Using Genetics to Understand Diseases

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Using Genetics toUnderstand Diseases Tailor
Treatment
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Dr. Ameet Patki M.D., D.N.B., F.C.P.S.,
F.R.C.O.G. (U.K.) Medical Director ReGenesis,
Reliance Life Sciences Centre for Assisted
Reproduction, Endoscopy and Fetal Medicine
Mumbai Consultant Obstetrician Gynecologist
Sir Harkisondas Hospital Research Centre,
Mumbai Hon. Assoc. Prof. K.J. Somaiya Medical
College and Hospital, Mumbai
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Genetics - Scientific Karma?
I have my father's laugh but my mother's sense of
humor! My mother's temperament and my father's
temper! My father's mouth, with my mother's taste!
We are who we are when we're born!

• A concoction of the family that came before us
• Hopefully a little more than just the sum of
  their parts

While we all make decisions that define our
future, we always make them
based on who we were born.
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Understanding Genetics
The foundation level is the molecule called DNA
The information in DNA is organized into Genes
Genes, in turn, make up Chromosomes - Genome

Every cell in an individual contains the genome
Genetics is the science of hereditary variation.
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What is the Human Genome?
The genome is a genetic instruction book for
human biology

• There are twenty-three chapters, called
  chromosomes
• Each chapter contains several thousand stories,
  called genes
• Each paragraph is made up of words, written in
  letters called bases
• There are 1 billion words and 3 billion letters
  in the book which makes it longer than 800
  Bibles."

The order of these letters will dictate whether
an organism is a human or another species, such
as a fruit fly or mouse.
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What is the Human Genome?
This book of 3 billion DNA letters is actually
three
books in one

• It is a history book that contains a record of
  our relatedness to other members of our species
  and to all other living things on the planet
• It is a parts list for developing a human body
  from sperm and egg
• It is a medical textbook that contains the clues
  for the understanding, prevention and cure of
  disease

  
All the information in every book ever written if
translated into DNA, could fit easily in a
teaspoon.
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DNA Unlocked the Key to Life
The discovery of the double helix by James
Watson(25) and Francis Crick (36) on Feb. 28,
1953, unveiled The Secret
of Life."

• DNA is the molecule that makes and maintains all
  life
• It enables life to re-create itself
• It contains the blueprints and the toolbox for
  understanding how humans work

DNA is both bricks and blueprint - an engineer's
dream.
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50 Years on The Double Helix Twists Turns

• The three billion rungs are made up of chemical
  units, called "base pairs,
• A, T, C and G
• Particular combinations of these DNA base pairs
  (or genes) constitute coded instructions for the
  formation of proteins, which make up the body and
  govern its biological functioning (examples of
  proteins include insulin, collagen, digestive
  enzymes)

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50 Years on The Double Helix Twists Turns

• Human DNA looks like a twisted ladder with three
  billion rungs
• If unwound, your DNA would stretch over five
  feet, but it is only 50 trillionths of an inch
  wide
• The total amount of DNA in the 100 trillion or so
  cells in the average-sized human body laid end to
  end would run to the sun and back some twenty
  times

The DNA in a chromosome is so densely packed that
it can be upto 100,000 times longer than the
chromosome itself.
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50 Years on The Double Helix Twists Turns

• The three billion rungs are made up of chemical
  units, called "base pairs,
• A, T, C and G
• Particular combinations of these DNA base pairs
  (or genes) constitute coded instructions for the
  formation of proteins, which make up the body and
  govern its biological functioning (examples of
  proteins include insulin, collagen, digestive
  enzymes)

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50 Years on The Double Helix Twists Turns
The strands of life are like a sticky zip.
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What The Double Helix Cannot Tell You
DNA may be the script of life, but some scenes
are largely improvised

• A pregnant mother's influence on her baby
• The differences between identical twins
• The fleeting liaisons of proteins in a cell

All seem to arise without direction from DNA's
sequence
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9 of the WHOs 10 leading causes of global
deaths have genetic components

• Heart disease (13.7)
• Stroke (9.5)
• Pneumonia (6.4)
• HIV/AIDS (4.2)
• COPD (4.2)
• Diarrhea (4.1)
• Perinatal (4.0)
• Tuberculosis (2.8)
• Trachea/bronchus/lung cancer (2.3)
• ? Traffic accidents (2.2)

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How to Conquer a Genetic Disease
Nearly 4,000 genetic diseases afflict human beings

• Which altered gene causes the disease?
• What protein does this gene normally produce?
• Can the altered protein or gene be fixed or
  replaced?

