The document presents a research protocol examining the role of caveolin1 and its regulation by specific miRNAs (miRNA 133a, miRNA 7-5p, miRNA 383) in head and neck cancer, a type of cancer prevalent in the north-east of India. The study aims to assess whether caveolin1 acts as a tumor suppressor and whether these miRNAs can serve as potential biomarkers for diagnosis. Methodology includes a case-control study with tissue and blood sample collection, and statistical analysis using SPSS to validate findings.

1. PROTOCOL
PRESENTATION

2. TITLE
Expression of Caveolin1 in Head and
Neck Cancer and its regulation by
miRNA 133a, miRNA 7-5P, miRNA 383
Candidate: Dr.Jyoti Sharma
Guide: Dr Mithu Banerjee

3. INTRODUCTION
• Head and neck cancers are the cancers of upper aerodigestive tract
which includes oral cavity, mucosal part of lip, oropharynx,
nasopharynx, hypopharynx, larynx, salivary gland and thyroid
• North-east region reports the highest incidence of the total cases .
• More prevalent in males
• 1/4th
of the total cases in the country
• Risk factors- - most common - tobacco and alcohol consumption and
HPV virus infection.
• Anatomical access difficulty of cancer leads to late diagnosis of the
cancer , hence poor prognosis and survival

4. Contd..
• Caveolin1-is an integral protein of flask shaped non
clathrin invagination in plasma membrane called caveolae.
• It regulate cholesterol distribution, signal transduction,
cell migration, and endocytic vesicular trafficking.
• Caveolin1 is one of the three isoforms (caveolin2 and
caveolin3) present in all cells –
• abundance in adipocytes, endothelial cells, muscle cells,
and fibroblasts.
• MicroRNAs (miRNA) are small noncoding RNAs,
around 22 nucleotides in length, that mediate post-
transcriptional regulation of gene expression.

5. • Scanty data present on role of caveolin1 in head and
neck cancer,
• Its regulation by miRNA 133a already established as
a tumor suppressor.
• Regulation of caveolin1 by miRNA7-5p and
miRNA383 in other cancer has been studied.
• Assessment of miRNA 133a, miRNA7-5p, miRNA
383 as potential biomarker in head and neck
cancer[19]

6. REVIEW OF LITERATURE
• The role of Caveolin in cancer progression is not
clearly understood
• Its role as a tumor promoter or tumor suppressor
gene depends on stage and type of tumor in
different organs.

7. • Low levels of Caveolin1 enable head and neck cancer cell lines
to undergo EMT and enhances prometastatic properties by
inducing the expression of α5β1 integrins in HNSCC- oncogene
• Caveolin1 reduces anchorage-independent growth, induces
G0 /G1 arrest in a cell cycle - increasing the apoptotic cell
fraction in HNSCC-tumor supressor
• Caveolin1 negatively regulates the EGFR–MAPK signaling
pathway in HNSCC.-tumor supressor
• Caveolin1 overexpression- abnormal expression of
E-cadherin/α-β catenin complex which correlates with lymph
node metastases in HNSCC-oncogene
.

8. •Hypothesis
• caveolin1 as a tumor suppressor in head and neck cancer and,
• It is regulated by miRNA 133a, miRNA7-5p, miRNA 383
• miRNA 133a, miRNA7-5p, miRNA 383 as potential biomarker in
head and neck cancer
•Research question
• 1.Does caveolin1 act as a tumor suppressor in head and neck cancer?
• 2.Do miRNA133a, miRNA7-5p, miRNA383 regulate level of
caveolin1 in head and neck cancer?
• Can miRNA133a, miRNA7-5p, miRNA383 be used as potential
biomarker in head and neck cancer?

9. Aims :
1.To study role of caveolin1 in head and neck cancer.
2. To study association of caveolin1 with miRNA133a, miRNA 7-5p,
miRNA383 in head and neck cancer.
3. To asses miRNA133a, miRNA 7-5p, miRNA383 as potential
biomarkers in head and neck cancer.
Objectives:
1.To study expression and level of caveolin1 in head and neck cancer.
1. 2.To study expression and levels of miRNA 133a, miRNA 7-5p,
miRNA 383 in head and cancer.

10. MATERIALAND METHODS
• Study setting: Department of Biochemistry AIIMS , Jodhpur.
• Type of study: Case-control study
• Study participants: Departments of ENT and Oncosurgery AIIMS
Jodhpur.
Inclusion criteria:
Newly diagnosed HPE
confirmed cases of head
and neck cancer will be
taken in this study.
Exclusion criteria:
1. History of recurrence of head and
neck/ any other cancer.
2. History of treatment for head and
neck cancer.

11. Contd..
• Sample size: 50 biopsy diagnosed cases and 50 control.
• 15 paired tissue samples
• Duration of study: 1.5 years
• Methodology: Blood samples will be collected from departments of
Oncosurgery, ENT.
• Blood will be collected in EDTA and Yellow vacutainer.
• 15 paired tissue sample will be collected

12. 15 paired tissue sample of head and
neck cancer
In EDTA for
plasma
In yellow vial for
serum
RNA isolation of miRNA and mRNA
by trizol method
Cdna conversion of miRNA and
Mrna by rt PCR
ELISA done for caveolin1
levels
Expression of miRNA and Mrna
By RT qPCR using SYBR GREEN DYE
50 cases and 50 controls
WORKFLOW

13. DATA COLLECTION AND ANALYSIS
• Statistical analysis will be performed using SPSS.
• Descriptive statistics will be carried out to determine mean, standard
deviation (SD), median and range.
• To assess normal distribution of the data, Shapiro Wilk Test will be
carried out.
• Parametric data using Student t test and non- parametric data using
Mann Whitney U test.
• To assess the correlation, Pearson or Spearman correlation test will
be performed accordingly.
• Expresssion of miRNA and mRNA will be studied by 2-
method
• P-value<0.05 were considered statistically significant.
• Sensitivity specificity auc roc will for potential biomarker

14. ETHICAL CONSIDERATION
• Informed consent will be taken from subjects.
• Subject can withdraw consent at any stage of study.
• Confidentiality of the subject will be maintained

15. Jul-Sep 2021
Oct-Dec 2021
Jan-Dec 2022
Jan-Dec 2022
Jan-Feb 2023
Feb-June 2023
Protocol submission and ethical
approval
Procurement of reagent and
standardization
Sample collection
Statistical analysis
Thesis writing and
submission
miRNA and mRNA analysis
and expression
GANTT CHART

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20. Thank you