CANINE LYMPHOMA Johnny D. Chretin, DVM, DACVIM(O) VCA West Los Angeles Animal Hospital, 2014

Transcription

1 CANINE LYMPHOMA Johnny D. Chretin, DVM, DACVIM(O) VCA West Los Angeles Animal Hospital, 2014 Introduction Malignancy of the immune system Monoclonal expansion of lymphocytes May arise anywhere more commonly lymph nodes, spleen and bone marrow 20% (range 7-24%) of all dog malignancies Etiology Genetic 1. Gain chromosome 13, Loss chromosome 14 (golden retrievers) 3. Clustering, familial, increased risk a. Boxer, bull mastiff, bulldog, Rottweiler 4. Increased risk for specific age a. Basset hound, St. Bernard, Scottish terrier, Airedale, Labrador retriever, golden retriever 5. Phenotype prevalence a. T-cell: boxer, Shih Tzu, Welsh corgi, Siberian husky b. B-cell: cocker spaniel, Doberman pinscher Molecular 1. Proto-oncogene (e.g. N-ras) a. Promotes cell growth and survival in healthy state b. When mutated constitutively promotes growth and cell survival in absence of normal signals 2. Tumor suppressor genes (e.g. p53, p16 cyclin) a. In normal state induces cell cycle arrest, apoptosis in response to cellular damage b. In mutated state no inhibition of cell cycle (no repair) and lead to immortalization of cells 3. Telomerase a. Telomeres are nucleoprotein chromosome cap that regulates number of replications and protects from abnormal chromosome fusion, recombination and DNA degradation. b. Telomerase replenishes telomere cap and is not active in most cells c. In malignancy telomerase is activated leading to unlimited cell division and chromosome instability. Infectious 1. Epstein Barr a. Herpes virus can lead to Burkett s and Hodgkin s lymphoma in humans b. Antibodies detected in 30-60% dogs with lymphoma c. Epstein Barr like virus discovered in some canine lymphomas but not consistent and significance unknown 2. Helicobacter

2 a. Associated with human gastric lymphoma b. Associated with gastric lymphoid follicle formation in research dogs which may be a pre-malignant condition for MALT lymphoma Environment 1. 2,4-dichlorophenoxyacetic acid herbicide a. Human: NHL, Nebraska farm workers b. Dog: 2x more likely to develop in households applying the herbicide >4x/year i. Questionable study but urine levels are elevated for 7 days after environmental exposure 2. Magnetic fields, Paints & Solvents, Industrial area, Proximity to waste a. Modest increased risk b. Studies not credible or reproducible Altered immune system 1. ITP regardless of age and gender are at a higher risk of LSA 2. Immunosuppressed renal transplant patients at higher risk of LSA 3. Secondary to cyclosporine immunosuppression single case 4. Secondary to lymphocytic plasmacytic enteritis (seen in Basenji s) 5. Secondary to atopic dermatitis single case Signalment Middle aged to older (median 6-9 years) Possibly decreased risk in intact females High risk breeds boxers, basset hounds, Scottish terriers, bulldogs Low risk breeds dachshund, Pomeranian Genetic predisposition reported Presentation Multicentric>gastrointestinal>cranial mediastinal>cutaneous>other extra nodal sites Multicentric 1. 80% 2. Peripheral lymphadenopathy 3. Up to 40% are clinically sick 4. Up to 75% will have radiographic intra thoracic changes 5. 20% will have a cranial mediastinal mass 6. Up to 37% will have ocular changes 7. 50% will have radiographic intra-abdominal changes Gastrointestinal 1. 7% of all 2. Male more commonly 3. Most = T-cell 4. Shar-pei, Boxer 5. Multifocal and diffuse 6. Submucosal and lamina propria ulceration common 7. Poor prognosis especially if diffuse Mediastinal 1. 5% overall

