백혈병에 골수대신 제대혈 줄기세포를 배양하여 이식?

골수이식을 대체할 수단으로 제대혈 줄기세포 배양을 주장하는 회사들이 많지요.

백혈병에 골수이식을 대신해서 더 좋은 결과를 얻을 수 있다고 합니다.

StemEx라는 제대혈 확장기를 개발한 회사도 그 중하나로,
이미 동물실험은 성공했다고 주장합니다.

그런데, 아마도 동물실험은 생생한 제대혈로 했을 것이고,
사람의 경우에는 얼린 것을 쓸 수 밖에 없으니까,
동물실험하고 같은 결과가 나올까요?

한번 얼린 줄기세포를 배양한다는 것이.... 쉽지는 않지요.
또한, HSC를 배양에 성공했다는 거 같은데? 정말일까요?

정말, 동물에서 제대혈의 HSC를 배양하는 데 성공했다면,
사람에서도 희망은 보입니다.
이 회사 주장대로라면, 현재 미국, 유럽에서 3상 임상실험중이랍니다.


아이를 낳는 병원에서 회사로 질소가스로 냉동해서 옮기지 말고,
그 자리에서 바로 충분한 양을 배양해서 , 그 배양한 것을
아주 천천히 얼려서, 세포가 최대한 보존되도록 한다면,
나중에 백혈병에 쓸 수 있을 것으로 보입니다.

관련 기사
관련 회사
관련회사2

관련 논문
 725] Nicotinamide Modulates Ex-Vivo Expansion of Cord Blood Derived CD34+ Cells Cultured with Cytokines and Promotes Their Homing and Engraftment in SCID Mice. Session Type: Oral Session, ASH December 12, 2006 Tony Peled, Sophie Adi, Iddo Peleg, Noga G. Rosenheimer, Yaron Daniely, Arnon Nagler, Eitan Fibach, Amnon Peled R&D, Gamida Cell Ltd, Jerusalem, Israel; Hematology, Chaim Sheba Medical Center, Tel Hashomer, Israel; Hematology, Hadassah-Hebrew University Medical Center, Jerusalem, Israel Nicotinamide (NA) is a non-competitive inhibitor of NAD(+)-dependent ADP-ribosyl transferases, of CD38 NADase (a major regulator of cellular NAD levels) and of Sir2 histone-deacetylase. These enzymes are playing a pivotal role in regulation of signal transduction pathways and gene expression. In the present study, we evaluated the effect of NA on the ex-vivo expansion of cord-blood (CB) derived CD34+ cells and their bone-marrow (BM) homing and engraftment potential. Culturing of CD34+ cells for three weeks in the presence of cytokines (SCF, TPO, IL-6, FLT3-ligand) only or cytokines + NA (5mM) resulted in similar expansion of CD34+ cells (40-fold relative to input). However, a remarkable increase in the fraction of CD34+ cells displaying an early progenitor cell phenotype (CD34+Lin-) was observed in the NA-treated cultures as compared with cytokines-only treated cultures (18.6±3% and 0.7±0.06%, n=6, p<0.05, respectively). Tracking the cell-cycle history by PKH2 staining showed fewer division cycles of CD34+ cells cultured with NA. These results may suggest a direct correlation between the rate of proliferation and expansion of CD34+Lin- cells. NA-treated CD34+ cells express similar levels of CXCR4 but display increased migratory activity in response to CXCL12 over CD34+ cells treated with cytokines only (36±19% and 11±4%, n=4, p<0.05, respectively). In order to test their homing potential, similar number of mononuclear cells (MNC), before or following expansion with or without NA, were labeled with CFSE and transplanted into irradiated NOD/SCID mice. Twenty-four hours later the numbers of human cells (CD45+CFSE+) and human progenitor cells (CD34+CFSE+) in the BM were counted. Homing of CD45+CFSE+ cells was comparable in the three groups tested. However, CD34+CFSE+ cells with BM homing potential were 3-fold more numerous in NA-treated cultures relative to cytokines-treated cultures, and 6-fold more than in non-cultured CB cells (n=14, p<0.05). To evaluate engraftment, SCID mice were transplanted with 3x103, 6x103 and 12x103 non-cultured CD34+ cells or their entire progeny
following 3-week expansion with cytokines only or cytokines + NA (n = 63). The frequency of SCID repopulating cells (SRC) was estimated by limiting dilution analysis as 1/ 36,756 (non-cultured), 1/19,982 (cytokines), 1/ 2,620 (NA) (SCID engraftment was considered as ≥0.5% human CD45+ cells). We found that, in correlation with homing, NA-treated cells have a 14- and 7.6-fold more SRC than non-cultured cells or cytokine-treated cells, respectively. The marked increase in SCID engraftment potential following culturing with NA may be attributed to both improved homing of CD34+ cells as well as higher proportion of early progenitor cells within the CD34+ cell compartment. Despite their numerical expansion, progenitor cells generated in cytokine-supplemented cultures have reduced homing and engraftment capacity. Our study demonstrates that NA modulates in-vitro expansion and augments the in-vivo homing and engraftment of CB-derived CD34+ cells cultured with cytokines. Abstract #725 appears in Blood, Volume 108, issue 11, November 16, 2006 Keywords: Engraftment|Ex vivo expansion|Homing

CD34 세포에 cytokine과 동시에  Nicotinamide를 가했더니 cytokine만 준 그룹은 물론, 배양을 안한 그룹보다
생착률과  homing율이 올라갔다고 합니다. 그것도 10배 이상!!!

