dyskeratosis congenita fibroblasts are abnormal and have - blood

Dyskeratosis Congenita Fibroblasts Are Abnormal and Have Unbalanced Chromosomal Rearrangements

By lnderjeet Dokal, Julie Bungey, Peter Williamson, David Oscier, Jill Hows, and Lucio Luzzatto

Dyskeratosis congenita (DC) is a rare inherited disorder characterized by bone marrow failure, dystrophic changes in the skin and mucous membranes, and a predisposition to malignancy. The DC locus has been mapped to Xq28. The primary defect responsible for this disease remains unknown. We have studied four patients with this disease, three from one family and one from another. In all four patients, primary skin fibroblast cultures were abnormal both in morphology (polygonal cell shape, ballooning, and dendritic-like projections) and in growth rate (doubling time about twice normal). Fibroblast survival studies using four clastogens (bleomycin, diepoxybutane, mitomycin-c, and 4-nitroquinoline-I-oxide) and gamma radiation showed no significant difference between DC and normal fibroblasts. Cytoge-

YSKERATOSIS congenita (DC) is a rare inherited D disorder that, like Fanconi’s anemia (FA), is characterized by bone marrow (BM) failure and a predisposition to malignancy, but differs from FA in other clinical features such as reticulate skin hyperpigmentation, mucosal leuco- plakia, and nail Although X-linked recessive inheritance is believed to apply to the majority of families, some families have been described in whom autosomal inheritence was likely. Linkage in one large family by using X chromosome-specific restriction fragment length polymor- phism (RFLP) markers has assigned the gene for DC to Xq2K6 The primary defect(s) responsible for this disease remains unknown.

Spontaneous chromosome instability has been reported in some studies of DC,7-9 but not in ~ t h e r s . ~ J ~ - l ~ Excessive spontaneous13 and cla~togen-induced~~ sister chromatid exchange (SCE) have also been reported in DC lymphocytes. However, this finding has not been confirmed in other ~ t u d i e s . ~ ~ J ~

Previous clastogenic stress studies of DC cells (lymphocytes and fibroblasts) that have used the DNA alkylating agents mitomycin-c and diepoxybutane have not shown any difference between DC and normal Excessive chromosome breakage was induced by 4-nitroquinoline-l- oxide in the lymphocytes of one DC patient,” but not by other clastogens such as mitomycin-c. Excessive chromatid damage in DC fibroblast cell lines exposed to X-irradiation during the G2 phase of the cell cycle has been reported by

From the Department of Haematology, Royal Postgraduate Medical School and Hammersmith Hospital, London; and the Department of Haematology, Royal Victoria Hospital, Boumemouth, UK.

Submitted May 11,1992; acceptedAugust 17, 1992. I.D. was supported by the Medical Research Council. Address reprint requests to Lucio Luzzatto, MD, Department of

Haematologv, Royal Postgraduate Medical School and Hammersmith Hospital, 150 Du Cane Rd, London W12 ONN, UK.

The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. section I734 solely to indicate this fact.

0 I992 by The American Society of Hematology. 0006-4971 l92/8012-0009$3.00 10

netic studies performed on peripheral blood lymphocytes showed no difference between DC and normal lymphocytes with or without prior incubation with clastogens. However, bone marrow metaphases from one of three patients and fibroblasts from two of four patients (who were the eldest of the 4) showed numerous unbalanced chromosomal rearrangements (dicentrics, tricentrics, and translocations) in the absence of any clastogenic agents. Cell-specific differences and a higher rate of chromosomal rearrangements in the older patients appear to correlate with the clinical evolution of the disease. These findings suggest that the DC defect predisposes DC cells to developing chromosomal rearrangements. 0 1992 by The American Society of Hematology.

DeBauche et aI18; and Pai et all2 found that DC cells (fibroblasts and lymphocytes) were hypersensitive to bleomycin.

In light of the inconclusive reports in the literature regarding the spontaneous chromosomal instability and response of DC cells to clastogens, we have undertaken cytogenetic and survival studies on cells from DC patients with and without prior incubation with clastogens. The work was prompted initially by the need to design a BM transplant (BMT) protocol for the index case. We found that the DC defect predisposes DC cells to developing chromosomal rearrangements in the absence of exposure to clastogens, and that fibroblasts from all patients were abnormal.