3 2. Hypercalcemia most commonly seen with mediastinal LSA 3. T-cell most common phenotype Cutaneous 1. 3% of all 2. Epithelotropic (mycosis fungoides) a. Associated with epithelial tissues b. Superficial epidermis, peradnexal, mucous membrane i. cell surface adhesion molecules (integrins) that are specific for epithelial tissue ii. Sezary syndrome a leukemic variant that is rare and does not affect prognosis c. T-cell d. Older (mean 11 years) e. Boxer, English cocker spaniel f. Skin = mycosis fungoides, but can occur on mucosa and mucocutaneous tissue g. Prognosis guarded but can be good in early stages 3. Nonepithelotropic a. Dermal, subcutaneous b. Most B-cell but can be T-cell c. Prognosis guarded unless indolent (low grade) 4. Splenic a. Rare b. Localized c. Indolent, low grade = good prognosis i. B-cell ii. Marginal zone (MZL), Mantle cell (MCL) d. Aggressive, high grade = guarded prognosis i. T or B-cell 5. Hepatosplenic a. Rare b. Aggressive, high grade c. T-cell most common d. Liver, spleen, bone marrow involvement Grading, Phenotype and Classification Grade and accurate classification can only be assessed via histology High grade (most B-cell) 60-70% Large cell, non-cleaved nucleus ( lymphoblastic, immunoblastic, etc.) Most diffuse Mitotic rate >10 mitotic figures per 400x field Intermediate grade 10-20% cases Most are large cells and diffuse but can be follicular Mitotic rate 6-10 mitotic figures per 400x field Low grade

4 Indolent 5-29% B and T-cell Most small cell, can have large cells concurrent ( mixed cell ) Considered to be follicular (old term = nodular) with exception to T-zone lymphoma 0-5 mitotic figures per 400x field Phenotyping Histology gold standard T-cell = CD3 B-cell = CD79a Natural killer cell (NK) = negative for T and B cell markers Flow cytometry (see below) B-cell: CD20, CD21, CD22, MHCII (antigen presenting cell too) T-cell: CD4, CD5, CD8, CD25 (activated) Pan leukocyte: CD45, CD18 Myeloid cells: CD14 Hematopoietic Stem/progenitor: CD34 B-cell most common Up to 38% are T-cell - mediastinal or cutaneous presentation most commonly Historical Classification 1. Many classifications systems a. Working Formulation b. Keil classification 2. Only histologic (small, intermediate, large cell; diffuse and follicular) +/- phenotype 3. Anatomic parameters not included 4. Plasma cell, epitheliotropic, leukemia considered separately 5. Most are diffuse large B-cell (immunoblastic/centroblastic), intermediate to high grade I. Indolent lymphoma a. T and B-cell, low grade ( lymphocytic ) b. RARE II. Aggressive lymphoma a. T and B-cell, intermediate and high grade ( lymphoblastic ) b. COMMON Current Classification (World Health Organization Revised European American Lymphoma) 1. A work in progress 2. Based on of histologic, anatomic and clinical behavior 3. Grade determined by combination of histologic, molecular and phenotype parameters 4. Includes architectural pattern (diffuse, follicular/nodular, marginal zone, mantle zone, paracortical) 5. Incorporates all lymphoid malignancies into one system B-cell I. Precursor (lymphoblastic, intermediate to large cell)

5 a. Acute leukemia and large lymphomas II. Mature (lymphocytic and small to large cell) a. Chronic leukemia and lymphocytic lymphomas b. Lymphoplasmacytic lymphoma c. Splenic marginal zone d. Follicular cell (indolent, typically small cell) e. Marginal zone (indolent, spleen, GI or node, intermediate to large cell) f. Follicle center (affects lymph nodes) g. Mantle cell (indolent, affects only spleen?, intermediate to large cell) h. Plasmacytic (variable size) i. Well differentiated, indolent plasmacytoma ii. Aggressive anaplastic plasmacytoma iii. Myeloma i. Diffuse large B-cell lymphoma i. T-cell rich, b-cell lymphoma (indolent) ii. Large cell immunoblastic iii. Mediastinal (thymic) b-cell T-cell and natural killer cell (NK) cell I. Precursor (lymphoblastic, intermediate to large cells) a. Acute leukemia and large lymphomas II. Mature (lymphocytic and small to large cell) a. Chronic leukemia and lymphocytic lymphomas b. Large granular lymphocyte lymphoma i. Aggressive natural killer cell leukemia ii. Intestinal T-cell lymphoma c. Cutaneous i. Epitheliotropic (mycosis fungoides, indolent and aggressive) ii. Nonepithelioltropic (indolent and aggressive) d. Anaplastic large cell lymphoma e. Hepatosplenic f. Extranodal or peripheral T-cell lymphoma unspecified i. Diffuse small cell (follicular cell) ii. Diffuse mixed cell (follicular cell) iii. T-zone (variable size, indolent, affects lymph nodes)??? Clinical Symptoms 1. Variable a. Stage, anatomic form, phenotype, grade, paraneoplastic syndromes b. Most asymptomatic and present with generalized lymphadenopathy c. 40% clinically ill 2. Anorexia, weight loss, gastrointestinal symptoms, dyspnea, uveitis, fever, etc. 3. Cutaneous a. Epithelotropic i. Can occur on mucus and mucocutaneous tissues as well ii. Premycotic: Eczema, erythema, alopecia, pigment changes

