Tumors of the Mammary Gland

Ch26-W0558

11/7/06

7:10 PM

Page 619

CHAPTER

26

Tumors of the Mammary Gland Susan E. Lana, Gerard R. Rutteman, Stephen J. Withrow

CANINE MAMMARY TUMORS Mammary neoplasms are among the most common tumors of the female dog. Estimates of lifetime risk for malignant tumors vary from 2% to more than 20%,1,2 a risk assessed to be exceeded twofold to fivefold by that of benign mammary tumors. Two studies of large populations of dogs have recently reported incidence information. The first study, involving a defined population of insured dogs in the United Kingdom, reported a standardized incidence rate for mammary tumors of 205 per 100,000 dogs per year.3 The second study surveyed a population of more than 80,000 insured female dogs in Sweden and found an overall incidence for any mammary tumor of 111 per 10,000 dog years at risk.4 This study also found that the incidence increased with age; at age 6 years, it was 1%; at age 8 years, 6%; and at age 10 years, 13%.4 The incidence of both benign and malignant tumors in specific populations of dogs correlates with life expectancy and is strongly reduced by the practice of ovariohysterectomy in young dogs, which is common in the United States but not encouraged in some European countries. The incidence is increased by the use of injectable progestins to prevent estrus. The risk in male dogs is 1% or less of that in female dogs. The median age of tumor manifestation is 10 to 11 years of age, with rare occurrence in dogs younger than 4 years old. Several spaniel breeds and, according to some studies, the poodle and dachshund seem to be predisposed to the condition.5-7 A recent report, however, suggested that the incidence of malignant tumors was different in small breed dogs compared with large breed dogs.8 In this study, of 101 tumors (60 small breed, 40 other), 25% of the small breed dogs had histologically malignant tumors compared with 58% of the large breed patients. The development of mammary tumors in the dog is clearly hormone dependent. Compared to the risk in intact dogs, the risk for malignant tumors in dogs spayed before the first estrus is 0.05%; after the first estrus, it is 8%; and it rises to 26% if the dog is spayed

after the second estrus.9 Later spaying does not reduce the risk for malignant tumors, although the risk for benign tumors seems to be reduced by ovariectomy even at a later age.10 In a retrospective study of 137 dogs with malignant mammary carcinomas, dogs spayed within 2 years of the development of the malignant tumor showed a survival advantage over dogs intact or spayed 2 years or longer before mastectomy.11 A protective effect of (early) pregnancy, well-known in humans, has not been demonstrated.9 Most likely, the tumor-enhancing effect of ovarian steroids, predominantly progesterone, is brought about by their mitogenic activity in mammary cells, exerted after they bind to their respective receptors. Normal mammary gland tissue contains both estrogen receptors (ERs) and progesterone receptors (PRs) receptors (as do most benign tumors), often at increased level. In contrast, carcinomas, devoid of remnants of normal mammary epithelium, contain ERs and PRs in a decreased number of cases, with rare occurrence in metastases.12-16 In a study by Millanta and colleagues,17 85 mammary lesions (28 dysplasias, 21 benign lesions, and 36 carcinomas [4 in situ, and 32 invasive]) from 47 dogs were evaluated by means of immunohistochemistry for ER and PR expression. Ten normal mammary tissues were used for comparison. The researchers found that 100% of the normal and dysplastic tissues and 95% of the benign tissues tested positive for ERs. Of the invasive carcinomas, 92% showed some ER staining, although only 62% had strong nuclear staining. No significant difference in quantitative ER expression was found between normal, dysplastic, benign, and in situ lesions, whereas the ER expression in invasive carcinomas was significantly lower. In this same study, PR expression was significantly lower in both benign and malignant lesions compared with healthy tissue.17 In another report, 228 tumors (155 malignant, 73 benign) from 100 dogs were evaluated by immunohistochemistry for ER and PR expression.18 In all tumors, one or both receptors were detected in 76% of samples (96% of benign samples, 66% of malignant 619

Ch26-W0558

11/7/06

620

7:10 PM

Page 620

Part IV • Specific Malignancies in the Small Animal Patient

samples). In seven cases of lymph node metastasis, both the primary tumor and the node were considered ER and PR negative. Overall, this pattern suggests a loss of steroid dependency for malignant mammary tumors. Prolactin, an important hormone for the development of mammary tissue, has been examined in tissue and serum from canine patients with both benign and malignant mammary tumors.19 Prolactin levels were significantly higher in the tissues and serum from malignant tumors than in normal mammary tissue. It is well known that in the female dog, progesterone or synthetic progestins such as chlormadinone acetate (CMA) or medroxyprogesterone acetate (MPA) induce full lobuloalveolar development of the mammary gland, with hyperplasia of secretory and myoepithelial elements, whereas estradiol stimulates ductal growth. However, prolonged administration of estrogens has not been shown to increase the incidence of mammary tumors in dogs. Conversely, progesterone administered to young female beagles mainly leads to the development of benign nodules. The risk for malignant tumors becomes higher after long-term experimental administration of estrogens combined with progestins at high dosage or if drugs are used that have a combined progestagenic-estrogenic activity.20 Injectable progestins used to prevent estrus in dogs have been shown to increase the incidence of benign tumors but not malignant tumors,10 although an increased incidence of the latter also has been reported.21 Administration of progestins to dogs also has been shown to increase the secretion of growth hormone (GH). This progestin-induced GH gene expression was found to originate from normal mammary gland epithelium.22 It is not known whether the progestin-induced GH acts as an intermediate in the progestin-stimulated development of canine mammary tumors. Parallel to the increase in GH production induced in the dog by progestins, a rise in the blood levels of insulin-like growth factor I (IGF-I) and IGF-II has been shown to occur,23 which may stimulate mammary cell proliferation. Hormonally controlled and autonomous expression of growth factors and their receptors also may influence mammary tumorigenesis, making it a highly complex process. Many genes are involved in the normal transmission of growth-promoting signals from the cell surface to the nucleus (e.g., ras). The ras proteins tend to be overexpressed in many human tumors, but studies of canine mammary tumors using polymerase chain reaction (PCR) analyses have failed to reveal expression of ras genes.24,25 Regarding oncogenes expressing for the receptor of growth factors, one study found overexpression of messenger ribonucleic acid (mRNA) for c-erbB-2 (also called c-neu) in most canine malignant (but not benign) mammary tumors,26 whereas another study found that its protein was expressed more often in

benign tumors (50%) than in malignant ones (19%).27 Recently Dutra and coworkers28 reported on c-erbB-2 protein expression in 48 malignant canine mammary tumors using the commercially available HercepTest system, which is an immunohistochemistry-based assay approved for use in human breast cancers. They found that although none of the 22 benign tumors overexpressed c-erbB-2, 35.4% of malignant tumors did.28 Standardization of methodology is necessary to allow proper interpretation of results and to elucidate the role of c-erbB-2 in canine mammary neoplasia. The p53 tumor suppressor gene is the most frequently mutated gene in human cancer. Recent studies in canine mammary cancer found that three of 1029 and six of 4030 primary cancers contained p53 mutations, with one dog carrying a germ-line mutation.29,30 In another study, 17% of 69 canine mammary carcinomas assayed showed a p53 gene mutation, and multivariate analysis indicated that this mutation conferred an increased risk of recurrence and death from mammary tumor.31 Alterations of a second tumor suppressor gene, BRCA1, which is responsible for part of hereditary human breast cancers, has been seen in some canine mammary cancers.32 However, the extent to which specific genetic alterations contribute to the pathogenesis of canine mammary tumors still needs to be resolved. Gross abnormalities in the nuclear deoxyribonucleic acid (DNA) content (DNA aneuploidy), as detected with flow cytometry, have been found in 50% to 60% of primary cancers, occurring as either an increase or a decrease in DNA content.33–35 Also, 15% to 25% of benign mammary tumors were aneuploid, possibly reflecting the potential to progress to malignancy.33-35 Other factors that may play a role in the progression of canine mammary tumors have been evaluated, including expression of adhesion and gap junction proteins such as E-cadherin, connexins, and paxillin. In general, the more invasive, proliferative, and aggressive a tumor was histologically, the less localized and intense the protein expression was, indicating a shift that favored increased cell mobility.36-38 Apoptotic pathways also have been explored as a contributor to mammary carcinogenesis. In a recent study, human and canine tumors were found to have increased patterns of expression of antiapoptotic proteins (bcl-2, bcl-Xl) and a significantly decreased expression of proapoptotic proteins (Bax, caspase 8 and 3).39 Induction of cyclooxyegenase-2 (COX-2) enzymes, thought to be involved in human breast carcinogenesis, also has been evaluated in normal, benign, and malignant canine mammary tissue. Dore and colleagues40 found that COX-2 was not expressed in normal tissues but showed increased expression in benign (24%) and malignant (56%) tumors, indicating a possible role in tumorigenesis.40 An association also has been shown between COX-2 expression and tumor histologic subtype.41

Ch26-W0558

11/7/06

7:10 PM

Page 621

Chapter 26 • Tumors of the Mammary Gland Experiments in rodents and epidemiologic studies in humans have shown that a high-fat diet and obesity increase the risk of mammary cancer. A recent study in pet dogs in the United States found that among spayed dogs, the risk of developing mammary cancer was reduced if the dogs were thin at 9 to 12 months of age.42 A study in Spain made a similar observation, that obesity at 1 year of age was a risk factor for the development of benign and malignant mammary tumors without consideration of ovariectomy.43 Somewhat surprisingly, the intake of homemade meals (compared with commercial foods) also was related to an increased risk. Nutritional factors, therefore, may play a role in canine mammary tumor development.

