**5. Diagnosis aids of multiple myeloma in veterinary medicine**

#### **5.1. Hematology**

cardium and anemia may exacerbate the severity of this condition [109]. Approximately 50% of the cats with MM in one study had idiopathic heart murmur [4]. Recently, three cases of cats evaluated for congestive heart failure with acute collapse, tachypnea, increased respira‐

Neurological manifestations often complicate the course of MM, but direct involvement of the central nervous system in rare in human patients, although there are some cases report‐ ed [110-113]. Similarly, neurological signs are uncommon manifestations of MM in animals. However, it is well recognized that MM may lead to abnormalities of the central nervous system either as a result of spinal cord compression by the neoplasm arising within a verte‐ bra or due to pathological fracture of a vertebra weakened by tumor infiltration [114-115]. It could also be due to the HVS where sludging of blood within the vasculature results in cen‐ tral nervous system hypoxia [116]. Neurological signs associated with MM have been re‐ ported in horses [1,7], cats [31] and dogs [22,114]. Edwards et al. [1] described the cases of three horses with rear leg paresis and/or ataxia. Spinal cord compression by an extradural tumor mass was observed in one of the two horses in which the spinal canal was examined. McConkey et al. [7] observed a horse with hind ataxia progressing to paralysis, with dys‐ phagia and ptyalism. These clinical signs were attributed to neoplastic involvement of the trigeminal nerve. Similarly, Appel et al. [31] presented the case of a cat with hind limb loco‐ motor difficulties, signs of pain along the lumbar portion of the vertebral column, with al‐ tered motor function and moderate muscle atrophy. In this case, survey radiographs

In MM patients, peripheral neuropathy has for a long time been considered as mainly secon‐ dary to the tumor, following a direct compression (radicular or medullar), light chain depos‐ its (amyloidosis), cryoglobulinemia or an autoimmune mechanism [117-121]. A paraneoplastic polyneuropathy is seen in association with IgM monoclonal gammopathy as‐ sociated with MM, Waldenstrom's macroglobulinemia, primary amyloidosis and lympho‐ ma, as well as monoclonal gammopathy of undetermined significance. IgM-M proteins have autoantibody activity and have been shown to bind to myelin associated glycoprotein re‐ sulting in a demyelinating peripheral neuropathy [121]. However, increased levels of cyto‐ kines are thought to cause the paraneoplastic neuropathy rather than an immune-mediated mechanism [122-123]. Furthermore, with the use of new drugs, the iatrogenic neurotoxicity has become the leading cause of peripheral neuropathy in people [124-126]. The commonest nerve involvement appears to be in the form of sensory-motor axonal neuropathy followed

In companion animals, paraneoplastic neuropathies have been reported sporadically in ma‐ lignant tumors, and include bronchogenic carcinoma, insulinoma, leiomyosarcoma, heman‐ giosarcoma, undifferentiated sarcoma, synovial sarcoma and adrenal adenocarcinoma [114,127-129]. Viviers and Dobson [22] described the case of a 12-year-old female German Shepherd dog that developed progressive hindlimb followed by forelimb ataxia with tetra‐ plejia. Neurological examination suggested lower motor dysfunction. MM was diagnosed

tory effort, and pulmonary crackles, have been reported secondarily to HVS [54].

**4.7. Other clinical signs**

294 Multiple Myeloma - A Quick Reflection on the Fast Progress

revealed osteolytic lesions in lumbar vertebras [31].

by sensory-motor demyelinating neuropathy [117].

Anemia is a prominent feature of MM, it is commonly associated with clinical progres‐ sion in human patients and occurs in more than two thirds of all patients [130-132]. Sim‐ ilarly, approximately 30% of dogs and 75% of cats with MM have a normocytic normochromic non-regenerative anemia [4,10,26-27,32-33] and anemia is also invariably present in horses with MM.

The pathogenic mechanisms involved in the anemia are chronic inflammation, tumor hemorrhage and/or hemostatic abnormalities, myelophthisis, increased red blood cell de‐ struction induced by the HVS, plasma expansion secondary to the osmotic effect of the paraproteins and red cell destruction by neoplastic cells [4,27,29]. In cats, erythrophago‐ cytosis by neoplastic plasma cells [32], mast cells [133]), lymphocytes [134-135] and histo‐ cytes [136] has been observed. In the same way, there are several reports of phagocytic plasma cells in people with MM [137]. Although erythrophagocytic plasma cells in hu‐ mans with MM are rare, neoplastic plasma cells have been observed with phagocytosed platelets and granulocytes [138]. The mechanism of hemophagocytosis in MM is unclear, as plasma cells have not phagocytic function under normal circumstances. Results of the direct antiglobulin test in humans are almost always negative, suggesting hemophagocy‐ tosis by neoplastic cells is not an autoimmune function [137]. It has been speculated that phagocytic plasma cells may arise as an expansion of a rare B-cell clone with innate phagocytic potential [137]. A single case of phagocytic plasma cells aberrantly expression CD15 (normally found on neutrophils and monocytes and involved in phagocytosis) has been reported in human beings [139].

Nevertheless, anemia of chronic disease appears to be of utmost importance in MM. Inter‐ leukin-1 and tumor necrosis factors are capable of suppressing erythropoiesis [130]. Anemia

has broad implications in these patients. First, the low hemoglobin concentration and packed cell volume have been associated with poor quality of life in people, and affect daily activity. Second, anemia has an impact on the cardiovascular system. In fact, anemia has been shown to induce or aggravate hypoxia and ischemic complications. Third, anemia has been shown to be a poor prognostic factor in MM [140-142].

recognized, the term biclonal gammopathy is often used. Monoclonal gammopathies are as‐ sociated with production of a single clone of immunoglobulin owing to clonal expansion of neoplastic lymphoid cells, such as plasma cells in MM and B cells in lymphoma [9]. In con‐ trast, a polyclonal gammopathy is characterized by a broader peak or multiple peaks in the γ- or β-γ regions. Polyclonal gammopathies are associated with chronic antigenic stimula‐ tion that occurs in chronic infections and other inflammatory conditions [148,150-152]. The term oligoclonal gammopathy refers to an electrophoretic pattern that is similar to a mono‐ clonal one, but in which the globulin peak is slightly wider than the albumin peak [9,149]. Oligoclonal gammopathy may occasionally occur in animals with chronic inflammation or

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Although monoclonal gammopathy is the laboratory landmark of MM, other conditions oc‐ casionally can induce a monoclonal gammopathy in animals, such as chronic inflammation (leishmaniosis, ehrlichiosis, chronic pioderma, feline infectious peritonitis) [149-150,153-155], amyloidosis [156-157], B-cell lymphoma [149], Waldenströms macroglo‐ bulinemia [158-159] and monoclonal gammopathy of undefined significance (MGUS) [160]. The inclusion criterion for MGUS are M-protein and <10% bone marrow plasmacytosis, with no evidence of lytic lesions, light chain proteinuria or other clinical, hematologic, and bio‐ chemical abnormalities [161-162]. MGUS occurs in 1-2% of people over the age of 50 and 3% of people over the age of 70 [162]. A significant proportion (25%) of these will evolve within 20 years into MM, primary amyloidosis, macroglobulinemia or another lymphoproliferative

In dogs affected by MM, the incidence of IgA and IgG is comparable, whereas in cats and horses IgG is most commonly involved [1,3-4,27]. In fact, of the 25 published feline MM with immunoelectrophoresis results, 20 had IgG gammopathies, and 5 had IgA gammopathies [2,13-14,85]. Similarly, there are some reports of IgA gammopathies in horses with MM [3,8].

