Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 https://doi.org/10.1186/s13287-021-02301-0 RESEARCH Open Access Evaluation of the Spanish population coverage of a prospective HLA haplobank of induced pluripotent stem cells Belén Álvarez-Palomo1,2*, Iris García-Martinez1, Jorge Gayoso3, Angel Raya4, Anna Veiga4, María Luisa Abad5, Adolfo Eiras5, María Guzmán-Fulgencio6, Mar Luis-Hidalgo6, Cristina Eguizabal7,8, Silvia Santos7,8, Antonio Balas9, Raquel Alenda9, Francisco Sanchez-Gordo10, Laura Ponce Verdugo10, Juliana Villa11, Enric Carreras11, Francisco Vidal1,12,13, Alejandro Madrigal14,15, María José Herrero1, Francesc Rudilla1 and Sergi Querol1,2* Abstract Background: iPSC (induced pluripotent stem cells) banks of iPSC lines with homozygous HLA (human leukocyte antigen) haplotypes (haplobanks) are proposed as an affordable and off-the-shelf approach to allogeneic transplantation of iPSC derived cell therapies. Cord blood banks offer an extensive source of HLA-typed cells suitable for reprogramming to iPSC. Several initiatives worldwide have been undertaken to create national and international iPSC haplobanks that match a significant part of a population. Methods: To create an iPSC haplobank that serves the Spanish population (IPS-PANIA), we have searched the Spanish Bone Marrow Donor Registry (REDMO) to identify the most frequently estimated haplotypes. From the top ten donors identified, we estimated the population coverage using the criteria of zero mismatches in HLA-A, HLA-B, and HLA-DRB1 with different stringencies: high resolution, low resolution, and beneficial mismatch. Results: We have calculated that ten cord blood units from homozygous donors stored at the Spanish cord blood banks can provide HLA-A, HLA-B, and HLA-DRB1 matching for 28.23% of the population. Conclusion: We confirm the feasibility of using banked cord blood units to create an iPSC haplobank that will cover a significant percentage of the Spanish and international population for future advanced therapy replacement strategies. Keywords: Induced pluripotent stem cells, Haplobank, HLA matching, Homozygous Introduction There has been extensive research and development in Induced pluripotent stem cells (iPSC) hold great promise the field to create safe iPSC and protocols to differenti- in the field of regenerative medicine due to their cap- ate them into clinically relevant cells for cell therapy ap- acity to both self-renew and differentiate into any cell- plications. This research effort culminated in 2015 with type of the human body. Unlike embryonic stem cells, the first successful clinical trial in Japan that used iPSC iPSC are free of ethical concerns and allow the autolo- derived retinal pigmented epithelial cells (RPE) in an gous application of cell replacement therapies [1, 2]. age-related macular degeneration (AMD) patient (RIKE N trial) [3]. However, the high cost and lengthy process * Correspondence: abalvarez@bst.cat; squerol@bst.cat of iPSC production could make the use of iPSC prohibi- 1 Banc de Sang i Teixits, Edifici Dr. Frederic Duran i Jordà, Passeig Taulat 116, tive for many applications. Consequently, the idea of 08005 Barcelona, Spain Full list of author information is available at the end of the article using clinically matched iPSC for HLA-A, HLA-B, and © The Author(s). 2021 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data. Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 2 of 8 HLA- DRB1 as an allogeneic treatment became more blood donors are already a reality in South Korea and widespread [4]. Japan [16, 19, 20]. Although MHC (major histocompatibility complex) The HLA haplotype landscape in Spain has been in- class I and II molecules are potentially immunogenic, vestigated before with small cohorts of patients and HLA-A, HLA-B, and HLA–DRB1 are the strongest de- healthy individuals [21–23] and a larger cohort of 5458 terminants of rejection of an allogeneic transplant [5]. units of cord blood from the Barcelona Cord Blood The effect of HLA-A, HLA-B, and HLA-DRB1 mismatch Bank, HLA-typed in high resolution [24]. on solid organ transplant rejection has been extensively The Spanish project IPS-PANIA aims at creating an documented [6]. It is important to note that HLA iPSC haplobank of at least seven clinical-grade lines to matching for solid organ transplantation mainly takes provide maximum coverage to the Spanish population. into account HLA-A, HLA-B, and HLA-DRB1 loci at a To identify the haplotypes providing maximum coverage low-resolution level [7]. (probability of zero mismatches in