Pathology and Natural Behavior Based on material from veterinary practices submitted for histologic diagnosis, 41% to 53% of mammary tumors that occur in the bitch are considered malignant.44,45 Histologic evidence of malignancy does not invariably imply a malignant clinical course. Also, marked variation in histologic appearance can occur within the same tumor mass. Most malignant mammary tumors are classified as epithelial tumors or carcinomas. Pure sarcomas (fibrosarcoma, osteosarcoma, sarcoma of another type) represent a minority. Whether sarcomas arise from myoepithelial tissue that has undergone neoplastic change or from the intralobular connective tissue is unclear. No evidence indicates that sarcomas arise from pre-existing, benign mixed cell tumors. Some malignant tumors are composed of cells that morphologically resemble the epithelial and connective tissue components (both of which are malignant); these tumors, which are uncommon, are called carcinosarcomas (in some systems, malignant mixed tumor). Benign tumors include simple/ complex adenomas, fibroadenomas, and the relatively common benign mixed tumor, which has an epithelial component and a mesenchymal component of cartilage and/or bone and/or fat, both components possibly originating from a pluripotent stem cell.46,47 Some histologic classification systems are based on recognition of histologic tissue patterns thought to identify the histogenetic origin of the tumor cells. However, this may have little reference to disease behavior. The revised classification established by the World Health Organization (WHO) (Box 26-1), in an attempt to obtain a division of prognostic weight, is descriptive and subdivides carcinomas further into noninfiltrating, complex, or simple carcinomas (simple carcinomas are the tubulopapillary, solid, or anaplastic type). This subdivision is believed to rank the tumors by increasing potential for malignancy. In a recent study covering the full life span of 672 intact female beagles, the multifocal nature of

Box 26-1

621

Histologic Classification of Canine Mammary Tumors

Malignant tumors Noninfiltrating (in situ) carcinoma Complex carcinoma Simple carcinoma Tubulopapillary carcinoma Solid carcinoma Anaplastic carcinoma Special types of carcinomas Spindle cell carcinoma Squamous cell carcinoma Mucinous carcinoma Lipid-rich carcinoma Sarcoma Fibrosarcoma Osteosarcoma Other sarcomas Carcinosarcoma Carcinoma or sarcoma in benign tumor Benign tumors Adenoma Simple adenoma Complex adenoma Basaloid adenoma Fibroadenoma Low-cellularity fibroadenoma High-cellularity fibroadenoma Benign mixed tumor Duct papilloma From Misdorp W, Else R, Hellman E et al: Histologic classification of mammary tumors of the dog and cat. In World Health Organization international histological classification of tumors of domestic animals, series 2, vol 7, no 2, Washington, DC, 1999, Armed Forces Institute of Pathology.

mammary neoplasia was illustrated, with 71% having at least one mammary neoplasm and 61% more than one tumor, accumulating to a total of 4755 neoplasms, including 1639 carcinomas.48 Based on a histogenetic classification system, the investigators found that of the carcinomas, ductular carcinomas represented 19% and adenocarcinomas of other histogenetic origin accounted for the rest. Only 73 carcinomas were fatal, and ductular carcinomas were eight times more likely to result in fatalities than adenocarcinomas, even though the difference in total (regional plus distant) metastasis rate (46% versus 32%) was less. In this same study, true carcinosarcomas were diagnosed in only five dogs, all of which died of metastatic disease. Another study of 356 female beagles determined a lifetime risk of 63% for the development of any mammary hyperplasia or tumor, and 23% for the development of a malignant tumor.2 In this study the

Ch26-W0558

11/7/06

622

7:10 PM

Page 622

Part IV • Specific Malignancies in the Small Animal Patient

metastatic rate was higher, reaching 77% in 72 dogs with carcinoma. Infiltration of the carcinoma into adjacent tissues was considered of greater prognostic importance than the histologic subtype.2 A different study applied a classification system adopted from a prognostic human pathologic staging system in 158 pet dogs with mammary cancer.45 Carcinomas were assigned to one of four histologic grades: (1) lesions that were noninfiltrating (i.e., in situ carcinoma [grade 0]); (2) lesions that invaded the surrounding stroma but without identifiable vascular or lymphatic invasion (grade I); (3) lesions that showed vascular or lymphatic invasion and/or metastasis to regional lymph nodes (grade II); and (4) lesions that showed pathologic evidence of distant metastasis (grade III). The tumors were categorized further according to the degree of nuclear differentiation (poorly differentiated, moderately differentiated, or well differentiated). Both the grade and the degree of differentiation were found to be significantly related to tumor aggressiveness.45 Another grading scheme recently was reported to have prognostic significance by Karayannopoulou and colleagues.49 In this study, a grade was determined for each case by scoring three different criteria: tubule formation, nuclear pleomorphism, and mitotic counts. Each criterion received a score ranging from 1 to 3, and the total score was used to describe a tumor as well differentiated (grade I), moderately differentiated (grade II), or poorly differentiated (grade III). Eighty-five cases of carcinoma were followed for 2 years after mastectomy. The authors found that patients with grade III tumors had a significantly worse survival rate than those with grade I or grade II tumors. They also found a 21-fold increased risk of death from the tumor with grade III lesions compared to grade I or grade II tumors.49 A specific designation is reserved for inflammatory carcinoma. In one study, 17% of 186 malignant mammary tumors were classified as inflammatory carcinoma.50 Histologically, there is evidence of a poorly differentiated carcinoma with extensive evidence of both mononuclear and polymorphonuclear cellular infiltrates and often edema. Dermal lymphatic invasion also can be seen histologically. Clinically, these neoplasms grow and metastasize extremely rapidly and invade lymphatics in the skin, resulting in marked edema and inflammation (Figure 26-1).51 In a study of 21 cases, all were found to be estrogen receptor (ER) negative.52

Tumors may be associated with the nipple or, more often, the glandular tissue itself. The dog has five pairs of glands, all of which can develop one or more benign or malignant tumors. Roughly 65% to 70% of canine tumors occur in glands 4 and 5, probably because of the greater volume of mammary tissue in these glands. In animals with benign mammary tumors, the tumor is small, well circumscribed, and firm on palpation. Clinical signs of malignancy include rapid growth, illdefined boundaries, fixation to the skin or underlying tissues, and ulceration or inflammation (Figure 26-2). The presence of one or more of these signs may indicate an increased risk of an underlying malignant growth.

Figure 26-1 Bilateral inflammatory carcinoma with vaginal edema and edema of the leg in a dog. Vaginal cytology and fine-needle cytology of the popliteal lymph node were positive for cancer.

History and Signs Mammary tumors manifest clinically as single or (in more than half of cases) multiple nodules within the mammary gland, developing simultaneously or subsequently. Multiplicity may be less common in bitches with a limited duration of exposure to ovarian steroids.

Figure 26-2 Inflammatory carcinoma in a dog, showing ulceration and nodular plaque formation.

Ch26-W0558

11/7/06

7:10 PM

Page 623

Chapter 26 • Tumors of the Mammary Gland As previously mentioned, the inflammatory carcinomas (ICs) have a unique clinical presentation. This tumor type should be suspected if a tumor is rapidly growing, affects multiple mammary glands and overlying skin, and is characterized by firmness, warmth, edema, erythema, thickening, and signs of pain. All or part of one or both mammary chains may be involved. Extensive lymphedema of a limb or limbs adjacent to this type of mammary cancer also may occur. Such edema is the result of occlusion of affected lymphatics, with accompanying retrograde growth down the limb. It is important to differentiate this type of malignancy from inflammatory mastitis, although the latter is an unusual condition in dogs. Inflammatory carcinomas tend to be quite firm and have a diffuse swelling, whereas mastitis tends to be more localized and usually is seen after estrus, pregnancy, or false pregnancy. Systemic signs often can accompany the diagnosis of IC. In one study, 94% of dogs with IC had generalized weakness, described as lack of energy, and decreased activity compared to only 18% of dogs with other mammary tumors.50 Pain also was seen in all 33 dogs with IC (100%), compared to only 16% of dogs with other mammary tumors. Carcinoma that metastasizes to the inguinal lymph nodes may enter the pudendal lymphatics and spread to the internal iliac nodes. Metastasis from the internal iliac nodes may be palpable and may cause pressure on and compression of the colon. The other common metastatic sites include the lungs, liver, kidneys and, less frequently, bone.53

Diagnostic Techniques and Workup The diagnostic workup should include a thorough physical examination and a routine hematologic and serum chemistry profile to assess general health. A coagulogram may be indicated in dogs suspected of having inflammatory carcinoma because of the concurrent association with disseminated intravascular coagulation.51,54 The first evaluation should reveal whether local or loco-regional disease is amenable to complete resection. The presence of an inflammatory carcinoma invariably is associated with an intravascular tumor, (microscopic) or metastatic disease, and rapid local or distant recurrence after attempted surgery. If malignancy cannot be excluded, thoracic radiographs in both the right and left lateral and ventrodorsal planes should be taken before surgery to evaluate the lungs and sternal lymph node for possible metastasis. In dogs, if the mammary tumors involve the caudal two glands, the sublumbar region should be evaluated for metastatic lymphadenopathy using caudal abdominal radiographs or ultrasonography. A rectal examination may reveal palpable evidence of internal iliac lymphadenopathy. Fine-needle aspiration with cytology to differentiate

623

benign from malignant tumors has been reported to be an insensitive method;55 however, cytologic evaluation of the mass can rule out other lesions, such as inflammatory lesions or mast cell tumors. Fine-needle aspiration for cytologic evaluation may also be beneficial in the diagnosis of inflammatory carcinomas. If lymph node metastasis is suspected, cytology should be used to assess suspect nodes. The most definitive way to obtain a diagnosis is through tissue biopsy. Practitioners must keep in mind that a presurgical distinction between a benign and a malignant mammary tumor does not necessarily change the extent of the operative procedure.55

Clinical Staging Accurate, precise staging is important before treatment is initiated (Box 26-2). The most important requirements of staging are to (1) evaluate the primary tumor, (2) evaluate the regional lymph nodes, and (3) attempt to identify any distant metastatic sites, including distant lymph nodes and the lungs. The most important features to note with the primary tumor are recent rapid growth, size, clinical evidence of invasiveness (fixation to skin or fascia), ulceration, and clinical evidence of inflammatory carcinoma. The most common sites of distant metastasis are the lungs; sublumbar, sternal, and prescapular lymph nodes; liver; and, rarely, bone.

