Summary of doctoral thesis in medicine: Study of clinical and paraclinical, histopathology and immunohistochemical characteristics of hepatocellular carcinoma with portal vein thrombosis

The project with objectives: Assessment of clinical characteristics, paraclinical tests, imaging diagnoses of HCC pateints with PVT. 2- analysis of imaging, histopathology and immunohistochemistry characteristics of PVT, and safe biopsic technic, supplemental survival time and involving factors in pateins with PVT.

AND TRAINING DEFENCE

108 INSTITUTE OF CLINICAL MEDICAL AND

PHARMACEUTICAL SCIENCES

-

TRINH XUAN HUNG

STUDY OF CLINICAL AND PARACLINICAL, HISTOPATHOLOGY AND IMMUNOHISTOCHEMICAL CHARACTERISTICS OF HEPATOCELLULAR CARCINOMA WITH PORTAL VEIN THROMBOSIS

Speciality: Internal Gastroenterology

Code: 62.72.01.43

SUMMARY OF DOCTORAL THESIS IN MEDICINE

HANOI – 2019

108 INSTITUTE OF CLINICAL MEDICAL AND

PHARMACEUTICAL SCIENCES

Supervisor:

1 Professor Mai Hong Bang MD., PhD

2 Associate professor Trinh Tuan Dung MD., PhD

The thesis can be founded in:

- National library

- Scientific Research Institute of Clinical Medicine and Pharmacy108’s library

THE LIST OF PUBLISHCATION RESULTS OF THE THESIS

1 Trịnh Xuân Hùng, Mai Hồng Bàng, Trịnh Tuấn Dũng, Nguyễn Tiến Thịnh (2019), “Results of portal vein thrombus

biopsy in patients with advanced hepatocellular carcinoma”,

Journal of 108 – Clinical medicine and pharmacy, volume 14 (N

5/2019), 1-7

2 Trịnh Xuân Hùng, Mai Hồng Bàng, Trịnh Tuấn Dũng, Nguyễn Tiến Thịnh (2019), “Study on clinical and laboratory

features of advanced hepatocellular carcinoma in patients with

vein thrombosis”, Vietnam medical journal, volume 480, 165-169

THE LIST OF ACRONYMS

AFP Alpha -Fetoprotein

ALT Alanine amino transferase

AST Aspartate amino transferase

BCLC Barcelona Clinic Liver Cancer

PVT Portal vein thrombosis

WHO World Health Organization

INTRODUCTION

Hepatocellular carcinoma (HCC) is a very popular disease in the world According to GLOBOCAN data in 2018, HCC is the fifth and the ninth most commonly diagnosed cancer in men and women, respectively There will usually be an estimated 841,000 new diagnosed cases of HCC and 782,000 died cases In Vietnam, According to GLOBOCAN data in

2018, the prevalence and mortality of HCC is one of the first commonly cancer The mortality of HCC has their equivalence to compare with new diagnosed cases, this show that HCC were often diagnosed at advance stages, so the prognose and control of the disease still have difficulties

The prognose of HCC is often very poor, especially, they are advance HCC with portal vein thrombosis (PVT) Previous studies show that HCC with PVT acount for 30-62.2%, the PVT also is signal point out that primary HCC is a advance HCC with poor prognosis, risk of oesophagus variceal bleeding, and hepatic failure (e.g thrombosis in trunk

of portal vein) Mean survival time of pateints with HCC having PVT without treatment is very short (2.7-4 months), meanwhile that of HCC having not PVT without treatment is 24.4 months Detection of PVT, especailly benign or neoplastic PVT in new diagnosed HCC has key meaning for prognosis as well as making choice of treatment methods

There are no any work to study systematicaly characteristics of histopathology and immunohistochemistry of PVT in patients with HCC in Vietnam as well as in the world From the mention above, we carried out the project with objectives:

1- assessment of clinical characteristics, paraclinical tests, imaging

diagnoses of HCC pateints with PVT

2- analysis of imaging, histopathology and immunohistochemistry characteristics of PVT, and safe biopsic technic, supplemental survival time and involving factors in pateins with PVT

* Thesis layout:

The thesis has 123 pages, including: Introduction (2 pages), Chapter 1: Overview (36 pages), Chapter 2: Subjects and methods (20 pages), Chapter 3: Results (28 pages), Chapter 4: Discussion (34 pages), Conclusion (2 pages), Recommendations (1 page) The thesis has 34 tables, 26 images The thesis has 141 references: 21 Vietnamese references and 127 English references

