Epithelial Mesenchymal Transition Markers in OSMF

NCT03732872 | Completed

• 1 other identifier: Oxford DC
• Study Type: observational
• Target: 185
• Locations: 0 countries
• Sites: N/A

Brief Summary

Oral Sub mucous Fibrosis (OSMF) is essentially an imbalance between collagen metabolism and wound healing mechanism induced by arecanut chewing habit. Clinically the disease progresses in stages with patients presenting with burning sensation, intolerance to spicy food, vesicles particularly on the palate, ulceration and dryness of the mouth , fibrosis of the oral mucosa, leading to lips, tongue, and palate rigidity and finally trismus. As the disease is progressively debilitating and has potential to turn in to malignant cancer a study was designed to assess if there any tissue or saliva markers that can be assessed for early diagnosis and indicate malignant transformation if any. Participants who had OSMF and habit history, patients without OSMF but habit history formed the case group where as normal patients without OSMF and no habit history were in control group. Eligible candidates who consented to participate in study were subjected to biopsy procedure and also their saliva samples were collected. Biopsy samples were subjected to immunohistochemistry (IHC) and polymerase chain reaction (PCR) to assess the EMT markers like vimentin, e-cadherin and collagen IV. miRNA copies were extracted from saliva and were subjected RT-PCR. Research question was:

1. 1.Is EMT a positive signature in OSMF.
2. 2.Does histopathological grading and dysplasia in OSMF have any correlation with EMT.
3. 3.Can aberrant EMT markers be a reliable indicator for risk assessment of early malignant transformation.
4. 4.Can expression of mi RNA 21 in saliva predict the disease severity and more importantly assess risk of early malignant transformation in OSMF.

Conditions

Oral Submucous Fibrosis

Keywords

OSMF; EMT; miRNA 21; vimentin; e-cadherin

Outcome Measures

Primary Outcomes (6)

• Change in Expression of E-cadherin levels in all the groups in IHC

  For IHC: The criteria used to define E-cadherin positive cells was brown staining in cell membrane of epithelial cells; the percentage of positively stained cells were scored as - Percentage of POSITIVE CELLS SCORE No cells 0 ≤ 10% 1 11% -50 % 2 51%-80% 3 ≥81% 4 Staining Intensity was also assessed in comparison with the positive control tissue and scored accordingly INTENSITY SCORE Negative 0 Weak 1 Moderate 2 Strong 3

  10 days from sample collection

• Change in Expression of vimentin levels in all the groups in IHC

  vimentin positive cells was brown staining in cytoplasm of epithelial cells; the percentage of positively stained cells were scored as - Percentage of POSITIVE CELLS SCORE No cells 0 ≤ 10% 1 11% -50 % 2 51%-80% 3 ≥81% 4 Staining Intensity was also assessed in comparison with the positive control tissue and scored accordingly INTENSITY SCORE Negative 0 Weak 1 Moderate 2 Strong 3

  10 days from sample collection

• Change in Expression of collagen IV levels in all the groups in IHC

  collagen IV positive cells was brown staining in basement membrane cells; the percentage of positively stained cells were scored as - Percentage of POSITIVE CELLS SCORE No cells 0 ≤ 10% 1 11% -50 % 2 51%-80% 3 ≥81% 4 Staining Intensity was also assessed in comparison with the positive control tissue and scored accordingly INTENSITY SCORE Negative 0 Weak 1 Moderate 2 Strong 3

  10 days from sample collection

• Change in Expression of miRNA 21 in all three groups using Real Time-PCR

  Considering the expression of miRNA21 in normal sample (N) as 1, the expression of all the samples were calculated The formula used is: number of copies = (amount \* 6.022x1023) / (length \* 1x109 \* 650)

  10 days from sample collection

• CHange in E cadherin levels in PCR

  After extraction of DNA from biopsy samples the primers for E-cadherin were designed using Primer Express software, synthesized and HPLC purified for real time PCR for quantification.

