Disclosure

Marcos MeseguerValencian Infertility Institute (IVI)Valencia, Spain

Declared no potential conflict of interest.

How we can use recent technology in the IVF cycles to improve IVF

Marcos Meseguer

Spain

✓ What makes a difference? technologies?

Excellence in IVF?

IVF Lab

Vitrification

GeneticScreening

IMSI

Macs

PolarizedMicroscopy

Time-Lapse

Meseguer M. Fertil Steril. 2016;105:295-6.

Time-lapse the remaining questions

Time-lapse set-up; IVI Valencia

Less disturbancebetter development

Time-lapse set-up; IVI Valencia

Less disturbancebetter development

Time-lapse set-up; IVI Valencia

More observations better selection

Time-lapse set-up; IVI Valencia

Less disturbancebetter development

INDIVIDUAL CHAMBERS & BENCHTOP

Fast Tº & CO2 recovery

Small volume

Undisturbed culture

Continuous monitoring

Time lapse incubator: Exploring Biomimetics Concepts

D5+D6 D5

Blastocyst Rate

20%

30%

40%

50%

60%

70%

80%69,00%

64,70%

75,00%69,60%

Dry Conditions

Humid Conditions

Albert C and Meseguer M Alpha 2018

Improved Cultured conditions Dry vs Humid conditions

**

10%

20%

30%

40%

50%

60%

70%

80%

90%

Abortion Pregnancy

Blastocyst Rate

26,50%

66,70%

16,00%

83,30%

Dry Conditions

Humid Conditions

n=1366 EmbryosN=84 patients

Time lapse incubator: Exploring Biomimetics Concepts

Albert C and Meseguer M Alpha 2018

Improved Cultured conditions Dry vs Humid conditions

n=1366 EmbryosN=84 patients

0

20

40

60

80

100

120

Dry Conditions

Humid Conditions

Time lapse incubator: Exploring Biomimetics Concepts

Albert C and Meseguer M Alpha 2018

Improved Cultured conditions Dry vs Humid conditionsn=1366 Embryos

N=84 patients

40,0

50,0

60,0

70,0

80,0

90,0

100,0

110,0

120,0

130,0

H1sm H2sm H3sm Ratiosm AVEHsm

Co

un

ts P

er

Seco

nd

Embryo Spent Culture Media Oxidation-ThermoChemoluminiscence (TCL)

Humid

Dry

Time lapse incubator : Exploring Biomimetics Concepts

The evolution of TL incubators

ESD+; Precision Embryology

HIGHEST RESOLUTION VIDEOS

ESD+; Precision Embryology

Accurate Annotations

Accurate Annotations

Accurate Annotations

CLINICAL RESULTS JANUARY 2016–JUNE 2017

All blastocysts 90% SET

Standard incubator (no evaluation on cleavage stage) vs time-lapse

More observations Better selection

Less disturbanceBetter development

41.1%58.9%

Number of cyclestime-lapse vs conventional

incubator

CI TIME-LAPSE

Conventional incubators vs time-lapse incubators(n = 5574)

CONVENTIONALINCUBATOR

Ongoing pregnancy

*

*p < 0.05

More observations Better selection

Less disturbanceBetter development

PGS, preimplantation genetic screening. Meseguer M, personal data.

Fresh or deferred first-transfer time-lapse - NO PGS

Age

% O

ngo

ing

pre

gnan

cy

*

Ongoing pregnancy

Meseguer M, personal data.

PGS first-transfer trophoectoderm biopsy time-lapse vs standard incubator

More observations Better selection

Less disturbanceBetter development

Age

% O

ngo

ing

pre

gnan

cy

*

56,2

0,8 0,7

41,3

1

55

1,5 0,3

42,4

0,8

52,8

0 0

47,2

00

10

20

30

40

50

60

ABNORMAL ABNORMAL/DESEQ AMPLIFICATION FAILURE NORMAL NORMAL/EQUIL

CI

ESD

GERI

*

*p = 0.022

n = 163

n = 1,832

n = 3,307

Meseguer M, personal data.