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The Old Genetics
Was about conditions wholly caused by

• An extra or missing complete chromosome or part
  of a chromosome
• Down Syndrome (Trisomy -21)
• Turner Syndrome (XO)

• A mutation in a single gene
• Cystic Fibrosis (mutations in CFTR gene)
• Thalassemia
• Sickle cell disease

Downs Syndrome
Karyotyping is used for studying changes in
number and structure of chromosomes.
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The New Genetics
A New Genetics" has emerged driven by knowledge
gained at the DNA level

• Diagnostic DNA testing
• Prenatal DNA testing
• Predictive DNA testing

Diagnostic tools- Fluorescence In Situ
Hybridization (FISH), Polymerase Chain
Reaction(PCR)
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Genetic Diagnostic Tests
Karyotyping Detection of chromosome
abnormalities after culture FISH Rapid test
for detection of certain chromosomal
abnormalities PCR Detection of Gene defects at
the DNA level
Three techniques for diagnosis of genetic
abnormalities.
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Diagnostic DNA testing
A patient presents with clinical features of a
disorder, and a DNA test is undertaken to confirm
a diagnosis
- Infectious Disease Testing -

• Human Immunodeficiency Virus (HIV)
• Hepatitis C Virus (HCV)
• Hepatitis B Virus (HBV)
• Cytomegalo Virus (CMV)
• Tuberculosis (TB)

- Chronic Myeloid Leukemia
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CML
Orange Chr. 9 Green Chr. 22
Philadelphia chromosome t(922)(q34q11)
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Newer Diagnostic testing

• Comparative Genomic Hybridization (CGH)
• Used in cancer research
• In solid tumors chromosome quality is poor
• DNA of tumor normal DNA labeled with 2
  fluorescent colors are co-hybridized on a normal
  metaphase
• Detects amplification deletions
• Spectral Karyotyping (SKY)
• Multicolor FISH (m-FISH)
• Used to detect complex translocations

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Prenatal DNA Testing
Amniocentesis

• Amniocentesis
• (15-17 wks)
• Chorionic villus sampling
• (9-11 wks)
• Fetal blood sampling
• (18-20 wks)

CVS
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Preimplantation Genetic Diagnosis

• Early form of prenatal diagnosis bringing the
  hope of healthy babies to couples at risk of
  transmitting genetic disorders to their offspring
  by NEGATIVE SELECTION of the affected embryos
  prior to implantation.
• No longer a boutique medicine (Simpson).

Embryology met Genetics and so was born this new
adjunct to IVF.
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Present Day Scope for PGD/PND

• Wide variety of disorders
• Chromosomal abnormality
• Structural (Translocation, Inversion, Deletion)
• Numerical (Trisomy, Monosomy)
• Single gene disorders
• Autosomal Disorders
• Dominant (Neuroblastoma)
• Recessive (?-Thalassemia, Cystic Fibrosis)
• Sex linked disorders (Haemophilia, Muscular
  Dystrophy)

From single gene abnormality to extra
chromosomes.
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PGD Facility
The Micromanipulator
FISH -Trisomy 18, X, Y
PCR - Cystic Fibrosis ? F 508 Mutation
Cleavage stage Embryo Biopsy
FISH - Polyploidy
Polar Body Biopsy
PCR - ? Thalassemia
PGD help patients From Infertility to healthy
family.
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Indication for PGD/PND