Biclonal gammopathy, with two M-components has been reported in humans [163-165], even though it was found to be very rare, occurring in about 1% of human beings with MM [166]. Biclonal gammopathies have been described in lymphoproliferative disorders in dogs and cats [2,9,19,23,167-168] including MM. The term biclonal is applied to the electrophoret‐ ic pattern and does not always correlate with true biclonal expansion because the biclonal electrophoretic may arise from a single clone of B-cells, usually mature plasma cells that produce one type of immunoglobulin with different dimerization patterns [9]. The biclonal pattern may also occur from production of two different classes of immunoglobulins, usual‐ ly IgG and IgA, by two separate cell clones [23]. However, production of separate heavy chain isotypes by a single clone of neoplastic cells, may result from isotype switching, which

The prevalence of biclonal gammopathy in companion animals is unknown, but it could be higher than reported. In many clinical veterinary laboratories, serum protein electrophoresis is performed using cellulose acetate as the support medium. However, better separation of protein fractions may be obtained using agar cell electrophoresis or capillary zone electro‐ phoresis. Facchini et al. [9] reported two cases (dog and cat) with gammopathies associated

infectious disease [9].

disease [162].

occurs normally during B-cell maturation [9].

Data concerning white blood cell count (total and different subpopulations) are not consis‐ tent in animals with MM. It has been found lymphopenia [24,30,32], leukocytosis due to neutrophilia [4,8], neutropenia with lymphocytosis [31], leukopenia [1,25,37], leukocytosis with neutrophilia and lymphocytosis [31], neutropenia [16], neutrophenia with lymphope‐ nia [18], pancytopenia [10] and absence of white blood cell abnormalities [3,6,22].

Thrombocytopenia is reported in approximately one third of all canine patients of MM [10,25,27]. It has also been described in cats [4,29] and horses [18,37]. However, there are many other reports of MM in veterinary medicine with patients that show normal number of platelets [3,6-8,16,22,24,31-32]. Thrombocytopenia that could promote bleeding disorders, is proposed to result from infiltration of bone marrow by malignant plasma cells, consump‐ tion of platelets as part of thrombohemorrhagic syndrome, such as disseminated intravascu‐ lar coagulation, shortened platelet half-life or immune-mediated destruction, even though the latter 2 mechanisms have yet to be verified in veterinary medicine [10,27,30,64,142-143].

#### **5.2. Blood clinical biochemistry**

#### *5.2.1. Serum protein concentrations and serum protein electrophoresis*

Hyperproteinemia, specifically hypoalbuminemia and hyperglobulinemia is very common in MM, but not an invariable feature. Hypoalbuminemia has been described consistently in MM in dogs [9,15-16,22-24], cats [3,9,29,32] and horses [1,3,18]. However, in the three animal species, there are some reports that reported serum albumin concentrations within the refer‐ ence range in animals [6-8,31].

The mechanisms of the hypoalbuminemia are unknown, but in human beings is primarily related to the extent of the proliferation of the MM and it is therefore of diagnostic and prog‐ nostic importance [144]. Several studies have suggested that low serum albumin concentra‐ tions correlate with increased serum concentrations of interleukin-6, a potent myeloma cell growth factor, reflecting disease severity and cell proliferation [145-146]. Interleukin-6 is a multifunctional, pro-inflammatory cytokine that stimulates B cell maturation and prolifera‐ tion and overproduction has been demonstrated in a variety of B-cell malignancies [147].

The neoplastic plasma cells are responsible for an overproduction of a homogeneous or monoclonal immunoglobulin product, known as paraproteins o M-component. The para‐ proteins may be complete immunoglobulin, free light chains, light chains fragments or poly‐ mers, or partial immunoglobulins missing one or both chains [148]. The term monoclonal gammopathy is commonly used to define hyperglobulinemia characterized by an electro‐ phoretic pattern with a sharply defined peak that is usually in the β- or γ- region and is nar‐ rower than the albumin peak [9,149]. When 2 narrow peaks with these features are recognized, the term biclonal gammopathy is often used. Monoclonal gammopathies are as‐ sociated with production of a single clone of immunoglobulin owing to clonal expansion of neoplastic lymphoid cells, such as plasma cells in MM and B cells in lymphoma [9]. In con‐ trast, a polyclonal gammopathy is characterized by a broader peak or multiple peaks in the γ- or β-γ regions. Polyclonal gammopathies are associated with chronic antigenic stimula‐ tion that occurs in chronic infections and other inflammatory conditions [148,150-152]. The term oligoclonal gammopathy refers to an electrophoretic pattern that is similar to a mono‐ clonal one, but in which the globulin peak is slightly wider than the albumin peak [9,149]. Oligoclonal gammopathy may occasionally occur in animals with chronic inflammation or infectious disease [9].

has broad implications in these patients. First, the low hemoglobin concentration and packed cell volume have been associated with poor quality of life in people, and affect daily activity. Second, anemia has an impact on the cardiovascular system. In fact, anemia has been shown to induce or aggravate hypoxia and ischemic complications. Third, anemia has

Data concerning white blood cell count (total and different subpopulations) are not consis‐ tent in animals with MM. It has been found lymphopenia [24,30,32], leukocytosis due to neutrophilia [4,8], neutropenia with lymphocytosis [31], leukopenia [1,25,37], leukocytosis with neutrophilia and lymphocytosis [31], neutropenia [16], neutrophenia with lymphope‐

Thrombocytopenia is reported in approximately one third of all canine patients of MM [10,25,27]. It has also been described in cats [4,29] and horses [18,37]. However, there are many other reports of MM in veterinary medicine with patients that show normal number of platelets [3,6-8,16,22,24,31-32]. Thrombocytopenia that could promote bleeding disorders, is proposed to result from infiltration of bone marrow by malignant plasma cells, consump‐ tion of platelets as part of thrombohemorrhagic syndrome, such as disseminated intravascu‐ lar coagulation, shortened platelet half-life or immune-mediated destruction, even though the latter 2 mechanisms have yet to be verified in veterinary medicine [10,27,30,64,142-143].

Hyperproteinemia, specifically hypoalbuminemia and hyperglobulinemia is very common in MM, but not an invariable feature. Hypoalbuminemia has been described consistently in MM in dogs [9,15-16,22-24], cats [3,9,29,32] and horses [1,3,18]. However, in the three animal species, there are some reports that reported serum albumin concentrations within the refer‐

The mechanisms of the hypoalbuminemia are unknown, but in human beings is primarily related to the extent of the proliferation of the MM and it is therefore of diagnostic and prog‐ nostic importance [144]. Several studies have suggested that low serum albumin concentra‐ tions correlate with increased serum concentrations of interleukin-6, a potent myeloma cell growth factor, reflecting disease severity and cell proliferation [145-146]. Interleukin-6 is a multifunctional, pro-inflammatory cytokine that stimulates B cell maturation and prolifera‐ tion and overproduction has been demonstrated in a variety of B-cell malignancies [147].

