❳#LetsTalkMensHealth❲❳#LetsTalkMensHealth❲10 Clinical Tips on the Care of Male Patients AUGUST 2017 ∣ Urology Times

Despite the ongoing controversies

regarding the accuracy and pre-

dictive power of routine semen

analysis, it continues to be used

by many clinicians worldwide as the de facto

test for male infertility (Int Braz J Urol 2014;

40:443-53). In its best

practice statement for the

evaluation of the infertile

male, the AUA has pro-

posed the use of advanced

tests of sperm function

in certain patients to

enhance the diagnostic

accuracy of semen analy-

sis, specifically in cases

of unexplained infertility,

recurrent pregnancy loss,

or failure of intrauterine

insemination (IUI) and in

vitro fertilization (IVF)

and intracytoplasmic

sperm injection (ICSI)

(The Optimal Evaluation

of the Infertile Male. AUA

Best Practice Statement,

2010).

As explained in this

article, these new tests

have the potential to

improve our ability to

better diagnose and treat

complicated male infertil-

ity patients.

Oxidative stress

Oxidative stress is thought

to contribute to 40%-80% of male infertility

(Fertil Steril 2003; 79:829-43) and arises as a

consequence of excessive production of reactive

oxygen species (ROS) and impaired antioxi-

dant defense mechanisms (figure 1) (Curr Med

Chem 2001; 8:851-62). Although small amounts

of ROS are important for normal sperm func-

tion, an excess of these highly reactive mole-

cules can cause damage to the lipid-rich plasma

membranes and the integrity of DNA within

the sperm nucleus, as well as impaired motility

and spermatozoa apoptosis (Fertil Steril 2003;

79:829-43).

Antioxidants such as some vitamins and

minerals combat these overproduced ROS. In

addition to excess ROS, oxidative stress can be

due to insufficient concentrations of antioxi-

dants as well. Over the last decade, research has

provided growing support for the fact that oxi-

dative stress leads to abnormal semen param-

eters. In addition, more and more laboratory

tests are now available to measure this oxida-

tive stress. Therefore,

it would be reasonable

to potentially screen

all infertile men for the

presence of increased

ROS levels. Specialized

training and equipment,

the lack of cost-effective

and efficient assays, and,

perhaps most important-

ly, the lack of a univer-

sally accepted analytical

methods have prevented

ROS testing from being

included as part of the

routine infertility work-

up.

ROS can be mea-

sured both directly and

indirectly. Chemilumi-

nescence is probably

the most common way

to directly measure

ROS in sperm currently

and can quantify both

intracellular and extra-

cellular ROS. It uses a

luminometer in conjunc-

tion with a chemilumi-

nescent probe such as

luminol, which can also

be used to measure a

total antioxidant capac-

ity (TAC). The results

can be expressed as a

ROS-TAC score, which can give an indication

of the combined oxidant and antioxidant activi-

ties of seminal constituents. Unfortunately, this

test requires special equipment, training, and is

costly for widespread clinical use.

Due to these limitations, nitroblue tetrazoli-

um (NBT) has been put forth as a cost-effective

alternative. NBT interacts with free radicals

and is converted to a blue pigment that can then

be measured with light microscopy. This test

suffers from a lack of standardization and low

inter- and intra-observer reliability.

The new MiOXSYS Analyzer (figure 2) used

at our institution measures the so-called oxida-

tion-reduction potential (ORP). ORP measures

New diagnostic tests for male infertilityTwo technologies will help clinicians direct therapiesfor complicated infertility patients

Nicholas N. Tadros, MD, MCR ▪ Ashok Agarwal, PhD

SECTION EDITOR

Steven A. Kaplan, MD, is professor of urology at the Icahn School of Medicine at Mount Sinai and director of benign urologic diseases, Mount Sinai Health System, New York. Follow him on Twitter at @MaleHealthDoc.

Dr. Tadros is assistant

professor and director of

male infertility and sexual

health at Southern Illinois

University, Springfield, and

Dr. Agarwal is professor

and director of the Andrology

Center at Cleveland Clinic’s

American Center for

Reproductive Medicine and

on staff in the Glickman

Urological and Kidney

Institute, Cleveland.