6 iii. Mycotic: Raised, erythematous plaques +/-ulceration iv. Tumor: Large nodules often with evidence of systemic involvement/metastasis. b. Non-epitheliotropic i. Can involve both epithelial and subcutaneous tissue ii. Variable c. Secondary pyoderma common Staging and Diagnosis Diagnosis 1. FNA vs. Biopsy a. For most, cytology adequate for diagnosis b. Immunophenotyping easier and more reliable with biopsy c. Biopsy more sensitive and specific test d. Biopsy/nodal architecture needed for grading (especially if entire lymph node) e. With immunophenotype, biopsy only way to definitively classify subtypes f. Complete lymph node excision best, good quality (3-5) needle core samples or wedge biopsy adequate. g. If cytology, save unstained slides for re-submission, immunocytochemistry 2. Clonality (PCR TCR gene, PCR Ig gene, PARR) a. Antigen receptor gene rearrangement (T-cell receptor and immunoglobulin gene) b. 80% sensitive and 94% specific for diagnosis i. Cross linage rearrangement ii. Lineage infidelity iii. Reactive clonal lymphocytes (Ehrlichia) iv. Failure to obtain enough malignant cells in areas of inflammation c. No formalin ideal, older samples okay d. Need sample (fluid, tissue, blood, bone marrow) that contains lymphocytes e. Primarily used for diagnosis f. Immunophenotyping but not as accurate as other techniques (67% sensitive, 98% specific) g. Grading not possible 3. Flow Cytometry a. Fresh <72 hours old lymphocyte rich whole blood, fluid, tissue, bone marrow b. Can be used for diagnosis (with cytology or biopsy) and immunophenotyping c. Large number or antibodies available for detecting numerous types, subtypes and maturity of hematopoietic cells d. For most subtypes cannot differentiate benign versus malignant e. Grading not possible f. Immunophenotyping 91% sensitive 98% specific Staging 1. Purpose a. Accurate treatment recommendation b. Assess overall health of patient along with any comorbid disease c. Impart an accurate prognosis d. Establish baseline information on patient

7 2. Physical exam (cutaneous, rectal, ocular ), Measure all peripheral lymph nodes 3. Full staging = CBC, serum biochemistry, urinalysis, thoracic radiographs, abdominal imaging, bone marrow aspirate, histopathology/cytology, immunophenotyping a. Thoracic radiographs i. 2-view sufficient for most ii. 75% will have radiographic changes iii. 20% will have cranial mediastinal mass prognostic b. Abdominal imaging i. Radiographs appropriate for most (organ changes) ii. Abdominal ultrasound more sensitive 1. Should be performed in patients that are debilitated, demonstrate gastrointestinal changes or have evidence of abdominal organ insufficiency 2. Gastrointestinal involvement highly prognostic c. Bone marrow aspiration i. Positive in 29% of cases in spite of no circulating changes ii. Should always be performed especially if anemia, lymphocytosis or cytopenia iii. Bone marrow core biopsy more sensitive (55% versus 34%) 1. Only focal involvement in 11% cases 2. 89% patients will have diffuse or obliterating infiltration hence core biopsy not necessary for most d. Cytology and/or histology of all affected organs i. Not necessary for most ii. Consider if atypical or concern for non-diagnostic sample. e. Other staging tests may be indicated but not part of core testing i. Pleural/Abdominal Fluid, CSF, serum protein electrophoresis, skin biopsy, coagulation profile World Health Organization Clinical Staging System (WHO) Stage I Single lymph node Stage II Regional lymph nodes +/- tonsils Stage III Generalized peripheral lymphadenopathy Stage IV Live and/or spleen involvement Stage V Blood, bone marrow or any other non-lymphoid organ Substage a Substage b Without systemic signs With systemic signs Treatment 1. Lymphoma, in general, is a systemic disease requiring systemic therapy (i.e. chemotherapy) 2. Rare subtypes of indolent lymphomas do occur where local therapy alone may be the treatment of choice. 3. Single agent chemotherapy lower remission rate and shorter survival compared to multiple agent chemotherapy

