Karen Dun

A second transmissible cancer in Tasmanian devils

Significance Transmissible cancers are somatic cell lineages that are spread between individuals via the transfer of living cancer cells. Only three transmissible cancers have been reported in nature, suggesting that such diseases emerge rarely. One of the known transmissible cancers affects Tasmanian devils, and is threatening this species with extinction. Here we report the discovery of a second transmissible cancer in Tasmanian devils. This cancer causes facial tumors that are grossly indistinguishable from those caused by the first-described transmissible cancer in this species; however, tumors derived from this second clone are genetically distinct. These findings indicate that Tasmanian devils have spawned at least two different transmissible cancers, and suggest that transmissible cancers may arise more frequently in nature than previously considered. Clonally transmissible cancers are somatic cell lineages that are spread between individuals via the transfer of living cancer cells. There are only three known naturally occurring transmissible cancers, and these affect dogs, soft-shell clams, and Tasmanian devils, respectively. The Tasmanian devil transmissible facial cancer was first observed in 1996, and is threatening its host species with extinction. Until now, this disease has been consistently associated with a single aneuploid cancer cell lineage that we refer to as DFT1. Here we describe a second transmissible cancer, DFT2, in five devils located in southern Tasmania in 2014 and 2015. DFT2 causes facial tumors that are grossly indistinguishable but histologically distinct from those caused by DFT1. DFT2 bears no detectable cytogenetic similarity to DFT1 and carries a Y chromosome, which contrasts with the female origin of DFT1. DFT2 shows different alleles to both its hosts and DFT1 at microsatellite, structural variant, and major histocompatibility complex (MHC) loci, confirming that it is a second cancer that can be transmitted between devils as an allogeneic, MHC-discordant graft. These findings indicate that Tasmanian devils have spawned at least two distinct transmissible cancer lineages and suggest that transmissible cancers may arise more frequently in nature than previously considered. The discovery of DFT2 presents important challenges for the conservation of Tasmanian devils and raises the possibility that this species is particularly prone to the emergence of transmissible cancers. More generally, our findings highlight the potential for cancer cells to depart from their hosts and become dangerous transmissible pathogens.

A Murine Xenograft Model for a Transmissible Cancer in Tasmanian Devils

The number of Tasmanian devils in the wild is rapidly declining owing to a transmissible cancer, devil facial tumor disease (DFTD). Although progress has been made to understand the spread of this disease, crucial research on the pathogenesis of DFTD has been limited because of the threatened status of the host species. Here, the authors describe the development of a NOD/SCID (nonobese diabetic / severe combined immunodeficiency) mouse model that reproduces DFTD and provides a much-needed model to undertake studies into this intriguing transmissible cancer. Histologically, the DFTD produced in NOD/SCID mice (xenografted DFTD) was indistinguishable from the DFTD identified in Tasmanian devils. At the protein level, all xenografted DFTD tumors expressed periaxin, a marker that confirmed the diagnosis of DFTD. The karyotype of DFTD in NOD/SCID mice reproduced similar chromosomal alterations as seen in diseased devils. Furthermore, each NOD/SCID mouse inoculated with cultured DFTD tumor cells developed tumors, whereas DFTD did not develop in any of the inoculated immune-competent BALB/c mice.

BCR-ABL1 gene rearrangement as a subclonal change in ETV6-RUNX1–positive B-cell acute lymphoblastic leukemia

We report here on a case of ETV6-RUNX1-positive B-cell acute lymphoblastic leukemia (B-ALL) that has acquired a BCR-ABL1 gene rearrangement as a subclonal change. The 19-year-old female patient presented with B symptoms, pancytopenia, and circulating blasts. The bone marrow aspirate was hypercellular and was infiltrated by an immature blast population that was confirmed as B-ALL by flow cytometry. Sequential fluorescent in situ hybridization was performed on the patients leukemic cells, which were shown to contain both ETV6-RUNX1 and BCR-ABL1 gene rearrangements. The majority of nuclei (85%) showed only the ETV6-RUNX1 gene rearrangement; however, an additional 10% also showed a variant BCR-ABL1 gene rearrangement, indicating the ETV6-RUNX1 gene rearrangement was the primary change. A review of the literature has shown that acquisition of a BCR-ABL1 gene rearrangement as a secondary change in B-ALL is a very rare occurrence, and the effect it may have on prognosis is uncertain in the modern therapy age.