• Advanced Maternal Age
• Familial Association to specific diseases (Downs
  syndrome, Muscular dystrophy, Cytic fibrosis,
  Hemophilia)
• Medical history/ conditions associated with
  genetic conditions
• Multiple miscarriages
• History of previously affected baby

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Predictive DNA testing
Predictive or presymptomatic DNA testing allows
genetic disorders to be detected in advance of
clinical presentation

• HNPCC (Colon cancer)
• Hereditary Nonpolyposis Colorectal Cancer
• BRCA1 and 2 (Breast Ovarian Ca)
• MODY 1,2,3 (Diabetes)
• Maturity Onset Type Diabetes in Young
• Alpha-synuclein (Parkinsons disease)

An ounce of testing could mean a pound of
prevention.
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The Human Genome Project (HGP)
The HGP was an international research effort to
decode the human genome

• Initiated in 1990 completed two years prior to
  deadline in 2003
• The human genome consists of over three billion
  chemical base pairs
• Approximately 30,000-40,000 genes

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Beyond the Human Genome Project

• Now that all genes are known, we will start
  understanding their function PATHWAYS
• We will then be able to correlate disease states
  to certain genes (Pathobiology)
• DISEASE GENE (S) GENE (S) DISEASE

We will find ways for rational treatment,
rational prevention rational diagnostics.
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Promise of the Human Genome Project
Improve diagnosis treatment through the
application of genetic information Technologies

• Predictive medicine
• Pharmacogenomics
• Population screening

Genomic Medicine is here..
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Transition from Genetics to Genomics
If genetics has been misunderstood, genomics is
even more mysterious
Genetics is the study of hereditary variation
Genomics," a term coined only 15 years ago, is
the study of the functions and interactions of
all the genes in the genome
Genomics is the study and identification of genes
and gene function.
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Genomic Medicine
The science of genomics rests on direct
experimental access to the entire genome
applies to conditions like

• Colon cancer
• Breast cancer
• Alzheimer disease
• HIV infection
• Tuberculosis

• Atherosclerosis
• Inflammatory bowel disease
• Diabetes
• Parkinsons disease

These disorders are due to the interactions of
multiple genes and environmental factors. They
are thus known as multifactorial disorders.
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Impact of Genomic Medicine
Genomic medicine will change health care by

• Providing knowledge of individual genetic
  predispositions
• Creating pharmacogenomics
• Allowing population based screening for certain
  Mendelian disorders

The focus is not the treatment of disease but the
eradication of the genetic problems that cause
disease.
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Impact of Genomic Medicine
Knowledge of individual genetic predisposition
will allow

• Individualized screening
• Individualized lifestyle changes
• Presymptomatic medical therapies

• Anti-colon cancer agents before colon cancer
  develops
• Antihypertensives before hypertension develops
• Tamoxifen as a prophylactic for CA breast

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Impact of Genomic Medicine
Pharmacogenomics will allow

• Use of Individualized medication based on
  genetically determined variation in effects and
  side effects
• More powerful/safer medicines the first time
• More accurate methods for determining medication
  doses
• New medications for specific genotypic disease
  subtypes

Drugs tailored to our individual genetic profiles
would reduce overall medical costs.
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How Does Society Prepare for Genomics?
Education to achieve understanding of

• The basics of the science of genetics
• The eventual use of genetics in health care
• How to deal with risk predisposition
• The personal impact of genetic information
• The social impact of genetics

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Concerns with Genomics

• Discrimination against individuals
• Discrimination against groups
• Genes run in families
• Confidentiality/privacy

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Tomorrows Medicine
Genomics opens up new therapeutic options

• Drugs derived from genetic engineering
• When genes acts as drugs gene therapy
• Medicine made to measure pharmacogenomics
• Predicting susceptibility to disease diagnostic
  tests
• Vaccines the treatment of infectious diseases
• Designing babies

The aim is to detect target specific diseases
develop more efficient and selective treatment.
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Genetics 25 Years Hence
By the double helix's 75th birthday, your genome
might be as familiar as your shoe size