HLA-A, HLA-B, and The level of HLA-matching requirements ranges de- HLA-DRB1), we have searched a large cohort of 32,000 pending on the type of transplant, with a minimum adult bone marrow donors and calculated the estimated score of 9/10 (HLA-A, HLA-B, HLA-C, HLA-DRB1, coverage for a study population of 418,981 individuals HLA-DQB1) for bone marrow transplantation of unre- including cord blood donors plus bone marrow donors lated donors, a minimum score of 6/8 (HLA-A, HLA-B, from REDMO registry. We have concluded that a haplo- HLA-C, HLA-DRB1) for cord blood [8] to kidney trans- bank of seven lines would cover 23.69% of the Spanish plants in which several degrees of “beneficial match” are population and ten lines would cover 28.23%. considered, from one mismatch in HLA-A or HLA-B to just matching HLA-DRB1 alone [9]. Currently, for Materials and methods hematopoietic progenitor transplantation, the common Study cohort and ethics approval use is high-resolution sequencing (two fields) of five The REDMO includes the HLA typing of all cord blood HLA loci [10] using Next Generation Sequencing (NGS) donations and all the adult bone marrow donors of the for typing. Spanish population. For population coverage studies, the The relevance and stringency of HLA-matching for target population consisted of all the cord blood plus all iPSC-derived cells for clinical transplantation has been the adult bone marrow donors in REDMO. The consult- discussed extensively [11]. Unlike transplantation of ation of the HLA data was approved by the Ethics Com- hematopoietic progenitors, iPSC derivatives do not mittee for Research with Medicines from Vall d’Hebron have contaminating immune cells and therefore the Hospital (Barcelona, Spain) and the Transplantation and level of compatibility required is measured only for Regenerative Medicine Commission of the Spanish Na- host vs. graft because the graft does not have in this tional Health System. case the ability to react against the host mismatched HLA antigens. HLA-A, HLA-B, and HLA-DRB1 matching have shown to confer a clear advantage HLA typing and haplotype frequency determination over totally allogeneic transplant, with different de- High-resolution typing was performed by Sanger se- grees of immunological responses observed indicating quencing in an ABI PRISM 3130xl Genetic Analyzer the need for immunosuppression [4, 12]. (Thermo Fisher) and/or NGS in a MiSeq platform (Illu- Banking of HLA-typed pluripotent cells for matching a mina) or in Ion GeneStudio S5 System (Thermo Fisher) wide proportion of a population was first proposed for for HLA-A, HLA-B, HLA-C, HLA-DRB1, HLA-DRB3/4/5, embryonic stem cells [13]. The authors also proposed the HLA-DQB1, and HLA-DPB1 genes. The resulting se- use of homozygous donors for common HLA-A, HLA-B, quences were analyzed using Assign 4.7.1 (CareDX), and HLA-DRB1 haplotypes as a way to provide HLA Type Stream Visual (One Lambda), and NGSengine match for a reasonable percentage of the target population (GenDX, 2.16), depending on the used procedure. Low with a limited number of cell lines. Later, with the appear- resolution was performed using a PCR-SSO (Luminex) ance of iPSC, Nakatsuji and colleagues proposed the use based method for HLA-A, B, C, DRB1, and DQB1 genes. of banked cord blood as a source of HLA-typed cells for The expectation-maximization algorithm implemented the construction of HLA homozygous iPSC banks (haplo- in the Arlequin software (version 3.5.2.2) [25] was used banks) [14]. It was estimated that a haplobank with only to estimate maximum-likelihood haplotype frequencies, 30 iPSC lines would be able to cover 82.2% of the Japanese considering the high-resolution (4-digit) allelic frequen- population and 50 lines, 90.7% [14]. Other studies have cies of three (A, B, and DRB1) and five HLA genes (A, calculated the coverage of haplobanks for the UK popula- B, C, DRB1, and DQB1), from 30,000 and 27,000, re- tion [15], South Korea [16], China [17], and the USA [18]. spectively, randomly selected adult subjects of the iPSC haplobanks created from cord blood and peripheral REDMO project. Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 3 of 8 Screening and selection of HLA haplotype homozygous donors All the cord blood units in REDMO were studied for the identification of potential HLA-A, HLA-B, and HLA- DRB1 homozygous cord blood donors. Selection and classification of homozygous units were performed by simple counts with Microsoft Excel. Calculation of match coverage To estimate the Spanish population HLA matching coverage of a 10 iPSC haplobank, we counted the num- ber individuals with zero mismatches in HLA-A, HLA-B, and HLA-DR when compared with the top ten haplo- types selected by frequency. The population cohort was composed of the combined data of the adult bone mar- row donors and cord blood donors in the REDMO col- lection. The calculation was done either in two digits HLA typing (low plus high resolution originally) on 418.980 individuals or in four digits (only high reso- lution) on 56,798 individuals. The haplotype match benefit (coverage) in the whole sample (including adult bone marrow donors and cord blood donors) was esti- mated with an in-house, iterative algorithm in R. Briefly, in each iteration, the estimated haplotype that matched the highest number of subjects was identified. Then, the best haplotype and all matched individuals were ex- tracted from the dataset and coverage was recomputed in the remaining data. The matching was based on the concordance of the A, B, and DRB1 alleles of the esti- mated homozygous haplotypes with at least one of the two alleles of these loci in each individual. To calculate the coverage allowing for mismatch in one or two alleles (beneficial match), the same strategy was applied, but one or two mismatches in any HLA loci was tolerated. In all cases, the cumulative percentage of coverage was calculated by dividing the number of matched individ- Fig. 1 a Top 30 ranking HLA-A, HLA-B, and HLA-DRB1 estimated uals in each iteration by the total sample size, multiplied haplotypes by % of frequency at the Spanish Bone Marrow Donor by 100. Registry. b Graphic representation of the % of frequency of the top 100 HLA-A, HLA-B, and HLA-DRB1 estimated haplotypes Results Haplotype frequencies with a 27,000 cohort from the REDMO registry. Only A cohort of 30,000 randomized high-resolution HLA the top six positions corresponded with those haplotypes typing adult bone marrow donors from the REDMO identified in the three genes analysis and only four had a identified a total of 8478 different haplotypes for HLA- frequency above 1%: A*29:02~ C*16:01~B*44: A, HLA-B, and HLA-DRB1 (Fig. 1a and Supplementary 03~DRB1*07:01~DQB1*02:02 (2.95%), A*01:01~C*07: Excel file for the complete list). Five of them presented a 01~B*08:01~DRB1*03:01~DQB1*02:01 (2.52%), A*30: frequency 1% or higher, namely A*29:02~B*44: 02~C*05:01~B*18:01~DRB1*03:01~DQB1*02:01 (1.89%), 03~DRB1*07:01 (3.12%), A*01:01~B*08:01~DRB1*03:01 and A*03:01~ C*07:02~B*07:02~DRB1*15:01~DQB1*06: (2.48%), A*30:02~B*18:01~DRB1*03:01 (1.99%), A*03: 02 (1.34%) (Sup. Figure 1). 01~B*07:02~DRB1*15:01 (1.34%), and A*33:01~B*14: 02~DRB1*01:02 (1.00%). Haplotype frequencies show a Identification of homozygous donors steep decline after the first five (Fig. 1b). To identify homozygous cord blood units available in We performed the same analysis for five genes, namely the Spanish banks that can be candidates as source cells HLA-A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1 to create the iPSC haplobank, we looked into the Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 4 of 8 Spanish registry, which comprises 52,220 cord blood do- Recipients matching coverage nations in Spain. HLA was typed in low resolution (sero- We tested the first 10 most frequent haplotypes for three logical and PCR based) for 42.801 units and high genes, HLA-A, HLA-B, and HLA-DRB1, and we found resolution (NGS) for 9419 units. We identified 322 cord that the cumulative coverage was 27.84% of the popula- blood units homozygous for HLA-A, HLA-B, and HLA- tion in high resolution and 31.27% in low resolution DRB1 (0.62% of the units) representing 111 different (Fig. 3a, b). We also explored the “maximum coverage” haplotypes (Sup. Figure 2). So far, 109 of the homozy- approach for 10 haplolines selecting not the most fre- gous units were verified by high-resolution HLA typing quent haplotypes but those that would optimize the and represented 43 different haplotypes. Not to deplete coverage. The choice of haplotypes with this approach the cord blood banks of any haplotype for hematopoietic did not change for the top four positions, and the fol- stem cell transplantation, we only considered those hap- lowing six, although in a different order, only one new lotypes with two or more units banked as candidates for haplotype was considered: A*02:01~B*18:01~DRB1*03: the iPSC bank (Fig. 2). Thirty-one haplotypes were rep- 01. The cumulative coverage in high resolution was resented at least twice in low resolution and 11 in high slightly increased to 28.23% in high resolution