Ring chromosome with deletion 7q in acute myeloid leukaemia

Cytogenetic abnormalities can be detected in approximately 50–60% of newly diagnosed adult patients with acute myeloid leukaemia (AML). Monosomy of the chromosome 7 (−7) and deletion of the long arm of the chromosome 7 (7q–) are considered as high cytogenetic-risk AML with a poor prognosis. These abnormalities can occur, as a single chromosomal aberration, in approximately 8% of newly diagnosed AML. We report an elderly patient with AML who had deletion 7q (7q–) along with ring chromosome, which was demonstrated in conventional cytogenetics and fluoresecent in-situ hybridisation techniques.

Atypical features in a patient with acute promyelocytic leukaemia: a potential diagnostic pitfall

Acute promyelocytic leukaemia (APML) is a malignancy with a high cure rate; however, delay in diagnosis or treatment can result in morbidity and mortality. APML has characteristic clinical, morphological, immunophenotypic and molecular features. In patients with acute leukaemia, a high index of suspicion is required to exclude APML. Very rarely APML patients at diagnosis can demonstrate atypical features. We reported a patient whose bone marrow features resembled acute myeloid leukaemia with predominantly agranular blasts, devoid of Auer rods and expressing CD34 and HLA-DR on flow cytometry. APML was not suspected initially but after cytogenetic and molecular genetic studies demonstrated t(15;17), appropriate therapy with ATRA+ chemotherapy was instituted and the patient showed remarkable and sustained response to treatment. This case highlights the fact that morphology and immunophenotyping are useful but not infallible indicators for these malignancies and, ultimate diagnoses will require detection of the characteristic molecular markers.

DSP30 and interleukin-2 as a mitotic stimulant in B-cell disorders including those with a low disease burden

Chromosome abnormalities detected during cytogenetic investigations for B‐cell malignancy offer prognostic information that can have wide ranging clinical impacts on patients. These impacts may include monitoring frequency, treatment type, and disease staging level. The use of the synthetic oligonucleotide DSP30 combined with interleukin 2 (IL2) has been described as an effective mitotic stimulant in B‐cell disorders, not only in chronic lymphocytic leukemia (CLL) but also in a range of other B‐cell malignancies. Here, we describe the comparison of two B‐cell mitogens, lipopolysaccharide (LPS), and DSP30 combined with IL2 as mitogens in a range of common B‐cell disorders excluding CLL. The results showed that DSP30/IL2 was an effective mitogen in mature B‐cell disorders, revealing abnormal cytogenetic results in a range of B‐cell malignancies. The abnormality rate increased when compared to the use of LPS to 64% (DSP30/IL2) from 14% (LPS). In a number of cases the disease burden was proportionally very low, less than 10% of white cells. In 37% of these cases, the DSP30 culture revealed abnormal results. Importantly, we also obtained abnormal conventional cytogenetics results in 3 bone marrow cases in which immunophenotyping showed an absence of an abnormal B‐cell clone. In these cases, the cytogenetics results correlated with the provisional diagnosis and altered their staging level. The use of DSP30 and IL2 is recommended for use in many B‐cell malignancies as an effective mitogen and their use has been shown to enable successful culture of the malignant clone, even at very low levels of disease.

Acute myeloid leukaemia with t(8;21)(q22;q22.3) and loss of the X chromosome

Cytogenetic abnormalities occur in approximately 60% of newly diagnosed patients with acute myeloid leukaemia (AML) and are useful in the risk stratification of AML. Translocation between chromosomes 8 and 21—t(8;21)—(q22;q22.3), which carries a favourable prognosis, is found in approximately 5% to 10% of all patients with AML. Additional chromosomal abnormalities have been described in patients with AML with t(8;21), which may impact on the favourable prognosis. We report a patient who had AML with t(8;21)(q22;q22.3) and loss of the X chromosome.

Complex hypodiploid acute myeloid leukaemia secondary to chemotherapy for hyperdiploid multiple myeloma