Less disturbanceBetter development

% ploidy of the results in each incubatorN = 5,311 blastocysts

*

CONVENTIONAL INCUBATOR

ESD

% p

loid

yOUTCOME FROM DIFFERENT TIME-LAPSE SYSTEMS

GERI

n = 4,573

n = 3,614n = 366

0

5

10

15

20

25

30

35

40

CI EEVA ESD GERI

Less disturbanceBetter development

Meseguer M, personal data.

% of good-quality blastocysts (A/B) on Day 5 in each incubator

**

CONVENTIONAL INCUBATOR

% o

f go

od

-qu

alit

y b

last

ocy

sts

(A/B

)

OUTCOME FROM DIFFERENT TIME-LAPSE SYSTEMS

Evolution of the data; exponential growth

1653 2120 363011699

44800

78000

124000

189000

243000

288000

0

50000

100000

150000

200000

250000

300000

350000

YEAR 2009 2010 2011 2012 2013 2014 2015 2016 2017

EMBRYOS RECORDED

Manual annotations

Database registration

Exponential growth of time required for data acquisition/annotation…

Next evolutionary steps of automated software analysis for CEM

27

The evolution of the analysis in DANA: V2.0. Machine vs Embryologists

Del Gallego ASRM 201828

Phase 4

Prospective Automatic

DANA vs Embryologist

320 embryos

Event detection

V.2% Total %V.1%

Precision embryology

92%97%

90%86%

91%

9%13%

87%

The evolution of the analysis: Software vs embryologist

Del Gallego, ASRM 201829

Phase 4

Prospective Automatic

DANA vs Embryologist

320 embryos

Accuracy

Detected events % of Events in Range

Precision embryology

Meseguer and Hickman, Eshre 2018

The evolution of the a analysis; artificial Intelligence vs embryologist

Grade of

Expansion

Cat. 1

2

Cat. 2

3

Cat. 3

4

Kappa

Average

Kappa 0.751

(0.620-0.883)

0.652

(0.521-0.783)

0.790

(0.659-0.921)

0.729

(0.632-0.826)

P-value < 0.001 < 0.001 < 0.001 < 0.001

Inner Cell MassCat. 1

1

Cat. 2

2

Cat. 3

3

Kappa

Average

Kappa da

categoria

0.779

(0.363-0.446)

0.688

(0.137-0.220)

0.681

(0.551-0.811)

0.705

(0.606-0.804)

P-value < 0.001 < 0.001 < 0.001 < 0.001

TrophoectodermCat. 1

1

Cat. 2

2

Cat. 3

3

Kappa 0.382

(0.256-0.507)

0.402

(0.273-0.531)

0.501

(0.370-0.631)

0.438

(0.332-0.544)

P-value < 0.001 < 0.001 < 0.001 < 0.001

Meseguer, Hickman Rocha. Eshre 2018

The evolution of the a analysis; artificial Intelligence vs embryologist

Grade of

Expansion

Cat. 1

2

Cat. 2

3

Cat. 3

4

Cat. 4

5

Kappa

Average

Kappa 0.422

(0.464-0.381)

0.222

(0.181-0.264)

0.508

(0.466-0.549)

0.114

(0.072-0.155)

0.371

(0.342-0.400)

P-value < 0.001 < 0.001 < 0.001 < 0.001 < 0.001

Inner Cell MassCat. 1

1

Cat. 2

2

Cat. 3

3-

Kappa

Average

Kappa da

categoria

0.404

(0.363-0.446)

0.178

(0.137-0.220)

0.285

(0.243-0.326)-

0.267

(0.236-0.298)

P-value < 0.001 < 0.001 < 0.001 - < 0.001

TrophoectodermCat. 1

1

Cat. 2

2

Cat. 3

3

Kappa 0.353

(0.312-0.395)

0.209

(0.167-0.250)

0.376

(0.334-0.417)-

0.299

(0.268-0.331)

P-value < 0.001 < 0.001 < 0.001 - < 0.001

Embryologist vs artificial Intelligence

Meseguer M, personal data.