and to resolution. The top ten positions in the most frequent 31.87% in low resolution (Fig. 3a, b). Since we did not haplotypes from the adult bone marrow donor study identify any homozygous cord blood unit in the Spanish were all represented in at least two units genotyped in banks with A*02:01~B*18:01~DRB1*03:01, we estimated low resolution and nine of them have confirmed units the optimized coverage considering only the available by high resolution. units and then the accumulated coverage in high reso- lution is 27.95% and 32.58% in low resolution. To calcu- late how many haplolines would be needed to cover close to 100% of the Spanish population, we repeated this iterative process with all the estimated haplotypes in our study population. We found that 100 haplolines would cover 65.94% of the Spanish population and that 630 haplolines would be necessary to cover 90% of the potential recipients (Fig. 3c). We also did this calculation with 5 genes: HLA-A, HLA-B, HLA-C, HLA-DRB1, and HLA-DQB1, and then the numbers shift to 100 units to cover 50% of the population and 897 units needed to cover 90% (Sup. Figure 3). Beneficial match As different iPSC-derived cell types and different en- graftment sites might present different HLA matching requirements, we introduced the approach of “beneficial match” and estimated the coverage in the case that one or two of the alleles studied did not match as the most immunogenic of the alleles (Fig. 4). The cumulative coverage for the top 10 HLA-A, HLA-B, and HLA- DRB1 haplotypes in the “maximum coverage” list showed a wide population coverage of 65.9% when allowing for one mismatch in any of the three alleles considered and a coverage of 94.81% when allowing to mismatch in two of the three alleles. Discussion Currently, clinical-grade iPSC haplobanks are already available in South Korea and Japan [19, 20, 26] and simi- lar initiatives are being undertaken in other countries Fig. 2 HLA haplotypes and number of units (N.) of HLA-A, HLA-B, such as Australia [27]. The effectiveness of the approach and HLA-DRB1 homozygous found in the Spanish registry of public will come from a global collaborative to share haplolines cord blood banks and present at least in two units. LR, low and this implies an effort to standardize the production resolution; HR, high resolution and quality control by the different banks [28, 29]. Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 5 of 8 Fig. 3 Estimated percentage and cumulative percentage of HLA-matched individuals in the Spanish population with a panel of 10 homozygous donors in high resolution (a) and low resolution (b). Estimated numbers of iPSC lines homozygous for HLA-A, HLA-B, and HLA-DRB1 (haplolines) to cover the Spanish population (c) The IPS-PANIA project aims at developing a haplo- bank that can serve a significant percentage of the Span- ish population with HLA-matching iPSC lines that can be used as starting material for iPSC-derived cell therap- ies. In the initial phase of the project, we have set a goal developing seven haplolines. A previous study of the estimated most frequent hap- lotypes in the Spanish bone marrow donor database identified five haplotypes with a frequency ranging from higher than 1% to the 6th position in the list were the same as previously described for the Barcelona cord blood bank study [24]. The differences in the lower posi- tions might reflect the higher accuracy of the larger database or geographical or generational differences. The haplotype with the top frequency is A*29:02~B*44: 03~DRB1*07:01 and the third one A*30:02~B*18: 01~DRB1*03:01 are very common in the western Medi- Fig. 4 Estimated cumulative percentage of matched individuals in terranean region. These haplotypes are less common in the Spanish population considering a “beneficial match” with a Italy and the rest of south-eastern Europe [30]. A*29: panel of 10 homozygous donors from Spanish cord blood Banks in 02~B*44:03~DRB1*07:01 is also a common haplotype in high resolution allowing for one mismatch in any of the alleles HLA- the Hispanic population in the USA, but less frequent A, HLA-B, or HLA-DRB1 (1 MM) or two mismatches (2 MM) for European ancestry and extremely rare for Asian Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 6 of 8 ancestry [17, 31]. The second and fourth most fre- determined by the type of cell and the transplantation quent haplotypes, A*01:01~B*08:01~DRB1*03:01 and sites. Studies performed in non-human primates match- A*03:01~B*07:02~DRB1*15:01 respectively, are among ing MHC antigens equivalent to