• In 25 years babies might have genetic identity
  cards
• This would include information from an
  individual's genome sequence
• Genetic ID cards could offer benefits for
  medical diagnosis, treatment and prevention

The complete genetic makeup of individuals could
soon be scanned and recorded on a smart card.
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The Year 2028
There's no room for technophobes in the medical
profession

• As a doctor swipes her patient's genetic ID card,
  the information downloads in seconds
• A string of letters scrolls down her computer
  screen

These are the raw data of DNA, the code of life
It seems reasonable to imagine genome sequencing
happening as a matter of routine in 25 years time.
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The Year 2028
For the patient, waiting anxiously, the news will
be mixed

• The results reveal that he carries a gene that
  increases his risk of cancer
• But should he fall ill, his genes also predict
  the best therapy -

A new tailor-made drug that brings with it an 80
chance of remission
Could this be the future of medicine?
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Better Drugs in The Year 2028?

• The drug is designed using genetic knowledge
• It targets an abnormal piece of DNA in cancerous
  cells, killing them but sparing healthy cells
• After a year's treatment, 90 of patients are
  free of disease, and 50 show a complete or near
  disappearance of the abnormal gene that triggers
  the disease

Mapping our DNA may help doctors to prescribe the
safest, most effective medicine.
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Microarrays/ DNA Biochips in Medical Practice
The DNA microchip/ microarray is a revolutionary
new tool used to identify mutations in genes

• The chip, which consists of a small glass plate
  encased in plastic, is manufactured somewhat like
  a computer microchip
• On the surface, each chip contains thousands of
  immobilized DNA sequences
• Microarrays allow thousands or tens of thousands
  of specific DNA or RNA sequences to be detected
  simultaneously

Monitoring the Genome on a Chip.
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Applications of Microarray Technology

• Gene discovery
• Disease diagnosis
• Drug discovery Pharmacogenomics
• Toxicological research Toxicogenomics
• Gene expression profiling
• Genotyping
• DNA sequencing

Spotted DNA microarray
Currently only used as a research tool.
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The Future for Alzehimer Disease - 2010

• 5 or 6 genetic variations identified that
  strongly predispose for Alzheimer disease
  another 10 or 12 with weaker association
• Chip-based genetic test gives personal likelihood
  of developing the condition
• Chip-based genetic test identifies the drug most
  likely to be effective for given individual
• Chip-based genetic test determines individual
  likelihood of drug side effects

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Gene Therapy-The Next Twist in the Genome Tale
Put most simply, it introduces a "good" gene into
a person who has a disease caused by a "bad" gene.
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Gene Therapy for Parkinson's disease

• Genetically modified virus carries the DNA into
  the body
• It passes into the nucleii of some brain cells
• These cells can then produce dopamine

Gene therapy can be used to treat cancer,
heart disease, Alzheimer's Parkinson's disease.
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DNA Based Vaccines

• Technique involves the direct injection of
  plasmids-loops of DNA that contain genes for
  proteins produced by the organism being targeted
  for immunity
• Once injected into the host's muscle tissue, the
  DNA is taken up by host cells, which then start
  expressing the foreign protein
• The protein serves as an antigen that stimulates
  immune responses
• If the body later encounters the organism
  carrying this antigen, its defenses are ready to
  launch a protective attack

The DNA vaccines are an offshoot of gene therapy.
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Clinical Trials with DNA Based Vaccines

• The first clinical trials using injections of DNA
  began for HIV in 1995
• Four other clinical trials using DNA vaccines
  against influenza, herpes simplex virus, T-cell
  lymphoma, and an additional trial for HIV were
  started in 1996
• A DNA vaccine for Malaria is being developed

Expected to be particularly useful to prevent and
treat infectious diseases such as Herpes,
Hepatitis, and AIDS.
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Benefits of DNA Based Vaccines

• Traditional vaccines are expensive and take a
  long time to produce
• DNA is relatively inexpensive and is easier to
  produce
• DNA vaccines are much more stable, allowing them
  to be easily transported
• With a live vaccine (Polio vaccine), there is
  always a danger of it reverting and becoming
  infective. DNA cannot become infective

