Acute Myelomonocytic Leukemia with inv(16)(p13q22) Complicating Philadelphia Chromosome Positive Chronic Myeloid Leukemia

Sverre Heim, Bjarne Egelund Christensen, Thoas Fioretos, Anne-Grethe Sarensen, and Niels Tinggaard Pedersen

A B S T R A C T : The reciprocal translocation (9:22)(q34:ql 1) is highly characteristic of chronic myeloid leukemia (CML) and the pericentric inversion inv(16)(p13q22) is almost only f lmnd in acute nonlym- phocytic leukemia nf the myelomonacytic subtype (ANLL M4). Only twice before have an inv(16) and a t(9:22) been found in the same (:ells, and bath times the patients seemed to have de nova ANLL M4. We describe the case of a 21-year-old man who in July 1986 presented with a clinically and hematologi- cally classic chronic phase CML. Treatment with busulfan led to no improvement; instead in September 1986 he developed blast crisis with ANLL M4Eo morphology. He was now cytogenetically examined and the karyotype 45,X,-Y,t(9:22)(q34;q11),inv(16)(p 13(t22 ) was flmnd. Southern blot analysis of the bone marrow DNA sampled at this time reveah;d a standard rearrangement in the 3' end of the M-bcr. Intensive cytostatic treatment caused cytapenia fl)llawed by complete hematologic, clinical, and cyto- genetic reversal to chronic phase CML, so that in January 1987 the bone marrow karyotype was 46,XY,t(9:22)(q34;q11). Persistent splenomegaly was treated with splenectomy, and a chloroma of the skin was remow;d by irradiation. In March 1987 he rece.ived an allogeneic bone marrow transplant. Since then his only medical problem has been mild graft-versus-host disease: he is well and is working full time as a blacksmith.

I N T R O D U C T I O N

The balanced t ranslocat ion (9;22)(q34;qll) gives rise to the Phi lade lphia (Ph) chromosome [1] and is the cytoge- netic hal lmark of chronic myeloid leukemia (CML). The translocat ion is found in 85-90% of patients with CML, and at least some of the remaining, seemingly Ph negative, CML patients in fact have submicroscopic rearrangements leading to BCR/ABL fusion genes indis t inguishable from those arising through the classic t(9;22) [2-4]. During the accelerated and blastic phases of CML, 75-80% of the pa- tients develop secondary aberrat ions in addi t ion to the 9;22 t ranslocat ion [2, 4, 5]. The most common secondary changes are +8, i(17q], +Ph, +19, and -'-21.

Among the various chromosomal abnormali t ies associ- ated with the different acute non lymphocyt ic leukemia (ANLL) subtypes (4, 6-8] , the inv(16)(p13q22) seen in acute myelomonocyt ic leukemia with abnormal eosino-

From the Department of Medical (;enetics (S. ILL Odense Uni- versity; the Departments of Pathology (A-G. S., N. T. P.) and He- matology (B. E. C.), Odense University Hospital, Odense, Den- mark; and the Department of Clinical Genetics (S. H., T. F.), University Hospital. Lund, Sweden.

Address reprint requests to: Dr. S. Helm, Department of Clini- cal Genetics, University Hospital, S-221 85 Lund, Sweden.

Received July 16, 1991; accepted August 15, 1991.

~¢; 1992 Elsevi~r Science Publishing Co. Inc. 655 Avenue of the Americas. New York, NY 10010

phil ia (ANLL M4Eo) is one of the most sensit ive and spe- cific [9-13[. Because inv(16)(p13q22) is frequent in M4 but rare in other ANLL subtypes and in non-ANLL bone mar- row neoplasms, the finding has a very high predict ive power with regard to the M4 diagnosis [14].