8 4. Multiple agent chemotherapy (CHOP based) = standard of care 5. Incorporation of half body radiation into multiple agent chemotherapy protocols still investigational, encouraging results but not reproducible 6. Bone marrow/hematopoietic Stem Cell Transplant (HSCT) 7. Others immunotherapy, antiangiogenic therapy Chemotherapy 1. Fundamentals a. Induction i. Induce a durable remission ii. Intensive portion of protocol where patients receiving weekly single or combinational chemotherapy b. Maintenance i. Maintain remission ii. Chemotherapy administered every 2-3 weeks as once in remission each chemotherapy drug should be able to maintain remission for 3-4 weeks. c. Reinduction i. After relapse from chemotherapy discontinuation of prolonged delay ii. If >3-4 weeks from last treatment use same induction agents iii. If <3-4 weeks, last drug given has failed (i.e. chemotherapy resistance), if other drugs in protocol still active continue protocol otherwise proceed to rescue therapy d. Rescue i. When induction or reinduction fail ii. When induction or reinduction fail to induce a complete remission iii. Use drugs not previously given Chemotherapy options 1. Single agent a. Decreased cost, risk for toxicity and more convenient scheduling b. Decreased remission rate and duration compared to combination protocol c. Early onset resistance common d. Prednisone i. 46% complete remission rate ii. Median survival 2-3 months iii. Do not give if owner still undecided on staging tests. iv. Multiple drug resistance common once resistant to prednisone e. Doxorubicin (Adriamycin) i. 30mg/m 2 IV Q14-21 days ii. <1 m 2 or 15kg treat at 1mg/kg or 25mg/m 2 IV Q days iii. 70% complete remission iv. Median survival 6 months f. Lomustine (CCNU) i mg/ m 2 orally Q days (pending protocol) ii. 35% complete remission iii. Median survival 3-4 month 2. Multiple agent

9 a. COP (cyclophosphamide, vincristine, prednisone) i. Less intensive induction, longer maintenance phase ii. Minimum of 1 year protocol (1-2year) iii. 70% CR (median 4 month) iv. Median survival 6 month, 20% survival 1 year b. CHOP based (cyclophosphamide, doxorubicin, vincristine, prednisone) i. Standard of care ii. 5-6 month protocol standard of care (e.g. Wisconsin-Madison 19 or 25 week, VELCAP-S, others ) iii % CR, median 6-9month iv. Median survival (overall) 1 year, 10-25% survival 2 year v. Longer protocols not definitively demonstrated to improve outcome vi. At relapse short protocol associated with higher second remission vii. Lower cost and risk for cumulative toxicity compared to long protocol viii. After reinduction long COP maintenance based protocol recommended once in remission (total duration typically years) 3. General recommendations a. Low intensity/palliation i. Prednisone alone ii. Prednisone + lomustine b. Moderate intensity i. Doxorubicin +/-prednisone ii. Doxorubicin +/- cyclophosphamide c. Standard of care i. CHOP based short protocol (5-6 months) ii. If owner is not likely to pursue reinduction after relapse proceed directly to long phase maintenance (COP based) d. High intensity i. CHOP + High dose cyclophosphamide 1. Improved remission duration and survival 2. Not consistently demonstrated ii. CHOP + half body irradiation 1. Improved remission duration and survival 2. Still investigational 3. Not reproducible iii. CHOP + hematopoietic stem cell transplant 1. Only true chance at cure 2. High cost and limited availability Rescue chemotherapy 1. No CR on protocol proceed to rescue a. First remission duration and survival longest if a complete remission is obtained 2. Relapse after achieving a complete remission and still receiving protocol a. Relapse <3-4 week from last treatment consider drug to have failed i. If possible continue protocol until resistant to all drug ii. Proceed with rescue