Box 26-2

Staging of Canine Mammary Tumors

Modified system T—Primary tumor <3 cm maximum diameter T1 T2 3-5 cm maximum diameter T3 >5 cm maximum diameter N—Regional lymph nodes N0 Histologic or cytologic—No metastasis N1 Histologic or cytologic—Metastasis present M—Distant metastasis M0 No distant metastasis detected M1 Distant metastasis detected Stages I: II: III: IV: V:

T1 T2 T3 Any T Any T

N0 N0 N0 N1 Any N

M0 M0 M0 M0 M1

Modified from Owen LN: Classification of tumors in domestic animals, Geneva, 1980, World Health Organization.

Ch26-W0558

11/7/06

624

7:10 PM

Page 624

Part IV • Specific Malignancies in the Small Animal Patient

Therapy Surgery remains the treatment of choice for all dogs with mammary gland tumors except those with inflammatory carcinomas or distant metastasis. The type of surgery depends on the extent of disease.

Surgical technique The theoretical and practical pros and cons of radical versus local excision have been extensively debated.56 In one prospective clinical trial of 144 dogs that compared simple mastectomy (i.e., removal of the affected gland or glands only) to chain mastectomy (i.e., chain resection on the affected side), no difference in recurrence rate and survival time was noted.57 Proponents of chain mastectomy argue that it is the most likely procedure to remove all tumor (known or occult) and that it reduces future risk by reducing the volume of breast tissue at risk. Some also argue for more aggressive surgery based on the observation that some benign lesions, namely those with atypia, are linked with an increased risk for the development of invasive cancer.45 These arguments may have more weight in relatively young, intact bitches. Those opposed to routine chain resection argue that it is too much surgery, when more than 50% of canine breast masses are benign; a more aggressive procedure can always be performed later on the 40% to 50% of patients with a malignant histology (especially if margins were incomplete); and chain mastectomy increases the morbidity, time, and expense of the treatment. The primary goal of surgery is to remove all tumor by the simplest procedure, which should take into consideration possible extension of malignant lesions through mammary lymphatics to regional nodes. More radical surgery for localized lesions may lead to a lower risk for the development of new tumors in only a subset of dogs, mentioned previously; however, it does not inhibit the outgrowth of occult metastases of the tumor to be treated. A variety of procedures can be used to remove canine breast tumors, and the choice of procedure is determined by the size of the tumor, whether it is fixed to surrounding tissue, the number of lesions, and the probability that a local cure can be achieved. Lumpectomy (Nodulectomy). Lumpectomy, also called nodulectomy, is indicated for small (less than 0.5 cm), firm, superficial, nonfixed nodules, which usually are benign. This procedure is not to be used for known malignant tumors. The skin is incised, and the nodule is bluntly dissected from the breast tissue with a small rim of normal tissue surrounding the tumor nodule. After resection the tumor is classified as benign or malignant, and the completeness of removal is evaluated. For benign lesions, even close and incomplete removal probably is adequate. If the lesion is small,

well circumscribed, and malignant, close but clean margins (1 to 2 cm) are acceptable. Incomplete resection of malignant tumors warrants more aggressive removal of the entire gland. Mammectomy. Removal of one gland is indicated for lesions that are centrally located within the gland and larger than 1 cm and that show any degree of fixation to the skin or fascia. Skin and/or abdominal wall fascia should be removed with the mass, if involved. Based on the individual patient’s gland confluency, removing glands 4 and 5 as a unit or glands 1, 2, and 3 as a unit may be easier than extensively dividing normal mammary tissue. In other words, if the individual gland is a distinct anatomic unit, single gland removal is acceptable. Regional Mastectomy. Regional mastectomy originally was proposed based on the known venous and lymphatic drainage of the mammary tissue.58,59 In the dog, the mammary glands have been identified, depending on their position, as the cranial thoracic gland (gland 1), caudal thoracic gland (gland 2), cranial abdominal gland (gland 3), caudal abdominal gland (gland 4), and inguinal gland (gland 5). Lymphatic drainage from the mammary glands has been documented to the axillary, superficial inguinal, sublumbar, and cranial sternal nodes, depending on the gland involved. The lymphatic drainage of canine mammary tumors is complex. Although lymphatic drainage can occur between all glands, a recent report indicated that major connections exist between glands 1 and 2 and between glands 4 and 5.59 Mammary glands 1, 2, and 3 and occasionally 4 drain to axillary and cranial sternal lymph nodes. The superficial inguinal lymph nodes provide lymphatic drainage to mammary glands 3, 4, 5, and occasionally 2. The superficial inguinal lymph nodes have efferent drainage to the medial iliac lymph nodes, which then continue to the lumbar trunks and finally to the cisterna chyli. Based on the premise of lymphatic drainage, tumors involving glands 1, 2, or 3 should be removed en bloc. Similarly, tumors involving gland 4 or 5 should be removed en bloc, including the adjoining lymph nodes, which is always possible. The axillary lymph node is removed only if enlarged (and mobile) or cytologically positive for malignant cells. Unilateral or Bilateral Mastectomy. Glands 1 through 5 can be removed as a unit if multiple tumors or several large tumors preclude rapid, wide removal by lesser procedures. Simultaneous bilateral mastectomy also has been proposed and can be accomplished in dogs and cats with pendulous mammae, although staged, single chain mastectomies are better tolerated.56 These procedures are done because they may be faster than multiple lumpectomies or mammectomies, not because they improve survival in the dog. Lymph Node Removal. The axillary lymph nodes are rarely involved with mammary cancer in the dog

Ch26-W0558

11/7/06

7:10 PM

Page 625

Chapter 26 • Tumors of the Mammary Gland and should not be removed prophylactically. Fixed, adherent, and large axillary nodes can only rarely be removed completely. The inguinal lymph node should be removed when enlarged and cytologically positive for cancer or whenever gland 5 is removed, because it is intimately associated with this gland. In summary, mammary cancer in the dog should be removed by the simplest procedure that will remove known cancer in the mammary gland. This does not mean that incomplete resection or debulking surgery is acceptable. In young, intact dogs with multiple lesions, the risk for future multiple operations would suggest consideration of more extensive surgery.

Chemotherapy Although adjuvant chemotherapy often is the standard of care for breast cancer in humans, little information exists about its efficacy for mammary tumors in dogs. Based on established canine mammary tumor cell lines, doxorubicin has been shown to have antitumor activity using in vitro clonogenic assays; however, these assays rarely recapitulate in vivo correlates.60 In one controlled study in a small number of dogs (n = 16), half received cyclophosphamide and 5-fluouroucil (5-FU) postoperatively, and half had regional mastectomy alone.61 All dogs had stage III disease, and two in each group had histologically confirmed regional lymph node metastasis. When the chemotherapy-treated and control groups were compared by means of Kaplan-Meier analysis, statistically superior differences were seen in both the disease-free interval (24 months versus 2 months) and survival (24 months versus 6 months). Although the power of this study is low, it may support the use of adjuvant chemotherapy in certain cases.61 Additional studies are needed to determine the optimal chemotherapeutic agent or agents for the treatment of canine malignant mammary tumors. Future clinical trials should concentrate on adjuvant chemotherapy of dogs with poor prognostic factors (e.g., large, lymph node–positive, invasive, high-grade tumors) after complete surgical removal.

Radiation therapy As with chemotherapy, no reliable information on the value of radiation is yet available. Radiation therapy, like surgery, is mainly limited by the extent of the tumor and may only be considered useful in dogs with tumors that are too extensive for surgery. Patients with malignant tumors and “complete” resection do not need radiation. The role of radiation for malignant tumors with incomplete margins has not been defined in dogs. Radiation therapy (8 Gy in 2 or 3 fractions) has been tried in inoperable patients and patients with inflammatory carcinoma, resulting in anecdotal reports of palliation. More carefully designed studies are needed before the efficacy of irradiation for canine mammary tumors can be stated.

625

Biologic response modifiers Studies of nonspecific immunomodulation using levamisole56 and Corynebacterium parvum with bacillus Calmette-Guérin (BCG) combined with surgery show little effectiveness over surgery alone.62 Based on an early study showing a favorable effect of intravenous BCG on survival, the WHO comparative oncology group performed a double-blind, prospective study on 130 bitches with confirmed mammary cancer treated by chain resection. The dogs were treated with intravenous BCG (live Pasteur strain), C. parvum vaccine, or intravenous (IV) saline. No statistical differences were reported in the 1-, 2-, and 3-year survival rates for the three treatment groups.63 In another prospective study, no difference in recurrence rates was seen between dogs operated on for invasive cancer that received liposome muramyl-tripeptide phosphatidylethanolamine (L-MTP), a derivate from the mycobacterium cell wall that is encapsulated in liposomes, and dogs that received empty liposomes.64 No validated, clinically applicable immunomodulating approach is available that can be considered to have proven therapeutic effectiveness.