The s imul taneous occurrence of t(9:22)(q34;q11) and ANLL subtype-specif ic abnormal i t ies is exceptional , and to the best of our knowledge only two patients with t(9:22) and inv(16) have been publ ished [15, 16]. We here de- scribe the cl inical , hematologic, (:ytogenetic, and molecu- lar genetic features of a CML patient whose blast crisis was an M4 with inv(16), but who hematological ly and cytoge- netical ly reverted to chronic phase CML after cytostatic therapy and today is cured of his bone marrow neoplasia after having received an al logeneic bone marrow trans- plantat ion 4 years ago.

CASE HISTORY

A 21-year-old man sought medical attention in the sum- mer of 1986 because of fatigue and a bleeding tendency of 2 months durat ion. When admit ted to hospital in July 1986, the cl inical and hematologic picture was that of a typical CML in chronic phase. He had pronounced spleno- megaly. His per ipheral blood values were WBC 333 × 10'q L, platelets 235 × a09/L, and Hb 42 g/L. The bone marrow

35 Cancer Genet Cytogenet 59:35-3,~ [1992) [} 165-4608:92.'$05.00

36 S. Heim et al.

was maximal ly hyperplasti(: . It was dominated by mature myelocytes and graimlocytes; only 5% were lnyeloblasts. No cytogenetic analysis was performed.

Treatntent was started with busulfan (6 mg/day), but with no measurabh: effe(:t. Instead, impairment of the blood values set in after 1 month, and a bone marrow smear froln the end of Septentber showe(l CML ntetam()r- phosis. The byperplasti( : bone marrow now (:ontaine(t 70% blasts. The morphol()gy (:orresponde(t to that of an ANLL M4Eo, with 45% myeloblasts , 25% monoblasts, and 4% abnormal eos inophi ls with big basoI)hili(: granules. Only a few mature (:ells were seen.

The therapy was changed to ANLL induct ion treatment with amsacr ine and high-dose (:ytosine arabinosi(te. This had a profound effect and after a period of bone marrow and peripheral cytopenia , the hematologic picture had by January 1987 (:hange(t back to that of chronic phase CML, with less than 5% blasts in the marrow. Because of persis- tent st)lenomegaly, a splene(:tomy was performed. The or- gan showed hematopoies is but no ex(:essive a(:cumula- tions of blasts. The patient also developed a localized skin infiltration diagnosed as a chloronta, which was remo~,ed by irra(liation.

In Mar(:h 1987 the patient re(:eived a bone marrow transplant from his HLA-identi(:al brother. He has sint:e made a remarkable re(:overv. Ex(:ept f()r mild graft-versus- host disease he has had no medi(:al problems. Todav. 4 years after the t ransplantat ion, he is feeling well and has a full-time joIt as a bla(:ksmith.

CYTOGENETIC AND MOLECULAR GENETIC INVESTIGATIONS

Cytogeneti(: analyses of the pat ient 's bone lnarrow (:ells were performed on two occasions: in September 1986, when the M4Eo blast crisis was diagnose(t, attd in January 1987, when reversal to CML (:hroni(: phase was diagnosed. The bone marrow cultures were synchronized with methotrexate (10 7 M) for 17 hours followed by thynfidine (10 -r' M) before Colcemid exposure an(t s tandard harvest- ing. The chromosomes were G-banded with Leishman's stain. In Septemher 1986, 29 of 30 analyzed (:ells had the karyotype 45,X,-Y,t(9:22)(q34:qll) , iIr¢(16)(p13q22) (Fig. 1); one (:ell with the same structural changes had retained the Y chromosome. After hematologic reversal to (:hroni(: phase CML it) January 1987, the kary()type (25 cells were examined) had changed to 46,XY.t(9;22)(q34:ql 1 ).