Machine Learning Embryo selection Software

360 videos and corresponding metadata (morphokinetics and clinical data); LiveBirth Prediction

1. Image Processing

2. Learning Embeddings of Images

3. Training Prediction Model using Image

4. Creating features from embryo meta-data

5. Training Prediction Model using embryo meta-data

6. Combining two prediction models (image + meta-data) to produce final prediction for the embryo

RCT, randomized controlled trial. Pribenszky C, et al. Reprod Biomed Online. 2017;35:511-20.

The last metanalysis reported More observations Better selection

Less disturbanceBetter development

Outcomes

Assumed risk(conventional

incubation, median-risk population)

Corresponding risk(time-lapse,Median-riskpopulation)

Relativeeffect

(95% CI)

No. of participants

(studies)

Quality ofevidence (GRADE)

Ongoing pregnancy 410/1,000517/1,000(457–577)

OR 1.542 (1.211–1.965)

1,637 (5 RCTs) Moderate 1.5

Early pregnancy loss 196/1,000139/1,000(103–186)

OR 0.662 (0.469–0.935)

904 (5 RCTs) Moderate 2.5

Live birth 321/1,000441/1,000(349–537)

OR 1.668(1.134–2.455)

481 (3 RCTs) Moderate 3.5

Stillbirth 29/1,00069/1,000(23–188)

OR 2.483(0.794–7.759)

481 (3 RCTs) Low 4.5

GRADE Working Group grades of evidence• High quality: further research is very unlikely to change our confidence in the estimate of effect• Moderate quality: further research is likely to have an important impact on our confidence in the estimate of effect and may

change the estimate• Low quality: further research is very likely to have an important impact on our confidence in the estimate of effect and is

likely to change the estimate• Very low quality: we are very uncertain about the estimate

Vitrification in IVF

Vitrification as an adjunct tool to PGT programs

1 hour

Vitrification

NGS

Euploid

blastocysts

Embryo transfer

0,00%

20,00%

40,00%

60,00%

80,00%

100,00%

SV IR CPR OPR

93,60%

56,20% 60,90%53,90%

N=3552 embryos

6% Euploid Embryos do not Survive

and ~40% do not implant

Overall outcomes in PGT cycles: TE biopsy + blastocyst vitrification

What are the reasons for the failure

Implantation failure of Euploid Embryos

✓Synchrony embryo-endometrium✓Age: Older patients are more likely

to have “slow” embryos✓Immunology and uterine receptivity

✓ Harmful effect of vitrification?✓ Embryo quality?

Survival Failure of Euploid Embryos

Implantation rate according to embryo quality

Own oocytes Ovum donation

IVI data 2016-2017 (Unpublished)

The biopsied blastocysts…o SV and IRo TE scoreo Expansion

Evaluation of Survival. Post-warming observations

2. Clearly Dead

T0 post-warmingPost-biopsy 6h post-warming 6h post warming

1. Clearly Alive

T0 post-warming

PGT Day 5-6 blastocyst. Embryo quality according to ASEBIR

22,40%

56,90%

19,90%

0,60%

A

B

C

D

N=3974 embryos

Embryo quality and PGT outcomes.

*= P<0.05N=3974 embryos

97,8

63,7 63,357,1

92,9

56,7 59,751,8

92

45,4 44,936,7

0

20

40

60

80

100

120

SV rate (%) IR (%) CPR (%) OPR (%)

ASEBIR PGS D5+D6

A B C

*

*

* ** *

*

Survival and Clinical outcomes for PGT embryos according to embryo quality (ASEBIR)