MATERIALS AND METHODS

Cell Lines and Culture Conditions

Primary skin fibroblast cultures were established from punch biopsy specimens from four DC patients (3 from family A and 1 from family B) and four non-DC members of family A. Fibroblasts were grown at 37°C in a 5% carbon dioxide environment in Dulbecco’s Modified Essential Medium (DMEM) supplemented with 20% fetal calf serum. Two Simian virus (SV)-40 immortalized fibroblast lines (GM6914I9 and Ch RuZ0) established from patients with Fanconi’s anemia were obtained from Dr Duckworth-Rysiecki and Dr Chaganti, respectively.

Fibroblast Survival Studies

Fibroblast survival studies using clastogens (bleomycin, diepoxybutane, mitomycin-c, and 4-nitroquinoline-1-oxide) and gamma radiation were performed on early passage fibroblasts. Skin fibroblast cultures were subcultured in 24-well plates containing growth medium. Twenty-four hours later, when they were in the logarith- mic growth phase, they were exposed to the appropriate concentra- tion of clastogen either as a pulse (bleomycin and diepoxybutane) or continuously (mitomycin-c and 4-nitroquinoline-1-oxide). The cells were then washed twice with phosphate-buffered saline and reincubated in fresh medium for a further 10 days. Fibroblasts were then harvested with trypsin and stained with trypan blue. Viable cells (unstained cells) were counted using a hemocytometer.

Cytogenetic Studies

without prior exposure to clastogens. Chromosomal analyses were performed on BM and fibroblasts

3090 Blood, Vol80, No 12 (December 15). 1992: pp 3090-3096

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DC FIBROBLASTS ARE ABNORMAL 3091

Fbl . flbroMntmorphology. (A) Flbroblasts from a normal control (original magnlfication ~400). (B) Fibroblasta from a patient witt, DC (original magnHiw- tion x400) showing abnormal morphology (polygonal shape, ballooning, and dendritic-like projections).

Cytogenetic studies on peripheral blood (PB) lymphocytes were performed with and without prior exposure to clastogens.2'

RFLP Genetic Linkage Study An RFLP genetic linkage study was undertaken in family A

using the DXSS2(St14-I) probe and Tu9 I digestion as described by Connor et aLL

Patients

Fami!y A Patient I (the index case in family A) first presented

at the age of 29 years in August I989 with symptoms of anemia. He Patient 1.

was found to have oral leukoplakia, nail dystrophy. and skin pigmentation. Investigations showed he had severe pancytopenia (hemoglobin [Hb]. 3.2 gldL: white blood cell count [WBC]. 1.2 X

lO"/L; platelets, 6 x lO"/L). A hiopsyof the RM showed that i t was very hypocellular, with approximately IOr& myeloblasts. I n January 1990. he was referred to the Hammersmith tlospital for matched unrelated donor (MUD) RMT. at which time his HLA-identical younger brother (patient 2) was also diagnosed to have DC. Recause the fibroblast survival studies had shown that his cells were not hypersensitive to clastogens, he was conditioned with the same dose (SO mglkg x 4) of cyclophosphamide as that used for patients with acquired aplastic anemia rather than the low d m (5 mglkg x 4) used in FA and total body irradiation. He did not develop severe mucosal toxicity, but failed to engraft. As a salvage




3092 DOKAL ET AL

procedure, he received a second BMT from his younger brother (patient 2), who had a normal blood count but poor in vitro growth in long-term BM cultures.22 He showed signs of early PB engraft- ment, but died from respiratory failure 3 weeks after his second BMT (March 21, 1990). The cause of the respiratory failure remained unknown. No pathogen was isolated from the bronchial lavage and he had no evidence of graft-versus-host disease (GVHD) or mucosal toxicity. It is possible that the underlying DC may have predisposed this patient to developing pulmonary complications posttransplant.’

Patient 2 was diagnosed when he was 26 years old while he was being assessed as a potential BM donor for his elder brother (patient 1). Like his brother, he had nail dystrophy, skin pigmentation, and oral leukoplakia. He was asymptomatic with a normal PB count. However, his BM culture studies showed a significant reduction in hematopoietic progenitors and long-term BM culture studies suggested a functional stem cell defect with normal stromal functionJ2 These studies suggest that he may develop severe BM failure in the future similar to that in his elder brother (patient 1).