The neoplastic plasma cells are responsible for an overproduction of a homogeneous or monoclonal immunoglobulin product, known as paraproteins o M-component. The para‐ proteins may be complete immunoglobulin, free light chains, light chains fragments or poly‐ mers, or partial immunoglobulins missing one or both chains [148]. The term monoclonal gammopathy is commonly used to define hyperglobulinemia characterized by an electro‐ phoretic pattern with a sharply defined peak that is usually in the β- or γ- region and is nar‐ rower than the albumin peak [9,149]. When 2 narrow peaks with these features are

nia [18], pancytopenia [10] and absence of white blood cell abnormalities [3,6,22].

been shown to be a poor prognostic factor in MM [140-142].

296 Multiple Myeloma - A Quick Reflection on the Fast Progress

*5.2.1. Serum protein concentrations and serum protein electrophoresis*

**5.2. Blood clinical biochemistry**

ence range in animals [6-8,31].

Although monoclonal gammopathy is the laboratory landmark of MM, other conditions oc‐ casionally can induce a monoclonal gammopathy in animals, such as chronic inflammation (leishmaniosis, ehrlichiosis, chronic pioderma, feline infectious peritonitis) [149-150,153-155], amyloidosis [156-157], B-cell lymphoma [149], Waldenströms macroglo‐ bulinemia [158-159] and monoclonal gammopathy of undefined significance (MGUS) [160]. The inclusion criterion for MGUS are M-protein and <10% bone marrow plasmacytosis, with no evidence of lytic lesions, light chain proteinuria or other clinical, hematologic, and bio‐ chemical abnormalities [161-162]. MGUS occurs in 1-2% of people over the age of 50 and 3% of people over the age of 70 [162]. A significant proportion (25%) of these will evolve within 20 years into MM, primary amyloidosis, macroglobulinemia or another lymphoproliferative disease [162].

In dogs affected by MM, the incidence of IgA and IgG is comparable, whereas in cats and horses IgG is most commonly involved [1,3-4,27]. In fact, of the 25 published feline MM with immunoelectrophoresis results, 20 had IgG gammopathies, and 5 had IgA gammopathies [2,13-14,85]. Similarly, there are some reports of IgA gammopathies in horses with MM [3,8].

Biclonal gammopathy, with two M-components has been reported in humans [163-165], even though it was found to be very rare, occurring in about 1% of human beings with MM [166]. Biclonal gammopathies have been described in lymphoproliferative disorders in dogs and cats [2,9,19,23,167-168] including MM. The term biclonal is applied to the electrophoret‐ ic pattern and does not always correlate with true biclonal expansion because the biclonal electrophoretic may arise from a single clone of B-cells, usually mature plasma cells that produce one type of immunoglobulin with different dimerization patterns [9]. The biclonal pattern may also occur from production of two different classes of immunoglobulins, usual‐ ly IgG and IgA, by two separate cell clones [23]. However, production of separate heavy chain isotypes by a single clone of neoplastic cells, may result from isotype switching, which occurs normally during B-cell maturation [9].

The prevalence of biclonal gammopathy in companion animals is unknown, but it could be higher than reported. In many clinical veterinary laboratories, serum protein electrophoresis is performed using cellulose acetate as the support medium. However, better separation of protein fractions may be obtained using agar cell electrophoresis or capillary zone electro‐ phoresis. Facchini et al. [9] reported two cases (dog and cat) with gammopathies associated with MM that were interpreted as oligoclonal by standard cellulose acetate electrophoresis but were determined to be biclonal con capillary zone electrophoresis.

The typical clinical signs of cryoglobulinemia are purpura, cold intolerance, acrocyanosis and ulceration, necrosis and gangrene of the skin of the distal extremities [184]. However of the cases of cryoglobulinemia reported in the veterinary literature, only 1 dog [183] and 2 horses [180,182] had typical lesions. Necrosis of the pinnae occurred in the dog [183] and in 1 of the horses [180] and distal limb swelling and ulceration in other horse [182]. Similarly, <50% of human patients with cryoglobulinemia have typical clinical signs even in cold con‐ ditions [184]. The lesions develop as a result of precipitation of cryoglobulin in small-diame‐ ter blood vessels, which causes vascular occlusion and tissue ischemia. Subsequently, inflammation may develop at the site of precipitation secondary to complement fixation by immunoglobulin G [185]. Despite extensive investigation, the physical and chemical charac‐ teristics accounting for the temperature-dependent solubility of cryoglobulins have not been determined. Proposed mechanisms include altered amino acid or carbohydrate content of the cryoglobin, leading to abnormal interactions between water and the protein [185].

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Azotemia is present in half of MM human patients when first evaluated [78-79]. Similarly, many animals with MM show azotemia when presented [1,4,10,24,27,29], whereas there are other cases with serum urea and creatinine concentrations within the reference limits [3,6,22]. Probably these differences depend on the existence of renal kidney and on the de‐

Hypercalcemia has been reported in approximately 15-20% of dogs and 20-25% of cats af‐ fected by MM [4,29,65]. Similarly, hypercalcemia seems to be common in horses with MM

Hypercalcemia is a complication of uncontrolled osteolysis, influenced in part by osteoclast activation factor and in human patients has been associated with extensive osteolytic disease [86]. However, MM-associated hypercalcemia is not reported as frequently as bone lysis [4,27,35,64], perhaps because disease progression is usually slow, allowing for appropriate metabolic controls. Other mechanisms of hypercalcemia are the release of an osteoclast acti‐ vating factor by either the bone marrow microenvironment or by neoplastic cells located in bone [186-190]. In humans, interleukin-1, interleukin-6, tumor necrosis factor and the recep‐ tor activator of the nuclear factor kappa B ligand (RANKL) all modulated osteoclast activity and may contribute to hypercalcemia [191]. Paratyroid hormone related peptide (PTH-rP) also may contribute to the pathogenesis of MM-related hypercalcemia. It seems that essen‐ tially every cell of the body makes PTHrP under normal conditions [192-193]. It has a broad range of physiological functions, including stimulation of bone resorption, vasorelaxation, and cell proliferation, regulation of placental calcium transport, organogenesis, parturition, lactation, and vascular smooth muscle proliferation and development of the skeletal system [193]. Despite its wide distribution of the body, PTHrP is normally present in minute amounts in the circulation [193-194] and high serum PTHrP concentrations have been found in conjunction only with pathologic conditions, principally malignancy [194-195]. PTHrP

[1,3,8], although there is a report of hypocalcemia in one horse with MM [18].

*5.2.2. Serum urea and creatinine concentrations*

*5.2.3. Serum total and ionized calcium concentrations*

gree of hydration.

MMs in veterinary patients lacking hyperglobulinemia have also been described [10]. The authors propose that the lack of hyperglobulinemia resulted from either M-protein associat‐ ed secondary hypogammaglobulinemia or the IgA nature of the M protein. Secondary hypo‐ gammaglobulinemia with an immunosuppressive phenomenon associated with MM is reported to occur in about 10% of human MM patients, and in a report, is most commonly seen in secretory immunoglobulin A- MM [169-172]. The mechanism underlying MM-asso‐ ciated hypogammaglobulinemia is unclear, but recent work suggest that appropriate B-cell maturation and immunoglobulin production are impaired by defects in CD4+, CD45R+, na‐ ïve T cells and increases in CD8+, CD11b1+ memory T cells [173-174]. The depression of nor‐ mal immunoglobulin production associated with exuberant M protein production has been described anecdotally, but not specifically described in dogs with MM [64]. Seelig et al. [10] supported their hypothesis in the immunoglobulin quantification data, which indicate mas‐ sive production of the immunoglobulin A M-protein and mild to moderate decreases in im‐ munoglobulins G and M in a dog.