10 iii. Recommend consult with oncologist as highly resistant cases can be challenging b. Relapse after achieving complete remission and discontinuing protocol i. Relapse >4months after protocol completion reinduce with same protocol and follow with long COP maintenance (minimum 1.5 years total) 1. 89% CR, median duration 7 months ii. Relapse <4months after protocol completion use alternate reinduction protocol or proceed with rescue agents 1. 67% CR, median duration 4 months if use same protocol 3. Single agent a. Mitoxantrone: 6mg/ m 2 IV Q 21 day i. 41% response rate ii. Median response 127 day duration b. Actinomycin-D (cosmegen, dactinomycin): 0.7mg/ m 2 IV Q21 day i. 41% response rate ii. Median response 129 day duration c. Lomustine (CCNU) 70-90mg/ m 2 PO Q28 day i. 27% response rate ii. Median response 86 day duration 4. Multiple Agent a. MOPP (28 day cycle) i. Mustargen 3mg/m 2 IV + VCR 0.7mg/m 2 IV d0 and 7 + Procarbazine 50mg/m 2 + Prednisone 40mg/m 2 PO d0-14 ii. 65% response rate, median 61 day iii. 20% dogs will have SD longer than first CR duration b. Anthracycline + Dacarbazine i. Dacarbazine 800mg/m 2 IV CRI 8 hours Q21d + Doxorubicin 30mg/m 2 IV or Actinomycin 0.6mg/m 2 IV Q21d ii. 70% response, median 50day c. Lomustine + Dacarbazine (28 day cycle) i. Dacarbazine 600mg/m 2 IV CRI 5 hour + CCNU 40mg/m 2 PO Q28d ii. 35% response, median 62 days Assessing Response Complete Remission (CR) Partial Response (PR) Stable Disease (SD) Progressive Disease (PD) 100% resolution of clinical and measurable abnormalities >50% resolution 50% resolution but not more than 25% progression >25% progression Treatment flow-sheet Quick reference especially for patients that have become resistant or on prolonged therapy Include date, drug, dose prescribed, dose given, location given and response Include tests performed and list any toxicities and recommended dose reductions Management Patient goals

11 1. Enhance and prolong quality of life 2. Slow onset of drug resistance 3. Maximize tumor death while minimizing treatment related toxicity Three Golden Rules 1. Treat them small 2. Don t be shy 3. Don t be late General Recommendations 1. Blood parameters should be within normal limits before proceeding with the next dose. During intensive portions of protocols (induction, rescue) minimum neutrophil count >2000cells/ul 2. Be knowledgeable of the drugs you are giving (what are the side effects, limitations). Do not give drugs that you or your staff are uncomfortable with. 3. Use a protocol in use by a local oncology center. a. Easier for advice b. Facilitates collaboration/sharing of cases 4. Dose on lean body or ideal body weight 5. Be aware of comorbid disease and unique side effects a. ABCB1 gene (formerly MDR1) mutations i. P-glycoprotein (export pump) is normally found in many cells and tissues. If functions to limit absorption and facilitate removal of substrate drugs from the body. ii. Mutations lead to toxic accumulations of P-glycoprotein substrate drugs iii. Substrate drugs 1. Acepromazine, butorphanol, ivermectin, erythromycin, loperamide, selamectin, milbemycin, moxidectin, others 2. Vincristine, vinblastine, doxorubicin, actinomycin-d, etoposide taxanes, vinorelbine, others iv. Herding breeds but any dog susceptible to mutation Breed Frequency Collie 70% Long-haired whippet 65% Australian shepherd 50% Silken windhound 30% McNab sheepdog 30% Shetland sheepdog 15% English shepherd 15% German Shepherd 10% Herding breed mix 10% Boarder collie <5% v. Mutations and chemotherapy substrate drugs 1. Consult with pharmacologist/laboratory running test for dose recommendations (VCPL@vetmed.wsu.edu) 2. Heterozygous ABCB1 (mutant/normal) a. Safest to avoid substrate drugs but in general can be used b. 25% reduction of substrate drugs