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Genetically Engineered Drugs
Protein Indication Year
Insulin Diabetes 1982
Human growth hormone Growth deficiency 1985
alpha-interferon Viral infectionsCancer 1985
Hepatitis B vaccine Vaccine against Hepatitis 1986
Tissue plasminogen activator Cardiovascular disease 1987
Erythro-poietin Anaemia 1988
Factor VIII Haemophilia 1993
Herceptin Breast Cancer 1999
)
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Designer Babies ?
In the Brave New World predicted, scientists will
do more than screen for disease free embryos
Parents will be able to reject their own genetic
heritage, and have instead of plump -
Beautiful, Clever or Sporty genes implanted into
their embryonic children
The babies of the future could be protected from
diseases.
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Que Sera Sera
Whatever will be will be, The futures not ours
to see, Que Sera Sera, What will be will be
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The Past 25 Years

• Significant advances in ART techniques have
  revolutionized both male female factor
  infertility

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History of ICSI

• 1992 Palmero et al., responsible for the
  worlds first baby conceived with ICSI.

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ICSI

• Bypasses the effective biologic mechanism of
  sperm selection.
• Application to human reproduction has not been
  preceded by extensive research trials in mammals.
  
• Human experience with ICSI is the experimental
  record.

Cause of concern with ICSI Promotion of
transgenerational transmission of genetic defects
to the offspring causing gametogenic failure.
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ICSI

• Male factor infertility
• should be considered
• as a
• potentially heritable
• condition
• (Meschede et al., 2000)

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Incidence of Chromosomal Aberration in Infertile
oligozoospermic and azoospermic males

(Liebaers et al., In Textbook of Assited
Reproductive Techniques Chapter 24 2001)
Aberrations Infertile Males (n 7876) Oligozoosermia (n 1701) Azoospermia (n 1151)
Autosomes 1.3 3.0 1.1
Sex Chromosomes 3.8 1.6 12.6
Total 5.1 4.6 13.7
Chromosome aberrations increase as sperm counts
decrease
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First trimester losses/major congenital
malformations in ICSI derived pregnancies
Ejaculate sperms Epididymal sperms Testicular Sperms
First trimester loss 24.6 31.2 33.3
2.2 had variety of major congenital
malformations.
Wisano et al., Hum. Reprod. 1996. Borduelle,
Devroey et al., Hum. Reprod. 1996. Survey of
1455 children Largest American and European
study.
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Areas of Concern

• 13.7 Azoospermic males and 4.6 oligozoospermic
  males have abnormal karyotypes.
• Further, meiotic disturbances limited to
  spermatogenic cells are found in 6 of males with
  severe OAT
• Newer research indicates that children born
  with ICSI for male infertility have twice the
  rate of major congenital malformations.

Hansen et al., N.Engl. J. Med. 2002 346 725-30.

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Genetic concern associated with Male Factor
Infertility
Genetic Abnormality Resultant Reproductive Abnormality
Y Chromosome Microdeletion (Multiplex PCR) Azoospermia, severe oligozoospermia
Cystic Fibrosis Gene Mutation (PCR) Congenital absence of Vas Deferens
Structural defects or Aneuploidies (Blood Karyotype and Sperm FISH) Variable (Klinefeleters syndrome, XYY Males)
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Genetic concern associated with Male Factor
Infertility
Genetic Abnormality Resultant Reproductive Abnormality
Y Chromosome Microdeletion (Multiplex PCR) Azoospermia, severe oligozoospermia
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Y Microdeletion

• The how and Y of Male infertility
• Y chromosome microdeletion common cause of
  spermatogenic failure.

• Incidence
• 15-20 of men with idiopathic azoopermia
• 7-10 of men with idiopathic severe
  oligozoospermia

Y chromosome microdeletion first described in
1976 by Tiepolo and Zuffardi.
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• Y Chr is paternally inherited from father to
  son

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Y Chr DNA

• Genic DNA production of proteins
• Junk DNA has little apparent function
• 98 of Y Chr is Junk DNA
• 50-70 of Junk DNA on Y Chr contain highly
  repetitive sequences.