Hormonal therapy Although early ovariohysterectomy (OHE) clearly has a preventative role in the development of mammary tumors in dogs, the issue of OHE for therapeutic benefit remains unresolved. In one study of 154 dogs treated with concurrent mastectomy and OHE, the mean survival time was reported to be 8.4 months.65 This offers no survival advantage compared to the mean survival times of 10 and 8 months in other studies for dogs treated by mastectomy alone.44 More recent retrospective studies in mammary cancer did not show any benefit of OHE on the tumor-related66 or overall death rate.67 One study reported improved survival in dogs if the animal was ovariohysterectomized less than 2 years before or at the time of mammary cancer resection, compared to those that remained intact or were ovariohysterectomized more than 2 years before mammary cancer resection.11 However, incomplete staging in this study and the presence of favorable prognostic factors (including tumor type) were more common in the first group of dogs. There have been no well-designed prospective studies that clearly address the issue of OHE as an adjunct treatment for dogs with malignant mammary tumors. Antiestrogens, such as tamoxifen, have been tested both in vitro for antiproliferative activity in canine mammary tumor cell lines60 and to a limited degree in clinical cases.68,69 Postulated adjuvant antitumor activity by tamoxifen could not be documented in a study of limited size in 18 dogs after mastectomy and OHE because of frequent (10 of 18 dogs) estrogen-like signs related to tamoxifen side effects. These side effects include vulvar swelling, vaginal discharge, incontinence,

Ch26-W0558

11/7/06

626

7:10 PM

Page 626

Part IV • Specific Malignancies in the Small Animal Patient

Box 26-3

Summary of Canine Mammary Tumor Prognostic Factors*

Good

Poor

Indifferent

<3 cm in diameter

>3 cm in diameter

Age

Well circumscribed

Invasive, ulcerated

Breed (small breeds may have more benign tumors8)

Lymph node: Negative

Lymph node: Positive

OHE status at time of surgery

ERs or PRs: Positive

ERs: Negative

Weight

Histologic subtype (carcinoma—well differentiated, complex, tubular/papillary)

Histologic subtype (carcinoma—poorly differentiated, simple, solid, anaplastic; inflammatory carcinoma; sarcomas)

Type of surgery (simple or radical)

Tumor grade I

Tumor grade III

Number of tumors

Index of proliferation AgNOR: Low count Ki-67: Low PCNA: Low

Index of proliferation AgNOR: High count Ki-67: High PCNA: High p53 gene mutation DNA aneuploidy

Glands involved

*These factors should serve as relative indicators of the prognosis; individual variation occurs. OHE, Ovariohysterectomy; ERs, estrogen receptors; PRs, progesterone receptors; AgNOR, argyrophilic nucleolar organizer region, PCNA, proliferating cell nuclear antigen.

urinary tract infection, stump pyometra, and signs of estrus.69 At this time, tamoxifen therapy is not advised in dogs.

Prognosis Based on both univariate and multivariate studies, the following factors have been determined to be prognostic: tumor size, lymph node involvement, presence of distant metastasis, histologic type, malignancy grade, degree of nuclear differentiation, evidence of lymphoid cellular reactivity in the tumor vicinity, degree of invasion, intravascular growth, steroid hormone receptor activity, S-phase fraction as a measure of proliferation, deoxyribonucleic acid (DNA) aneuploidy, and number of silver-staining nucleolar organizer regions (AgNORs) (Box 26-3).* Factors that do not seem to be associated with the prognosis are the tumor location, number of tumors present, type of surgery (as long as histologically adequate resection is achieved), and OHE at surgery, although controversy exists regarding the importance of age at diagnosis.9,57,70-74 The influence of breed according to the patient’s size or weight also has been evaluated. One study found that small breed was associated with longer survival, but also that small breed dogs had

*References 13, 14, 34, 45, 57, 62, 67, and 70-72.

a higher incidence of benign tumors.8 In dogs with malignant tumors, breed and weight have not been shown to influence survival.74 One study found a correlation between the histologic grade (as described previously) and the disease-free interval after mastectomy in 158 dogs with mammary cancer.45 Only 19% of the dogs with grade 0 (in situ or noninvasive) carcinomas had recurrence or metastasis within 2 years after initial mastectomy compared to 60% of dogs with grade I disease (stromal invasion) and 97% of dogs with grade II disease (invasion into vascular or lymphatic vessels). The prognosis is very good for dogs with noninvasive carcinoma (Figure 26-3). The degree of nuclear differentiation (i.e., poorly, moderately, or well-differentiated tumors) also has been shown to be an important factor in the prognosis. The risk of developing recurrent or metastatic carcinoma in less than 2 years after mastectomy was 90% for dogs with poorly differentiated tumors, 68% for those with moderately differentiated tumors, and only 24% for those with well-differentiated carcinoma.45 A report using a different grading scheme also showed prognostic significance for nuclear differentiation; patients with grade III tumors had a shorter survival time and an greater likelihood of tumor-related death.49 In this study, tumor size did not influence grade and was not associated with survival, but lymph node metastasis worsened the prognosis.

Ch26-W0558

11/7/06

7:10 PM

Page 627

Chapter 26 • Tumors of the Mammary Gland

627

Figure 26-3

Figure 26-4

Kaplan-Meier disease-free interval curve comparing histologic grade after mastectomy in 158 dogs with malignant mammary tumors. Grade 0, n = 35; grade I, n = 76; grade II, n = 47. The three stages are significantly different; p <0.0001. (From Kurzman ID, Gilbertson SR: Semin Vet Med Surg 1:25-32, 1986.)

Kaplan-Meier disease-free interval curve comparing tumor size after mastectomy in 54 dogs with locally invasive, malignant mammary tumors. Dogs with tumors smaller than 3 cm in diameter (n = 16) had significantly longer disease-free intervals than dogs with tumors 3 to 5 cm (n = 20) or larger than 5 cm (n = 18); p <0.04. (From Kurzman ID, Gilbertson SR: Semin Vet Med Surg 1:25-32, 1986.)

Lymphoid cellular reactions in the tumor vicinity are another factor that has been correlated with the prognosis. Lymphoid cellular activity may indicate morphologic evidence of an antitumor immune response. In one study, dogs with mammary cancer that did not have evidence of lymphoid cellular activity at the time of initial mastectomy had a significantly higher risk of developing recurrence within 2 years than those with reactivity.45 In this study, dogs with histologic grade I tumors showing lymphoid cellular reactivity had a 45% recurrence rate within 2 years, and dogs without cellular reactivity had an 83% recurrence rate within 2 years. Mammary sarcomas are considered to have a poor prognosis.72,75 Most dogs with sarcomas die of the disease within 9 to 12 months. With carcinomas, the histologic subtype has been shown to influence survival. In one study of 99 dogs, those with anaplastic carcinoma had a worse postoperative survival in univariate analysis (median survival, 2.5 months) than dogs with adenocarcinoma (median survival, 21 months), solid carcinoma (median survival, 16 months), and other types of tumors (median survival, 24 months).74 However, this finding did not hold up in multivariate analysis. In another report, the tumor grade was associated with

the histologic subtype, with simple carcinomas having a higher percentage of grade III undifferentiated tumors (50%) than grade I tumors (18%). Inflammatory carcinomas also have a poor prognosis.51,54 Most cannot be resected surgically, and if resected, they tend to recur within weeks to 1 month after surgery. These patients also often have some degree of disseminated intravascular coagulation, and excessive bleeding frequently occurs at the time of surgery. In one study, 33 dogs with IC had a mean survival time of 25 days with palliative care.50 Tumor size is an important prognostic factor (Figure 26-4). The WHO clinical staging system categorizes dogs according to the diameter of the largest malignant tumor (see Box 26-2). In dogs with locally invasive disease, significant differences have been found between T1 and T2, and T1 and T3 tumors, and in some but not other studies between T2 and T3 tumors.67,70,74 Dogs with invasive cancer that have tumors smaller than 3 cm in diameter have a significantly better prognosis than dogs with malignant tumors 3 cm in diameter or larger (Figure 26-4). In one study of a group of dogs with invasion of lymphatic vessels or lymph node metastasis, no significant differences were found among T1, T2, and T3 tumors.70 In another study, dogs with a

Ch26-W0558

11/7/06

628

7:10 PM

Page 628

Part IV • Specific Malignancies in the Small Animal Patient

T3 carcinoma (over 5 cm in diameter) had a median survival time of 40 weeks, compared to 112 weeks for animals with smaller tumors.76 In yet another study, dogs with tumors smaller than 3 cm in diameter survived significantly longer (22 months) than dogs with tumors larger than 3 cm (14 months).74 In univariate analyses, lymph node involvement predicted poorer disease-free survival, with 80% of the dogs with lymph node involvement having recurrence within 6 months.45,70,73 In contrast, dogs with mammary carcinoma (excluding sarcomas) with negative lymph nodes usually have a recurrence rate of 30% or lower by 2 years after surgery70 (Figure 26-5). Karayannopoulou and colleagues49 also reported that lymph node metastasis had a negative impact on survival. During a 2-year follow-up period, 24 of 28 patients (86%) with positive lymph nodes died of disease, compared to 8 of 38 patients (21%) that did not have lymph node metastasis. The presence of metastatic disease at the time of diagnosis also has a worse prognosis. In one study, the median postoperative survival was 5 months, compared to 28 months for dogs without metastasis at presentation.74 In another study, the presence of ERs or PRs (or both) above a certain threshold (10 fmol/mg cytosolic protein) correlated with improved survival in

45 dogs with mammary cancer after surgery.13 Whether the presence of steroid receptors is an independent prognostic factor is questionable, because the presence of these receptors has been correlated with well-differentiated tumors, and they have been found more often in complex than simple carcinomas.77 A study involving 84 patients looked at receptor status as a prognostic indicator for disease-free progression (DFP). Although univariate analysis indicated that ER+ or PR+ or both had a favorable influence on DFP, this did not hold up in multivariate analysis.18 In a study of 136 dogs with mammary cancer and adequate follow-up, both the presence of DNA aneuploidy and a high S-phase percentage (a measure of the proliferation rate) were associated with reduced survival time.72 Multivariate analyses diminished the value of DNA ploidy status, leaving a high S-phase fraction (SPF), sarcoma, and age as important prognostic factors that increased the hazard of death from tumor or unknown cause.72 Other immunohistochemical techniques detect the presence of antigens expressed during the cell cycle, including Ki-67 and proliferating cell nuclear antigen (PCNA). Of these, the Ki-67 labeling index recently was indicated to be of prognostic value.78 Additional investigation has been done using cytologic preparations instead of routine histologic samples. In a study of 31 dogs, the Ki-67 proliferation indices were lower for nonmalignant tumors, and high Ki-67 index values correlated with metastasis, death from disease, and a low overall disease-free interval (DFI) and survival rate.79 In addition, a high AgNOR count (another proliferation marker) was implicated as a prognosticator in canine mammary cancer.80 In one study, multivariate analysis indicated that a mutation in the p53 gene conferred an increased risk of tumor recurrence and death from mammary tumor.31 Continued investigation into these and other molecular markers hopefully will lead to clinically useful predictors of the prognosis or, more important, will define potential targets for therapeutic strategies.

FELINE MAMMARY TUMORS

Figure 26-5 Kaplan-Meier disease-free interval curve comparing lymph node metastasis after mastectomy in 45 dogs with malignant mammary tumors. Dogs in which the lymph nodes tested negative (n = 26) had a significantly longer disease-free interval than those in which the lymph nodes tested positive (n = 19); p <0.001. (From Kurzman ID, Gilbertson SR: Semin Vet Med Surg 1:25-32, 1986.)