DNA analysis was performed on the September 1986 bone marrow sample. The (:ell pellet, originally intended for cytogenetic analysis, had been stored at -20°C in methanol :acet ic acid fixative. After a brief rinse in 10 mM Tris pH 7.8 buffer, the DNA was extracted by standard methods. The DNA thus obtained was digeste(t with Bglll. BamH1. and /-/indIII, ele(:trophoresed on a 0.5% agarose gel, and blotted it) denatur ing solut ion (0.4 M NaOH, 0.6 M NaCI) according to Southern {17]. The DNA was then transferred onto filters, nentral ized for 30 rain in 25 mM NazHPO4, pH e.5, and hybr idized to 2 kb BglIl/ttindIII 5' and tlin(tIII/BglII 3' major breakpoint cluster regi()n (M-

Figure 1 crisis. Arrowheads indicate breakpoints.

The 45,X,-Y,t(9:22)(q34:qll) , inv(16)(p13q22) bone marrow karyotype fotmd during blast

-11 -II-+l 1 2 3 4 5

II--+I-- IS 7 8 9

- I I - - h - - M - 13 14 1S

10 11 12

- tZ -'-- t X - - i ' i - 16 17 18

:bJt 111 20 21 22

I XY

AML with inv(16)(p13q221 in Ph* CML 37

bcr) probes (kindly provided by Professor Bartram, [Jim, Germany). The Southern blot analysis revealed an M-her rearrangement in subregion 4 [18], i.e., in the 3' end of the M-bet.

DISCUSSION

The simultaneous occurrence of t(9;221 and inv(16) in bone marrow cells has been reported twice before. Mecucci et al. [15] described a patient with acute myelomonocytic leukemia with bone marrow eosinophilia and basophilia and the karyotype 46,XY,t(9:22) (q34;q11),inv(161(p13q221. Li and Hayhoe [16] described another ANLL M4 patient whose complicated leukemia karyotype was interpreted as 47,XY,-18,~22,t(9;22) (q34;q11),der(l 6)inv(16)(p13q22)t(16:?18){q24;q21), der(20}del(20)(p 12pl 3)del(20)(q 12ql 3)del(q 12ql 3), + mar. DNA analyses of bcr rearrangements were not performed. Both patients seemed to have de novo ANLI~I no clinical disease reminiscent of CML had been suspected prior to the development of fully blown ANLL, and the patient in [16] entered full remission, not reversal to CMI, chronic phase, after induct ion treatment. The case we describe is therefore the first report of an inv(16)(p13q22) occurring in a chronic phase CML patient and apparently causing the transformation of the disease into a blastic phase ANLL. No similar importance can be attributed to the loss of the Y chromosome during metamorphosis: this change is not clearly associated with any particular FAB subtype except when occurring in ANLL M2 as part of the clnnal evolu- tion in patients with t(8;21)(q22:q22) [19]. There is also direct evidence that the loss of the Y took place after the inv(16), inasmuch as one cell was found in the September 1986 investigation that had t(9;22) and inv(16) but no -Y.

The fact that all three patients with t(9:22) and inv(16), both the two de novo leukemias and the blast crisis case, developed acute myelomonocytic leukemia with eosino- philia further underscores the strong genotype-phenotype association that exists between this leukemia morphology and inv(16)(p13q22) [4, 6, 7, 9-14]. In this respect the situatkm parallels that seen when t(15;17)(q22;qll), which is normally confined to ANLL M3 (4, 6, 7, 14]. is found as a secondary aberration in CML patients. The five cases in which this has been described had M3 blast crisis morphology [20-23].

The CML blast crisis is the most malignant of all acute leukemias, with a complete remission rate with intensive chemotherapy of less than 30% and a median survival of 3 -6 months [24-27]. The remarkable recovery of the pa- tient we describe is therefore noteworthy. It seems reason- able to attribute this to the type of ANLL-associated chro- mosomal aberration he developed in the blastic phase, inv(16)(p13q221, which is associated with the best re- sponse to treatment and the most favorable prognosis of all the cytogenetic subtypes of ANLL [28]. Dangerous though it is to extrapolate from a single case, it seems that patients with inv(16)(p13q221 have a relatively good prognosis not only when the disease is de novo ANLL, but also in those rare instances when the inversion occurs during CML metamorphosis.

This work was supported by grants from the Danish and Swedish Cancer Societies.

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