Dr. Agarwal

Dr. Tadros

Source: Illustration courtesy of Cleveland Clinic Foundation, 2016

Pathogenesis of oxidative stress

in male infertility❳UT Figure 1 ❲

Primary pathologies of male reproductive system

Environmental lifestyle factors

Systemic pathologies

Male accessory gland infections

Prolonged stasis of spermatozoa in the epididymis during

transit

Immature/abnormal

spermatozoaVaricocele

Diabetes

Cancer

Drugs

Smoking

Pollution and radiation

Sperm DNA fragmentation

¥ Infertility¥ Impaired reproductive outcomes after IUI, IVF, and ICSI¥ Increased risk for genetic/birth defects?

Evaluation of sperm DNA

fragmentation

¥ Avoiding factors promoting ROS and SDF¥ Treatment of underlying pathology¥ Recurrent ejaculations alone or combined with micromanipulation-based sperm

selection techniques¥ Testicular sperm for ICSI

©CCF 2016

Systemic infection

Oxidative stress

O2

H2O2

OH

ES949499_UT0817_010.pgs 07.25.2017 23:11 ADV blackyellowmagentacyan

11www.urologytimes.com ∣ AUGUST 2017

the balance of all oxidants and antioxidants in

the specimen and gives a complete picture of

the oxidative stress. This test can be performed

in less than 5 minutes. It requires no special-

ized training and may possibly replace the more

complex and traditional oxidative stress tests

without sacrificing the reliability. Elevated

ORP levels correlate well with infertility, with

a significantly higher ORP seen in infertile

patients than in fertile controls.

In a recent study, ORP was able to detect

at least one abnormal sperm parameter with a

sensitivity of 70.4% and a specificity of 88.1%.

It had an 88% sensitivity and 91.2% specific-

ity when detecting oligozoospermia (Urology

2017; 104:84-89). Given the increased recogni-

tion that oxidative stress plays an important role

in male infertility, development of reproduc-

ible and cost-effective techniques in measur-

ing oxidative stress may help in tailoring our

treatments for infertile couples.

DNA fragmentation

Researchers have turned their attention to the

genetic contents of sperm, as embryo develop-

ment depends in part on the inherent integrity

of sperm DNA. DNA integrity testing is rela-

tively new to the armamentarium for fertility

specialists. While originally described in 1993,

it failed to gain traction as a clinical test due to

lack of availability and standardization. Sperm

DNA is highly compacted by binding tightly

to protamine. A certain degree of sperm DNA

damage can often be repaired by the oocyte’s

antioxidant enzymes. When damage exceeds

the repair capability of the oocyte, deleterious

effects of sperm DNA fragmentation (SDF) may

result, such as miscarriage and poor embryo

development (Hum Reprod 1999; 14:1039-49).

Because of this, more attention has turned

to testing for SDF. This test does not evaluate

the actual genetic code of the DNA within the

sperm, but rather the overall superstructure of

the DNA strands. There are many different

ways to test for SDF, each with pros and cons.

We will focus our discussion on the most com-

monly used tests as well as what is considered

the gold standard, terminal deoxynucleotidyl

transferase dUTP nick end labeling (TUNEL)

(Asian J Androl 2016; 18:205-12).

The acridine orange (AO) test uses a fluo-

rescent dye that emits a different wavelength

of light based on whether it is bound to double-

strand DNA (normal) or single-strand DNA

(abnormal). This test is fast, simple, and inex-

pensive but limited by inter-laboratory varia-

tions and lack of reproducibility. Another rela-

tively simple test, the “halo” test, has similar

pros and cons as the AO test, but evaluates the

characteristic halo of dispersed DNA loops

around sperm with non-fragmented DNA.

The latest technology using TUNEL detects

Please see INFERTILITY TESTS, page 12

Source: Illustrations courtesy of Cleveland Clinic Foundation, 2016

A. MiOXSYS Analyzer

❳UT Figure 2 ❲

Sensor Socket

Socket insertion end

Referencecell

Sampleport

Sensor Module

B. MiOXSYS sensor strip used in the analyzer

A B

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12 AUGUST 2017 ∣ Urology Times

How prostate cancer genetics will change front-line care

A: The completion of the Human Genome Proj-

ect in 2003 opened the door for not only basic

science advances but drove the clinical appli-

cations of genomic and genetics. Urologists

have recently become very familiar with the

genomics of prostate tumors studying somatic

mutations to help guide treatment decisions.

The area we are now becoming interested in

is known as germline testing or the study of

inherited genetics. We’ve been able to identify

more and more inherited genetic alterations in

medicine. The traditional ones that we have

the most familiarity with are the BRCA1 and

BRCA2 abnormalities associated with heredi-

tary breast and ovarian cancer. But it turns out

that a significant number of men can also have

BRCA1 or BRCA2 genetic alterations that can

confer an increased risk of prostate cancer.