Mammary tumors are the third most common tumor in the cat, after hematopoietic neoplasms and skin tumors.1,81-83 The incidence of mammary tumors in the cat is less than half that seen in humans and dogs; however, these tumors account for 17% of neoplasms in female cats.1,82-84 Mammary tumors also have been reported in male cats, although less often (1% to 5% of feline mammary tumors).85,86 In contrast to humans and dogs, at least 85% of feline mammary tumors are malignant.76,81-83 Some evidence indicates a breed-associated predilection for mammary tumors; domestic shorthair and

Ch26-W0558

11/7/06

7:10 PM

Page 629

Chapter 26 • Tumors of the Mammary Gland Siamese cats appear to have higher incidence rates.82,87,88 Siamese cats may have twice the risk of any other breed of developing mammary tumors.86 Mammary neoplasia has been reported to occur in cats from 9 months to 23 years of age (mean age, 10 to 12 years).82-90 One study suggests that the disease occurs at an earlier age in Siamese cats and that the incidence in this breed reaches a plateau at 9 years of age.82 The mean age reported for a group of 39 male cats (12.5 years) was slightly higher than that seen in females.85 Hormonal influences probably are involved in the pathogenesis of mammary tumors in the cat. Dorn and coworkers1 found that cats ovariectomized at 6 months of age had approximately a sevenfold reduced risk of mammary cancer than intact cats. More recently, in a case control study by Overly and colleagues,91 cats spayed before 6 months of age had a 91% reduction in their risk of mammary carcinoma compared to intact cats, and cats spayed before 1 year of age had an 86% reduction in risk. A strong association also has been documented between previous use of drugs containing synthetic progestins or estrogen-progestin combinations and the development of benign or malignant mammary tumors in cats; in both cases, the risk was more than threefold over that in untreated cats.92 A benign fibroepithelial hyperplasia may be seen in some cats recently exposed to sex steroids. Evidence that sex steroids may interact with mammary cells and thereby influence mammary tumorigenesis comes from the observation that both normal tissue and benign proliferative lesions frequently express low levels of ERs and moderate levels of PRs.13,93,94 A less frequent expression of ERs and PRs in feline mammary carcinomas suggests loss of steroid dependence during malignant progression.13,93,95 Because of this and other similarities (e.g., histopathologic appearance and pattern of metastasis), feline mammary carcinoma has been proposed as a useful comparative model for hormone-independent human breast carcinomas.96 To this end, recent molecular studies have evaluated HER2/neu overexpression in feline mammary carcinomas. When amplified and overexpressed in human breast cancer, HER2/neu (c-erbB-2) has been associated with clinically aggressive tumors and an overall poorer prognosis. This overexpression is reported to occur in 10% to 40 % of human breast carcinomas.97,98 Millanta and colleagues97 reported the same overexpression in 59% of 47 cases of feline mammary carcinoma studied and found it to be significantly associated with a shorter overall survival. A second study reported that the feline HER2 gene kinase domain had 92% homology to its human counterpart; it also found HER2 overexpression in 39% of mammary carcinomas tested.98 Other molecular targets have been investigated in feline mammary tumors to elucidate the prognosis or

629

the pathways of tumorigenesis. Many of these targets suggest an analogy between feline mammary carcinoma and aggressive human breast cancer. In one study, cyclin A, which acts as a regulatory protein in cell cycle progression, was found in 46% of tumors tested but was absent in benign mammary tissue.99 Immunohistochemical staining for vascular endothelial growth factor (VEGF) also has been investigated in malignant tissues in cats, and a higher percentage of cells that stained positive for VEGF has been correlated with an unfavorable prognosis.100 E-cadherin, a cell adhesion molecule, has been shown to be reduced or absent in 70% of feline mammary carcinomas compared to normal tissue.101 Reports on the importance of p53 overexpression or mutations are conflicting with respect to feline mammary carcinomas.102,103

Pathology and Natural Behavior Mammary tumors and dysplasias Between 85% and 93% of feline mammary tumors are histologically malignant. Many of the tumors, especially the large, more invasive neoplasms, adhere to the skin and are ulcerated. Lymphatic and lymph node invasion is common.84,87,90 In several studies, more than 80% of cats with mammary malignancy had metastases to one or more of the following organs at the time of death: lymph nodes, lungs, pleura, liver, diaphragm, adrenal glands, and kidneys.82,84,87,89,104 More than 80% of feline mammary tumors are classified histologically as adenocarcinomas.83-87 The frequency of diagnosis of the specific types of adenocarcinomas differs slightly among pathologists, but most agree that tubular, papillary, solid, and cribriform carcinomas are the most common, and some carcinomas show a combination of histologic types in one lesion. Sarcomas, squamous cell carcinomas, and mucinous carcinomas are less common malignancies. Inflammatory mammary carcinoma recently was reported in three cats based on the rapidly progressing clinical signs and on the associated inflammation and dermal lymphatic invasion.105 Approximately 15% of mammary masses are benign neoplasms or dysplasias, including simple/complex adenomas and fibroadenomas.81,84,87,89,104 In addition, there are three types of noninflammatory hyperplasia of the feline mammary gland: ductular hyperplasia, lobular hyperplasia, and fibroepithelial hyperplasia.

Fibroepithelial hyperplasia Fibroepithelial hyperplasia81 may occur in young cats shortly after a (silent) estrus from about 6 months of age, or during pregnancy until 2 years of age. This has also been reported in both male and female cats treated with exogenous progestins.94,106-110 One or (more often) several glands may be enlarged, and sometimes massive, two-sided enlargement is seen (Figure 26-6).

Ch26-W0558

11/7/06

630

7:10 PM

Page 630

Part IV • Specific Malignancies in the Small Animal Patient the hormonal influence may be difficult, and clinical signs may not resolve with OHE.109

History and Signs Feline mammary cancers often are in an advanced state of development by the time the cat is brought to the veterinarian.88 The neoplasm may adhere to the overlying skin or the underlying abdominal wall. The tumor usually is firm and nodular. At least 25% of affected patients have ulcerated masses. The involved nipples may be red and swollen and may exude a tan or yellow fluid. The tumor may involve any or all of the mammary glands without left-right or cranial/caudal predeliction.87,88,90,104 More than half of affected cats have multiple gland involvement.84,85,87,88 Metastatic lung and thorax involvement may be extensive and may cause respiratory insufficiency as a result of a pleural carcinomatosis with an effusion, often containing malignant cells.

Diagnostic Techniques and Workup

Figure 26-6 Gross appearance of fibroepithelial hyperplasia in a young cat.

This is thought to result from hormonal stimulation of the glandular tissue. The mass may become erythematous, ulcerated, and necrotic, leading to bleeding and localized infection. Edema of the skin and subcutis is common and occasionally may extend to the rear legs. Fibroepithelial hyperplasia can also resemble acute mastitis. Treatment involves removal of the hormonal influence, if possible; regression has been seen after termination of pregnancy in some affected animals. Ovariohysterectomy was found effective in most cases, with gradual regression of the masses over weeks to months. If an OHE is to be performed and the glands are still greatly enlarged, a flank incision should be considered. In animals experiencing the influence of long-acting, injectable progestins, removal of

Before any diagnostic or therapeutic steps are taken, the cat’s health status must be fully assessed. A serum chemical profile, urinalysis, and complete blood count should be done to identify any presurgical abnormalities. The number, site, and size of primary tumors should be recorded, together with possible signs of fixation. Alterations in the size and consistency of lymph nodes should be assessed, and any suspected node should be aspirated or biopsied. Thoracic radiographs should be taken in both the right and left lateral and ventrodorsal planes to search for pulmonary, lymph node, and pleural metastases. Mammary tumor pulmonary metastases appear as interstitial densities. They range from those that are faintly seen, to those that are several centimeters in diameter, to miliary pleural lesions than can produce significant effusion. Sternal lymphadenopathy occasionally is seen. Because these tumors so often are malignant, an aggressive approach should be taken to confirm the diagnosis. Except for cases in which fibroepithelial hyperplasia is suspected based on the history and clinical signs, a preliminary biopsy usually is not recommended, because 80% to 85% of the masses in a mammary gland will be malignant. However, cytology may be helpful for ruling out nonmammary malignancies. Tissue for histopathology is taken at the time of mastectomy.

Clinical Staging The most important goals of staging of malignant tumors (Box 26-4) are to (1) evaluate the primary tumor or tumors, (2) evaluate the regional lymph nodes, and (3) identify any distant metastatic sites.

Ch26-W0558

11/7/06

7:10 PM

Page 631

Chapter 26 • Tumors of the Mammary Gland

Box 26-4

Staging of Feline Mammary Tumors

Modified system T—Primary tumor T1 <2 cm maximum diameter T2 2-3 cm maximum diameter T3 >3 cm maximum diameter N—Regional lymph nodes No histologic/cytologic metastasis N0 N1 Histologic/cytologic metastasis M—Distant metastasis M0 No evidence of metastasis M1 Evidence of metastasis Stages I: II: III: IV:

T1 T2 T1,2 T3 Any T

N0 N0 N1 N0,1 Any N1

M0 M0 M0 M0 M0

Modified from Owen LN: Classification of tumors in domestic animals, Geneva, 1980, World Health Organization.

The size of the primary tumor and clinical evidence of invasiveness (fixation to skin or fascia) lead to the proper T category in the tumor-node-metastasis (TNM) system. The regional lymph nodes should be examined carefully, and fine-needle aspiration or surgical removal may be necessary to determine metastasis and to help categorize the N status. In the M category, any sign of distant metastasis establishes M1 status; this includes involved distant lymph nodes, pleural effusion proven to contain tumor cells or radiographic signs of metastatic disease in the lungs, or radiographic or ultrasonographic signs of lymph node involvement in the thorax or abdomen.

