Molecular biology seminar

1. NathaliaPineda Martinez
Molecular biology
UniversidadPontificia Bolivariana
Third Semester
2018-2

2. INTRODUCTION
CANCER → Rapid
multiplication of abnormal
cells that extend beyond
their usual limits and may
invade adjacent parts of
the body or spread to other
organs, in a process termed
as “metastasis”. We can
also talk about of
“malignant tumors” or
“malignant neoplasms”.
→Colorectal cancer is one of
the most life-taking in a year,
after lung and liver cancer.
→There are some non-
modifiable risk factors such as
age, race and ethnicity, genetic
predisposition, several
diseases and modifiable
factors such as red meat,
processed meat, obesity,
smoking and alcohol.

3. NKP608
→It is a non-peptidic
derivative of 4-
aminopiperidine which
acts as a selective,
specific and potent
antagonist of SP at the
neurokinin-1 (NK-1)
receptor.
→It has been
characterized mainly by
its anxiolytic and
antidepressant effects.
INTRODUCTION
NK1 Receptor
→It is a receptor coupled
to protein G (GPCR).
NK1R is the main receptor
of tachykinins with major
affinity for substance P
(SP), which is associated
with hematopoiesis,
wound healing,
permeability of the
microvasculature, cell
survival, among others.
→ The complex between
SP/NK1R is associated
with tumorigenesis and
cancerous cell growth.
WNT signaling
pathway
→It is an evolutionarily
conserved pathway that
use some glycoproteins to
regulates crucial aspects of
cell fate determination and
regulation of cell growth.
→This signal stimulates
several transduction
cascades.

4. OBJECTIVE
INHIBITION OF WNT SIGNALING
PATHWAY IS SIGNIFICANT IN THE
SEEK FOR POTENCIAL
ANTITUMOR DRUGS

5. METHODS
USE
It is used because the cell culture
allows to have prepared cells to be able
to make other types of tests oriented
to the study performed, such as cell
migration and invasion assay, detection
of apoptosis, cell proliferation and
western blot.
CELL CULTURE
FUNDAMENT
Process in which cells are grown under controlled
conditions (necessary nutrients, ideal temperature, gases,
pH) generally outside their natural environment.

6. METHODS
CELL
PROLIFERATION
ASSAY
CELL MIGRATION
AND INVASION
ASSAY
FUNDAMENT
Measure the number of
cells or the change in the
proportion of cells that are
dividing under specific
conditions
FUNDAMENT
To measure the capacity of
cell motility and
invasiveness towards a
chemo-attractant gradient
(such as lipids, chemokines,
etc) or due to
over-expression of a
receptor.
USE
To identify the
concentration for the
experiment by measuring
the absorbance.
USE
Evaluate under
microscope the cell
migration and invasion
after a 24h NKP608
incubation.

7. METHODS
DETECTION OF
APOPTOSIS
FUNDAMENT
Using flow cytometry with apoptosis markers it is possible to
detect the cells that make apoptosis through several
mechanisms such as loss of membrane asymmetry, Caspase
activation or Mitochondrial transmembrane potential
decrease, differentiating them from other processes such as
necrosis or autophagy.
USE
In the study, it was used to evaluate the way in
which cells incubated with NKP608 could
significantly induce apoptosis.

8. METHODS
WESTERN BLOT
FUNDAMENT
Study to detect a specific protein according to
its size through gel electrophoresis, subjected to
reaction with antibodies.
USE
Detect concentration of proteins using
antibodies against Wnt-3a, -catenin, E-
Cadherin, VEGF, Cyclin D1, Bcl-2, Bax,
Active-caspase-3 and GAPDH.

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11. R
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12. DISCUSSION
AUTHOR CONCEPT YES OR NO
Gonzalez-Ortega A, et al.
It is known that the SP/NK1R system
enhances the migration and
invasion of cancerous cells.
Nusse R, et al.
It is well established that the Wnt
signaling involves in controlling cell
proliferation, migration, and cell
differentiation.
Abraha AM, et al.
Apoptosis, commonly called
programmed cell death, is a
prominent hallmark of human
cancers.

13. The inhibition of the Wnt
signaling pathway is the main
objective in the realization of
anticancer drugs. Those drugs
could bring benefits for cellular
control, especially for abnormal
cell growth in colorectal tissue.
1
CONCLUSIONS
The stimulation of pro apoptotic
molecules such as Bax and caspase-3 and
decreasing Bcl-2 improves the condition
of malignant growths, preventing their
proliferation and migration to other
organs.
2

15. ¡THANK YOU!