Reversal of isolated 20q deletion with vitamin B₁₂ replacement in a patient with pernicious anaemia

Abstract

Severe vitamin B₁₂ deficiency is well known to cause morphological alterations in bone marrow. In rare instances, these myelodysplastic and megaloblastic changes can coexist with cytogenetic abnormalities. Here, we report a case of a 38-year-old African-American woman with pernicious anaemia, who was found to have an isolated 20q deletion and which resolved after vitamin B₁₂ replacement. We also discuss various mechanisms in which vitamin B₁₂ deficiency can lead to chromosomal abnormalities. A literature review is also performed to evaluate various other chromosomal aberrations associated with B₁₂ deficiency.

Background

Folic acid and vitamin B₁₂ (B₁₂) play an important role in DNA synthesis and gene expression. Although there are numerous in vivo and in vitro studies linking folate to chromosomal abnormalities and mutagenesis, the data on B₁₂ is however surprisingly limited to few case studies. To the best of our knowledge, this is the first reported case of an isolated 20q deletion secondary to vitamin B12 deficiency. Our report adds new findings to the current existing literature of various other cytogenetic abnormalities seen in the bone marrow from severe B₁₂ deficiency. This manuscript also highlights the importance to exclude B₁₂ deficiency in patients with new diagnosis of myelodysplasia with cytogenetic abnormalities.

Case presentation

A 38-year-old African-American woman was initially referred to our haematology clinic for evaluation of macrocytic anaemia. The patient was in a normal state of health until 6 months prior to this presentation. She then experienced progressively worsening fatigue, heart burn, nocturnal cough and weight loss over 20 lbs. Pertinent physical examination and vital signs were grossly unremarkable. Initial laboratory findings were as follows: haemoglobin (Hb): 10 g/dL, mean corpuscular volume: 110 fL, white cell count (WCC): 3.92×10³/μL, platelets: 281 000/μL, reticulocyte count: 1.4%, serum B₁₂: 4187 pg/mL, serum folate: 33 ng/mL, iron: 84 μg/dL, total iron binding capacity: 251 μg/dL, ferritin: 119 ng/mL, transferrin saturation: 33%, lactate dehydrogenase: 205 U/L, haptoglobin: 32 mg/dL, thyroid stimulating hormone: 0.650 μIU/mL, copper: 166 μg/dL. Hepatitis and Human Immunodeficiency Virus 1 and 2 serology was negative. Peripheral blood smear revealed macrocytosis, anisocytosis, tear drop cells, adequate platelets and no evidence of dysplastic cells. Secondary to peripheral blood examination revealing macrocytic red blood cells, tear drop cells in the context of normal B₁₂ levels, a bone marrow biopsy was suggested, which was declined by the patient. Six months later after being lost to follow-up, she presented to the emergency room with bilateral leg weakness and her physical examination revealed marked skin hyperpigmentation, glossitis, moderately increased muscular tone, decreased sensation to touch and pressure in the lower extremities, decreased reflexes in the ankle and knee joints and wide based gait.

Investigations

Repeat laboratory findings were: Hb: 8.6 g/dL, WCC: 2.06×10³/μL, platelets: 193 000/μL, folate: 19.9 ng/mL. Bone marrow biopsy revealed hypercellular bone marrow with megaloblastic and dysplastic changes, 6% CD34+ blast cells and rare ringed sideroblasts. Metaphase cytogenetic studies on bone marrow aspirate showed del 20 q (figure 1). B₁₂ this time was noted to be at 83 pg/mL, serum homocysteine: 218 μmol/L, serum methyl malonic acid: 23246 nmol/L. Antiparietal antibody and anti-intrinsic factor antibody were strongly positive. The initial finding of a falsely elevated B₁₂ levels can be perceived as an erroneous laboratory finding.

Figure 1.

This figure demonstrates cytogenetic analysis revealing 20 q deletion before vitamin B₁₂ replacement (chromosome 20 magnified (not to scale) to demonstrate the abnormality).

Differential diagnosis

At patient's initial presentation, as all the work up for macrocytic anaemia was negative, myelodysplasia was considered in the differential diagnosis. But, the differential diagnosis was soon narrowed to B₁₂ deficiency when the patient presented later with severe ataxia along with classic neurological manifestations in the presence of macrocytic anaemia. The primary diagnosis of pernicious anemia was confirmed when the serology was positive for both anti-parietal and anti-intrinsic factor antibodies.

Treatment

The patient was subsequently treated with intramuscular B₁₂ supplementation (1000 µg every day for 1 week, every 1 week for 4 weeks and monthly thereafter) with rapid resolution of all haematological parameters in a week and more gradual improvement of neurological symptoms.

Outcome and follow-up

Seventy days after B₁₂ replacement, a repeat bone marrow biopsy was performed and revealed a normocellular bone marrow with no evidence of myelodysplastic features and 3% CD34+ blast cells. Repeat cytogenetic studies on bone marrow aspirate demonstrated normal female karyotype and reversal of prior noted del 20q (figure 2).

Figure 2.

This figure demonstrates normal female karyotype and the reversal of the previously seen 20 q deletion after vitamin B₁₂ replacement.