12 3. Homozygous ABCB1 (mutant/mutant) a. Avoid substrate drugs unless no other drug can be substituted. If used significant risk of toxicity and/or lack of efficacy may occur. b. In general 50% reduction can slowly escalate if no toxicity vi. Mutations and non-chemotherapy substrate drugs 1. Different then chemotherapeutics 2. Guidelines and safety variable 3. Refer to individual drug guidelines b. Hepatic i. Doxorubicin & Vinca Alkaloids Cleared through the biliary system ii. Consider a 15-25% reduction in dose given iii. Lomustine (CCNU) % 2. Boxers may be predisposed 3. Denamarin protective but will not prevent c. Cardiac i. Echocardiogram (minimum ECG) should be performed on high risk breed and if underlying cardiac disease is suspected ii. Doxorubicin 1. Cumulative cardiotoxicity in dogs (>180mg/m 2 ) 2. Cardiomyopathy is irreversible (MST days to weeks) d. Renal i. Doxorubicin 1. Renal toxicity in cats (>150mg/ m 2 ) 2. Do not give to cats with renal insufficiency or failure ii. Cisplatin 1. Fatal pulmonary edema in cats 2. Renal toxic in all species e. Myelosuppression i. Vincristine/L-asp combination 1. Severe 2. L-asparaginase competitively inhibits the clearance of vincristine ii. CCNU (lomustine) 1. Severe 2. Always give prophylactic gram (-) antibiotic until nadir known f. Cystitis i. Cyclophosphamide ii. Sterile hemorrhagic cystitis iii. Give early in day, instruct owner to walk frequently, give with furosemide iv. May take months to resolve 6. Special Considerations a. T-cell lymphomas i. CHOP presently still standard of care but may change ii. CHOP associated with lower remission rate, duration and survival compared to dogs with B-cell

13 iii. MOPP and CCNU may be better alternatives but still unproven 1. Doxorubicin 50% ORR with T-cell, 100% ORR with B-cell 2. Dogs that receive MOPP (in addition to other treatment) and have a T-cell lymphoma appear to survival longer 3. CCNU effective in rescue setting even if T-cell (30% ORR) 4. CCNU effective for epitheliotropic lymphoma (70-80% ORR) b. Indolent lymphoma i. 5-29% of all lymphomas ii. Localized 1. Surgery and or irradiation may be all that is necessary 2. Careful and complete staging required 3. Chemotherapy as adjunctive treatment has shown no survival advantage iii. Systemic or advanced 1. Chlorambucil and prednisone may be just as effective as more aggressive protocols. 2. Safflower oil (mycosis fungoides) 3. Caution as many indolent lymphomas can transform into lymphoblastic/aggressive form iv. Epitheliotropic lymphoma 1. Must treat on a case by case basis a. Grade, stage, patient clinical status critical 2. Localized a. Surgery +/- radiotherapy b. Careful and complete staging required c. Adjunctive therapy benefit for this form unknown i. If indicated safflower oil? Chlorambucil/prednisone? 3. Systemic, Metastatic, Advanced, Aggressive a. CCNU and MOPP b. CHOP should be considered but may be less effective Radiotherapy Lymphocytes and all hematopoietic cells sensitive Gastrointestinal cells sensitive as well Penetrates into protected environments (CNS) No cross resistance with chemotherapy Indications 1. CNS lymphoma a. Must be given in conjunction with systemic therapy unless localized b. Limited survival information available 2. Localized a. Nasal, stage 1, bone lesion, etc. b. Requires careful and complete staging 3. Palliation