Patient 3 is the nephew of patients 1 and 2. He was in- vestigated at the age of 12 years because the RFLP genetic linkage study had shown that his mother (the younger sister of patient 1) was a carrier, as she had inherited the same allele (a 4.5-kb Taq I fragment identified with St14-1 probe) as that segregating with DC (see Fig 4). Further investigations showed that patient 3 had inherited this allele (4.5 kb) from his mother and had early changes of nail dystrophy and skin pigmentation. He was otherwise clini- cally asymptomatic and had a normal PB count. However, his BM culture studies showed a significant reduction in hematopoietic progenitors: granulocyte-macrophage colony-forming units (CFU- GM) were 16.4 per lo5 mononuclear cells plated (normal range, 40 to 240); burst-forming units-erythroid (BFU-E) were 4.5 per lo5 mononuclear cells plated (normal range, 30 to 330).

Fami& B Patient 4 was referred to the Hammersmith Hospital

at the age of 15 years in May 1990 for consideration for MUD BMT .because of a deterioration in his blood count (WBC, 2 x 109/L; platelets, 30 x 109/L). His BM cultures showed a significant reduction in hematopoietic progenitors: CFU-GM were 5.0 per 1@ mononuclear cells plated; BFU-E were 7.5 per 1@ mononuclear cells plated. Similar to the three patients in family A, he had nail dystrophy, leukoplakia, and skin pigmentation. In addition, he also had an esophageal stricture, partial deafness, hypogonadism, and

Patient 2.

Putient3.

Patient 4.

A

- a 2 - I

100 r, Patient 1 (JB) - patient 4 (LC) 1 Normal-father 80

Mother - E.slster I Y.Slster - Fanconl Ch Ru

60

40

20

0 0 2 4 6 8 1 0

Diepoxybutane pglml (1 hour pulse)

mild mental retardation. No suitable MUD donor was found and he died in July 1991.

RESULTS

Fibroblast Morphology

In all four patients, primary skin fibroblasts were abnormal in morphology (polygonal cell shape, ballooning, and dendritic-like projections; Fig 1A and B). The different fibroblast cultures were of similar ages because the skin biopsies from which they were established were all taken simultaneously. The growth rate was also abnormal for all DC fibroblasts. The doubling time for normal fibroblasts was 2 to 3 days, whereas for DC fibroblasts it was 6 to 10 days. In one obligate carrier (the mother of patients 1 and 2), the fibroblasts looked normal and the doubling time was 2 days (ie, indistinguishable from normal).

Sensitivity to Clastogenic Agents

Fibroblast survival studies using bleomycin (0 to 120 kg/mL for 2 hours), diepoxybutane (0 to 10 kg/mL for 1 hour), mitomycin-c (0 to 25 nmol/L continuously administered), 4-nitroquinoline-1-oxide (0 to 25 nmol/L continuously administered), and gamma radiation (0 to 250 rad as a pulse) showed no significant difference between DC and normal fibroblasts (Fig 2). This is in sharp contrast to FA fibroblasts, which, as expected, were hypersensitive to diepoxybutane (Fig 2A) and mitomycin-c (Fig 2B).

Cytogenetic Analyses

PB PB lymphocytes showed no difference (gaps or breaks)

between DC and normal lymphocytes with or without prior incubation with diepoxybutane (0.1 p,g/mL for 48 hours), mitomycin-c (5 pg/mL for 72 hours), or 4-nitroquinoline-l- oxide mol/L for 48 hours). This is in sharp contrast to cells from patients with FA, which show increased chromosomal gaps and breaks upon exposure to clastogens.

In one of four patients (the eldest of the 4), a triradial configuration (Fig 3A) was seen in 1 of 30 metaphases without prior exposure to clastogens.

100

80

60

40

- Patient 4 (LC) 1 Father-normal - Normal (GPD)

I Fanconl GM69

2 0 1 \ 3

0 0 1 0 2 0 3 0

nM MMC (continuous)

Fig 2. Sensitivity to ciastogen agents. (A) Diepoxybutane survival curves showing no difference in sensitivity between DC and normal fibroblasts. The Fanconi fibroblast line (Ch Ru) is hypersensitive to diepoxybutane compared with normal fibroblasts. (6) Mitomycin-c survival curves showing no difference in sensitivity between DC and normal fibroblasts. The Fanconi fibroblast line (GM6914) is hypersensitive to mitomycin-c compared with normal fibroblasts.




DC FIBROBLASTS ARE ABNORMAL 3093

RM BM from one of thrcc paticnts (the cldcst of the 4)

showcd (Fig 3R) numcrous unbalanced chromosomal rcar- rangcments without prior incubation with any clastogcns.

