UNIVERSITY
of TORONTO
Robert F Casper MD, Dennis de la Cruz BSc, Frederic Mitri MD, Anat Hershko Klement MD, Yaakov Bentov MD, Paul Chang MD, Navid Esfandiari PhD
As women age, oocyte mitochondrial activity
declines leading to lower ATP levels and less
likelihood of blastocyst development. In
animal and human studies, the transfer of
young donor ooplasm including mitochondria
into oocytes of women previously producing
abnormal embryos led to improved embryo
development and pregnancy. Unfortunately,
these procedures result in mitochondrial
heteroplasmy. Discovery of oocyte or egg
precursor cells in the ovarian cortex now
allows preparation of potentially healthy
autologous mitochondria for injection without
creating heteroplasmy. The oocyte precursor
cell mitochondria appear morphologically
similar to oocyte mitochondria (Figure 1)
We hypothesized that injection of autologous
mitochondria derived from the patient’s own
oocyte precursor cells would improve
embryo development in ICSI cycles and
result in clinical pregnancies.
Two cycles were not completed; one because of no oocytes retrieved and one with all oocytes vitrified because no sperm could be
obtained. Of 26 treated cycles in which mitochondria were injected, there were 214 MII oocytes and 147 2PN zygotes for a 69%
fertilization rate (similar to previous ICSI cycles without mitochondrial injection; 184 2PN/271 MII=68%). There were 3 cycles with no
normal fertilization, 3 cycles with arrested embryo development, and 3 in which all blastocysts were vitrified (2 for OHSS risk, and 1 for
thin endometrium). Seventeen women (Table 1) completed cycles with embryo transfer (ET) including 3 transfers on day 3 (poor
embryo quality, 0 pregnant) and 14 blastocyst transfers (11 pregnant; Table 2). After previous failed IVF cycles, there were 9 clinical
pregnancies in 17 ET cycles (53%) and two chemical pregnancies. Two spontaneous abortions occurred at 6 weeks. Embryo
development was compared and there was no difference in blastocyst formation rate between pre- and post-mitochondrial injection
cycles (Table 3) similar to the results of published cytoplasmic transfer cycle outcomes.
Figure 1. Oocyte precursor cell mitochondria are similar
to oocyte mitochondria
Figure 2. Methodology
1
ICSI
1. White et al., 2012
2. Harvard Medical School
A. Bannwarth et al., 2014
B. Nottola et al., 2007
C. Harvard Medical School
Table 2
Table 1
We evaluated our preliminary clinical
experience in 28 women ≤ age 40 who had
1 to 3 failed IVF cycles often with poor
embryo quality including fragmentation and
arrest and no previous live births. Patients
had laparoscopic ovarian cortex biopsy with
scissors and oocyte precursor cells were
identified using flow cytometry with a
specific antibody and cryopreserved (Figure
2). On the day of oocyte retrieval for IVF,
the oocyte precursor cells were thawed,
membranes disrupted and differential
centrifugation was used to obtain an
enriched mitochondrial preparation. About
1-2 pL of the mitochondrial preparation was
injected into MII oocytes together with a
spermatozoon at the time of ICSI.
1
Fresh Embryo
Transfer
Frozen Embryo Transfer
N=10
N=7
Pregnancy = 5
(1 twin pregnancy)
(1 spontaneous
abortion, 2 chemical)
Pregnancy = 6
(1 spontaneous
abortion)
Patient
001
002
003
004
005
006
007
008
009
010
011
Age
37
33
28
30
31
35
29
40
35
39
30
Outcome
Ongoing
Ongoing
SAB
Ongoing twins
Ongoing
SAB
Ongoing
Chemical
Ongoing
Ongoing
Chemical
Table 3
Previous
IVF cycles
2
1
2
2
3
2
1
1
3
3
1
Pre-treatment
cycles
Mitochondrial injection cycles
184 2PN zygotes
147 2PN zygotes
Cav Mor 17/184
9.2%
Cav Mor 20/147
13.6%
Blastocysts 30/184
16.3%
Blastocysts 28/147
19.0%
Our clinical experience with autologous egg precursor cell mitochondrial injections at the time of ICSI appears promising in women with
previously poor embryo development and failed IVF cycles. The pregnancy rate is impressive in these women despite no difference in
blastocyst formation rate. Consequently, rather than moving to donor oocytes, it may be possible to obtain live births with a patient's own
oocytes. In addition, further research in our lab is exploring whether this procedure may be beneficial for women with failed IVF cycles over
the age of 40 years.