HLA-A, HLA-B, and the most common in the northern and central Euro- HLA-DRB1 have revealed from mild immune infiltration pean population. From the top 10 most frequent hap- in iPSC-RPE implanted in the retina [4] to a significant lotypes in Spain, seven are in the top 30 described reaction to the allograft requiring immunosuppression for the UK [15]. As expected, they are not identified in the central nervous system [10]. Unlike hematopoietic with the most frequent haplotypes identified for progenitors and organ transplants, iPSC-derived cells Korea and Japan [14, 16]. will be free of contaminating T cells, except for a small The identification of homozygous units confirmed that possibility in the case of hematological lineage deriv- the Spanish public cord banks contained enough homo- ation, and no graft-versus-host disease is expected, mak- zygous units that could be used to build an iPSC haplo- ing sense to consider less HLA-match stringent bank to cover the top haplotypes without compromising scenarios closer to solid organ transplants like the kid- the availability of any lifesaving unit needed for ney. The population coverage when considering haplo- hematopoietic progenitor transplantation. We found a types in two-field resolution was not much increased as 0.62% of the cord blood units were homozygous, a sig- compared to four-field resolution, highlighting the pre- nificantly lower number as compared for that reported dominance of certain subgroups of haplotypes. When for the South Korean population (0.79%), which might the beneficial match was considered, as expected, the reflect the wider heterogeneity of the HLA genetics in gains were much greater when allowing for one or two the Spanish population. mismatches in any of the three alleles. The choices of The estimated population coverage of the most fre- HLA matching stringency requirements will be deter- quent haplotypes seems to also reflect a higher HLA mined by future evidence for the different cell types de- genetic variability in the Spanish population as com- rived from iPSC and the different transplantation sites. pared to other populations: the top haplotype in Spain Taken together, a 30% population match for a ten covers 6.3% and together with the second most frequent cell line haplobank is a significant proportion of po- haplotype, the cumulative coverage is 11.2%, while for tential patients who may benefit from the cell bank. Korea, the percentages are 9.2 and 14.5, and for UK, This justifies the construction of the Spanish haplo- 16.9% and 26.4%. A ten cell line haplobank would cover bank, more so when considering a worldwide effort 28% in four-field resolution and 32% in two-field reso- to share cells internationally with other banks that lution, while in UK or Japan, the reported coverages for might contain less frequent haplotypes. Also, as the ten cell lines are closer to 50% of the population. The production of clinical-grade iPSC lines is optimized “maximum coverage” approach for the selection of the and streamlined, it will become easier and more af- top ten candidates, opposed to following just the higher fordable to increase the number of haplolines in the haplotype frequency order, improved the coverage for bank to reach several dozens and cover a much wider the lower positions but the effect was almost lost when proportion of the population. With this study, we the ten haplolines were considered. When calculating have investigated the feasibility to provide homozy- how many haplotypes would be needed for almost gous cord blood units to create an iPSC bank of a re- complete coverage of the Spanish population, we found duced number of haplolines that will serve a that 631 would be needed to cover 90%, again revealing significant percentage of the Spanish and international a much wider variety of existing haplotypes as compared population. Besides RPE cells for the treatment of to other populations analyzed in other studies, such as AMD, several other iPSC-derived cells are presently Korea that found 90% covered with less than 200 cell being tested in clinical trials to treat conditions such lines [16]. as spinal cord injury, Parkinson’s disease, graft-versus- Wider coverage of iPSC haplolines by lowering the host-disease, heart failure, or cancer. Clinical-grade HLA stringency might be worth consideration for an iPSC are intended to be used as starting material for iPSC bank. It is still early days to know what the real future clinical trials and cell therapy products, accel- HLA matching requirements of iPSC derived