Several newer genes such as HOXB13 and

ATM have also been identified as being asso-

ciated with prostate cancer. Importantly, we’re

recognizing that not only can prostate cancer

run in families but it also can be related to

breast cancer, ovarian cancer, pancreatic can-

cer, melanoma, and Lynch syndrome in other

family members. This area of research is giving

us some direction on how urologists can think

about approaching our patients concerning the

need for more detailed family histories.

Lastly, genetic panels are now being offered

by commercial laboratories specifically for

prostate cancer. Urologists need to be aware

that these panels are out there, and the best way

to utilize these genetic testing panels is some-

thing we’re all going to have to learn.

Q: You recently served as co-chair for the

Prostate Cancer International Consensus

Conference on the role of genetic testing

for inherited prostate cancer risk. Could

you talk about the rationale for this

meeting and what was discussed?

A: With all the recent advances in genomics

and genetic testing, we realized that there was

❳Q&A ❲❳Q&A ❲LEONARD G. GOMELLA, MD

INHERITED PROSTATE CANCER RISK

Dr. Gomella was interviewed by Urology Times Editorial Coun-cil member Stacy Loeb, MD, MSc, assistant professor of urol-ogy and population health at New York University Langone and the Manhattan VA, New York.

Please see PCA GENETICS, page 13

Ongoing genetic discoveries continue to enhance our knowledge of

conditions such as prostate cancer. In this interview, Leonard G.

Gomella, MD, provides an update on prostate cancer genetics, discusses

the recent Prostate Cancer International Consensus Conference,

and outlines why urologists should conduct more extensive family

histories of their prostate cancer patients. Dr. Gomella is chairman

of the department of urology and senior director for clinical affairs,

Jefferson Sidney Kimmel Cancer Center, Thomas Jefferson University,

Philadelphia.

Q: What percentage of prostate cancer

cases are caused by genetics?

A: Most cases of prostate cancer are caused by

genetic alterations. The problem is that when

you break it down to very specific, identifiable,

inherited prostate cancer risk genes, we have

very few at the present time. All tumors are

driven by genetics, but when you look at spe-

cific inherited risk, our current level of under-

standing is that about 10% to 15% of patients

can have a clearly identifiable inherited com-

ponent to their prostate cancer.

Q: This is a very active area of research.

Please talk about what’s new and exciting

in the world of prostate cancer genetics.

“nicks” or free ends of DNA by utilizing fluores-

cent nucleotides with a flow cytometer (or fluo-

rescence microscopy) to quantify the incorpora-

tion of dUTP into DNA breaks. This test is sen-

sitive, reliable, and has minimal inter-observer

variability. It can also be performed on samples

with severe oligozoospermia. Many now con-

sider this the gold standard for DNA fragmenta-

tion testing. More importantly, SDF can provide

valuable information for both the infertile couple

and fertility specialists in guiding treatment.

In men with infertility and varicoceles, some

will not see any improvement in semen param-

eters after varicocelectomy, and SDF testing

may help to identify those in whom varicocele

ligation would be of most benefit. SDF can also

be used in couples with recurrent pregnancy

loss or who have failed IUI as a predictive tool

to identify and effectively stratify patients based

on elevated SDF and potentially offer IVF or

ICSI sooner rather than later. In patients who

have already failed IVF or ICSI with ejaculated

sperm, SDF testing can help determine the next

steps in treatment.

Because DNA fragmentation increases dur-

ing sperm transit through the epididymis, these

patients may benefit from IVF/ICSI with sperm

extracted from the testis instead of repeated IVF

cycles with ejaculated or epididymal sperm.

SDF may also help identify environmental

exposures and lifestyle choices that affect male

fertility. Factors like smoking, obesity, and

occupational exposures can all worsen SDF.

In these patients, SDF testing can help predict

their fertility potential and monitor response to

lifestyle modification.

Conclusion

Both oxidative stress testing and DNA fragmen-

tation testing are relatively new technologies

that will help clinicians choose the most appro-

priate therapies for their patients. While no

large-scale randomized studies have been per-

formed yet, emerging evidence on both are

promising. We believe they will eventually

become commonly used in clinical practice for

the management of male infertility.

INFERT IL I T Y TESTScontinued from page 11