Diagnosis of CUP - carcinoma of unknown primary

Definition of Carcinoma of Unknown Primary (CUP):

• No obvious identifiable primary site, despite a careful clinical history, physical examination, radiologic imaging, and biochemical or histologic investigations.

• Immunohistochemistry plays a vital role in diagnosis and classification of CUP lesions.

• Most common cases of CUPs --> carcinoma.
• Adenocarcinoma accounts for ~70% cases of CUPS.
• Poorly differentiated carcinoma: 15-20%
• Squamous cell carcinoma 5%
• Neuroendocrine carcinoma 5%

Steps:

1. To find the line of differentiation - lineage markers --> keratins (Pancytokeratin/ CAM 5.2), lymphoid (LCA), melanoma (S100), germ cell (OCT 3/4), and sarcoma markers (Vimentin).

2. To determine the type of CK distribution in the tumor cells - some subsets of CKs are seen in certain tumours.

3. To see if there is coexpression of vimentin.

4. To see if there is expression of supplemental antigens of epithelial or germ cell derivation, that

is, carcinoembryonic antigen (CEA), epithelial membrane antigen (EMA), or placental alkaline

phosphatase (PLAP).

5. Finally look for expression of cell-specific products, cell-specific structures, and transcription

factors or receptors that are unique identifiers of cell types.

Ref: Dabb's Diagnostic immunohistochemistry

Endomyocardial Biopsy

EMB  - Endomyocardial Biopsy

• Widely used for surveillance of cardiac allograft rejection and unexplained ventricular dysfunction.
• Should be done in young patients with myocarditis and older patients with infiltrative cardiac disease.

Approach:
Through femiral vein or artery
Right internal jugular vein (preferred)

Biopsy samples are taken from IVS (interventrucular septum) - since RV wall is thin and can lead to perforation.

Can be guided by fluoroscopy or 2D echocardiography.
Transthoracic echo guidance is preferred.

Commonly used bioptomes:
1. Novatome
2. Argon EMB forceps
3. Bipal 7 bioptome.

Five biopsies are taken and put in isotonic saline.

Transferred to
1. 10% neutral buffered formalin - rejection, cardiomyopathy, myocarditis abd tumours

2. Zeus fixative - for IF studies for rejection  (C4d)

3. 4% glutaraldehyde - electron microscopy for drug toxicity, metaboloc or storage disease and LCDD (light chain deposition disease)

4. Snap frozen tissue in liquid nitrogen - PCR (viral myocarditis) Dystrophin (muscular dystrophy)

EMB is absolutely necessary for diagnosis of the following  conditions:
1. Anthracycline induced cardiomyopathy  - loss of myofilaments and vacuolar degeneration.
2. Cardiac allograft rejection
3. Sarcoidosis
4. Giant cell myocarditis
5. Hylereosinophilic syndrome

Complications of EMB:
IMMEDIATE:
Site hematoma
Transient RBBB
Transient arrhythmias
Tricuspid regurgitation
RV perforation -- cardiac tamponade and pericardial effusion
Occult pulmonary embolism

LATE:
Coronary artery to RV fistula formation
Severe tricuspid regurgitation

TENSINS - A short note

TENSINS:

• Tensin is a cytoplasmic phosphoprotein.
• Localized to integrin-mediated focal adhesions.

Integrins are a family of proteins which form important connections between the cell and the ECM(extracellular matrix).

Focal adhesions are generated following interaction of integrins with ECM.

• They come to the site of action through integrins.
• Include numerous proteins (approximately 60) and include:

1. Talin
2. Tensin
3. Vincullin
4. Paxillin
5. Src
6. Focal adhesion kinase (FAK)
7. Protein Kinase C 

Functions of focal adhesions:

• Important in the cytoskeletal organisation and signal transduction.
• These focal adhesions in general are important in cell adhesion, migration, differentiation, proliferation, tissue development, apoptosis and genesis of disease.

Coming back to Tensins:

• Tensins act as a molecular bridge between integrins and actin cytoskeleton.
• It is a 220KD cytoplasmic phosphoprotein.
• Localised at focal adhesins.
• Phosphorylates on tyrosin, serine and threonine residues.

Tensin gene: Has 4 members
1) TNS1 - Normally localised to Heart, skeletal muscle, kidney and lung
2) TENC1
3) TNS3 - Normally localised to Placenta, kidney
4) TNS4/ CTEN - Normally localised to Placenta, prostate
------------------------------------------------------------------------------------------------------------
Structure of Tensin: 
Has 2 domains: Cytoskeletal domain (N-terminus) and signal transduction domain (C-terminus).

I) Cytoskeletal domain (N terminus)
Composed of two Actin Binding Domains (ABD) and PTEN
The cytoskeletal domain is absent in cten.

• ABD interacts with actin filaments 

II) Signal transduction domain (C terminus)
Composed of Src-homology2 (SH2 domain) and PTB (Phosphotyrosin binding domain)

• SH2 interacts with tyrosine phosphorylated and non-phosphorylated proteins and leads to signal transduction.
• PTB interacts with the cytoplasmic tails of β integrin.

------------------------------------------------------------------------------------------------------------

Role of tensins in health and disease:

Play an important role in 

1. Cell adhesion and motility

2. Cell survival and apoptosis

1. Cell adhesion and motility

Cten and tensin 3 control the migratory phenotype by direct interaction with integrin.

Resting state -- tensin 3 stabilises the actin-beta integrin complex.

On EGF stimulation -- cten competes with tensin 3 to bind to integrin resulting in loss of interaction with integrin, weakening of adhesion and cell migration.

Cell migration --> promotion of metastasis in cancers.
Invasion of cancer cells is thus facilitated by EGF dependant switch in isoforms of tensin.

It is also important in the pathogenesis of wound healing, gastrulation etc where migration of cells occur.

Mutations of KRAS/BRAF/MAPK (as seen in many tumours)  -> upregulation of cten --> metastasis.

Actin is also required for cell migration. But after a certain point, more than the strength of the actin pull, it is the strength of adhesion that is inhibitory to movement.

Cancer cell dissemination occurs by 2 strategies - 1. Amoeboid movement as single cells  2. Movement in cell clusters.

Amoeboid movement is facilitated by cten.

2. Cell survival and apoptosis:

Detachment of integrins --> Loss of contact with the extracellular matrix --> Cell death

Tensin and its downstream signaling molecules may be targets for therapeutic interventions in wound healing and cancer.