Supplementary Matterial
Annals of Hematology
Identification of a Potential Topoisomerase II ‘‘Hotspot’’ DNA Region in the DEK Gene in two t(6;9)-positive Therapy-Related myeloid neoplasms
Maria Liliana Piredda1,2, Gianfranco Catalano2, Claudia Ciardi3, Mariadomenica Divona3, Laura Cicconi2, Paola Panetta3, Paola Curzi3, Eduardo Garza1,2,4, Carmen Martínez-Losada1,2,5, Massimiliano Postorino2, Francesco Lo-Coco1,2, Nélida I. Noguera1,2.
1- Laboratory of Neuro-Oncoematology, Santa Lucia Foundation, Rome, Italy.
2- Department of Biomedicine, "Tor Vergata" University, Rome, Italy.
3- Policlinico di tor Vergata, Rome, Italy
4- Laboratorios Dr. Moreira, Monterrey, Mexico.
5-Alfonso Martin Escudero Foundation, Madrid, Spain.
Contact information for correspondence:
Dr. Nelida I.
Supplementary Patients
Patient 1
A 65 year old woman with a five year history of previous breast cancer treated with quadrantectomy and 4 cycles of cyclophosphamide, doxorubicin, plus tamoxifen was admitted to our center. She was still on tamoxifen when a t(6;9), jak2- (V617F) and FLT3-ITD-positive AML, was diagnosed. She underwent the 3+7 classical induction chemotherapy (daunorubicin plus cytosine-arabinoside), obtaining partial remission. Two months later she received 9 high-dose ARAC cycles with no response, and died eight months later with disease progression.
Patient 2
A 63 year old woman, with a two year history of infiltrating breast cancer treated with mastectomy, chemotherapy (Epirubicin), radiotherapy and tamoxifen (at the time still under treatment with tamoxifen), was diagnosed with a t(6;9)-positive RAEB-1 (10% bone marrow blasts). She underwent 10 cycles of hypomethylating therapy with azacytidine, as a bridge to hemopoietic stem cell transplantation from a familiar donor. Ten months later she was admitted to our center, with disease progression (BM-blasts: 67%) and a phenotype compatible with t(6;9), FLT3-ITD and Jak2(V617F) –positive AML. She was administered mitoxantrone, etoposide, and cytarabine (MICE) chemotherapy with no response. She then received salvage therapy with fludarabine, cytarabine and idarubicin, obtaining partial remission (BM blasts: 16%). Less than one month later the disease progressed and the patient received high dose ARA-C, but the disease was resistant. She died six months later with disease progression.
Supplementary Figure 1
Supplementary Fig 1 : Schematic representation of the position of primers for long template PCR. F1-6 are forward (DEK) primers and R7-14 are (NUP214) reverse primers.
Supplementary Figure 2
Supplementary Fig 2: Trasnlocation t (6;9) DEK-NUP214 of t-AML. Bone marrow G-banded metaphase showing a 46,XX t (6;9) (p23;q34) karyotype. The arrows show the rearranged chromosomes.
Supplementary Table 1
Primer List
Number / NAME / SEQUENCE / APPLICATIONF1 / DEKA / GCAGCACCACCAAGAATCA / LPT and RT PCR of DEK-NUP214
F2 / DEKC / ACTGGCCAGTGCTAACTTGGAA / LPT and RT PCR of DEK-NUP214
F3 / DEK1 / CTGGCCAGTGCTAACTTGGAAGAA / LTP AND NESTED PCR
F4 / DEK2 / TAGAAGCTAGATGTGGTGGCTCAC / LTP AND NESTED PCR
F5 / DEK3 / AGAGGTCAGGTGAGAAGGA / LTP AND NESTED PCR
F6 / DEK11 / GGAGAAAGCAATTTGGTGGGAAAGGAGAAG / LTP and nested PCR
R7 / CANB / TTCCCAGAGCAGCATTCACAG / LTP and RT-PCR of DEK-NUP214
R8 / CAND / AAACAGACTTCCCTGTGGAGC / LTP and RT-PCR of DEK-NUP214
R13 / CANsec1 / ATCCCATGTTCATGGATGGATAG / SEQUENCING REACTION
R14 / CANsec2 / GAGCAAGACTCCGTCTCAAA / SEQUENCING REACTION
R9 / CAN8 / TAGGAAATTCGGCGCCTTCATCAG / LPT and nested PCR
R10 / CAN7 / AGGCAGCCATACCCATTCT / LPT and nested PCR
R11 / CAN6 / CCTCGAGTAATACACCCGTC / LPT and nested PCR
R12 / CAN4 / CCGGCTGAGCATGGGATGT / LPT and nested PCR
ABL-A2B-5 / GCATCTGACTTTGAGCCTCAG / RT-PCR for ABL
ABL-A3E-3 / TGACTGGCGTGATGTAGTTGCTT / RT-PCR for ABL
LTP: Long Template PCR