14 a. Apoptosis will begin within 6-12 hours of irradiation = rapid response b. Due to rapid response aggressive supportive care recommended if large tumor burden as tumor lysis like syndrome can occur 4. Half body irradiation a. As adjunct to chemotherapy benefit demonstrated by only one group b. Median survival 98 weeks, 44% survival at 3 years c. Not reproducible 5. Total body irradiation a. Only low dose can be given due to significant risk of toxicity b. No benefit demonstrated outside of hematopoietic stem cell transplantation and cutaneous lymphoma Immunologic and Biologic Therapy 1. Monoclonal antibody a. Canine CD20 in clinical trials (B-cell) b. Canine CD52 in development (T-cell) 2. Vaccine a. Murine CD20 DNA vaccine i. Promising results ii. Not commercially available b. Autologous whole cell vaccine i. Benefit unproven ii. Not commercially available 3. Adoptive immunotherapy a. CD8+ T-cell i. CD8 cells best line of defense against neoplasia ii. In tumor bearing dogs CD8 cell count abnormally low and cells loose ability to mount significant response iii. In dogs receiving chemotherapy CD8 cells are destroyed and require months following chemotherapy to return to baseline levels iv. Results promising but still not commercially available 1. Remission time 2x greater then control population 2. Remission duration 2x greater then control population Hematopoietic Stem Cell Transplantation 1. Peripheral blood stem cell collection (most) therefore, no longer bone marrow transplant 2. Allogeneic a. Donor and recipient different i. Requires DLA (dog leukocyte antigen testing)/matching ii. Limited to four generations of relatives iii. Higher risk for toxicity and complications but associated with higher cure rate due to graft versus tumor effect (GVT) 1. Cyclosporine = toxicity, infection, etc. 2. Failure to engraft or prolonged recovery 3. Graft versus host disease (GVHD)

15 iv. Not practical for most 3. Autologous a. Donor and recipient are same patient b. Less risk for toxicity and complications but associated with lower risk of cure i. No graft versus tumor ii. Radiation and high dose consolidation are what lead to cure iii. Higher risk of re-introducing neoplasia as harvested stem cells may contain undetectable malignant cells c. Less cost d. More practical for owners 4. Patient preparation, autologous a. Establish foot print of malignancy (clonality) b. Obtain clinical remission (CHOP based chemotherapy) c. Obtain molecular remission (clonality testing on peripheral blood) d. Consolidation/Conditioning i. High dose cyclophosphamide days prior to transplantation ii. Irradiate as much malignancy as possible iii. Stimulate rebound stem cell (CD34+) cell production e. Stem cell mobilization i. rhg-csf subcutaneously x 5 days ii. AMD 3100 (stem cell release from bone marrow into circulation) f. Intestinal sterilizing antibiotics i. Enrofloxacin ii. Neomycin iii. Polymyxin-B g. Apheresis h. Total body irradiation i. 10Gy in two fractions ii. Low dose rate i. Isolation i. Typical recovery white cells day 8-10 ii. Typical recovery platelets day Reasons for Treatment Failure 1. Death from treatment related toxicity a. Historical 20-25% b. Current 1-5% 2. Discontinuation of chemotherapy 3. Development of multiple drug resistance (MDR) a. Innate (Goldie Colman hypothesis) b. Acquired (increased drug export, inactivation or increased cellular repair) 4. Failure to achieve adequate concentrations in certain sites (CNS, renal) General Prognosis No treatment 4-6 weeks median survival Prednisone alone 8-12 weeks median survival

16 Combination chemotherapy (pending prognostic factors) % complete remission 2. Median survival 6-14 months 3. 2 year survival of 25% 4. Cure 3% Prognostic Factors The four big ones: Immunophenotype and Tumor subtype >Substage>Stage Immunophenotype 1. Remission a. B-cell 80-90% b. T-cell 50-70% 2. Survival a. B-cell median survival months b. T-cell median survival 4-7 months Substage 1. Remission duration a months b. 3-5 months 2. Survival a. Median survival 9-18 months b. Median survival 4-7 months Stage 1. Conflicting reports a. Indolent lymphomas current classification not appropriate b. Staging criteria not consistent among studies c. Different classifications being utilized 2. Most agree lower stage associated with best prognosis a. I-II median survival months, prolonged remission b. III-V median survival 8-11 months, shorter remission Tumor grade 1. Teske et al 1994 a. High grade MST 300 days b. Intermediate grade 600 days c. Low grade >1000 days d. Difficult to interpret as multiple systems for grading have been used and when using same system in multiple studies not always prognostic 2. Overall a. High grade 1. Rapid growth/mutation 2. Rapid response to treatment 3. Higher chance at cure b. Low grade 1. Slow response to treatment 2. Slow growth/mutation, prolonged preclinical period 3. Cure unlikely without hematopoietic stem cell transplantation