Fibroblasts Fibroblasts from two of four paticnts (thc 2 cldcst

patients) showcd a varicty of unbalanced chromosomal rcarrangcmcnts (dicentrics, tricentrics, and translocations) in thc abscncc of any clastogcns. Fibroblasts from thc othcr two paticnts showcd no chromosomal rcarrangcmcnts. The typc of abnormalities seen in the two paticnts (paticnts 1

and 2) arc shown in Fig 3C and listed in Table 1. In addition, inversion 8 [inv(X)(pl1.2q22)] was sccn in all the mctaphascs from patient 2. This abnormality was also obscrvcd in his mothcr and, thcrcforc, wc bclicvc it is unrclatcd to DC.

RFLP Linkage Analysis A linkage study was undcrtakcn in family A bccausc thc

two sistcrs of paticnts 1 and 2 wishcd to know whethcr they were carriers of thc DC allclc. Bccausc Connor et al" havc shown no recombination bctwccn the DC gene and the RFLPs idcntificd by St14-I in nine informativc meiotic cvcnts (maximal Lod scorc of 3.33 at zcro rccombination

ma. C y t ~ 0 b n o f m . C ltks. (A) Metaphase rpreod mado from PB of the eldoat potient (patknt 1) showing a triradial configuration. (B) Metaphase rpread made from the EM of patient 1 rhowing a ring chromosome.




3094 DOKAL ET AL

t r ic

die

a 9 I 2

9 I2

Fig 3. (Cont'd) (C) Partial kayotype showing chromosomal mar- mngements IdentMed In fibroblast culture from patient l. Two types of dicentric chromosomes and one hicentrlc resulting from rearrangements between chromosomes 9 and 12 are shown.

fraction), wc uscd this probc in family A. In our study, we found that thc fragmcnts idcntificd by thc St14-1 probc after Tuq I digcstion of gcnomic DNA wcrc informativc (Fig 4). Thc mothcr of paticnt 1 was found to havc thc 4.5-kb and 4.8-kb fragmcnts. Both paticnts 1 and 2 had inhcritcd thc 4.5-kb fragmcnt. This mcant that thc DC allclc was scgrcgating with the 4.5-kb fragment. Thc cldcr sistcr was found to havc inhcritcd thc 4.8-kb fragment and thc youngcr sistcr thc 4.5-kb fragmcnt from thc mothcr. This mcant (assuming tcro rccomhination) that thc cldcr sistcr was normal and thc youngcr was a carricr. This prompted us to study thc 12-ycar-old son (paticnt 3) of thc youngcr sistcr. Hc was found to havc inhcritcd thc 4.5-kb fragmcnt and thcrcfore (assuming zcro rccomhination) thc DC allclc from his mothcr. This was confrmcd on clinical cxamination, as hc had carly changcs of nail dystrophy and skin pigmcntation. Thc RFLP study was cxtcndcd to includc othcr mcmbcrs of family A and this allowcd us to idcntify furthcr carricrs (Fig 4) in this family. Unfortu- natcly, in gcncration IV, thc two fcmalc carricrs arc hoth homozygous for thc 4.5-kb fragmcnt. This makcs it impossiblc to say whcthcr thcir malc childrcn (gcncration V) arc normal or havc DC.

DISCUSSION

PB lymphocytcs from al l DC paticnts (and obligatc carricrs) did not cxhibit incrcascd chromosomal brcakagc with or without prior incubation with clastogcns. In addition, fibroblasts from all four paticnts wcrc not hypcrscnsi- tivc to blcomycin, dicpoxybutanc, mitomycin-c, 4-nitroquinolinc-I-oxidc, and gamma radiation. This confirms the findings of somc prcvious studics.ii.i?.".i" Howcvcr. somc studics havc shown incrcascd scnsitivity of DC cclls to ionising radiationix or to 4-nitroquinolinc-l-oxidci- and bleomycin.l? Thcsc discordant findings rcgarding thc rc- sponsc of DC cclls to clastogcns and radiation may rcsult from gcnctic hctcrogcncity in thc discasc. Thcrcforc, in a given paticnt it is impossiblc to prcdict thc sensitivity to clastogcns and whcthcr a paticnt is to rcccivc high-dosc chcmoradiothcrapy hcforc a BMT. It is important to cstablish thc scnsitivity of that individual's cclls to clastogcns to cnsurc that thc appropriatc dosc of chcmothcrapy is administcrcd. In paticnt I, thc fibroblast survival studics and thc lymphocyte cytogcnctic studics had shown normal scnsitivity to clastogcns. Hc was thcrcforc trcatcd with thc same dosc of cyclophosphamidc as that uscd for paticnts with acquircd aplastic ancmia rathcr than low-dosc cyclophosphamidc uscd in FA paticnts. This paticnt did not dcvclop scvcrc mucosal scnsitivity posttransplant and, dc- spitc high-dosc cyclophosphamidc, rcjcctcd thc first graft.