Discussion

Dietary B₁₂ (cobalamin) is absorbed in the distal ileum after binding to intrinsic factor secreted by the gastric parietal cells.¹ Cobalamin aids in the conversion of 5-methyl tetrahydrofolate (THF) to N 5,10 methylene THF, which is necessary for deoxy-thymidine monophosphate (dTMP) synthesis.² Cobalamin also mediates the conversion of homocysteine to methionine which is a precursor for S-adenosyl methionine, a methyl donor required for the maintenance of methylation patterns in DNA and the synthesis of formyl-THF.² Deficiency of B₁₂ also leads to accumulation of homocysteine. B₁₂ deficiency ultimately leads to trapping of folate, increased deoxy-uridine monophosphate (dUMP), which prevents incorporation of dTMP into the DNA. High intracellular ratio of dUMP/dTMP and subsequent increase in the deoxy-uridine triphosphate/deoxy-thymidine triphosphate (dUTP/dTTP) ratio results in misincorporation of uracil into the DNA strand.^3–5 Uracil DNA glycolase/apyrimidinic nuclease which corrects this mis-incorporation is overwhelmed in the continued absence of B12, and this lack of corrective process leads to accumulation in chromosomal fragile sites, chromosomal breaks and chromosomal deletions without any repair^6–7 and chromosomal deletions without any repair. Altered methylation of DNA is also hypothesised to play a role in mutagenesis and carcinogenesis.^8–10 Other plausible mechanisms for mutagenesis which have been studied in vitro include increased homocysteine levels,¹¹ and decreased telomere length¹² seen in patients with a B₁₂ deficiency that affects genomic integrity. B₁₂ deficiency can also cause micronucleus formation,^13–14 and accelerated apoptosis;¹⁵ however, their role in mutagenesis is unknown.

Studies attempting to characterise chromosomal changes associated with B₁₂ deficiency date back to as early as 1950s. But, a consistent demonstration of this finding was a challenge secondary to the limitations of laboratory techniques involved.¹⁶ As the molecular techniques improved, various groups demonstrated multiple chromosomal abnormalities: increase in nuclear size,¹⁷ hypodiploidy^{18 19} and chromosomal breaks.^20–23 Some of these reports also demonstrate reversal of these abnormalities on replacement of B₁₂.^{20 21 24 25} For example, B₁₂ deficient patients with isolated 7q and 3p chromosomal deletions had reversal of these chromosomal findings on replacement therapy.^{26 27} Table 1 summarises the various reports of chromosomal abnormalities which were associated with B₁₂ deficiency.

Table 1.

Other reports of B₁₂ deficiency associated with chromosomal abnormalities

	Reports	N*	Deficiency	Comments
1	Ford, 195916	1	B12	Hypodiploidy; subsequently reported as an error
3	Astaldi et al, 196218	1	B12	Aneuploidy and changes in chromosome morphology Less pronounced after B12 replacement
4	Forteza and Baguena, 196328	1	B12	Aneuploidy
5	Kiossoglou et al, 196519	3	B12	Aneuploidy, chromosomal and chromatid breaks Changes persisted in one after B12 replacement
6	Jensen et al, 196722	4	B12	Increased chromosomal breaks
7	Heath, 196621	14	B12 and folate	Chromosome breakage along with severe megaloblastic changes
8	Bottura et al, 196824	1	B12	Chromosomal breaks in metaphase; corrected after B12 replacement
9	Keller et al, 197020	3	B12	Chromosomal breaks resolved after B12 replacement in one patient
10	Lawler et al, 197123	15	B12 and folate	Chromosomal breaks. Reversal seen in 48 hours with B12 replacement
11	Goh, 198129	1	B12	Del (18p), persisted with replacement. Unlikely from B12 deficiency
12	Wollman et al, 199626	1	B12 and folate	Del (7q) with reversal after B12 replacement
13	Chintagumpala et al, 199625	3	B12	Spontaneous chromosomal fragility. Reversal with B12 replacement in one patient
14	Parmentier et al 201227	1	B12	Del (3p), with reversal of cytogenetic abnormalities after B12 replacement

*N, number of patients described in the case report.

B₁₂, vitamin B₁₂.

Our case involved an isolated 20q deletion, a non-specific, yet a recurrent chromosomal abnormality seen in myelodysplastic syndrome (MDS).³⁰ Del (20q) preferentially involves erythroid and megakaryocytic precursors and this isolated abnormality when associated with MDS portends a favourable prognosis in terms of its overt progression into acute myeloid leukemia (AML).^{31 32} It is to be noted that other conflicting reports about poor prognosis of del (20q) and higher conversion to AML have also been reported.³³ Del 20q is also known to be associated with myeloproliferative disorders, acute myeloid leukaemia, pure red blood cell aplasia and angioimmunoblastic lymphadenopathy with dysproteinaemia.³⁴ Morphologically bone marrow examination in a B₁₂-deficient individual can mimic those with frank myelodysplasia/leukaemia.^35–37 Cases have been reported where B₁₂ deficiency and MDS/AML coexist.^38–39 Das et al⁴⁰ demonstrated that reversal of chromosomal breaks and despiralisation can take up to 12 weeks to normalise after B₁₂ and folate supplementation.

Learning points.

• This case demonstrates reversal of haematological parameters and cytogenetic abnormality after vitamin B12 replacement in a patient with pernicious anaemia.

• Reversible clonal cytogenetic abnormalities and bone marrow changes mimicking myelodysplastic syndrome may be seen in patients with severe B₁₂ deficiency. It is prudent to identify and treat the B₁₂ deficiency before making a misdiagnosis of myelodysplastic syndrome.

• Although B₁₂ deficiency can cause chromosomal and hematopoetic abnormalities, its role in leukemogenesis is not clear. Further studies are needed to confirm if B₁₂ deficiency can lead to mutagenesis and carcinogenesis.