Diagnosis of parental balanced reciprocal

translocations by trophectoderm biopsy

and comprehensive chromosomal

screening

Lian Liu, MD

Co-Authors:

L. W. Sundheimer1, L. Liu2 , R. P. Buyalos1,3 , G. Hubert1,3 , Z. Al-Safi1 , M.

Shamonki1,3

1 Division of Reproductive Endocrinology and Infertility, Department of

Obstetrics and Gynecology, David Geffen School of Medicine,

University of California, 10833 Le Conte Avenue, Room 27-139 CHS, Los

Angeles, CA 90095- 1740, USA

2 PacGenomics, 28222 Agoura Road Suite 200, Agoura Hills, CA 91301,

USA

3 Fertility and Surgical Associates of California, 325 Rolling Oaks Drive,

Thousand Oaks, CA 91361, USA

Abstract

Purpose This study investigates a case series of eight couples in one IVF center

who underwent trophectoderm (TE) biopsy and comprehensive chromosomal

screening (CCS) for routine aneuploidy screening and were found to have CCS

results concerning for previously undetected parental balanced reciprocal

translocations.

Methods In each case, controlled ovarian hyperstimulation and in vitro

fertilization (IVF) yielded multiple blastocysts that each underwent CCS with

high-density oligonucleotide microarray comparative genomic hybridization

(aCGH).

Results Parental translocations were suspected based on the finding of identical

breakpoint mutations in multiple embryos from each couple. Confirmation of

these suspected translocations within blastocysts was performed with next-

generation sequencing (NGS). Subsequent parental karyotypic evaluation

resulted in a diagnosis of parental balanced reciprocal translocation in each

case.

Conclusions We demonstrated that high-resolution aCGH and NGS on TE

biopsies can accurately detect parental reciprocal translocations when

previously unrecognized.

Typical log2 ratio charts for PGS from

PacGenomics

(Euploidy)

46, XX (vs. both Male and Female reference)

46, XY (vs. both Male and Female reference)

46, XX (vs. Male reference)

46, XY (vs. Male reference)

Array-based PGS NGS-based PGS

Typical log2 ratio charts for PGS

from PacGenomics

(Aneuploidy)

Array-based PGS NGS-based PGS

46, XY

Del of 3q22.2q29;

Dup of 5q12.1q35.3

46, XY

Del of 6q14.1q27;

Del of 13q12.11q14.11

Case one

Questions:

• What could be the diagnosis?

• Is it a 46, XY?

1

2

3

4

5

NGS-based PGS of the other samples from Case one

Combined view of all 5 samples’ Log2 ratios

Prediction of parental karyotype:

t(3;5),

the breakpoints at chr-3: 3q22.2

the breakpoints at chr-5: 5q12.1

Answer to Case one 46, XY

Yes. It is a 46, XY in a sense of “molecular” genetics.

But it also could be a balanced translocation if we examine it with cytogenetic method.

We recommended the patient couple for a blood karyotyping (G-banding). The

results was the mother has a balanced translocation between chr-3 and chr-5,

with the breakpoint as we predicted.

Case two

2

3

Questions:

• What could be the

diagnosis?

• Are they 46, XX and

46, XY?

1

7

6

5

4

3

2

Array-based PGS of the other samples from Case two

Prediction of parental karyotype:

t(6;13),

the breakpoints at chr-6: q14.1

the breakpoints at chr-13: q14.11

Combined view of all 7 samples’ Log2 ratios

Answer to Case two

46, XX

46, XY

But they also could be balanced

translocations if we examine them

with cytogenetic method.

We recommended the patient

couple for a blood karyotyping (G-

banding). The results was the

mother has a balanced

translocation between chr-6 and

chr-13, with the breakpoint as we

predicted.

2

3

What we did

All reported cases were identified based on judgment calls. An inherited translocation was suspected when

(1) at least two embryos of partial aneuploidy shared the exact same breakpoint(s);

(2) the breakpoint call log2 values were close to + 0.59 (gain)/− 1.0 (loss) to show no or very low mosaicism and signifying that the likely inherited translocation was most likely non-mosaic; or

(3) multiple embryos shared the same whole chromosome loss/gain.

Once the laboratory identified a suspected translocation, the referring provider was notified for further parent evaluation.

Summary Eight patient couples from the study cohort were identified as having a recurring

identical breakpoint mutation in their embryos via CCS, raising suspicion for a parental balanced translocation.

A total of 1847 CCS cases (in this IVF center, 2013~2016) were performed during the same time period, excluding previously known translocation cases, indicating an incidence of just over 0.4% of cases undergoing PGS for routine aneuploidy screening. The patients ranged in age from 28 to 38 years and male partners from 35 to 39 years. Patients elected for CCS and had a history of infertility or RPL (Table 1). A translocation was suspected when a segmental abnormality was detected with the identical chromosomal breakpoint mutation in more than one embryo. Of these cases, a total of 87 embryos were evaluated from 14 IVF cycles. Twenty distinct breakpoint mutations were identified. When a recurring breakpoint mutation was identified suggestive of a translocation, NGS testing was performed to replicate and validate the aCGH findings. In all cases, the high-density oligonucleotide aCGH and NGS results were identical.

None of the patients had a known history of translocation of POC, blood or any types. PGS has been the first sign the patient couple could be balanced translocation carriers.

The presence of a parental balanced reciprocal translocation was confirmed in one parent in all cases.

In one case (patient 2), a conventional karyotype had been performed prior to IVF and was reported as normal in both the male and female. After the CCS results for this couple were reviewed, the karyotypes were repeated with special attention to the breakpoint noted in the embryos. The resultant karyotype, 46, XX t(5:17)(q32;q23), confirmed the female patient as a translocation carrier.

Discussion

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Potential Clinical Applications

For balanced translocation carrier, we can use this to identify the reason for

infertility / multiple miscarriage.

If the patients (otherwise non-significant) have long history of infertility /

multiple miscarriage (the reason to seek IVF treatment), after parental

translocation be identified, sperm/oocyte donation may be

recommended.

For IVF patient who use sperm/oocytes donations, we can potentially

identify if the translocation inherited from the donor.