  20 days after sample collection

• Change in vimentin levels in PCR

  After extraction of DNA from the biopsy samples the primers for Vimentin were designed using Primer Express software, synthesized and HPLC purified for real time PCR for quantification

  20 days after sample collection

Secondary Outcomes (8)

• Change in e-cadherin expression with respect to disease category clinically in OSMF patients with habits

  15 days from sample collection

• Change in e-cadherin expression with respect to disease category histologically in OSMF patients with habits

  15 days from sample collection

• Change in vimentin expression with respect to disease category clinically in OSMF patients with habits

  15 days from sample collection

• Change in vimentin expression with respect to disease category histologically in OSMF patients with habits

  15 days from sample collection

• Change in collagen IV expression with respect to disease category clinically in OSMF patients with habits

  15 days from sample collection

Study Arms (3)

Patient with habits and having OSMF

Patients who had history of arecanut chewing habit in any form and composition and who were not undergone any treatment for their current condition i.e, OSMF

Diagnostic Test: Biopsy tissue analysis; Diagnostic Test: Saliva analysis

Patients with habits and had no clinical symptoms of OSMF

Patients who had history of arecanut chewing habit in any form and composition and had no symptoms of OSMF clinically

Diagnostic Test: Biopsy tissue analysis; Diagnostic Test: Saliva analysis

Healthy human volunteers

patients who reported no history of areacnut chewing habits and had no clinical symptoms of OSMF

Diagnostic Test: Biopsy tissue analysis; Diagnostic Test: Saliva analysis

Interventions

Biopsy tissue analysis DIAGNOSTIC_TEST

biopsy samples for IHC staining and PCR

Healthy human volunteers; Patient with habits and having OSMF; Patients with habits and had no clinical symptoms of OSMF

Saliva analysis DIAGNOSTIC_TEST

Saliva samples for miRNA 21 expression analysis

Healthy human volunteers; Patient with habits and having OSMF; Patients with habits and had no clinical symptoms of OSMF

Eligibility Criteria

• Age: 18 Years+
• Sex: all
• Healthy Volunteers: Yes
• Age Groups: Adult (18-64), Older Adult (65+)
• Sampling Method: Probability Sample

Study Population

Subjects were selected from outpatient department of Dental College OPD

You may qualify if:

• Any patient who had history of arecanut chewing habit with clinical manifestations of OSMF.
• Patients who had arecanut chewing habit history of more than 1 year and had no clinical symptoms of OSMF.
• Healthy human volunteers who are indicated for extraction of tooth and had no clinical features of OSMF and had no habit history

You may not qualify if:

• Patients who had bleeding disorders
• patients who are diagnosed with salivary gland disorders that can alter salivary flow or composition

Sponsors & Collaborators

• The Oxford Dental College, Hospital and Research Center, Bangalore, India: lead
• Credora Life Sciences, India: collaborator

Related Publications (3)

• Yang Y, Li YX, Yang X, Jiang L, Zhou ZJ, Zhu YQ. Progress risk assessment of oral premalignant lesions with saliva miRNA analysis. BMC Cancer. 2013 Mar 19;13:129. doi: 10.1186/1471-2407-13-129.

  PMID: 23510112 BACKGROUND

• Sawant SS, Vaidya Mm, Chaukar DA, Alam H, Dmello C, Gangadaran P, Kannan S, Kane S, Dange PP, Dey N, Ranganathan K, D'Cruz AK. Clinical significance of aberrant vimentin expression in oral premalignant lesions and carcinomas. Oral Dis. 2014 Jul;20(5):453-65. doi: 10.1111/odi.12151. Epub 2013 Jul 19.

  PMID: 23865921 BACKGROUND

• Pinto GA, Vassallo J, Andrade LA, Magna LA. Immunohistochemical study of basement membrane collagen IV in uterine cervix carcinoma. Sao Paulo Med J. 1998 Nov-Dec;116(6):1846-51. doi: 10.1590/s1516-31801998000600004.

  PMID: 10349192 BACKGROUND

Biospecimen

Retention: SAMPLES WITH DNA

1. Patient tissue samples taken in biopsy from all three groups 2. Saliva samples taken from all three groups

MeSH Terms

Conditions

Oral Submucous Fibrosis

Condition Hierarchy (Ancestors)

Mouth Diseases; Stomatognathic Diseases

Study Design

• Study Type: observational
• Observational Model: CASE CONTROL
• Time Perspective: CROSS SECTIONAL
• Sponsor Type: OTHER
• Responsible Party: PRINCIPAL INVESTIGATOR
• PI Title: Senior lecturer

Study Record Dates

• First Submitted: October 27, 2018
• First Posted: November 7, 2018
• Study Start: January 15, 2015
• Primary Completion: December 28, 2017
• Study Completion: March 20, 2018
• Last Updated: November 7, 2018

Record last verified: 2018-11

Data Sharing

• IPD Sharing: Will not share