Cryoglobulinemia has also been described in human patients with MM [175-176]. Cryoglo‐ bulins are proteins, usually immunoglobulins that precipitate as serum is cooled to tempera‐ ture less than body temperature and dissolve upon rewarming. They are most commonly evident as a white gelatinous material but may sometimes appear crystalline or flocculent [177] Cryoglobulinemia is rare in animals and are limited descriptions in dogs with MM [114,178], in a dog with Waldenström's macroglobulenia [179], in a cat [28], in a horse with lymphoma [180], and in several horses with glomerulonephritis [181-182]. In human pa‐ tients, cryoglobulins are classified into 3 groups on the basis of their immunoglobulin com‐ position. In type-1 cryoglobulinemia a single monoclonal immunoglobulin, usually IgM is present. This is most commonly associated with lymphoproliferative disorders such as MM, Waldenström's macroglobulinemia, lymphoma, and lymphocytic leukemia, but occasionally can develop in conjunction with immune-mediated diseases [177]. In type II cryoglobuline‐ mia, a monoclonal immunoglobulin, usually IgM complexes with polyclonal IgG, whereas in type III cryoglobulinemia, polyclonal immunoglobulins, usually immunoglobulin M com‐ plex with polyclonal immunoglobulin G. Type II and III cryoglobulinemia and may develop secondary to infection, immune-mediated diseases or very rarely, lymphoproliferative dis‐ ease. In some instances, an underlying disease is not found and the cryoglobulinemia is de‐ scribed as essential. Using this classification system, type-I and mixed cryoglobulinemia in dogs, horses and cats have been described. Two dogs with MM had type-I immunoglobulin A cryoglobulinemia [114,178] and the dog with Waldenström's macroglobulinemia had type-I immunoglobulin M cryoglobulinemia [179]. Another dog had an essential mixed im‐ munoglobulin G-M cryoglobulinemia and cryofibrinogenemia [183], and although a thor‐ ough investigation for the underlying disease was not performed, the diagnosis was supported by resolution and lack of recurrence of clinical signs when the dog was main‐ tained in a warm environment [183]. A cat had type I immunoglobulin G cryoglobulinemia in association with MM [28].

The typical clinical signs of cryoglobulinemia are purpura, cold intolerance, acrocyanosis and ulceration, necrosis and gangrene of the skin of the distal extremities [184]. However of the cases of cryoglobulinemia reported in the veterinary literature, only 1 dog [183] and 2 horses [180,182] had typical lesions. Necrosis of the pinnae occurred in the dog [183] and in 1 of the horses [180] and distal limb swelling and ulceration in other horse [182]. Similarly, <50% of human patients with cryoglobulinemia have typical clinical signs even in cold con‐ ditions [184]. The lesions develop as a result of precipitation of cryoglobulin in small-diame‐ ter blood vessels, which causes vascular occlusion and tissue ischemia. Subsequently, inflammation may develop at the site of precipitation secondary to complement fixation by immunoglobulin G [185]. Despite extensive investigation, the physical and chemical charac‐ teristics accounting for the temperature-dependent solubility of cryoglobulins have not been determined. Proposed mechanisms include altered amino acid or carbohydrate content of the cryoglobin, leading to abnormal interactions between water and the protein [185].

#### *5.2.2. Serum urea and creatinine concentrations*

with MM that were interpreted as oligoclonal by standard cellulose acetate electrophoresis

MMs in veterinary patients lacking hyperglobulinemia have also been described [10]. The authors propose that the lack of hyperglobulinemia resulted from either M-protein associat‐ ed secondary hypogammaglobulinemia or the IgA nature of the M protein. Secondary hypo‐ gammaglobulinemia with an immunosuppressive phenomenon associated with MM is reported to occur in about 10% of human MM patients, and in a report, is most commonly seen in secretory immunoglobulin A- MM [169-172]. The mechanism underlying MM-asso‐ ciated hypogammaglobulinemia is unclear, but recent work suggest that appropriate B-cell maturation and immunoglobulin production are impaired by defects in CD4+, CD45R+, na‐ ïve T cells and increases in CD8+, CD11b1+ memory T cells [173-174]. The depression of nor‐ mal immunoglobulin production associated with exuberant M protein production has been described anecdotally, but not specifically described in dogs with MM [64]. Seelig et al. [10] supported their hypothesis in the immunoglobulin quantification data, which indicate mas‐ sive production of the immunoglobulin A M-protein and mild to moderate decreases in im‐

Cryoglobulinemia has also been described in human patients with MM [175-176]. Cryoglo‐ bulins are proteins, usually immunoglobulins that precipitate as serum is cooled to tempera‐ ture less than body temperature and dissolve upon rewarming. They are most commonly evident as a white gelatinous material but may sometimes appear crystalline or flocculent [177] Cryoglobulinemia is rare in animals and are limited descriptions in dogs with MM [114,178], in a dog with Waldenström's macroglobulenia [179], in a cat [28], in a horse with lymphoma [180], and in several horses with glomerulonephritis [181-182]. In human pa‐ tients, cryoglobulins are classified into 3 groups on the basis of their immunoglobulin com‐ position. In type-1 cryoglobulinemia a single monoclonal immunoglobulin, usually IgM is present. This is most commonly associated with lymphoproliferative disorders such as MM, Waldenström's macroglobulinemia, lymphoma, and lymphocytic leukemia, but occasionally can develop in conjunction with immune-mediated diseases [177]. In type II cryoglobuline‐ mia, a monoclonal immunoglobulin, usually IgM complexes with polyclonal IgG, whereas in type III cryoglobulinemia, polyclonal immunoglobulins, usually immunoglobulin M com‐ plex with polyclonal immunoglobulin G. Type II and III cryoglobulinemia and may develop secondary to infection, immune-mediated diseases or very rarely, lymphoproliferative dis‐ ease. In some instances, an underlying disease is not found and the cryoglobulinemia is de‐ scribed as essential. Using this classification system, type-I and mixed cryoglobulinemia in dogs, horses and cats have been described. Two dogs with MM had type-I immunoglobulin A cryoglobulinemia [114,178] and the dog with Waldenström's macroglobulinemia had type-I immunoglobulin M cryoglobulinemia [179]. Another dog had an essential mixed im‐ munoglobulin G-M cryoglobulinemia and cryofibrinogenemia [183], and although a thor‐ ough investigation for the underlying disease was not performed, the diagnosis was supported by resolution and lack of recurrence of clinical signs when the dog was main‐ tained in a warm environment [183]. A cat had type I immunoglobulin G cryoglobulinemia

but were determined to be biclonal con capillary zone electrophoresis.

munoglobulins G and M in a dog.

298 Multiple Myeloma - A Quick Reflection on the Fast Progress

in association with MM [28].

Azotemia is present in half of MM human patients when first evaluated [78-79]. Similarly, many animals with MM show azotemia when presented [1,4,10,24,27,29], whereas there are other cases with serum urea and creatinine concentrations within the reference limits [3,6,22]. Probably these differences depend on the existence of renal kidney and on the de‐ gree of hydration.

#### *5.2.3. Serum total and ionized calcium concentrations*

Hypercalcemia has been reported in approximately 15-20% of dogs and 20-25% of cats af‐ fected by MM [4,29,65]. Similarly, hypercalcemia seems to be common in horses with MM [1,3,8], although there is a report of hypocalcemia in one horse with MM [18].