Acquired clonal chromosomal abnormalities commonly occur in haematological malignancies and are known to have prognostic implications in such patients. We report a patient who developed complex hypodiploid acute myeloid leukaemia secondary to chemotherapy for hyperdiploid multiple myeloma (MM). A 70-year-old Caucasian male presented in October 2009 with anaemia (haemoglobin: 90 g/L), renal impairment, lytic bone lesions, and very high IgG-kappa paraprotein level (74 g/L). Bone marrow biopsy was markedly hypercellular with 50 % abnormal plasma cells consistent with multiple myeloma (Fig. 1). Bone marrow chromosomal analysis revealed a hyperdiploid male karyotype with gains of chromosomes 3, 5, 6, 7, 9, 11, 15, 17, and 19 in approximately 25 % of cells analysed. The karyotype was 54–56, XY, ?3, ?5, ?6, ?7, ?9, ?9, ?11, ?11, ?15, ?17, ?19[cp4]/46,XY[14] (Fig. 2). The fluorescent in situ hybridization (FISH) testing showed no evidence of deletions at 13q14 and 17p13.1 or IgH gene re-arrangement at 14q32.3, which excluded the poor prognostic myeloma markers. He was treated with melphalan and prednisolone (MP) regimen and underwent complete remission with undetectable IgG paraprotein after 13 months. At that stage, bone marrow biopsy showed a normal karyotype (46, XY) and no evidence of the previous hyperdiploid karyotype. He remained in complete remission of MM even after stopping melphalan and prednisolone 24 months after initiation. In September 2013, 4 years after the diagnosis of MM, his full blood counts showed pancytopenia (haemoglobin: 83 g/L, white cell count: 2.4 9 10/L and platelet count: 42 9 10/L) with 10 % blasts in the blood film. Bone marrow biopsy revealed tri-lineage dysplasia and increase in leukaemic blasts (40 %), which were minimally differentiated (Fig. 3). Flow cytometric analysis showed expression of surface markers CD34, CD117, CD13, CD33, and HLA-DR on the leukaemic blasts, consistent with a myeloid phenotype. The diagnosis according to WHO classification of tumours of haematopoietic and lymphoid tissues—2008 was therapy-related acute myeloid leukaemia (t-AML), which was deemed secondary to melphalan therapy for MM. Bone marrow chromosome analysis revealed a very complex hypodiploid karyotype of 41, XY,add(1)(p32),-5,del(7)(q22q36),der(8)add(8)(p11.2) add(8)(q24.3),del(12)(p11.2p12),-15, add(15)(p11.2),-16, -18,i(20)(p10),-21,[6]/42,idem, ?mar1[6]/42,idem,-12, del(19) (q13.1), ?20,i(20), ?mar2[cp2]/43,idem,-add(1),del(4) (q21q31), ?del(5)(q31q35),-del(7), ?dic(7;12)(q11.2;p12), ?add(8)(p11.2),-der(8), ?15, ?18,-19, ?20,-i(20), ?mar3 [cp3]/41-42,idem,add(12)(p11.2),-19, ?20,-i(20)[cp2]/46, XY[1] (Fig. 4). There were many structural and numerical abnormalities. This complex karyotype included a myelodysplasia-related abnormality of del(12)(p11.2p12), in addition to loss of chromosome 5 and del(7)(q22q36), which may be associated with morphologic multilineage dysplasia. However, the significance of the remaining cytogenetic abnormalities is uncertain. Since therapy-related AML with complex cytogenetic abnormalities augur a M. Mohamed (&) Haematology Department, Launceston General Hospital, Launceston, Australia e-mail: muhajirbm@gmail.com; muhajirbm@yahoo.com

Tetraploidy with double t(15;17)(q22;q21) in acute promyelocytic leukaemia

Acute promyelocytic leukaemia (APML) is a subtype of acute myeloid leukaemia characterised by the chromosomal translocation t(15;17)(q22;q21) with the PML-RARa fusion gene. Additional chromosomal abnormalities are occasionally found in patients with acute promyelocytic leukaemia. In this report, we describe of tetraploidy with two copies of the t(15;17)(q22;q21) in a patient with acute promyelocytic leukaemia which is an extremely rare occurrence.

Testing Hygrometers Used in Cytogenetics Laboratories for Metaphase Preparation

This protocol describes procedures for checking small laboratory hygrometers for accuracy at three relative humidity (rh) levels. The work arose out of the need to provide laboratory assessors with documentary evidence that the hygrometer used to monitor humidity in the vicinity of the laboratory where medical cytogenetics testing slides are prepared and dried in the ambient environment is reproducible and sufficiently accurate. The procedure is based upon the physicochemical principle that when water or certain saturated salt solutions are placed into a sealed environment, the humidity will equilibrate to well defined levels. We choose to check our hygrometers at three points: 95%, 75%, and 33% rh, using distilled water, saturated sodium chloride solution, and saturated magnesium chloride solution, respectively. Our results have demonstrated that the procedure is convenient and of sufficient accuracy to be fit for this annual hygrometer validation purpose. The procedure takes 24 hr per relative humidity point checked.