Y Chr is a genetic junkyard as well as a gold mine
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Y Microdeletion

• Multiplex PCR
• Simple, powerful and fast tool ideal for
  screening of idiopathic infertile male
• When used with rigorous procedures reaches the
  accuracy of other sophisticated techniques such
  as Southern blot.

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Y Chr microdeletions

• MDS are clustered in 3 main regions
• AZFa, AZFb, AZFc
• 15 novel genes identified on Y Chr
• DFFRY, RBM, DAZ genes associated with male
  infertility

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Effect of deletions on extent of spermatogenesis
AZF a Lack of germ cells or sertoli cells only syndrome (SOCS)
AZF b Spermatogenic Arrest
AZF c Maturation arrest of post meiotic germ cells
Krausz and McElreavey, 1999
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PCR for Y microdeletions
M A K V -A B K V -B C K V
-C D K V -D
M Molecular weight marker A,B,C,D Multiplex
MMX Normal control - Negative control
D/W 06 07 Test patients
Promega Y chromosome Deletion Detection System,
Version 1.1
Del at DYS240 at AZFc Region
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PCR for Y microdeletions
M Molecular weight marker A,B,C,D Multiplex
MMX 038 Test patient
The absence of bands (for 038) in MMX A, B, C
and D indicates deletion of loci DYS237, DYS236
at AZFd region alongwith deletion of entire DAZ
gene and locus DYS240 at AZFc region.
Promega Y chromosome Deletion Detection System,
Version 1.1
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Y-13 Translocation-FISH,Karyotyping
FISH
Green - X Orange - Y Blue - 18
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M Molecular weight marker A,B,C,D Multiplex
MMX 085 Test patient

• The absence of bands (for 085) in MMX A, B, C
  and D indicates deletion of loci
• DYS271, KALY at AZFa region
• DYS212, SMCY, DYS218, DYS219, DYS221 at AZFb
  region
• DYF51S1, DYS237, DYS236 at AZFd region and
• Entire DAZ gene and locus DYS240 at AZFc region.

Promega Y chromosome Deletion Detection System,
Version 1.1
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Our Data on Y chromosome Microdeletions
Indication Total Normal Deletion present in Abnormal
Azoospermia 25 19 7 28
Severe OAT 35 31 4 11.43
OAT 13 11 1 7.7
Total 73 61 12 16.44
Our data coincides with the data
reported by clinics
worldwide
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Y Microdeletion

• Newer research indicates no correlation between
  the severity of spermatogenic defect and the
  localization and extent of the Yq deletions.
  Genotype-phenotype correlations difficult to
  establish.

• Hence other genes outside the AZF and/or
  environmental factors may modulate the effects of
  AZF deletions.
• Possibility that affected males can produce
  normal counts during puberty and young adulthood.
  Hence sperms could be harvested for future use.

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Genetic concern associated with Male factor
Infertility
Genetic Abnormality Resultant Reproductive Abnormality
Cystic Fibrosis - CFTR Gene Mutation (PCR) Congenital absence of Vas Deferens
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Cystic Fibrosis (CF) male infertility

• CF is a autosomal recessive disorder
• Caused due to mutations in CFTR gene
• Current medical support has improved life
  expectancy

• These individuals
• have delayed puberty
• are generally infertile
• have Azoospermia
• Congenital absence of vas deferens (CAVD)

.
PCR detects 85 of all mutations
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Cystic Fibrosis

• Product of cystic fibrosis gene is known as
  cystic fibrosis transmembrane conductance
  regulator (CFTR) protein.
• ? F 508 prevents normal maturation of the CFTR
  protein and causes failure of its normal
  localization to the cell membrane.