Surgery Mammary neoplasms in the cat have been treated in a variety of ways. Surgery is the most widely used treatment. It may be used alone or in combination with chemotherapy or other modes of cancer therapy. The success of surgery is hindered by the invasive nature of the disease and its tendency for early metastasis. Chain mastectomy (i.e., removal of all four glands on the affected side) is the surgical method of choice because it significantly reduces the chance of local tumor recurrence.88,110-112 This procedure frequently is used regardless of the size of the tumor. The cat, unlike the dog, usually has four pairs of mammary glands. The cranial two glands on each side have a common lymphatic system and drain into the axillary lymph

631

nodes and then to sternal nodes. The caudal two glands drain to inguinal lymph nodes. In contrast to the dog, in which more conservative resections may be appropriate in carefully selected cases, most cats require a complete unilateral or bilateral mastectomy. Tumor fixation to skin or abdominal fascia necessitates en bloc removal of these structures. Complete unilateral mastectomy usually is performed if the tumor (or tumors) is confined to one side. Staged mastectomy (2 weeks apart) or simultaneous bilateral mastectomy is done when tumors are bilateral. The inguinal lymph node is almost always removed with gland 4, whereas the axillary lymph nodes are removed only if they are enlarged and cytologically test positive for tumor. Aggressive or prophylactic removal of axillary nodes is unlikely to have therapeutic benefit.88,110-112 Although OHE has not been shown to reduce the incidence of recurrence, some believe it is warranted.82,88,113 If the mammary mass was caused by a benign condition (e.g., fibroepithelial hyperplasia), OHE often results in regression of the hyperplastic tissue (see earlier discussion).

Radiation therapy Radiation therapy is not used routinely to treat feline mammary tumors. Currently, no evidence indicates that radiation improves the survival rate of patients with feline mammary tumors.

Chemotherapy Chemotherapy with doxorubicin (25 mg/m2 given intravenously slowly) every 3 weeks alone or with cyclophosphamide (50 to 100 mg/m2 given orally on days 3, 4, 5, and 6 after doxorubicin) has induced a short-term response in about half of cats with metastatic or nonresectable local disease.114-116 Partial responses (more than 50% regression) were noted with doxorubicin alone (nine of 14 cats) and with doxorubicin with cyclophosphamide (seven of 14 cats). The chemotherapy protocol can be repeated every 3 to 4 weeks. A recent retrospective study that evaluated single agent, adjuvant doxorubicin therapy (1 mg/kg given intravenously every 3 weeks for five total treatments) after surgical excision in 67 cats reported a Kaplan-Meier median survival time of 448 days, with 59% of the patients alive at 1 year, 37% at 2 years, and 17% at 5 years.117 The median DFI for the same study population was 255 days. This study suggests that the use of chemotherapy as an adjuvant holds promise for improved outcome, but controlled, randomized trials are needed to prove its true efficacy. In addition, doxorubicin can be nephrotoxic in cats,118 and careful evaluation of renal function is recommended.

Biologic response modifiers Studies using nonspecific biologic response therapy, such as levamisole111 and bacterial vaccines,112 have

Ch26-W0558

11/7/06

632

7:10 PM

Page 632

Part IV • Specific Malignancies in the Small Animal Patient

Figure 26-7 Kaplan-Meier survival curve comparing tumor volume after mastectomy in cats with malignant mammary adenocarcinoma. Statistically, tumor volume is highly significant as a prognostic factor. (From MacEwen EG, Hayes AA, Harvey HJ et al: J Am Vet Assoc 185:201-204, 1984.)

shown minimal effect on reducing the recurrence or prolonging the survival time in cats when combined with surgery. The use of L-MTP after mastectomy has not shown any significant reduction in local recurrence or survival compared to surgery alone.119 To date, no biologic response modifier has proved efficacious in cats with mammary cancer.

Prognosis Little progress has been made over the past 20 years in extending the survival time of patients with feline mammary tumors. Because stromal invasion almost always is present and metastases frequently are present at the time of surgery, a guarded to poor prognosis should always be given. With conservative surgery, two thirds of cats in which the tumors are surgically excised have a recurrence at the surgical site.88,110,112 Most studies state that the interval from tumor detection to the death of the cat is 10 to 12 months.82,83,88,104,110 The most significant prognostic factors with regard to recurrence and survival with feline malignant mammary

tumors are the tumor size,104,110,112 (Figure 26-7), extent of surgery110 (Figure 26-8), and histologic grade.104,117,120-122 Cats with tumors larger than 3 cm in diameter reportedly have a median survival time of 4 to 12 months.120,122 Cats with tumors 2 to 3 cm in diameter have a significantly better median survival time of 15 to 24 months, and cats with tumors smaller than 2 cm in diameter have a median survival time of longer than 3 years. Therefore both the owner and the practitioner should realize that early diagnosis and aggressive treatment are important prognostic factors with malignant feline mammary tumors. Few studies have reported on the significance of lymph node metastasis with regard to the prognosis; however, those that do validate the intuitive negative effect of nodal metastasis.122,123 A recent multiinstitutional trial in cats treated with surgery followed by doxorubicin found that in cats that developed metastatic disease, the location of the metastasis had prognostic significance.117 In this study, the survival time for patients with pulmonary metastasis was 331 days, and for those with nodal metastasis, it was longer than 1500 days,

Ch26-W0558

11/7/06

7:10 PM

Page 633

Chapter 26 • Tumors of the Mammary Gland

633

Figure 26-8 Kaplan-Meier disease-free interval curve comparing the results of conservative surgery to those of radical mastectomy in cats with malignant mammary adenocarcinoma. Cats that undergo radical mastectomy have a statistically significant reduced rate of local recurrence. (From MacEwen EG, Hayes AA, Harvey HJ et al: J Am Vet Assoc 185:201-204, 1984.)

although the number of patients that developed nodal metastasis was small. Very few studies have been performed to evaluate the effectiveness of the extent of local therapy in malignant feline mammary tumors. One study showed that radical mastectomy reduced local recurrence but did not increase the overall survival time (see Figure 26-7).112 More recently, Novosad and colleagues117 found that the extent of surgery had a significant effect on survival in a subset of cats treated at a single institution; those that underwent bilateral radical mastectomies survived for 917 days, those that had regional mastectomies survived for 428 days, and those that had unilateral mastectomies survived for 348 days. Another prognostic factor with malignant mammary tumors is the degree of nuclear differentiation. Well-differentiated tumors with few mitotic figures (indicative of a low proliferation rate) have correlated with an improved survival time.104 Unfortunately, this tumor type is rare compared to the more undifferentiated forms. In addition, a high proliferation index, as assessed by measurement of Ki-67–positive cells,

appeared to be related to reduced survival in cats that underwent surgery for mammary carcinoma.123 Other phenotypic factors that may be indicative of the prognosis include VEGF expression and HER2/neu overexpression.97,100

REFERENCES 1. Dorn CR, Taylor DON, Schneider R et al: Survey of animal neoplasms in Alameda and Contra Costa Counties, California. II. Cancer morbidity in dogs and cats from Alameda County, J Natl Cancer Inst 40:307-318, 1968. 2. Moulton JE, Rosenblatt LS, Goldman M: Mammary tumors in a colony of beagle dogs, Vet Pathol 23:741-749, 1986. 3. Dobson JM, Samuel S, Milstein H et al: Canine neoplasia in the UK: estimates of incidence rates from a population of insured dogs, J Small Anim Pract 43:240-246, 2002. 4. Egenvall A, Bonnett BN, Ohagen P et al: Incidence of and survival after mammary tumors in a population of over 80,000 insured female dogs in Sweden from 1995 to 2002, Prev Vet Med 69:109-127, 2005. 5. Priester WA, McKay FW: The occurrence of tumors in domestic animals, NCI Monograph 54, Bethesda, MD, 1980, National Cancer Institute.

Ch26-W0558

11/7/06

634

7:10 PM

Page 634

Part IV • Specific Malignancies in the Small Animal Patient

6. Gottwald D: Ein Beitrag zur Häufigkeit von Mammatumoren beim Hund: Statistiche Auswertung der Einsendungen einer Praxis für Tierpathologie aus den Jahren 1990 bis 1995, doctoral dissertation, Munich, 1998, University of München.

27. Rungsipipat A, Tateyama S, Yamaguchi R et al: Immunohistochemical analysis of c-yes and c-erbB-2 oncogene products and p53 tumor suppressor protein in canine mammary tumors, J Vet Med Sci 61:27-32, 1999.

7. Zatloukal J, Lorenzova J, Tichy F et al: Breed and age as risk factors for canine mammary tumours, Acta Vet Brno 74:103-109, 2005.

28. Dutra AP, Granja NVM, Schmitt FA et al: C-erbB-2 expression and nuclear pleomorphism in canine mammary tumors, Braz J Med Biol Res 37:1673-1681, 2004.

8. Itoh T, Uchida K, Ishikawa K et al: Clinicopathological survey of 101 canine mammary gland tumors: differences between small-breed dogs and others, J Vet Med Sci 67:345-347, 2005. 9. Schneider R, Dorn CR, Taylor DON: Factors influencing canine mammary cancer development and postsurgical survival, J Natl Cancer Inst 43:1249-1261, 1969. 10. Misdorp W: Progestogens and mammary tumors in dogs and cats, Acta Endocrinol (Copenh) 125(suppl):27-31, 1991. 11. Sorenmo KU, Shofer FS, Goldschmidt MH: Effects of spaying and timing of spaying on survival of dogs with mammary carcinoma, J Vet Intern Med 14:266-270, 2000.

29. Chu LL, Rutteman GR, Kong MC et al: Genomic organization of the canine p53 gene and its mutational status in canine mammary neoplasia, Breast Cancer Res Treat 50:11-25, 1998. 30. Veldhoen N, Watterson J, Brash M et al: Identification of tumour-associated and germ line p53 mutations in canine mammary cancer, Br J Cancer 81:409-415, 1999. 31. Wakui S, Muto T, Yokoo K et al: Prognostic status of p53 gene mutations in canine mammary carcinoma, Anticancer Res 21:611-616, 2001.