Hypercalcemia is a complication of uncontrolled osteolysis, influenced in part by osteoclast activation factor and in human patients has been associated with extensive osteolytic disease [86]. However, MM-associated hypercalcemia is not reported as frequently as bone lysis [4,27,35,64], perhaps because disease progression is usually slow, allowing for appropriate metabolic controls. Other mechanisms of hypercalcemia are the release of an osteoclast acti‐ vating factor by either the bone marrow microenvironment or by neoplastic cells located in bone [186-190]. In humans, interleukin-1, interleukin-6, tumor necrosis factor and the recep‐ tor activator of the nuclear factor kappa B ligand (RANKL) all modulated osteoclast activity and may contribute to hypercalcemia [191]. Paratyroid hormone related peptide (PTH-rP) also may contribute to the pathogenesis of MM-related hypercalcemia. It seems that essen‐ tially every cell of the body makes PTHrP under normal conditions [192-193]. It has a broad range of physiological functions, including stimulation of bone resorption, vasorelaxation, and cell proliferation, regulation of placental calcium transport, organogenesis, parturition, lactation, and vascular smooth muscle proliferation and development of the skeletal system [193]. Despite its wide distribution of the body, PTHrP is normally present in minute amounts in the circulation [193-194] and high serum PTHrP concentrations have been found in conjunction only with pathologic conditions, principally malignancy [194-195]. PTHrP may be synthesized by normal cells activated by the presence of a malignancy or by neo‐ plastic cells. There is a report of a horse with MM and high concentrations of PTHrP [8]). High levels of PTHrP have been also described in other neoplasms, such as thymoma [196], nasal carcinoma [197], squamous cell carcinomas [198], angiomyxoma [199], mammary car‐ cinoma [200], lymphoma [201-203] adenocarcinoma of the apocrine gland of the anal sac [203] and malignant melanoma [204].

*5.2.6. Alterations in serum cholesterol concentrations*

even is not always used in veterinary medicine.

**5.3. Urinalysis**

MM [1,3].

**5.4. Diagnostic ancillary aids**

nutrition, hepatic insufficiency or hyperthyroidism [213-215].

Hypocholesterolemia has been noted in approximately 69% of affected cats in one study [4]. Similarly, hypocholesterolemia was found in a horse with MM [1]. The incidence in cats is higher than in human patients [211-212]. Serum cholesterol concentrations are thought to be correlated inversely with globulin concentrations [35,211]. It has been postulated that the hypocholesterolemia is the result of a down-regulation of cholesterol production by the liver to maintain oncotic pressure in the face of hyperglobulinemia [78]). The main causes of hy‐ pocholesterolemia in veterinary medicine include protein-losing enteropathies, severe mal‐

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Proteinuria, detected by routine urinalysis is present in 90%of human patients with MM [216-221]. In the same way, proteinuria using dipsticks has been found in horses [1,3,6], dogs [10,22] and cats [4,28,30,32] with MM. However, dipsticks detect primarily albuminu‐ ria and therefore, the sulfosalicylic acid test (SSA) provides greater sensitivity for globulin detection but specificity is low due to the concomitant detection of albumin, globulin, Bence Jones proteins, proteases and polypeptides. False positive SSA results may occur with peni‐ cillin and its derivatives, tolbutamide or sulfisoxazole metabolites, or certain contrast media in the urine [222]. In people, false positive results for Bence Jones proteins detected by means of heat precipitation can occur due to excessive amount of polyclonal light chain pro‐ teins in patients affected by a variety of conditions, including connective tissue diseases, non-plasmacytic tumors and chronic renal failure [4,216,222]. Therefore, urine protein elec‐ trophoresis remains the preferred diagnostic modality to detect monoclonal proteinuria,

Bence Jones proteinuria has been estimated to occur in approximately 25-40% of dogs an ap‐ proximately 65% of cats with MM [4,27,29]. Similarly, it has been determined in horses with

Survey radiographies and echographies are required for screening of skeletal lesions and identification of abdominal organ neoplasia respectively. Furthermore, a funduscopic ex‐ amination should be carried out in patients suspicious of MM, mainly in small animals, in order to rule out ocular lesions associated with the HVS, such as retinal hemorrhages, retinal

As explained before, skeletal lesions vary from areas of osteopenias observed in early stages of the disease to lytic lesions typical of later stages. Biopsy and histopathology of a lytic le‐ sion may sometimes be necessary for a definitive diagnosis. Survey radiographies are com‐ monly performed now in small animals with MM, because increased clinician awareness of the lesions, better quality radiographs and increased knowledge about the disease. In horses with MM, survey radiographies are less used, probably because of the higher incidence of skeletal lesions of other origin (sport horses). Lung radiography is recommended in equine

detachment, venous tortuosity, dilation, sacculation, and blindness.

The measurement of ionized calcium is recommended to confirm hypercalcemia in these pa‐ tients, because binding of calcium by the M protein will increase total calcium concentration, while ionized calcium will remain within normal limits [205]. Therefore, increased ionized calcium concentrations supports true hypercalcemia. There is a recent report that present a case of MM in a dog with increased total serum calcium concentration, but with serum ion‐ ized calcium concentrations within normal limits [25]. The authors suggested that the major‐ ity of the calcium was protein-bound to serum M proteins. In fact, serum calcium exists in two major fractions, free and protein bound. A small portion of calcium is bound to other small anions such as citrate, lactate, and phosphate. Because ionized calcium is the physio‐ logically active species of blood calcium, it is ordinarily maintained within very narrow lim‐ its by rigidly controlled mechanisms [205-206]. Approximately half of normal total serum calcium is bound to negatively charged sites on albumin. Human MM has been reported to bind calcium on the Fab portion of the globulin molecules [148].

MM is frequently complicated by an increase in the concentration of ionized calcium, which if persistent leads to secondary nephrogenic diabetes insipidus and loss of the renal medul‐ lary concentration gradient causing polyuria and polydipsia [25].

#### *5.2.4. Alterations in the coagulation profile*

Approximately 50% of dogs affected by MM have abnormal prothrombin and partial throm‐ boplastin times [149,207] and these abnormalities have also been found in horses [6,18,37,86] and cats [4,29]. However, other animals with MM had normal bleeding times [3,22].

Coagulation defects can result from paraproteins interference with clotting factors, protein coating of platelets leading to thrombocytopenia and binding of the Fab fragment of the Mprotein to fibrin, preventing aggregation [116,208].

#### *5.2.5. Alterations in serum sodium concentrations*

Three horses with MM were hyponatremia [1]. The decreased concentrations could have re‐ sulted from displacement of the aqueous phase of plasma by the hyperglobulinemia. How‐ ever, true hyponatremia in human beings with MM has been described. Suggested mechanism include displacement of sodium by cationic paraproteins, decreased plasma wa‐ ter secondary to unusual hydration characteristics of paraproteins, and syndrome of inap‐ propriate antidiuretic hormone release [209-210]. Alterations in serum sodium concentrations do not appear to be common in small animals.

#### *5.2.6. Alterations in serum cholesterol concentrations*

Hypocholesterolemia has been noted in approximately 69% of affected cats in one study [4]. Similarly, hypocholesterolemia was found in a horse with MM [1]. The incidence in cats is higher than in human patients [211-212]. Serum cholesterol concentrations are thought to be correlated inversely with globulin concentrations [35,211]. It has been postulated that the hypocholesterolemia is the result of a down-regulation of cholesterol production by the liver to maintain oncotic pressure in the face of hyperglobulinemia [78]). The main causes of hy‐ pocholesterolemia in veterinary medicine include protein-losing enteropathies, severe mal‐ nutrition, hepatic insufficiency or hyperthyroidism [213-215].