* The new main contributions of the thesis

In Vietnam, this is the first study that biospy of PVT was carried out in HCC pateints, assessment of deffirentiation of PVT, staining

immunohistochemistry to assess angiogenesis level in PTV involving with

supplemental survival time The benign or malignant PVT are also standards for indication and contraindication of some HCC treating methods Therefore, the thesis has very reliable and scientific meanings in clinical practise The results of project shown that, the biospy of PVT in HCC patients is pretty safe technic with less complications (it is a problem that professionals are previously hesitant to mention)

CHAPTER I: OVERVIEW 1.1 Epidemiology of hepatocellular carcinoma in the world and Vietnam

HCC is the very common malignant disease, it is commonly detected

in late stage, very poor prognosis, and high mortality in short time The estimated 500.000-1.000.000 new cases of HCC, and estimated 600.000 died cases worldwide

Frome GLOBOCAN data in 2018, HCC is leading cancer in Vietnam, new HCC incidence as well as mortality are higher than lung cancer and stomach cancer, approcimately 25.335 new diagnosed cases and the mortality of HCC, acount for 15.4% and 21.5% (19.568 cases) of total of cancer, respectively Therefore, nowadays, the trend of HCC have been increasing according to the time and this is indeed a hude challenge for medicine in my country

1.2 Diagnosis of hepatocellular carcinoma

1.2.1.Clinical symptoms

The clinical symptoms of HCC are often present in late stage of the disease because of good functional balance of liver

1.2.2 Marker of hepatocellular carcinoma

Alpha-Fetoprotein is a tumer marker for HCC that is used most

in diagnose of HCC

1.2.3 Imaging diagnose of HCC

1.2.3.1 Ultrasound : This is the leading technic for screening

localized lesions in the liver, including: B-mode ultrasound, doppler ultrasound, Contrast - enhanced Ultrasound

1.2.3.2 Computed tomography scan: Nowadays, this is the most

widely used technique to diagnose HCC Multi-sequential 3-dimensional

CT scan (artery stage, portal vein and late stage) is considered a standard technique for HCC diagnosis

1.2.3.3 Magnetic resonance imaging: providing very good

characteristics of typical angiogenesis of HCC with sensitivity and specificity up to 90-95%

In addition, there are also somme technique such as: Digital Subtraction Angiography and positron emission tomography/computer

tomography (PET/CT)

1.2.4 Anapathology

The differentiation level of HCC cells: according to Edmondson and Steiner, Armed Forces Institute of Pathology as well as the World Health Organization, differentiation level of HCC cells was often divided into 4 different levels: high and medium , low and non-differentiated levels basing on ratio of nucleus/cytoplasm

1.2.5 Diagnostic criteria of hepatocellular carcinoma

- The diagnotic and follow-up process is more simplified in American Liver Foundation's guidelines in 2011

- According to the guidelines of Vietnam Ministry of public Health

in 2012:

• There are Anapathologic criteria of HCC

• Diagnosing images of abdominal CT scans with contrast agent or abdominal MRI with contrast agent + AFP concentration is higher than normal, but less than 400 ng/mL + B or C hepatitis

1.3 Portal vein thrombosis

Portal vein thrombosis can occur in many different diseases, but the most common is cirrhosis with HCC Other diseases, including: other cancer diseases, bone marrow disorders, bacterial infection, post-surgery, estrogen treatment, and portal hypertension without cirrhosis

1.3.1 Frequency of portal vein thrombosis

The PVT frequency of 10-25 % for solitary cirrhosis, a result of study with a large sample size of 701 solitary cirrhotic patients showed that the prevalence of PVT in solitary cirrhotic patient of 11%

The frequency of PVT in HCC is much higher than others, with the detected rate of PVT of advance HCC of 40 to 90.2% A result of

retrospective cohort study with 336 HCC patients in Thailand showed PVT rate of 50% The rate of 31% of another study with 194 patients was carried out in the United States

1.3.2 Pathogenesis of portal vein thrombosis

PVT is due to reducing flow rate of the portal vein system, vascular endothelial damage and coagulopathy in cirrhosis In addition to cirrhosis, HCC also directly invades the portal vein system, or metastasis of cancer cells into the portal system is considered a most essential factor for a formation of malignant PVT The development of cancer tissue with platelet aggregation inside the vein facilitate thrombosis to develop and spread In addition, the compression of the tumor in the portal vein system enhances to obstruct the flow rate, which is also a factor to motivate formation of thrombosis

1.3.3 Diagnosis of portal vein thrombosis

In order to detect and diagnose PVT, imaging diagnosis plays a extreme key role, however, the result of cell aspiration with small needle

or biospy of thrombosis are considered a gold standard of the diagnose

1.3.3.1 Imaging diagnosis

In order to diagnose malignant PVT basing on 3 signs, including: appearance of a clear invasive image of tumor into the lumen, a stretch of the portal vein of 23 mm or more and angiogenesis in PVT

