salt and sugar in the extract, so a greater amount allows more of its compounds to interact.
What does VP16 do?
It’s a DNA-targeting drug, the most common form of chemotherapy.
How did LGE and DRE compare to VP16 in lymphoma cancer cells?
Inflicting same toxicity as chemotherapy on cancer cell.
How did LGE and DRE compare to VP16 in human normal fibroblasts?
Greater number of healthy cells left alive (almost all), meaning these extracts selectively induced apoptosis in cancer cells.
How to measure mitochondria activity?
Oxygen consumption assay. If the mitochondria is not working, there will be less oxygen consumed.
What was the positive control of mitochondrial collaspe?
Antimycin A (AMA) as it’s an inhibitor of complex III.
What did LGE show in mitochondria in lymphoma cancer cells compared to DMSO?
DMSO had no effect (negative control), so almost 100% oxygen consumption; however, LGE lowered oxygen levels, indicating that they targeted cancerous mitochondria.
What can anti-oxidants do in cancer?
Prevent apoptosis and thus let cancer cells live longer.
How did LGE induce apoptosis in lymphoma cancer?
Triggers oxidative stress and mitochondrial depolarization.
How to show that LGE caused oxidative stress in lymphoma cancer?
Used with and without an anti-oxidant (NAC).
What did LGE show in lymphoma cancer when used with anti-oxidant agents?
An inhibition of its effects, showing an increase in cancer cells (no apoptosis going on).
What happened to human lymphoma cells xenographed mice fed with oral extracts of LGE and WT extracts?
Both inhibited the growth of human tumor xenografts and extracts were well tolerated (no weight changes).
What agent does greater burdock contain?
Arctigenin
What does greater burdock do in cnacer?
Reported to exert antitumor activity by reducing the tolerance of cancer cells to glucose deprivation.
What was greater burdock tested in?
Advanced pancreatic cancer.
What extract of greater burdock was used?
GBS-01.
What was the purpose of the phase 1 trial for pancreatic cancer?
Determine the dose standard and dose limited toxicity of GBS-01 in pancreatic cancer patients.
What dose was recommended for the next level of trial for pancreatic patients?
12gm/day of GBS-01.
What was the disease control rat and overall survival of pancreatic patients using GBS-01?
Increase disease control rate (33.3%) and overall longer survival (5.68 months) following treatment with the drug.
Did everyone recieve a good results in the pancreatic patient/GBS-01 study?
One partial response.
What is a blood marker for pancreatic cancer?
CA19-9
With the use of GBS-01, how did CA19-9 do?
There was a reduction in this blood marker in cells after 5 months, indicating less cancer cells.
Describe the prostate cancer study.
Men of 74 years old with prostate cancer were randomized to receive an oral capsule of polphenol-rich whole foods like pomegranate, green tea, broccoli, and tumeric, or an identical placebo for six months.
What shows the progression of prostate cancer?
Prostate specific antigen, so as PSA level increases, so does cancer stage.
What was the result of the prostate cancer study?
Food supplement group had a lower rise in the PSA levels (14.7%) as opposed to the placebo group (78.5%).
What was the clinical case report of a patient with acute monocytic leukemia who used DRE?
His blood cancer levels were very high (pallative care), and suggested DRE. Once used, within two years of diagnosis, there was almost a normal white blood count level.
What was the poor case report of DRE?
One person stopped talking for one month because it was bitter. White blood cells counts went back up after not using.
How to check blood cell count?
Using PCR technique.
What was the white blood cell count of a chronic myleoid leukemia patient before starting DRE?
BCR-AB/ABL was 0.0036% in leukocytes, which is still a high counts, millions of cancerous cells.
What was the white blood cell count of a chronic myleoid leukemia patient before starting DRE?
BCR-AB/ABL was 0, this was a clear pathological and clinical report. It proved non-toxic, should potentially take before other samples.
What are symptoms of Alzheimer’s disease?
Gradual progressive loss in memory, daily independent living functions, and the inability to learn new skills.
What are hallmarks of AD in the brain?
Neurofibrillary tangles and amyloid-beta plaques (mutated in this disease and oligermize).
What is the treatment of AD?
Symptomatic relief like increasing serotonin.
What is a new treatment of AD?
Monoclonal antibodies that binds and neutralises the amyloid-beta plaque.
What are risk factors of AD that you can change?
Depression, diabetes, high alcohol consumption, low levels of formal education, obesity, poor diet, etc.
What are risk factors of AD that you cannot change?
Age, gender (more female with AD), and genetics.
How can AD be caused by genetics?
Malfunction of neurons, which stays with you until you die.
What genes directly cause AD?
PS1, PS2, and APP. If both copes are mutated here, you will develop the disease early (40ys).
Why is there an increase in AD?