#### **5.3. Urinalysis**

may be synthesized by normal cells activated by the presence of a malignancy or by neo‐ plastic cells. There is a report of a horse with MM and high concentrations of PTHrP [8]). High levels of PTHrP have been also described in other neoplasms, such as thymoma [196], nasal carcinoma [197], squamous cell carcinomas [198], angiomyxoma [199], mammary car‐ cinoma [200], lymphoma [201-203] adenocarcinoma of the apocrine gland of the anal sac

The measurement of ionized calcium is recommended to confirm hypercalcemia in these pa‐ tients, because binding of calcium by the M protein will increase total calcium concentration, while ionized calcium will remain within normal limits [205]. Therefore, increased ionized calcium concentrations supports true hypercalcemia. There is a recent report that present a case of MM in a dog with increased total serum calcium concentration, but with serum ion‐ ized calcium concentrations within normal limits [25]. The authors suggested that the major‐ ity of the calcium was protein-bound to serum M proteins. In fact, serum calcium exists in two major fractions, free and protein bound. A small portion of calcium is bound to other small anions such as citrate, lactate, and phosphate. Because ionized calcium is the physio‐ logically active species of blood calcium, it is ordinarily maintained within very narrow lim‐ its by rigidly controlled mechanisms [205-206]. Approximately half of normal total serum calcium is bound to negatively charged sites on albumin. Human MM has been reported to

MM is frequently complicated by an increase in the concentration of ionized calcium, which if persistent leads to secondary nephrogenic diabetes insipidus and loss of the renal medul‐

Approximately 50% of dogs affected by MM have abnormal prothrombin and partial throm‐ boplastin times [149,207] and these abnormalities have also been found in horses [6,18,37,86]

Coagulation defects can result from paraproteins interference with clotting factors, protein coating of platelets leading to thrombocytopenia and binding of the Fab fragment of the M-

Three horses with MM were hyponatremia [1]. The decreased concentrations could have re‐ sulted from displacement of the aqueous phase of plasma by the hyperglobulinemia. How‐ ever, true hyponatremia in human beings with MM has been described. Suggested mechanism include displacement of sodium by cationic paraproteins, decreased plasma wa‐ ter secondary to unusual hydration characteristics of paraproteins, and syndrome of inap‐ propriate antidiuretic hormone release [209-210]. Alterations in serum sodium

and cats [4,29]. However, other animals with MM had normal bleeding times [3,22].

[203] and malignant melanoma [204].

300 Multiple Myeloma - A Quick Reflection on the Fast Progress

*5.2.4. Alterations in the coagulation profile*

protein to fibrin, preventing aggregation [116,208].

concentrations do not appear to be common in small animals.

*5.2.5. Alterations in serum sodium concentrations*

bind calcium on the Fab portion of the globulin molecules [148].

lary concentration gradient causing polyuria and polydipsia [25].

Proteinuria, detected by routine urinalysis is present in 90%of human patients with MM [216-221]. In the same way, proteinuria using dipsticks has been found in horses [1,3,6], dogs [10,22] and cats [4,28,30,32] with MM. However, dipsticks detect primarily albuminu‐ ria and therefore, the sulfosalicylic acid test (SSA) provides greater sensitivity for globulin detection but specificity is low due to the concomitant detection of albumin, globulin, Bence Jones proteins, proteases and polypeptides. False positive SSA results may occur with peni‐ cillin and its derivatives, tolbutamide or sulfisoxazole metabolites, or certain contrast media in the urine [222]. In people, false positive results for Bence Jones proteins detected by means of heat precipitation can occur due to excessive amount of polyclonal light chain pro‐ teins in patients affected by a variety of conditions, including connective tissue diseases, non-plasmacytic tumors and chronic renal failure [4,216,222]. Therefore, urine protein elec‐ trophoresis remains the preferred diagnostic modality to detect monoclonal proteinuria, even is not always used in veterinary medicine.

Bence Jones proteinuria has been estimated to occur in approximately 25-40% of dogs an ap‐ proximately 65% of cats with MM [4,27,29]. Similarly, it has been determined in horses with MM [1,3].

#### **5.4. Diagnostic ancillary aids**

Survey radiographies and echographies are required for screening of skeletal lesions and identification of abdominal organ neoplasia respectively. Furthermore, a funduscopic ex‐ amination should be carried out in patients suspicious of MM, mainly in small animals, in order to rule out ocular lesions associated with the HVS, such as retinal hemorrhages, retinal detachment, venous tortuosity, dilation, sacculation, and blindness.

As explained before, skeletal lesions vary from areas of osteopenias observed in early stages of the disease to lytic lesions typical of later stages. Biopsy and histopathology of a lytic le‐ sion may sometimes be necessary for a definitive diagnosis. Survey radiographies are com‐ monly performed now in small animals with MM, because increased clinician awareness of the lesions, better quality radiographs and increased knowledge about the disease. In horses with MM, survey radiographies are less used, probably because of the higher incidence of skeletal lesions of other origin (sport horses). Lung radiography is recommended in equine patients with MM in order to rule out pneumonia, one common finding in these cases [3,18]. In addition, hepatomegaly (58%), splenomegaly (25%), cardiomegaly (67%) and renomegaly (9%) have been detected in cats with MM [4,29].

ieved, chemotherapy is often effective in decreasing tumor burden, reducing serum immunoglobulin levels, promoting bone remodeling and providing symptomatic relief. Fur‐ ther, this treatment leads to improved quality and possible duration of life [27,225] and is

Multiple Myeloma in Horses, Dogs and Cats: A Comparative Review Focused on Clinical Signs and Pathogenesis

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303

Melphalan is an alkylating agent whose oral absorption is unpredictable, requiring adminis‐ tration to be made preferable on the empty stomach. In dogs, melphalan is initially adminis‐ tered at 0.1 mg/kg PO once a day for 10 days and then 0.05 mg/kg PO every other day. In cats, they are administered at 0.5-2.0 mg/kg PO once a day. Melphalan is combined with glucocorticoids, prednisone [22,24-25,42] or less common, prednisolone [25,30-31]. Glucocor‐ ticoids have been shown to induce apoptosis in vitro via inhibition of Iκβ activation and de‐ creased nuclear factor κB activity [231-232]. Prednisone in dogs with MM is generally administered at 1.0 mg/kg for the first 10 days of therapy and then, decreased to 0.5 mg/kg every other day [22,24-25,32,225]. This combination treatment is continued indefinitely, until

Other alkylating drugs including cyclophosphamide, chlorambucil and 1(2-chloroethyl)-3 cyclohexyl-1-nitrosourea (CCNU) have also been used to treat MM in small animals [10,233]. The addition of cyclophosphamide to a prednisone- melphalan regimen may be beneficial to patients with severe clinical signs and/or hypercalcemia. Due to its platelet-sparing effect, cyclophosphamide may be used in place of melphalan in thrombocytopenic patients, al‐ though this drug can have severe suppressive effects on other bone marrow lineages [234-237]. Chorambucil, administered at 0.2 mg/kg PO once daily has been used successful