1.3.3.2 Histopathology and immunohistochemistry tests

Histopathology test of PVT is a gold standard of diagnosis of benign PVT or malignant PVT Differentiation level of PVT as well as tumor in liver were devided into 4 separated levels, including: high, medium, and low, and nondifferentiation However, in some suspicious cases, especially poor or non-differentiated PVT immunohistochemistry can be used to determine

whether PVT derived from HCC Immunohistochemistry is also significant for assessment of vessels as well as level of angiogenesis in PVT

Basic methods for selecting PVT specimens:

* Cell aspiration using small needle according to the ultrasound guide

* Biopsy: selecting PVT specimens by using a biopsy gun (FAST gun)

* Surgery: dissection of the liver and selecting PVT specimens

1.3.4 Treatment of HCC with PVT

The standard drug for treatment in the stage with preserved liver function is Sorafenib Some recent clinical trials have shown that HCC with PVT can also consider for surgery, injection with 100% alcohol in thrombosis, Transarterial radioembolization (TARE) with yttrium-90-microspheres, stereotactic body radiation therapy using CyberKnife

+ General status of patient (ECOG): from 0 to 2 scores (WHO)

- Age: from 18 to 90 years old

- No gender inequality

- The patients agree to take part in the study in writing for assurance

2.1.2 Exclusive criteria

- Severe liver disease: Child - Pugh C, hepatic encephalopathy, ascites

- Bleeding esophageal varicose

- Tumors of HCC and PVT are in a position where biopsy cannot be performed

- Some internal diseases that increase the risk of bleeding, renal failure, liver failure, venous occlusion in the liver, Rendu - Osler disease

- Pregnant women and nursing mothers

- Uncooperative patients, do not obey the process of study

2.2.4 Steps of research process

2.2.4.1 Subject Selection: Clinical and paraclinical examinations

* Paraclinical examinations:

- Routine blood tests (Hematology tests, Common blood chemistry tests, immunological tests)

- imaging diagnosis: Ultrasound, Multi-sequential 3-dimensional CT scan

* The patient is explained about the study and making guarantee

2.2.4.2 Technical procedure

PVT Specimens was collected by biopsy though the skin according to

ultrasound guidance or surgery of liver tumor and removing whole PVT

* Techniques of PVT biospy

- Indication: Suspicion of benign or malignant PVT

- Contraindication: coagulation disorders; ascites; the patient has combined severe diseases

- Utilize of ultrasound examination before performence of the PVT biospy, after that choosing the position of the PVT biopsy through skin, put the patients on a favorable position

- Preparation of insertion of the needle into the biopsy gun (FAST gun)

- Determination of puncturable point on the skin, and then disinfection and making local anesthetization with 2% lidocaine

- Puncturing the needle biopsy through the skin, the needle should be passed the healthy parenchyma of liver and then stopping before reaching for the edge of the thrombus

- Turn on the trigger of FAST gun so that the tip of needle automatically go into the thrombus

- Pulling out the needle and bandaging

- The specimens were put into 10% formol, carrying them to department

of anapathology for histopathology and immunohistochemistry tests

- After biospy, the patients were motionlessly lied for 3-4 hours, they were monitored pulse, blood pressure, and abdominal state

* Procedure of histopathology and immunohistochemistry tests

- Specimens were processed by routine methods using synchronous system of Microm (Germany) Dyeing of specimens with automatic dyeing machine HMS-70 to assess the damage of liver cells, morphology and structure of liver tumors

- Immunohistochemistry test: The dyeing was performed by Ventana's

BenchMark Ultra automatic dyeing machine (United States) Depending

on specific cases, it could be dyed by markers, including: Hepar-1, Hepatocyte, AFP, CK7, CK 20, CDx2, Vimentin, Chromogranin, NSE, Synaptophysin and so on; assessment of angiogenesis in thrombosis using

2 additional markers such as CD31 and CD34

* Post-biopsy monitoring: Some common side effects were liver pain and fever Monitoring of severe complications (bleeding, perforation, ) mornitoring of extra-survival time, the patient could be whether or not interventionally treated according to the hospital cancer council

- Gender: men, women

- Risk factors, other diseases

- Clinical symptoms:

- Using The ECOG Performance Status as one such measurement to assess how a patient's disease was progressing, including: level 0, 1, and 2

* Paraclinical criteria:

- Hematology tests, biochemical tests, immunology tests

- AFP was divided 4 levels: AFP ≤ 25IU / ml, AFP> 25 - 200 IU / ml, AFP> 200-400 IU / ml, AFP> 400 IU / ml (1IU / ml = 0.92ng / ml)

- Gastroscopy: The classification of esophageal varices (normal esophageal varices, grade I, II, III)

2.2.5.2 Criteria of imaging diagnosis

* Characteristics of liver tumors: location, number, size, u properties