- Detection is non-existent (just see observations)
- Longevity 10x lower is where we see disease more often
Is there a higher diagnosis of AD in CAN or USA?
Actually in Canada because when you divide by population number it gives 2x higher rate.
What are holistic approaches to AD?
Physically active, reduce stress, make healthy choices, eat well, volunteer (socially active, physical activity, and pleasure).
ALL KEEP THE BRAIN ACTIVE
Why would you eat foods rich in anti-oxidants for AD?
Oxidative stress could be higher in the brain as it uses 10x more the normal oxygen.
What are the possible biochemical mechanisms of AD?
- Oxidative stress
- Mitochondrial dysfunction
- Accumulation of damaged proteins
- Inhibition of autophagy and proteasomes
- Inflammation
How does oxidative stress cause AD?
As you age, the ability to combat oxidative stress decreases.
How does mitochondrial dysfunction cause AD?
May not be recieving enough energy, which could cause it top open up, producing/releasing oxidative stress.
How does inhibition of autophagy lead to AD?
Over time this ability to clean up our cells worsens, thus there is a build up of damage proteins and mitochondria.
What to target in AD?
To protect neurons, preventing them from dying.
Prevention of AD
Behavioural, mental, physical, and diet level changes.
How can excitotoxicity lead to AD?
Neurons get excited by neurotransmitters, but extra production of neurotransmitters and their action potential leads to over-excitation, which is the influx of calcium (NOT good).
What should treatments of AD help with?
Inhibiting oxidative stress, activating autophagy, inhibiting inflammation, and stabilizing mitochondria.
How to create neuronal cells?
No one donates them, so you have to build them by:
- converting brain tumor cell lines to normal neurons
How do undifferentiated and differentiated neuronal cells differ?
Undifferentiated = Round, immature morphology with few projections
Differentiated = Polarized shape with long and many connections
If you increase the concentration of CO-Q10, what do you get?
Produces lots of ATP and also a good anti-oxidant.
What happened to NT2 cells pre-treated with CoQ10?
Cells took up the enzyme and there was near perfect protection against oxidative toxicity. This is because they began moving away from water.
What does COQ10 do?
Increase mitochondrial activity and can give and take electrons.
What happened to neural cells treated with hydrogen perioxide?
Died
What happened to neural cells with CoQ10 without hydogren perioxide?
Died
What happened to neural cells with CoQ10 in the presence if hydrogen perioxide?
All neurons remained in tact (we don’t really experience the oxidative molecule).
What happened to neural cells treated with hydrogen peroxide in the prescence of CoQ10?
Not much apoptosis.
What happened to neural cells treated with hydrogen perioxide in the absence of CoQ10?
Nuclei condensed, which is usually a manner of apoptosis.
Double transgenic mice in AD
Two genes (APP-PSI)
What does double transgenic AD mice lead to?
Overproduction of amyloid beta.
Triple transgenic mice in AD
Three genes (App-PS1-Tau)
What does triple transgenic AD mice lead to?
Phosphorylation defect
How do they test mice with AD?
They feed them with neuroprotective extract and follow up with a memory test. They then evaluate brain tissue for amyloid beta plaque, oxidative stress, and neuronal counts.
What does ubisol Q10 do in AD mice?
It prevented the decline of long-term memory usually exhibited in the transgenic/water mice. The wild-type/water group was almost similar to transgenic/UbQ10 one.
How did the measure ubisol Q10?
To exploration of Y-maze after a long rest.
How did the untreated transgenic mice explore the Y-maze?
After a long rest, they always explored it as it was new, bumping into certain areas, many times.
What was the second experiment of mice with AD?
Object has been moved, and they have to explore to see if they think it’s missing (spatial working memory).
What did the untreated transgenic mice exhibit in the spatial working memory?
They often didn’t realize that an object had been moved.
What did the wild-type and treated transgenic mice show in their spatial working memory?
Recoginze that an object has been moved by exploring it morw.
What does ubisol Q10 help with?
Preserves spatial short-term (working) memory that declines in AD mice.
What is an early indicator if human AD?
Decrease in spatial short-term (working) memory.
What did the wild type mice show from amyloid-beta serum level? Why?
No peptide as they don’t have the gene for it. It’s not mutated in them.
What did the treated mice show from amyloid-beta serum level?
A decrease in amyloid-beta plaques.
What confirmed plaque decrease in treated AD mice?
Congo red, so there was little to no red fluorescence in the treated mice.
Did ubisol-Q10 show to decrease plaque?
In the hippocampus, there was a decrease in amyloid plaque in the treated mice compared to the untreated ones.
What did Ashwagandha show in AD in middle aged?
Inhibition of memory loss and inhibition of plaque formation.
What did Ashwagandha show in AD in older aged?
Partially efficient in plaque formation.
What dosage of Aswagandha fed to mice?
1 gm/kg/day (too high).
How did Aswaganha reverse AD pathology?