Combination chemotherapy protocols incorporating vincristine, carmustine, melphalan, cy‐ clophosphamide and prednisone or vincristine, melphalan, cyclophosphamide and predni‐ sone have been used in human beings, but outcomes are essentially comparable to those of patients with melphalan and prednisone alone [238-241]. The administration of high dose dexamethasone in conjunction with vincristine and doxorubicin was investigated in humans with refractory MM and resulted in a response greater than 50%. Rapid tumor response, al‐ leviation of bone pain, resolution of hypercalcemia and absence of damage to bone marrow stem cells were remarkable advantages to this treatment combination [239]. Anecdotally, in dogs, responses of a few months duration have been achieved with a combination of doxor‐

The efficacy of inteferon for the treatment of MM is controversial [244-245]. While a re‐ sponse rate of approximately 20% was reported in humans with relapsed MM, the addition of interferon to standard chemotherapy approaches failed to provide a significant benefit to the overall survival time in a meta-analysis of 2286 patients [244]. However, there is other report that stated that even though most interferon benefits to MM patients are relatively small when viewed in the light of survival expectancies, they seem clinically relevant. Since median overall survival of conventionally treated MM patients ranges between 30 and 50

for the treatment of immunoglobulin M macroglobulinemia in dogs [159].

ubicin, vincristine and prednisone in lymphoma [242-243] and in MM [10].

months, 3-7 months gains of life amount to a increase of 10-25% [244].

also used in human patients with MM [226-230].

relapse or myelosuppression.

The most common ultrasonographic abnormalities involved the spleen and the liver and to a lesser extent the kidneys [4]. In cats, the most consistent finding in MM is splenic enlarge‐ ment and diffuse or nodular hypoechogenicity. The most consistent hepatic abnormality is diffuse hyperechogenicity and enlargement [4]. Bone marrow cytology should be done to confirm the diagnosis. A detailed description of the histopathological characteristics in bone marrow and in other organs after necropsy, consistent with MM is out of the scope of the present review.

#### **5.5. Diagnostic criteria**

Current published recommendations for determining a diagnosis in veterinary medicine in‐ dicate that the animal should have at least 2 of the following 4 criteria: 1) Bone marrow plas‐ macytosis with >20% plasma cells; 2) Monoclonal gammopathy based on serum protein electrophoresis; 3) Osteolysis and 4) Light chain (Bence-Jones) proteinuria (2). These criteria are unweighted for animal patients. In human patients, criteria are weighted as 'major' and 'minor' and accommodate lower plasma cell percentages (17). In people, confirmation of MM requires first that the patient be symptomatic (i.e. have bone pain) or have anemia, hy‐ percalcemia, azotemia, hypoalbuminemia, or bone demineralization. The diagnostic criteria of MM are then applied. Major criteria include: 1) plasmacytoma(s) with biopsy; 2) marrow plasmatocytosis >30%; 3) M-protein with >3.5 g/dl immunoglobulin G or 2.0 g/dl immuno‐ globulin A and 4) κ or λ chain excretion on 24-h urine protein electrophoresis. The 4 minor criteria include: a) marrow plasmacytosis with 10-30% plasma cells; b) M-protein at values less than indicated above; c) lytic bone lesions and d) >50% normal serum immunoglobulin concentration. If the diagnosis includes major criteria, then any 2 of the 4 will suffice, or ma‐ jor criterion 1 plus minor criterion b, c, or d; or major criterion 3 plus minor criterion a or c. If the diagnosis is based on only minor criteria, then it must include the first and second cri‐ teria (a and b), plus 1 of the remaining 2 criteria (c or d). This system has been recently incor‐ porated for the diagnosis in animals [4,10,24,31-32]. However, some modifications have been introduced. Plasma cell atypia has been included as a criterion when marrow plasmacytosis was between 10 and 20%. In humans, nuclear-cytoplasmic maturation asynchrony, nuclear immaturity, and pleomorphism are considered reliable markers for distinguishing neoplas‐ tic cells from reactive plasma cells [223]. In addition, reactive plasma cells usually do not ex‐ ceed 5% of all nucleated cells in marrow and are well differentiated [223-224].

#### **6. Treatment options in veterinary patients with mm**

Treatment of MM with oral melphalan and glucocorticoids (prednisone or prednisolone) is the standard of therapy due to its dual ability to reduce the bulk of the tumor and the symp‐ toms of the decrease [16,22-25,31-33,225]. Although complete eradication is only rarely ach‐ ieved, chemotherapy is often effective in decreasing tumor burden, reducing serum immunoglobulin levels, promoting bone remodeling and providing symptomatic relief. Fur‐ ther, this treatment leads to improved quality and possible duration of life [27,225] and is also used in human patients with MM [226-230].

patients with MM in order to rule out pneumonia, one common finding in these cases [3,18]. In addition, hepatomegaly (58%), splenomegaly (25%), cardiomegaly (67%) and renomegaly

The most common ultrasonographic abnormalities involved the spleen and the liver and to a lesser extent the kidneys [4]. In cats, the most consistent finding in MM is splenic enlarge‐ ment and diffuse or nodular hypoechogenicity. The most consistent hepatic abnormality is diffuse hyperechogenicity and enlargement [4]. Bone marrow cytology should be done to confirm the diagnosis. A detailed description of the histopathological characteristics in bone marrow and in other organs after necropsy, consistent with MM is out of the scope of the

Current published recommendations for determining a diagnosis in veterinary medicine in‐ dicate that the animal should have at least 2 of the following 4 criteria: 1) Bone marrow plas‐ macytosis with >20% plasma cells; 2) Monoclonal gammopathy based on serum protein electrophoresis; 3) Osteolysis and 4) Light chain (Bence-Jones) proteinuria (2). These criteria are unweighted for animal patients. In human patients, criteria are weighted as 'major' and 'minor' and accommodate lower plasma cell percentages (17). In people, confirmation of MM requires first that the patient be symptomatic (i.e. have bone pain) or have anemia, hy‐ percalcemia, azotemia, hypoalbuminemia, or bone demineralization. The diagnostic criteria of MM are then applied. Major criteria include: 1) plasmacytoma(s) with biopsy; 2) marrow plasmatocytosis >30%; 3) M-protein with >3.5 g/dl immunoglobulin G or 2.0 g/dl immuno‐ globulin A and 4) κ or λ chain excretion on 24-h urine protein electrophoresis. The 4 minor criteria include: a) marrow plasmacytosis with 10-30% plasma cells; b) M-protein at values less than indicated above; c) lytic bone lesions and d) >50% normal serum immunoglobulin concentration. If the diagnosis includes major criteria, then any 2 of the 4 will suffice, or ma‐ jor criterion 1 plus minor criterion b, c, or d; or major criterion 3 plus minor criterion a or c. If the diagnosis is based on only minor criteria, then it must include the first and second cri‐ teria (a and b), plus 1 of the remaining 2 criteria (c or d). This system has been recently incor‐ porated for the diagnosis in animals [4,10,24,31-32]. However, some modifications have been introduced. Plasma cell atypia has been included as a criterion when marrow plasmacytosis was between 10 and 20%. In humans, nuclear-cytoplasmic maturation asynchrony, nuclear immaturity, and pleomorphism are considered reliable markers for distinguishing neoplas‐ tic cells from reactive plasma cells [223]. In addition, reactive plasma cells usually do not ex‐

ceed 5% of all nucleated cells in marrow and are well differentiated [223-224].

Treatment of MM with oral melphalan and glucocorticoids (prednisone or prednisolone) is the standard of therapy due to its dual ability to reduce the bulk of the tumor and the symp‐ toms of the decrease [16,22-25,31-33,225]. Although complete eradication is only rarely ach‐

**6. Treatment options in veterinary patients with mm**

(9%) have been detected in cats with MM [4,29].