Table 1. Chromosomal Rearrangements Soen in Patients 1 and 2 in the Absence of Clastogens

Metaphases Examined

Fibroblasta PB BM

Patient 1 30 1 triradial

2946, XY

Patient 2 30 30 46.XY

inversion 8.

12 5 46,XY. der(22) 1

11;22)(q11;p11) 246,XY. -18. + R 5 46,XY

15 15 46,XY

inversion 8'

10 1 dicentric (9;12)

1 dicentric (9;12), 12p+ 112p+ 1 dicentric (12;22) 1 isochromosome (12q). 9q+ 1 dicentric (12;?:12),9q+,

1 Robertsonian translocation (14;15)

1 tricentric isochromosome (12:9;9:121 + isochromosome (9p)

1 translocation (9;12), 12p+, Robertsonian translocation (22;22)

1 46, XY

9q +

10 4 inversion 8' 4 inversion 8'. translocation

2 inversion 8'. der l l l ) t (1;ll) 3;12

Inversion 8. [inv(E)(pl1.2q22)1 was seen in a11 metaphases from patient 2. This was also observed in the patienrs mother, and we therefore infer that it is constitutional and unrelated to DC.

Abbreviation: R, ring chromosome.




DC FIBROBLASTS ARE ABNORMAL

4.5 4.1 4.8

I

4.5

3095

4.5 4.1

4.5 4.5 3.9

I V

V

Fig 4. Pedigree of family A. The RFLP analysis for the individuals studied using the St14-1 probe and Taq I digestion are indicated. (0) Female carriers; (W) male affected members. The index case in this family is indicated by the arrow. He was found to have inherited the 4.5-kb fragment from his mother. The disease was therefore segregating with this 4.5-kb fragment. This RFLP analysis was informative up to generation V and allowed us to identify female carriers and one individual (patient 3) with DC before he had presented with clinical abnormalities. Because the mother of the boy (age 2 years at time of analysis) in generation V was homozygous for the 4.5-kb fragment, it was not possible to say whether this boy had DC.

suggesting the presence of residual, functional host lymphocytes after therapy. In this case, therefore, the in vitro studies correlated well with the in vivo observation.

Fibroblasts from all four patients were abnormal in both morphology and growth rate. Furthermore, fibroblasts from two of the four patients (the 2 older patients) showed numerous unbalanced chromosomal rearrangements in the absence of any clastogens. The elder of these two patients (patient 1) also had similar chromosomal abnormalities (but less numerous) in the blood and BM, with morphologic evidence for leukemic transformation. The demonstration of unbalanced chromosomal rearrangements in the PB, BM, and fibroblasts in the oldest patient and in the fibroblasts alone in the second oldest patient suggests a primary defect that predisposes DC cells to developing chromosomal rearrangements. These findings of cell- specific differences and of a high rate of chromosomal rearrangements in the two older patients appears to correlate with the evolution of the disease in vivo.

In our linkage study, we found that the fragments

identified by the St14-1 probe after Tag I digestion of genomic DNA were informative and allowed us to correctly identify carriers and patient 3 in family A. Thus, this study confirms the findings of Connor et a16 and strengthens the linkage between the DC locus and the RFLPs identified by St14-1. Furthermore, this RFLP linkage analysis clearly shows that, in family A, DC is segregating as an X-linked recessive trait.

Although patients with FA and DC share some features in common (eg, BM aplasia), they appear to differ in two fundamental ways. Firstly, unlike Fanconi cells, in our four patients, DC cells were not hypersensitive to clastogens. Secondly, the primary defect in DC appears to predispose cells to developing chromosomal rearrangements rather than the chromosomal gaps and breaks seen in FA.

ACKNOWLEDGMENT

We are very grateful to M. Fitchett and Dr M. Crocker for performing extensive cytogenetic work on two of the patients.