By enhancing low-density lipoprotein receptor-related protein in liver.
What doe neurons use for ATP generation?
Oxidative phosphorylation.
Hypnotic effects
Producing sleep
Sedative effects
Calming effects, reducing agitation, and permitting sleep
Hops
Traditionally used for insomnia and excitability, sleep disorder, and restlessness
Lemon balm
Used for sleeping disorders
What can lemon balm do in dementia patients?
Clinical evidence for leak extract improving cognitive performance.
What can Kava help with?
Effective in relieving anxiety.
Valerian
Used for mild anxiety and sleep improvement.
How does valerian have a sedative effect?
It causes an increase in GABA concentration, which can inhibit brain activity.
Saffron
Used for anti-inflammatory, immunomodulatory, antioxidant, and depressive disorders
What part of saffron is used for major depressive disorders?
Stigma and petals.
St. John’s Wort
A nerve tonic and for moderate depression.
Lists some stimulants
Caffeine, analgesics (morphine), and cocaine.
How is caffeine a stimulant?
Inhibits degradation of cyclic AMP leading to hyper energy.
How is cocaine a stimulant?
Blocks dopamine transporters thus blocking the uptake of the molecule. More dopamine left in synapse, more reward signals in the midbrain VTA.
What does Parkinson’s disease result from?
Progressive loss of dopaminergic (DA neurons) in the SNc.
What are symptoms of Parkinson’s?
Resting tremours, postural instability, bradykinesia, and more than 50% of DA neurons.
How does oxidative stress come into play with Parkinson’s?
In the SNc oxidative stress if normally high, but even more elevated in PD. A rat model of PD showed a 30-40% decreases in complex I activity.
What three contributions did the drug addict show for PD research?
- MPTP contimination of taxan heroin produced the PD symptoms
- Treatment with leva-dopa caused hallucinations and dillusions
- Experimental deep brain stimulation helped to relieve symptoms and give better quality of life
What does MPP+ do?
Selectively targets dopaminergic neurons, inhibiting complex I of the mitochondrial electron transport chain, leading to ATP depletion and cell death.
What did they use paraquat for studies instead of MPP+?
It was a known environmental toxic that was easier to get, not detrimental, simialr structure to MPP+, and people showed that they had a 10x risk of developing Parkinson’s in this herbicide area.
What did paraquat do to neuronal SHSY-5Y cells?
Condense nuclei, leading to apoptosis of cells.
If the cells were treated with CoQ10, did paraquat affect them?
No, it inhibited its effects. No apoptosis, protecting the neuronal cells.
What happened with hydrogen perioxide was applied to normal human fibroblast cells?
The mitochondria collasped, showing oxidative stress due to decrease in red fluorescence.
What happened when hydrogen perioxide was applied to normal human fibroblast cells injected with CoQ10?
Protected the mitochondria, it neutralized the toxic material and prevented oxidative stress.
What does ubisolQ10 do to the mitochondrial membrane potential?
It stabilizes it.
Does ubisol Q10 enter the blood brain barrier?
Yes it can and over time it increases.
What did the ubisol-Q10 treated paraquat-induced rats show?
For a time period of 4 weeks, this enzymes protected the DA neurons in the SNpc from death and other abnormalities.
What is a neuromolecular marker? What does it measures?
Tyrosine hydroxlase enzyme (TS). An increase in it means there is more DA neurons.
What did ubisol Q10 show in MPTP-induced mice PD model?
Neuroprotection as there was an increase in TS/TH markers compared to the one shown in the untreated model.
What are DJ-1 mice?
A transgenic PD model where there is a homozygous knockout mouse strain, meaning they lack the DJ-1 gene.
What did ubisol-Q10 do in MPTP mice with DJ-1 mice transgenic PD model?
Neuroprotection, more neurons left as opposed to the MPTP/H20 model. This was a prolonged treatment.
What did aswangandha extract show in paraquat and manib induced PD in rats?
Demonstrated neuroprotective role in 3 weeks, but this protection was gone in 6 to 9 weeks.
What did parquat and water do to neurons?
Decreased the amount, almost entire loss.
What did paraquat and PTS (carrier of CoQ10) do to neurons in rats?
Still loss of neurons.
What did paraquat and ubisol-Q10 and ashwagandha do to neurons in rats?
Neuroprotection and extensions and synpatic connections protected. This are important as used in communication.
What does THC do?
Pyschotropic chemical that makes people feel “high.”
What does CBD do?
It’s a nonpyschotropic chemical.
What was the CBD study?
Evaluate the impacts of acute CBD adminstration at a dose of 300 mg on anxiety measures and tremours induced by a stimulated public speaking test (SPST) in individuals with PD.
What did CDB do for PD patients?
It reduced the anxiety experimentally induced by the SPST and decreased tremor amplitude in an anxiogenic situation.