302 Multiple Myeloma - A Quick Reflection on the Fast Progress

present review.

**5.5. Diagnostic criteria**

Melphalan is an alkylating agent whose oral absorption is unpredictable, requiring adminis‐ tration to be made preferable on the empty stomach. In dogs, melphalan is initially adminis‐ tered at 0.1 mg/kg PO once a day for 10 days and then 0.05 mg/kg PO every other day. In cats, they are administered at 0.5-2.0 mg/kg PO once a day. Melphalan is combined with glucocorticoids, prednisone [22,24-25,42] or less common, prednisolone [25,30-31]. Glucocor‐ ticoids have been shown to induce apoptosis in vitro via inhibition of Iκβ activation and de‐ creased nuclear factor κB activity [231-232]. Prednisone in dogs with MM is generally administered at 1.0 mg/kg for the first 10 days of therapy and then, decreased to 0.5 mg/kg every other day [22,24-25,32,225]. This combination treatment is continued indefinitely, until relapse or myelosuppression.

Other alkylating drugs including cyclophosphamide, chlorambucil and 1(2-chloroethyl)-3 cyclohexyl-1-nitrosourea (CCNU) have also been used to treat MM in small animals [10,233]. The addition of cyclophosphamide to a prednisone- melphalan regimen may be beneficial to patients with severe clinical signs and/or hypercalcemia. Due to its platelet-sparing effect, cyclophosphamide may be used in place of melphalan in thrombocytopenic patients, al‐ though this drug can have severe suppressive effects on other bone marrow lineages [234-237]. Chorambucil, administered at 0.2 mg/kg PO once daily has been used successful for the treatment of immunoglobulin M macroglobulinemia in dogs [159].

Combination chemotherapy protocols incorporating vincristine, carmustine, melphalan, cy‐ clophosphamide and prednisone or vincristine, melphalan, cyclophosphamide and predni‐ sone have been used in human beings, but outcomes are essentially comparable to those of patients with melphalan and prednisone alone [238-241]. The administration of high dose dexamethasone in conjunction with vincristine and doxorubicin was investigated in humans with refractory MM and resulted in a response greater than 50%. Rapid tumor response, al‐ leviation of bone pain, resolution of hypercalcemia and absence of damage to bone marrow stem cells were remarkable advantages to this treatment combination [239]. Anecdotally, in dogs, responses of a few months duration have been achieved with a combination of doxor‐ ubicin, vincristine and prednisone in lymphoma [242-243] and in MM [10].

The efficacy of inteferon for the treatment of MM is controversial [244-245]. While a re‐ sponse rate of approximately 20% was reported in humans with relapsed MM, the addition of interferon to standard chemotherapy approaches failed to provide a significant benefit to the overall survival time in a meta-analysis of 2286 patients [244]. However, there is other report that stated that even though most interferon benefits to MM patients are relatively small when viewed in the light of survival expectancies, they seem clinically relevant. Since median overall survival of conventionally treated MM patients ranges between 30 and 50 months, 3-7 months gains of life amount to a increase of 10-25% [244].

High-dose chemotherapy in association with autologous transplantations using bone mar‐ row or blood-derived stem cells is now widely accepted for the treatment of hematological malignancies including MM. This approach yielded to complete remissions in refractory hu‐ man patients, but mortality rate due to bone marrow suppression was high. Contamination of most bone marrow and blood stem cell samples with neoplastic cells within the auto‐ graph resulted in recurrence of disease, emphasizing the need of optimize purging techni‐ ques [246-250]. Autologous bone marrow transplant has also been added to chemotherapy in the treatment of some malignancies in companion animals, such as lymphoma and acute myeloid leukemia [251-253].

protein 8 weeks after commencement of treatment reflected a less aggressive form with a

Multiple Myeloma in Horses, Dogs and Cats: A Comparative Review Focused on Clinical Signs and Pathogenesis

In a study, a possible relationship between prognosis and immunoglobulin isotype was sug‐ gested in cats with MM [2], even though there are few detailed cases. Although immunoglo‐ bulin A appears to be less commonly produced than immunoglobulin G in cats, as described before, the published cases with immunoglobulin A paraproteins had visceral involvement and decreased survival time (ranging from a few days to 6 months) [12,65,267-268]. Either immunoglobulin has been associated with clinical signs of HVS, including cardiac insuffi‐ ciency, retinal hemorrhages and neurological signs [85,268]. This phenomenon relates to the size of the paraproteins and the degree of hyperglobulinemia. Since immunoglobulin A may assume a dimeric or multimeric form, it may be more commonly associated with hypervis‐ cosity than immunoglobulin G. HVS can contribute to decreased survival time in animals

The lifespan of horses diagnosed of MM usually does not exceed two years [1,3,6,8]. There is not any published case of equine MM that attempted chemotherapy and most horses are eu‐

, M. Gómez-Díez3

1 Department of Animal Medicine and Surgery, School of Veterinary Medicine, University

2 Department of Animal Medicine and Surgery, School of Veterinary Medicine, Cardenal

3 Equine Sport Medicine Centre, School of Veterinary Medicine, University of Córdoba, Cór‐

[1] Edwards DE, Parker JW, Wilkinson JE, Helman RG. Plasma Cell Myeloma in the Horse. A Case Report and Literature Review. Journal of Veterinary Internal Medicine

[2] Bienzle D, Silverstein DC, Chaffin K. Multiple Myeloma in Cats: Variable Presenta‐ tion and Different Immunoglobin Isotypes in Two Cats. Veterinary Pathology

and F.M. Castejón3

http://dx.doi.org/10.5772/54311

305

median survival of 387 days (range between 120 and 720 days) [29].

with MM [13,23,54,85].

**Author details**

, C. Riber1

of Córdoba, Córdoba, Spain

1993; 7(3) 169-176.

2000;37(4) 364-369.

A. Muñoz1

doba, Spain

**References**

thanized owing to the advance stage of the disease.

, K. Satué2

\*Address all correspondence to: pv1mujua@uco.es

Herrera-CEU University, Valencia, Spain

, P. Trigo3

In human patients with MM, biphosphonates such as pamidronate, have been used to pre‐ vent or to delay the onset of bone lesions and associated bone pain [254-257]. Bisphospho‐ nates have been administered in dogs with appendicular osteosarcoma [258-259] and with malignant histiocytosis [260], but they have not been used in veterinary patients with MM.

Treatment of patients affected by indolent MM with the anti-angiogenic agent thalidomide resulted in a 66% response rate and the drug appeared to have potential to delay the onset of clinical signs associated with the disease [261-262]. The efficacy of thalidomide for the treatment of refractory relapsed MM has also been confirmed [263-265]. Studies evaluating the possible efficacy of thalidomide for the treatment of MM in companion animals are lack‐ ing. Bortezomib, a proteasome inhibitor, induces apoptosis of MM cells and inhibits their binding to bone marrow stromal cells, which otherwise would trigger the transcription of interleukin-6 via an NFκB-dependent pathway. In different studies a 25% response rate was achieved in human beings with MM and an overall survival time of 16 months. Further‐ more, addition of dexamethasone to the treatment regimen improved responses in 19% of treated patients [227-228,264,266].

Additional, some patients experience severe clinical signs secondary to hypercalcemia, renal dysfunction, HVS or pathologic fractures will require palliative therapy specifically directed to the clinical complications of the disease.