17 4. MST from EARLY diagnosis better then high grade. However, if diagnosed in advanced or symptomatic stage survival regarded as similar to high grade lymphoma. c. To be most useful has to be evaluated in conjunction with anatomic site, full node architecture and immunohistochemistry Tumor Subtypes 1. Marginal Zone Lymphoma (MZL) a. Typically slowly progressive (long hyperplastic period) i. With spleen or spleen + regional node and only surgery 1. None of three studied cases died of disease ii. Median survival 21.2 months b. Nodal form and CHOP based chemotherapy i. Long term remissions >20month ii. Only one of 6 documented cases died of disease 2. Mantle Cell Lymphoma (MCL) a. Very rare, sudden onset b. Splenic form (only type documented in dogs) i. With splenectomy long term survival likely but should be followed closely ii. Caution with blastic form as most aggressive type in humans iii. CHOP if blastic or involving more than spleen 3. Follicular Lymphoma (FL) a. Slowly progressive b. Wait and watch approach reasonable if asymptomatic as treating early not proven to prolong survival c. Once extensive or symptomatic CHOP likely most appropriate otherwise could also consider chlorambucil d. With treatment survival >20 months 4. T-Zone Lymphoma (TZL, small clear cell) a. CD45-, CD5+, CD3+, CD21+, +/-CD4, CD8 b. Golden retrievers = 50% of reported cases c. Lymphocytosis common (50%, all dogs had aberrant phenotype even in absence of lymphocytosis) d. Slowly progressive e. Wait and watch f. Of 8 cases two received multiple drug chemotherapy but no difference in survival g. Median survival months h. If indicated pred/chlorambucil? CHOP?, CCNU? 5. Small lymphocytic, lymphoplasmacytic and T-cell rich B-cell lymphoma a. Unknown b. Transformation to more anaplastic type described c. If localized treatment indicated (surgery, radiotherapy) d. If systemic treatment decisions must be made based on grade and stage e. Wait and watch, CHOP or CP may be appropriate 6. Epitheliotropic / Mycosis fungoides a. In early stages indolent behavior months to years

18 b. In advanced systemic stage MST 3 months 7. Hepatosplenic and gastrointestinal a. Aggressive b. MST 3 months or less Tumor location 1. Gastrointestinal: MST 2-3 months 2. CNS a. Protected environment (BBB) b. MST 3-5 months 3. Cranial mediastinal lymphadenopathy (CML) a. T-cell = MST 101 days vs. no CML = 192 days b. B-cell = MST 191 days vs. no CML = 322 days Adjunctive therapy 1. Half body radiation with CHOP a. Median remission 58 week b. Median survival 98 week c. 44% (calculated) 3 year survival d. Caution results not reproducible and may lead to prolonged thrombocytopenia 2. CHOP+HDCTX+SC s a. Median remission 54 week b. Median survival 139 week c. 46% (calculated) 3 year survival d. Repeated study without stem cell infusion showed survival and remission advantage but much lower than initial study 3. Immunotherapy (vaccine, antibody, adoptive T-cell therapy) a. Most state of the art therapy being investigated. b. Very exciting results being demonstrated c. Still considered to be experimental for most forms of cancer and in dogs 4. Hematopoietic stem cell transplantation a. 40% cure expected b. Current results indicate 33% long term survival c. Too early to draw conclusions but benefit already being proven Other Prognostic factors 1. Response to therapy 2. AgNOR s, Bromodeoxyuridine (BrdU) short doubling time = better prognosis 3. MHC II expression low = bad 4. Neutered females = good 5. Thrombocytopenia = bad 6. Anemia = bad 7. Many others but not clinically relevant as same or more accurate prognostic information can be obtained from more standardized tests