12. MacEwen EG, Patnaik AK, Harvey HJ et al: Estrogen receptors in canine mammary tumors, Cancer Res 42:2255-2259, 1982.

32. Yuzbasiyan-Gurkan V, Cao Y, Venta P et al: Loss of heterogenozygosity at the BRCA1 locus in canine mammary carcinoma, Proc Vet Cancer Soc 19:7, 1999.

13. Martin PM, Cotard M, Mialot JP et al: Animal models for hormone-dependent human breast cancer, Cancer Chemother Pharmacol 2:13-17, 1984.

33. Rutteman GR, Cornelisse CJ, Dijkshoorn NJ et al: Flow cytometric analysis of DNA ploidy in canine mammary tumors, Cancer Res 48:3411-3417, 1988.

14. Inaba T, Takahashi N, Matsuda H et al: Estrogen and progesterone receptors and progesterone metabolism in canine mammary tumours, Jpn J Vet Sci 46:797-803, 1984.

34. Hellmén E, Lindgren A, Linell F et al: Comparison of histology and clinical variables to DNA ploidy in canine mammary tumors, Vet Pathol 25:219-226, 1989.

15. Rutteman GR, Misdorp W, Blankenstein MA et al: Oestrogen (ER) and progestin receptors (PR) in mammary tissue of the female dog: different receptor profile in nonmalignant and malignant states, Br J Cancer 58:594-599, 1988.

35. Pérez-Alenza MD, Rutteman GR, Kuipers-Dijkshoorn NJ et al: DNA flow cytometry of canine mammary tumours: the relationship of DNA ploidy and S-phase fraction to clinical and histological features, Res Vet Sci 58:238-243, 1995.

16. Donnay I, Rauïs J, Devleeschouwer N et al: Comparison of estrogen and progesterone receptor expression in normal and tumor mammary tissues from dogs, Am J Vet Res 56:1188-1194, 1995.

36. Sarli G, Preziosi R, De Tolla L et al: E-cadherin immunoreactivity in canine mammary tumors, J Vet Diagn Invest 16:542-547, 2004.

17. Millanta F, Calandrella M, Bari G et al: Comparison of steroid receptor expression in normal, dysplastic, and neoplastic canine and feline mammary tissues, Res Vet Sci 79:225-232, 2005.

37. Scibelli A, Angelo D, Pelagalli A et al: Expression levels of the focal adhesion-associated proteins paxillin and p130CAS in canine and feline mammary tumors, Vet Res 34:193-202, 2003.

18. Martin de las Mulas J, Millan Y, Dios R: A prospective analysis of immunohistochemically determined estrogen receptor α and progesterone receptor expression and host and tumor factors as predictors of disease free period in mammary tumors of the dog, Vet Pathol 42:200-212, 2005.

38. Torres LN, Matera JM, Vasconcellos CH et al: Expression of connexins 26 and 43 in canine hyperplastic and neoplastic mammary glands, Vet Pathol 42:633-641, 2005.

19. Queiroga FL, Perez-Alenza MD, Silvan G et al: Role of steroid hormones and prolactin in canine mammary cancer, J Steroid Biochem Mol Biol 94:181-187, 2005. 20. Rutteman GR: Contraceptive steroids and the mammary gland: Is there a hazard? Breast Cancer Res Treat 23:29-41, 1992. 21. Støvring M, Moe L, Glattre E: A population-based case control study of canine mammary tumours and clinical use of medroxy– progesterone acetate, APMIS 105:590-596, 1997. 22. Selman PJ, Mol JA, Rutteman GR et al: Progestin-induced growth hormone excess in the dog originates in the mammary gland, Endocrinology 134:287-292, 1994.

39. Kumaraguruparan R, Karunagaran D, Balachandran C et al: Of humans and canines: a comparative evaluation of heat shock and apoptosis associated proteins in mammary tumors, Clin Chim Acta 10:1-9, 2005. 40. Dore M, Lanthier I, Sirois J: Cyclooxygenase-2 expression in canine mammary tumors, Vet Pathol 40:207-212, 2003. 41. Heller DA, Clifford CA, Goldschmidt MH et al: Cyclooxygenase2 expression is associated with histologic tumor type in canine mammary carcinoma, Vet Pathol 42:776-780, 2005. 42. Sonnenschein EG, Glickman LT, Goldschmidt MH et al: Body conformation, diet and risk of breast cancer in pet dogs: a casecontrol study, Am J Epidemiol 133:694-703, 1991. 43. Pérez Alenza MD, Rutteman GR, Peña L et al: Relation between habitual diet and canine mammary tumors in a case-control study, J Vet Intern Med 12:132-139, 1998.

23. Mol JA, Selman PJ, Sprang EPM et al: The role of progestins, insulin-like growth factor (IGF) and IGF-binding proteins in the normal and neoplastic mammary gland of the bitch: a review, J Reprod Fertil Suppl 51:339-344, 1997.

44. Brodey RS, Goldschmidt MA, Roszel JR: Canine mammary gland neoplasms, J Am Anim Hosp Assoc 19:61-90, 1983.

24. Castagnaro M: RAS gene analysis in mammary tumors of dogs by means of PCCR-SSCP and direct genomic analysis, Annali dell Istituto Superiore di Sanita 31:337-341, 1995.

45. Gilbertson SR, Kurzman ID, Zachrau RE et al: Canine mammary epithelial neoplasms: biological implications of morphologic characteristics assessed in 232 dogs, Vet Pathol 20:127-142, 1983.

25. Mayr B, Dressler A, Reifinger M et al: Cytogenetic alterations in eight mammary tumors and tumor-suppressor gene p53 mutations in one mammary tumor from dogs, Am J Vet Res 59:69-78, 1998.

46. Hellmén E: The pathogenesis of canine mammary tumours, Cancer J 9:282-286, 1996.

26. Ahern TE, Bird RC, Church Bird AE et al: Expression of the oncogene c-erbB-2 in canine mammary cancers and tumorderived cell lines, Am J Vet Res 57:693-696, 1996.

47. Gartner F, Geraldes M, Cassali G et al: DNA measurement and immunohistochemical characterization of epithelial and mesenchymal cells in canine mixed mammary tumours: putative evidence for a common histogenesis, Vet J 158:39-47, 1999.

Ch26-W0558

11/7/06

7:10 PM

Page 635

Chapter 26 • Tumors of the Mammary Gland 48. Benjamin SA, Lee AC, Saunders WJ: Classification and behavior of canine mammary epithelial neoplasms based on life span observations in beagles, Vet Pathol 36:423-436, 1999. 49. Karayannopoulou M, Kaldrymidou E, Constantinidis TC et al: Histological grading and prognosis in dogs with mammary carcinomas: application of a human grading method, J Comp Pathol 133:1-7, 2005. 50. Perez Alenza MD, Tabanera E, Peña L: Inflammatory mammary carcinoma in dogs: 33 cases (1995-1999), J Am Vet Med Assoc 219:1110-1114, 2001. 51. Susaneck SJ, Allen TA, Hoopes J et al: Inflammatory mammary carcinoma in the dog, J Am Anim Hosp Assoc 19:971-976, 1983. 52. Peña L, Perez Alenza MD, Rodriguez-Bertos A et al: Canine inflammatory mammary carcinoma: histopathology, immunohistochemistry and clinical implications of 21 cases, Breast Cancer Res Treat 78:141-148, 2003. 53. Misdorp W, Hart AAM: Canine mammary cancer. I. Prognosis; II. Therapy and causes of death, J Small Anim Pract 20:385-394, 1979. 54. Stockhaus C, Kohn B, Rudolph R et al: Correlation of hemostatic abnormalities with tumour stage and characteristics in dogs with mammary carcinoma, J Small Anim Pract 40:326-331, 1999. 55. Allen SW, Prasse KW, Mahaffey EA: Cytologic differentiation of benign from malignant canine mammary tumors, Vet Pathol 23:649-655, 1986. 56. Ferguson RH: Canine mammary gland tumors, Vet Clin North Am Small Anim Pract 15:501-511, 1985. 57. MacEwen EG, Harvey HJ, Patnaik AK et al: Evaluation of effects of levamisole and surgery on canine mammary cancer, J Biol Response Mod 4:418-426, 1985. 58. Wilkinson GT: The treatment of mammary tumors in the bitch and a comparison with the cat, Vet Rec 89:13-16, 1971. 59. Sautet JY, Ruberte J, Lopez C et al: Lymphatic system of the mammary glands in the dog: an approach to the surgical treatment of malignant mammary tumors, Canine Pract 17:30-33, 1992. 60. Sartin E, Barnes S, Toivio-Kinnucan M et al: Heterogenic properties of clonal cell lines derived from canine mammary carcinomas and sensitivity to tamoxifen and doxorubicin, Anticancer Res 13:229-236, 1993. 61. Karayannopoulou M, Kaldrymidou E, Constantinidis TC et al: Adjuvant post-operative chemotherapy in bitches with mammary cancer, J Vet Med A Physiol Pathol Clin Med 48:85-96, 2001. 62. Parodi AL, Misdorp W, Mialot JP et al: Intratumoral BCG and Corynebacterium parvum therapy of canine mammary tumors before radical mastectomy, Cancer Immunol Immunother 15:172-177, 1983. 63. Rutten VPMG, Misdorp W, Gauthier A et al: Immunological aspects of mammary tumors in dogs and cats: a survey including own studies and pertinent literature, Vet Immunol Immunopathol 26:211-225, 1990. 64. Teske E, Rutteman G, Ingh T et al: Liposome-encapsulated muramyl tripeptide phosphatidyl-ethanolamine (L-MTP-PE): a randomized clinical trial in dogs with mammary cancer, Anticancer Res 18:1015-1020, 1998. 65. Fowler EH, Wilson GP, Koestner AA: Biologic behavior of canine mammary neoplasms based on a histogenetic classification, Vet Pathol 11:212-229, 1974.