REFERENCES 1. Zinsser F: Atrophia cutis reticularis cum pigmentatione,

dystrophia unguium et leukoplakia oris. Ikonogr Dermatol (Hyoto) 5:219, 1906

2. Engmann MF: A unique case of reticular pigmentation of the skin with atrophy. Arch Dermatol Syph 13:685,1926

3. Cole HN, Rauschkolb JC, Toomey J: Dyskeratosis congenita with pigmentation, dystrophia unguis and leukokeratosis oris. Arch Dermatol Syph 21:71,1930

4. Sirinavin C, Trowbridge AA: Dyskeratosis congenita: Clinical features and genetic aspects: Report of a family and review of the literature. J Med Genet 12:339, 1975

5. Davidson HR, Connor JM: Dyskeratosis congenita. J Med Genet 255343,1988

6. Connor JM, Gatherer D, Gray FC, Pirrit LA, Atfara N A Assignment of the gene for dyskeratosis congenita to Xq28. Hum Genet 72:348,1986




3096 DOKAL ET AL

7. Scoggins RB, Prescott KJ, Asher GH, Blaylock WK, Bright RW: Dyskeratosis congenita with Fanconi-type anemia: Investiga- tions of immunologic and other defects. Clin Res 19:409,1971

8. Morrison JG: Dyskeratosis congenita, two extremes. S Afr Med J 48:223,1974

9. Aguilar-Martinez A, Lautre-Ecenarro MJ, Urbina-Gonzalez F, Cristobal-Gil MC, Guerra-Rodreguez P, Garcia-Perez A Cyto- genetic abnormalities in dyskeratosis congenita-Report of five cases. Clin Exp Dermatol13:100,1988

10. Connor JM, Teague RH: Dyskeratosis congenita: Report of a large kindred. Br J Dermatol105:321,1981

11. Womer R, Clark JE, Wood P, Sabio H, Kelley TE: Dyskera- tosis congenita: Two examples of this multisystem disorder. Pediat- rics 71:603,1983

12. Pai GS, Yan Y, DeBauche DM, Stanley WS, Paul SR: Bleomycin hypersensitivity in dyskeratosis congenita fibroblasts, lymphocytes, and transformed lymphoblasts. Cytogenet Cell Genet 52:186,1989

13. Burgdorf W, Kurvink K, Cervenka J: Sister chromatid exchange in dyskeratosis congenita lymphocytes. J Med Genet 14256,1977

14. Carter DM, Pan M, Gaynor A, McGuire JS, Sibrack L: Psoralen-DNA cross-linking photoadducts in dyskeratosis congenita: Delay in excision and promotion of sister chromatid exchange. J Invest Dermatol73:97,1979

15. Kano Y, Fuziwara Y: Dyskeratosis congenita: Survival, sister-chromatid exchange and repair following treatment with crosslinking agents. Mutat Res 103:327,1982

16. Auerbach AD, Adler B, Chaganti RSK Prenatal and postnatal diagnosis and carrier detection of Fanconi anemia by a cytogenetic method. Paediatrics 67:128,1981

17. Schneider A, Mayer U, Gebhart E, Harma D, Gromball J, Glockell U, Beck JD: Clastogen-induced fragility may differentiate pancytopenia of congenital dyskeratosis from Fanconi anemia. Eur J Pediat 148:37,1988

18. DeBauche DM, Pai GS, Stanley WS: Enhanced G2 chromatid radiosensitivity in dyskeratosis congenita fibroblasts. Am J Hum Genet 46:350,1990

19. Duckworth-Rysiecki G, Toji L, Ng J, Clarke C, Buchwald M: Characterization of a simian virus 40-transformed Fanconi anemia fibroblast line. Mutat Res 166:207,1986

20. Shaham M, Adler B, Chaganti RSK Transformation of chromosome breakage syndrome fibroblasts by SV40 DNA trans- fection. Cancer Genet Cytogenet 20:137,1986

21. Auerbach AD, Rogatko A, Schroeder-Kurth TM: Interna- tional Fanconi Anemia Registry: Relation of clinical symptoms to diepoxybutane sensitivity. Blood 73:391,1989

22. Marsh CW, Will AJ, Hows JH, Sartori P, Derbyshire P, Williamson PJ, Oscier DG, Dexter TM, Testa NG: “Stem cell” origin of the hematopoietic defect in Dyskeratosis congenita. Blood 79:3138,1992

23. Ling NS, Fenske NA, Julius RL, Espinoza CG, Drake LA: Dyskeratosis congenita in a girl simulating chronic graft-vs-host disease. Arch Dermatol 121:1424, 1985




1992 80: 3090-3096

I Dokal, J Bungey, P Williamson, D Oscier, J Hows and L Luzzatto unbalanced chromosomal rearrangementsDyskeratosis congenita fibroblasts are abnormal and have

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