What are ayurvedic medicinal plants used for brain health?
Ashwagandha, tumeric, gotu kola, etc.
What is ashwagandha used in Ayruveda as?
Adaptogen, anti-stress agent, improves immunity, good anti-oxidant and inflammatory agent.
What did in-vitro studies indicate about ashwagandha?
- Inhibit the activation of NF-xB (pro-inflammatory)
- Block beta-amyloid production
- Reduce apoptotic cell death
- Restore synaptic function (connection between neuron to neuron)
- Produce strong antioxidant effects through the triggering of Nrf2 (transcription) to the nucleus, where it increases the expression of anti-oxidenat enzymes and neuroprotective genes
- Treatment of human neuroblastoma SK-N-Sh cells with methanolic extracts of Ashwagandha root results in dendrite extension, neurite outgrowth, and synapse formation.
What was the in vivo study of PD?
Injected amyloid beta into the brain.
What did in-vivo studies indicate about ashwagandha?
It restores amyloid beta-induced memory deficit in mice and recovers the decline of axons, dendrites, and synapses in the cerebral cortex and hippocampus. It lead to a decrease of amyloid plaques and improved cognition in double transgenic mice.
What did the preliminary clinical trial of bipolar patients with ashwagandha do?
Improved auditory-verbal working memory in those that had a dosage of 500 mg/day.
What did a combination of ASH and CoQ10 do in double transgenic mice for PD?
Reduced beta-amyloid plaques AND pro-inflammatory microglia activation. It was as good as the wild type.
How did ubisol-q10 reveal in AD double transgenic mice?
Activation of autophagy, rescuing cells that usually accumulate over time.
What did a combo of ASH and CoQ10 reveal in AD double transgenic mice?
Much better activation of autophagy.
What did ASH reveal in AD double transgenic mice?
Not much autophagy activation.
What are the mechanisms of ASH?
- Frees Nrf2 from its inhibitor where it travels to the nucleus and upregulates anti-oxidative genes, reducing oxidative stress
- Blocks NFkB which reduces inflammation levels
- Directly inhibits breaking process and production of amyloid beta
- Inhibits apoptosis
- Corrects and strengthens synpatic dysfuntion by dephosphorylating Tau
What is the active ingredient in tumeric?
Curcumin
How does tumeric have an anti-oxidative effect?
Curcumin dissociates Keap1 and Nrf2 which allows Nrf2 to move into the nuclei and start transcription of anti-oxidative genes.
How does tumeric show anti-inflammatory effect?
Inhibits NF kappa B, inhibition of PLA2, and COX-2 enzymes associated with the metabolism of neural membrane phospholipids to prostaglandins.
What does curcumin bind to in vitro and in vivo studies?
Binds to amyloid-beta and inhibits its aggregation.
Where can curcumin reach in the body in vivo studies?
Blood-brain barrier.
Besides plaque formation, what can curcumin inhibit?
Tau phosphorylation, which aggregates of this create neuronal triangles that are toxic.
What is the bioavailability and stability of curcumin improved by?
Black pepper
Are there studies with dementia patients?
Not clinical ones.
What was a dementia-like study using curcumin?
Healthy volunteers were given 400 mg/day of curcumin and there was an improvement on sustained attention and working memory tasks compared to placebo adults that are over 60ys.
Cognition and curcumin study.
Treatment of curcumin (1.5 g/day) results in no loss of cognition in the treatment group whereas loss of cognition was found in the placebo group, among community-dwelling older adults.
What does bacopa monnieri do?
Stimulates neutralization of ROS through enzymes involved in pathway like superoxide dismutase.
What does guggulu do?
Blocks amyloid beta plaque formation through any breaking and porcessing.
What does gotu kola do?
Blocks mitochondrial dysfunction.
What does CBD bind to?
CB receptor 2 that is mainly expressed in the cell of the immune system.
What does THC bind to?
CB receptor 1 that is mainly expressed in the central nervous system.
Is CBD addictive?
No
What are endocannabinoids?
Body makes THC-like compounds from brain cells like endorphins.
When are endocannbinoids made?
Under extreme events (sever the nerve system not letting the pain signal go to the brain).
What do endocannabinoids do?
Inhibit secretion of neurotransmitter so you feel calm or relaxed.
What does Health Canada say about AD and cannabis use?
Still alot to learn about the long-term effects of cannbis on the brain. Some studies have shown that long-term cannabis use is associated with memory problems.
What trials haven’t be done for AD and cannabis?
No pre-clinical work found on cells and animals.
What can cannabis help manage in dementia?
Few clinical trials identified that cannabis can help manage behavioural symptoms in people with dementia like agitation and agression.
Where do these endocannabinoids bind? What does this do?
Binds to pre-synaptic terminals and triggers G-linked protein signalling which eventually block the release of calcium.