또한 이렇게 배양을 하니까 early progenitor의 비율도 증가했답니다.
즉, 또리또리한 세포들로만 많이 배양이 되었다는 것입니다.

이 원리를 이용하여 gamida cell에서는 StemEx라는 CD34 세포 배양기기를 만들어 냈답니다.
이것을 이용하여, 제대혈의 HSC를 배양하여, 백혈병에 사용하겠다는 것입니다.
이론적으로는 , 배양 안하는 기존 골수이식보다 생착율이 올라갈 것이고,
 면역관용이 더 많으므로, 더 많은 사람들에게 쓰일 수 있을 것입니다.

정확히,


역시 궁금한것은,
냉동했던  제대혈의  HSC도 같은정도로 배양, 생착이 될까요?
면역 관용의 정도는 얼마만큼까지 일까요?

또한, MSC도 이러한 기기로 배양하면 더 생착율을 올릴 수 있지 않을까요?

이 회사가 홈페이지에 공개한 기술입니다.

Gamida Cell employs proprietary technologies to expand populations of functional hematopoietic progenitor cells (HPC).  These epigenetic technologies utilize small molecules to modulate differentiation, homing and engrafment abilities of cultured cells.  The Phase I/II study of the first developed technology showed preliminarily positive results which prompted a subsequent global, multi-center, pivotal phase III study to further investigate safety and efficacy.  www.clinicaltrials.gov

Successful ex vivo expansion of HPC depends on their ability to proliferate while maintaining their basic characteristics. Most protocols for ex vivo expansion utilize various combinations of cytokines that support extensive proliferation of cultured cells. Nonetheless, in vitro proliferation is tightly coupled with commitment and differentiation, thereby lessening the clinical utility of cultured cells. Thus, culture conditions attenuating in vitro differentiation were suggested to support more successful expansion of HPC and to increase their clinical applicability.

Gamida Cell has developed four proprietary technologies for ex vivo expansion of HPC utilizing low molecular weight compounds, which regulate physiological pathways of cell differentiation. Two of the four technologies have been presented in scientific forums. Gamida Cell is applying these expansion technologies to develop a pipeline of products for bone marrow transplantation and tissue regeneration.

Copper Chelator Based Technology

The first technology developed by Gamida Cell is based on the discovery that copper ions modulate self-renewal and differentiation of cultured hematopoietic progenitor cells (HPC). Copper deficiency delays differentiation and prolongs proliferation while increase in cellular copper level, accelerates differentiation and substantially reduce proliferation potential. (Exp Hematol. 2005; 33:1092.). Intriguing, copper deficiency in patients was shown to mimic refractory anemia manifested by excess of immature cells and reduced proportion of differentiated cells in the bone marrow. Oral copper replacement completely normalized the bone marrow findings (Leuk Res. 2008; 32(3):495). Therefore, this mechanism is used to modify the balance between self renewal and differentiation in vitro and in vivo.

The lead molecule of the copper-based technology is the high affinity copper chelator – Tetraethylpentamine (TEPA).  TEPA reduces the intracellular copper level, delays differentiation, and enables robust expansion of HPC during the first few weeks in culture. The resulting cell population contains increased proportions and absolute numbers of HPC displaying increased self-renewal potential, and shows high levels of engraftment as well as multi-lineage differentiation potential in pre-clinical in vivo models (Exp Hematol. 2004 Jun;32(6):547-55.)

즉, 다음의 내용이 중요하답니다.

Most importantly, thepercentage of engrafted human progenitor cells as well as
that of myeloid and lymphoid cells was reproducibly and significantly superior in SCID mice transplanted with TEPA-mediated ex vivo expanded cells than mice transplanted with similar numbers of cells expanded with the cytokine cocktail without TEPA, or with the corresponding cell fraction before expansion.

NAM Based Technology

Ongoing pre-clinical studies have enabled Gamida Cell to develop an additional propriety technology based on epigenetic modulation of NAD+ -dependent ADP ribosyl transferase enzymes. Nicotinamide (NAM), a form of vitamin B3, shown to be the most potent inhibitor of NAD+ -dependent ADP ribosyl transferase enzymes, is the lead molecule of this novel technology. NAM increases the therapeutic potential of stem cells. The pre-clinical results show the tremendous potential of NAM technology to improve the homing and rate of engraftment of cord blood derived stem cells to maximize their full therapeutic potential in bone marrow transplantation and tissue regeneration. In December 2006, Gamida Cell addressed the American Society of Hematology in an oral presentation on the recent results of a pre-clinical  study of its NAM technology.  Currently, the company is focused on applying these technologies in development of new products.

 

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