PGT. SV and IR according to embryo quality and day of biopsy/vitrification

94,8%95,7%

92,6%91,9%

87,3%85,7%

80,0%

82,0%

84,0%

86,0%

88,0%

90,0%

92,0%

94,0%

96,0%

98,0%

A B C

Survival according to ASEBIR

SV D5 SV D6

* *

Nº warmed D5 embryos

A

B

C

Number of warmed D6 embryos A

B

62,9%58,1%

51,1%

58,4%52,8%

41,0%

0,0%

10,0%

20,0%

30,0%

40,0%

50,0%

60,0%

70,0%

A B C

IR according to ASEBIR

IR D5 IR D6

**

* P<0.05

25,8%

61,3%

12,9%

16,8%

51,6%

31,5%

Day5 Day6

Nº warmed embryos 2159 1815

* P<0.05

PGT. SV and IR according to Blastocyst expansion and day of biopsy/vitrification

NS

98,9% 99,6%

92,9%

98,6% 99,6%

88,5%

80,0%

85,0%

90,0%

95,0%

100,0%

105,0%

BE BHi BH

Survival rate Day 5/6

SV D5 SV D6

73,7%

53,6%

43,7%

75,0%

43,0%37,7%

0,0%

10,0%

20,0%

30,0%

40,0%

50,0%

60,0%

70,0%

80,0%

BE BHi BH

Implantation rate Day 5/6

IR D5 IR D6

*

Nº embryos D5

BHi

83,8%

11%5,2%

Nº embryos D6

BHi

65,7%

30,8%3,5%

o Both survival and IR are closely related toembryo quality.

o Type A embryos: comparable resultsbetween day-5 and day 6 (IR).

o Type B and C embryos: higher outcomesfor day 5 vs. day 6 (SV and IR).

o Hatched Blastocysts yield the lowestoutcomes.

The hatched blastocyst.

Can we do better?

Evaluation of survival of biopsied blastocysts can be sometimes difficult, especially with hatched blastocysts….

t0

4.5h post warming

1.

**

t0

2h post warming

4.

**

2.

*

t0

3.

*

t0

T0 post-warming 2h post-warming

5.

TimingProgramming of biopsies and vitrification procedures in order to perform the biopsy at the HiB stage. RESCHEDULE.

If not sure of the quality of the blastocyst once biopsied: Do the tubbing and wait at least 2 hours before going on with the vitrification procedure.

Timing after warming: Make sure about survival: re-expansion.

Re-expansion of the he warmed biopsied blastocyst

ZP-Included: 3.2h ZP-Included: 4.3h Hatched Blastocysts: 5.3h

Initiation of re-expansion

Coello, A., et al. (2017) Fertil Steril 108(4): 659-666 e654.

Time-lapse evaluation of re-expansion of warmed blastocysts (No PGT)

Timing for cryotransfer after warming

64,80% 63,60%

50,00%

35,30%

0,00%

10,00%

20,00%

30,00%

40,00%

50,00%

60,00%

70,00%

up to 4 hours 4-5 hours 5-6 hours >6 hours

Included within the ZPCPR

60,90%68,20%

81,80%

50,00%

0,00%

10,00%

20,00%

30,00%

40,00%

50,00%

60,00%

70,00%

80,00%

90,00%

up to 4hours

4-5 hours 5-6 hours >6 hours

Hatching and Hatched Blastocysts

CPR

The Operator

Capalbo et al. (2016). Hum Reprod 31(1): 199-208.

Operator experience and outcome

This technology is the combination of:

- A single dish were the embryos remain meanwhile the media

are exchanged automatically.

- Automatic sealing “reduce cross contamination risk”

- Very thin dish walls to allow quick vitrification and warming

Fungible of individual use; Gavi Dish, Cassette, Gavi Cartridge

with media, Gavi Cartridge with pipette and seal.

4 steps

Automated Vitrification Instrument

Source: Web search

AUTOMATION IN THE IVF LAB

Acknowledgements:

Lucia Alegre (DANA)Raquel del Gallego (DANA)

Lorena Bori (ESD+)Ernesto Bosch (Data Analysis)

Belen Aparicio-Ruiz (Eeva)Carmela Albert (Geri)

Sonia Perez-Albala (Eeva)

Thank you