products erating the application of iPSC-based therapies soon. will be as the first clinical trial carried out at the Kobe City Medical Center in collaboration with Osaka Conclusion University, using allogeneic iPSC-derived retinal pigmen- With the present study, we have been able to confirm ted epithelial cells of a HLA-A, HLA-B, HLA-DRB1- that HLA homozygous cord blood units stored in Span- matching haplotype has not been reported yet. It is clear ish cord blood banks can provide for the construction of that HLA matching is advantageous and will reduce the an iPSC bank that is useful for a significant part of the degree of immunosuppression although this will be Spanish population (about 28%). The haplotypes Álvarez-Palomo et al. Stem Cell Research & Therapy (2021) 12:233 Page 7 of 8 identified as providing the greatest coverage, may also be Competing interests useful for other populations such as Europe and North The authors declare no competing interests. America. Author details 1 Banc de Sang i Teixits, Edifici Dr. Frederic Duran i Jordà, Passeig Taulat 116, Abbreviations 08005 Barcelona, Spain. 2Musculoskeletal Tissue Engineering Group, Vall AMD: Age-related macular degeneration; HLA: Human leukocyte antigen; d’Hebron Research Institute (VHIR), Passeig de la Vall d’Hebron 129-139, iPSC: Induced pluripotent stem cells; NGS: Next Generation Sequencing; 08035 Barcelona, Spain. 3Organización Nacional de Trasplantes, Ministerio de REDMO: Spanish Bone Marrow Donor Registry; RPE: Retinal pigmented Sanidad, C/ Sinesio Delgado 6 - Pabellón 3, 28029 Madrid, Spain. 4Programa epithelial cells de Medicina Regenerativa, Institut d’Investigació Biomèdica de Bellvitge. IDIBELL, Hospital Duran i Reynals, Gran Via de l’Hospitalet, 199-203, 08908 L’Hospitalet de Llobregat, Barcelona, Spain. 5Axencia Galega de Sangue, Supplementary Information Órganos e Tecidos, Rúa Xoaquín Díaz de Rábago 2, 15705 Santiago, Spain. The online version contains supplementary material available at https://doi. 6 Centro de Transfusión de la Comunidad Valenciana, Av. del Cid, 65-acc, org/10.1186/s13287-021-02301-0. 46014 Valencia, Spain. 7Basque Center for Blood Transfusion and Human Tissues, Barrio Labeaga, s/n, 48960 Galdakao, Spain. 8Cell Therapy, Stem Cells Additional file 1: Supplementary Figure 1. Top 30 ranking HLA-A, -B, and Tissues Group, Biocruces Bizkaia Health Research Institute, Células Madre -C, -DRB1 and -DQB1 estimated haplotypes by % of frequency at the y Tejidos, Cruces Plaza, 48903 Barakaldo, Spain. 9Centro de Transfusión de la Spanish Bone Marrow Donor Registry. Comunidad de Madrid, Avda. de la Democracia, s/n, 28032 Madrid, Spain. 10 Centro de Transfusión, Tejidos y Células de Málaga, Avda. Doctor Gálvez Additional file 2: Supplementary Figure 2. HLA types and number of Ginachero s/n, 29009 Málaga, Spain. 11REDMO / Fundació i Institut de units (N.) of HLA-A, HLA-B and HLA-DRB1 homozygous found in the Recerca Josep Carreras, C/Muntaner, 383 2n, 08021 Barcelona, Spain. Spanish registry of public cord blood banks. 12 Transfusional Medicine, Vall d’Hebron Research Institute, Universitat Additional file 3: Supplementary Figure 3. Estimated numbers of Autònoma de Barcelona (VHIR-UAB), Barcelona, Spain. 13CIBER de iPSC lines homozygous for HLA-A, -C, -B, -DRB1 and -DQB1 (haplolines) to Enfermedades Cardiovasculares (CIBERCV), Madrid, Spain. 14Royal Free cover the Spanish population. Hospital, Pond Street, Hampstead NW3 2QG, UK. 15UCL Cancer Institute, Additional file 4. Medical School, 74 Huntley St, Bloomsbury, London WC1E 6DE, UK. Received: 30 July 2020 Accepted: 22 March 2021 Acknowledgements We would like to acknowledge Dr. Emma Enrich Randé for discussion of the results, Mrs. Elisabet Tahull Navarro for administrative and management help, References and Dr. Michael J. Edel for proof reading. We would also like to thank the 1. Robinton DA, Daley GQ. The promise of induced pluripotent stem cells in cord blood donors, the donation programs professionals, and all the cord research and therapy. Nature. 2012;481(7381):295–305. https://doi.org/10.103 blood banks staff for their invaluable dedication to helping bring hope to 8/nature10761 Review. many patients in need. 2. Eguizabal C, Aran B, Chuva de Sousa Lopes SM, et al. Two decades of embryonic stem cells: a historical overview. Hum Reprod Open. 2019; Authors’ contributions 2019(1):hoy024. https://doi.org/10.1093/hropen/hoy024 eCollection 2019. BAP has participated in the design of the work; the acquisition, analysis, and 3. 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