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CBAVD and Cystic Fibrosis
Before performing ICSI with sperm from CBAVD
males with CFTR mutations, their partners should
also be tested to avoid the risk of having a
child affected with Cystic Fibrosis.
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Detection of Cystic Fibrosis mutation

• Common gene mutations in CAVD are
• ?F508
• W1282 X
• N1303 K
• G542 X
• 1717G-A
• R553X
• R117H

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Our Data on C.F.
Number Other Abnormalities
No. of Patients analyzed (Male Infertility with CAVD) 8
Normal 7 One patient Normal Karyotype, but presence of Y microdeletion
Carrier of ? F 508 mutation 1 XYY karyotype
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Genetic concern associated with Male factor
Infertility
Genetic Abnormality Resultant Reproductive Abnormality
Structural defects or Aneuploidies (Blood Karyotype and Sperm FISH) Variable (Klinefeleters syndrome, XXY Males)
81
Abnormal karyotype in men with severe male factor
infertility
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Karyotyping - Male contribution - 200 cases
Primary/Sec. infertility 71 (35.5) BOH/RSA in
wife 129 (64.5) Chromosome Abnormalities
(4.5) Numerical abnormalities
(1.5) 47,XXY 2 cases Primary
infertility 47,XYY 1 case CAVD Structural
Abnormalities - Reciprocal translocations
(3) 46,XY,t(713)(p11q34) OAT 46,XY,t(810)(
p23q24) BOH 46,XY,t(1013)(q23.2q34)
" 46,XY,t(1116)(q21p13.3)
" 45,X,t(Y13)(q23q11)
"
45,XY,t(1314)(p11q11) "
83
Partial karyotypes showing balanced reciprocal
translocations in 3 male partners of BOH cases.
84
Chromosome Variants (10)
Variants n Indication 46,X,inv(Y) 3
OAT / BOH 46,XY,inv(9) 2 Azoospermia /
BOH 46,X,Yqh 3 Polytailed sperm / BOH
(2) 46,X,Yqh- 3 Primary Infertility (2)/
BOH 46,XY,9qh 5 BOH 46,XY,21ps 2 Primary
infertility 46,XY,22ps 2 Primary infertility
Single cell mosaicism (2) 46,XY (1/50 cell with
47,XXY) BOH (2 cases) 46,XY 1 cell with
t(714)(p12q12) OAT 46,XY 1cell with trisomy
21 BOH (2 cases)
85
FISH on Sperm
Normal 13, 21
Disomy 18
XY Sperm
Disomy 18
XY Sperm
X and Y Sperm
X and Y Sperm
Y bearing Sperm (Normal)
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Should Y chromosome analysis and genetic
counselling be offered to infertile male?

• 96 of couples choose Y chromosome testing when
  offered
• Transmission of infertility to offspring is
  weighed carefully by infertile couples
• - Rucker et al., 1998

87

• Genetic testing should become an integral part of
  patient assessment and treatment planning

88
Genetic Counseling
All couples considering ICSI are offered
counseling to inform of the potential genetic
risks to their possible offspring's.


89
Concerns on doing ICSI in infertile males

• Y Chr microdeletions are passed to Sons
• As a result the Sons are also infertile since
  they have inherited the same genetic defect that
  rendered their father infertile
• These couples need to be counseled may opt for
• Using donor sperm
• Adoption
• To have only daughters through PGD
• Such male babies may produce normal amounts of
• sperm during puberty and as youths - their
  sperms can
• be harvested for future use

90
Package for workup of male infertility

• Karyotyping
• Fluorescence in-situ hybridization (FISH) on
  sperm
• Y Chr microdeletions of 18 loci
• Cystic Fibrosis mutation analysis

91
Genetic Counseling

• Karyotyping Strongly recommended. In addition
  testicular biopsy and chromosome content of
  individual spermatozoa.
• CF Mutation Screening Screen for large number of
  mutations and the new intron 857 variant.
• 3. AZF Microdeletion Possibility of transmitting
  from father to son.
• 4. Newborn Screening Karyotyping or DNA mutation
  analysis.

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Thank You