635

68. Novoson CA: Principles of treatment for mammary gland tumors, Clin Tech Small Anim Pract 18(2):107-109, 2003. 69. Morris JS, Dobson JM, Bostock DE: Use of tamoxifen in the control of mammary neoplasia, Vet Rec 133:539-542, 1993. 70. Kurzman ID, Gilbertson SR: Prognostic factors in canine mammary tumors, Semin Vet Med Surg 1:25-32, 1986. 71. Misdorp W, Hart AAM: Prognostic factors in canine mammary cancer, J Natl Cancer Inst 56:779-786, 1976. 72. Hellmén E, Bergstrom R, Holmberg L et al: Prognostic factors in canine mammary tumors: a multivariate study of 202 consecutive cases, Vet Pathol 30:20-27, 1993. 73. Yamagami T, Kobayashi T, Takahashi K et al: Influence of ovariectomy at the time of mastectomy on the prognosis for canine malignant mammary tumours, J Small Anim Pract 37:462-464, 1996. 74. Philibert JC, Snyder PW, Glickman N et al: Influence of host factors on survival in dogs with malignant mammary gland tumors, J Vet Intern Med 17:102-106, 2003. 75. Misdorp W, Cotchin E, Hampe JF et al: Canine malignant mammary tumors. I. Sarcomas, Vet Pathol 8:99-177, 1971. 76. Bostock DE: Canine and feline mammary neoplasms, Br Vet J 142:506-515, 1986. 77. Rutteman GR, Misdorp W: Hormonal background of canine and feline mammary tumours, J Reprod Fertil 47(suppl):483-487, 1993. 78. Peña L, Nieto A, Perez-Alenza D et al: Immunohistochemical detection of Ki-67 and PCNA in canine mammary tumors: relationship to clinical and pathologic variables, J Vet Diagn Invest 10:237-246, 1998. 79. Zuccari DA, Santana AE, Cury PM et al: Immunocytochemical study of Ki-67 as a prognostic marker in canine mammary neoplasia, Vet Clin Pathol 33:23-28, 2004. 80. Bostock DE, Moriarty J, Crocker J: Correlation between histologic diagnosis mean nucleolar organizer region count and prognosis in canine mammary tumors, Vet Pathol 29:381-385, 1992. 81. Carpenter JL, Andrews LK, Holzworth J et al: Tumors and tumor-like lesions. In Holzworth J, editor: Diseases of the cat: medicine and surgery, Philadelphia, 1987, WB Saunders. 82. Hayes HM Jr, Milne KL, Mandell CP: Epidemiological features of feline mammary carcinomas, Vet Rec 108:476-479, 1981. 83. Schmidt RE, Langham RF: A survey of feline neoplasms, J Am Vet Med Assoc 151:1325-1328, 1967. 84. Patnaik AK, Liu SK, Hurvitz AL et al: Nonhematopoietic neoplasms in cats, J Natl Cancer Inst 54:855-860, 1975. 85. Skorupski KA, Overly B, Shofer FS et al: Clinical characteristics of mammary carcinoma in male cats, J Vet Intern Med 19:52-55, 2005. 86. Kessler M, Vonbomhard D: Mammary tumors in cats: epidemiologic and histologic features in 2,386 cases (1990-1995), Kleintierpraxis 42:459, 1997. 87. Hayden DW, Neilsen SW: Feline mammary tumors, J Small Anim Pract 12:687-697, 1971. 88. Hayes A: Feline mammary gland tumors, Vet Clin North Am 7:205-212, 1977. 89. Moulton JE: Tumors in domestic animals, ed 2, Berkeley, 1978, University of California Press. 90. Anderson J, Jarrett WFH: Mammary neoplasia in the dog and cat. Part II, J Small Anim Pract 7:697-701, 1966.

66. Morris JS, Dobson JM, Bostock DE et al: Effect of ovariohysterectomy in bitches with mammary neoplasms, Vet Rec 142: 656-658, 1998.

91. Overley B, Shofer FS, Goldschmidt MH et al: Association between ovarihysterectomy and feline mammary carcinoma, J Vet Intern Med 19:560-563, 2005.

67. Yamagami T, Kobayashi T, Takahashi K et al: Prognosis for canine malignant mammary tumors based on TNM and histologic classification, J Vet Med Sci 58:1079-1083, 1996.

92. Misdorp W, Romijn A, Hart A: Feline mammary tumors: a casecontrol study of hormonal factors, Anticancer Res 11:1793-1797, 1991.

Ch26-W0558

11/7/06

636

7:10 PM

Page 636

Part IV • Specific Malignancies in the Small Animal Patient

93. Rutteman GR, Blankenstein MA, Minke J et al: Steroid receptors in mammary tumours of the cat, Acta Endocrinol 125: 32-37, 1991. 94. Hayden DW, Johnston JD, Kiang DT et al: Feline mammary hypertrophy/fibroadenoma complex: clinical and hormonal aspects, Am J Vet Res 42:1699-1703, 1981. 95. Hamilton JM, Else RW, Forshan P: Oestrogen receptors in feline mammary carcinoma, Vet Rec 99:477-479, 1976. 96. Cardazzo B, Zappulli V, Frassineti F et al: Full-length sequence and expression analysis of estrogen receptor alpha mRNA in feline mammary tumors, J Steroid Biochem Mol Biol 96:109-118, 2005. 97. Millanta F, Calandrella M, Citi S et al: Overexpression of HER-2 in feline invasive mammary carcinomas: an immunohistochemical survey and evaluation of its prognostic potential, Vet Pathol 42:30-34, 2005. 98. De Maria R, Olivero M, Iussich S et al: Spontaneous feline mammary carcinoma is a model of HER2 overexpressing poor prognosis human breast cancers, Cancer Res 65:907-912, 2005. 99. Murakami Y, Tateyama S, Rungsipipat A et al: Immunohistochemical analysis of cyclin A, cyclin D1 and p53 in mammary tumors, squamous cell carcinomas and basal cell tumors in dogs and cats, J Vet Med Sci 62:743-750, 2000. 100. Millanta F, Lazzeri G, Vanozzi I et al: Correlation of vascular endothelial growth factor expression to overall survival in feline invasive mammary carcinomas, Vet Pathol 39:690-696, 2002. 101. Pereira PD, Gartner F: Expression of E-cadherin in normal, hyperplastic and neoplastic feline mammary tissue, Vet Rec 153:297-302, 2003. 102. Mayr B, Blauensteiner J, Edlinger A et al: Presence of p53 mutations in feline neoplasms, Res Vet Sci 68:63-70, 2000. 103. Nasir L, Krasner H, Argyl DJ et al: Immunocytochemical analysis of the tumor suppressor protein (p53) in feline neoplasia, Cancer Lett 155:1-7, 2000. 104. Weijer K, Hart AAM: Prognostic factors in feline mammary carcinoma, J Natl Cancer Inst 70:709-710, 1983. 105. Perez-Alenza MD, Jimenez A, Nieto AI et al: First description of feline inflammatory mammary carcinoma: clinicopathological and immunohistochemical characteristics of three cases, Breast Cancer Res 6:R300-R307, 2004.

109. Loretti AP, Ilha MRS, Ordas J et al: Clinical, pathological and immunohistochemical study of feline mammary fibroepithelial hyperplasia following a single injection of depot medroxyprogesterone acetate, J Feline Med Surg 7:43-52, 2005. 110. MacEwen EG, Hayes AA, Harvey HJ et al: Prognostic factors for feline mammary tumors, J Am Vet Med Assoc 185:201-204, 1984. 111. MacEwen EG, Hayes AA, Mooney S et al: Evaluation of effect of levamisole on feline mammary cancer, J Biol Response Mod 5:541-546, 1984. 112. Hayes AA, Mooney S: Feline mammary tumors, Vet Clin North Am Small Anim Pract 15:513-520, 1985. 113. Wilson GP: Mammary glands, their development and diseases. In Bojrab MJ, editor: Pathophysiology in small animals, Philadelphia, 1981, Lea & Febiger. 114. Jeglum KA, DeGuzman E, Young K: Chemotherapy of advanced mammary adenocarcinoma in 14 cats, J Am Vet Med Assoc 187:157-160, 1985. 115. Mauldin GN, Matus RE, Patnaik AK et al: Efficacy and toxicity of doxorubicin and cyclophosphamide used in the treatment of selected malignant tumors in 23 cats, J Vet Intern Med 23: 60-65, 1988. 116. Stolwijk JAM, Minke JMHM, Rutteman GR et al: Feline mammary carcinomas as a model for human breast cancer. II. Comparison of in vivo and in vitro Adriamycin sensitivity, Anticancer Res 9:1045-1048, 1989. 117. Novosad CA, Bergman PJ, O’Brian MG et al: Retrospective evaluation of adjunctive doxorubicin for the treatment of feline mammary gland adenocarcinoma: 67 cases, J Am Anim Hosp Assoc, 42:110-120, 2005. 118. Cotter SM, Kanki PJ, Simon M: Renal disease in five tumor-bearing cats treated with Adriamycin, J Am Anim Hosp 21:405, 1985. 119. Fox L, MacEwen EG, Kurzman ID et al: Liposome-encapsulated muramyl tripeptide phosphatidylethanolamine for the treatment of feline mammary adenocarcinoma: a multicenter randomized double-blind study, Cancer Biother 10:125-130, 1995. 120. Ito T, Kadosawa T, Machizuki M et al: Prognosis of malignant mammary tumors in 53 cats, J Vet Med Sci 58:723-726, 1996.

106. Allen HL: Feline mammary hypertrophy, Vet Pathol 10: 501-508, 1973.

121. Castagnaro M, Casalone C, Bozetta E et al: Tumour grading and the postsurgical prognosis in feline mammary carcinomas, J Comp Pathol 119:263-275, 1998.

107. Center SA, Randolph JF: Lactation and spontaneous remission of feline mammary hyperplasia following pregnancy, J Am Anim Hosp Assoc 21:56-58, 1985.

122. Viste JR, Myers SL, Singh B et al: Feline mammary adenocarcinoma: tumor size as a prognostic indicator, Can Vet J 43: 33-37, 2002.

108. Hinton M, Gashell CJ: Non-neoplastic mammary hypertrophy in the cat associated with either pregnancy or with oral progestogen therapy, Vet Rec 100:277-280, 1977.

123. Castagnaro M, De Maria R, Bozetta E et al: Ki-67 index as indicator of the postsurgical prognosis in feline mammary carcinoma, Res Vet Sci 65:223-226, 1998.